EP4544052A1 - Traitement de maladies et de troubles liés au gpam - Google Patents

Traitement de maladies et de troubles liés au gpam

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Publication number
EP4544052A1
EP4544052A1 EP23827986.3A EP23827986A EP4544052A1 EP 4544052 A1 EP4544052 A1 EP 4544052A1 EP 23827986 A EP23827986 A EP 23827986A EP 4544052 A1 EP4544052 A1 EP 4544052A1
Authority
EP
European Patent Office
Prior art keywords
oligonucleotide
sense strand
modified
antisense strand
purines
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Pending
Application number
EP23827986.3A
Other languages
German (de)
English (en)
Inventor
Omri GOTTESMAN
Emma BRANDT
Shannon BRUSE
Brian CAJES
David JAKUBOSKY
David Lewis
Gregory Mcinnes
John VEKICH
David Rozema
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Empirico Inc
Original Assignee
Empirico Inc
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Empirico Inc filed Critical Empirico Inc
Publication of EP4544052A1 publication Critical patent/EP4544052A1/fr
Pending legal-status Critical Current

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A61P1/16—Drugs for disorders of the alimentary tract or the digestive system for liver or gallbladder disorders, e.g. hepatoprotective agents, cholagogues, litholytics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/7088—Compounds having three or more nucleosides or nucleotides
    • A61K31/7125—Nucleic acids or oligonucleotides having modified internucleoside linkage, i.e. other than 3'-5' phosphodiesters
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • A61P9/10—Drugs for disorders of the cardiovascular system for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
    • C12N15/113—Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing
    • C12N15/1137—Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing against enzymes
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/10—Type of nucleic acid
    • C12N2310/14—Type of nucleic acid interfering nucleic acids [NA]
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/30—Chemical structure
    • C12N2310/31—Chemical structure of the backbone
    • C12N2310/315—Phosphorothioates
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/30—Chemical structure
    • C12N2310/34—Spatial arrangement of the modifications
    • C12N2310/343—Spatial arrangement of the modifications having patterns, e.g. ==--==--==--
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/30—Chemical structure
    • C12N2310/34—Spatial arrangement of the modifications
    • C12N2310/344—Position-specific modifications, e.g. on every purine, at the 3'-end
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Y—ENZYMES
    • C12Y203/00—Acyltransferases (2.3)
    • C12Y203/01—Acyltransferases (2.3) transferring groups other than amino-acyl groups (2.3.1)
    • C12Y203/01015—Glycerol-3-phosphate O-acyltransferase (2.3.1.15)

Definitions

  • compositions that target GPAM Described herein are compositions comprising an oligonucleotide that targets GPAM and when administered to a subject in an effective amount reduces a GPAM mRNA or protein level.
  • a composition comprising an oligonucleotide that targets GPAM and when administered to a subject in an effective amount decreases circulating cholesterol, apolipoprotein B, bilirubin, alanine aminotransferase, aspartate aminotransferase, or aspartate aminotransferase in a subject.
  • the cholesterol comprises total cholesterol, low density lipoprotein cholesterol, or non-high density lipoprotein cholesterol.
  • the decreased by about 10% or more, as compared to prior to administration.
  • compositions comprising an oligonucleotide that targets GPAM and when administered to a subject in an effective amount decreases a liver fibrosis score, non-alcoholic fatty liver disease (NAFLD) activity score, or liver fat percentage in a subject. In some embodiments, the decrease is by about 10% or more, as compared to prior to administration.
  • compositions comprising an oligonucleotide that targets GPAM and when administered to a subject in an effective amount decreases a use of statin (HMG CoA reductase inhibitor) medication. In some embodiments, the decrease is by about 10% or more, as compared to prior to administration.
  • compositions comprising an oligonucleotide that targets GPAM and when administered to a subject in an effective amount decreases a measurement that reflects a phenotype of esophageal varices, portal hypertension, NAFLD, NASH, alcoholic liver disease, liver fibrosis, liver cirrhosis, hepatocellular carcinoma, hyperlipidemia, ischemic heart disease, or coronary heart disease in a subject.
  • the decrease is by about 10% or more, as compared to prior to administration.
  • compositions comprising an oligonucleotide that targets GPAM and when administered to a subject in an effective amount increases circulating ketone -1- Attorney Docket No.54462-742.601 bodies in a subject.
  • the increase is by about 10% or more, as compared to prior to administration.
  • the oligonucleotide comprises a modified internucleoside linkage.
  • the modified internucleoside linkage comprises alkylphosphonate, phosphorothioate, methylphosphonate, phosphorodithioate, alkylphosphonothioate, phosphoramidate, carbamate, carbonate, phosphate triester, acetamidate, or carboxymethyl ester, or a combination thereof.
  • the modified internucleoside linkage comprises one or more phosphorothioate linkages.
  • the oligonucleotide comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 modified internucleoside linkages.
  • the oligonucleotide comprises a modified nucleoside.
  • the modified nucleoside comprises a locked nucleic acid (LNA), hexitol nucleic acid (HLA), cyclohexene nucleic acid (CeNA), 2'-O-(2- methoxyethyl), 2'-O-alkyl, 2'-O-allyl, 2'-O-allyl, 2'-fluoro, or 2'-deoxy, or a combination thereof.
  • the modified nucleoside comprises an LNA.
  • the modified nucleoside comprises a 2’,4’ constrained ethyl nucleic acid.
  • the modified nucleoside comprises a 2'-O-methyl nucleoside, 2'-deoxyfluoro nucleoside, 2'-O-N-methylacetamido (2'- O-NMA) nucleoside, a 2'-O-dimethylaminoethoxyethyl (2'-O-DMAEOE) nucleoside, 2'-O-aminopropyl (2'-O-AP) nucleoside, or 2'-ara-F, or a combination thereof.
  • the modified nucleoside comprises one or more 2’fluoro modified nucleosides.
  • the modified nucleoside comprises a 2' O-alkyl modified nucleoside.
  • the oligonucleotide comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or 21 modified nucleosides.
  • the oligonucleotide comprises a hydrophobic moiety attached at a 3’ or 5’ terminus of the oligonucleotide.
  • the hydrophobic moiety comprises cholesterol, myristoyl, palmitoyl, stearoyl, lithocholoyl, docosanoyl, docosahexaenoyl, myristyl, palmityl, stearyl, or ⁇ - tocopherol, or a combination thereof.
  • the hydrophobic moiety comprises stearyl, t- butylphenyl, n-butylphenyl, octylphenyl, dodecylphenyl, phenyl n-dodecyl, octadecylbenzamide, hexadecylbenzamide, or octadecylcyclohexyl
  • the hydrophobic moiety comprises any , , or , wherein the dotted line indicates a covalent connection to an end of the oligonucleotide, n is 1-3, and R is an alkyl group containing 4-18 carbons.
  • the oligonucleotide comprises an N-acetylgalactosamine (GalNAc) moiety, an N-acetylglucosamine (GlcNAc) moiety, or a mannose moiety, attached at a 3’ or 5’ terminus of the oligonucleotide.
  • the oligonucleotide comprises a GalNAc moiety.
  • the oligonucleotide comprises a small interfering RNA (siRNA) comprising a sense strand and an antisense strand.
  • siRNA small interfering RNA
  • the sense strand is 12-30 nucleosides in length.
  • the antisense strand is 12-30 nucleosides in length.
  • compositions comprising an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises an siRNA comprising a sense strand and an antisense strand, each strand is independently about 12-30 nucleosides in length, and at least one of the sense strand and the antisense strand comprises a nucleoside sequence comprising about 12-30 contiguous nucleosides of SEQ ID NO: 12867.
  • any one of the following is true with regard to the sense strand: (i) all purines comprise 2’ fluoro modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines; (ii) all purines comprise 2’-O-methyl modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines; (iii) all purines comprise 2’ fluoro modified purines, and all pyrimidines comprise 2’-O-methyl modified pyrimidines; (iv) all pyrimidines comprise 2’ fluoro modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; (v) all pyrimidines comprise 2’-O-methyl modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; or (vi)
  • the sense strand comprises any one of modification patterns 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S .
  • any one of the following is true with regard to the antisense strand: (i) all purines comprise 2’ fluoro modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines; (ii) all purines comprise 2’-O-methyl modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified -3- Attorney Docket No.54462-742.601 pyrimidines; (iii) all purines comprise 2’-O-methyl modified purines, and all pyrimidines comprise 2’ fluoro modified pyrimidines; (iv) all pyrimidines comprise 2’ fluoro modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; (v) all pyrimidines comprise 2’-O- methyl modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and
  • the antisense strand comprises any one of modification patterns 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the oligonucleotide comprises an antisense oligonucleotide (ASO).
  • ASO antisense oligonucleotide
  • the ASO is 12-30 nucleosides in length.
  • compositions comprising an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises an ASO about 12-30 nucleosides in length and a nucleoside sequence complementary to about 12-30 contiguous nucleosides of SEQ ID NO: 12867.
  • Some embodiments include a pharmaceutically acceptable carrier.
  • Described herein are methods of treating a subject having liver disease, comprising administering an effective amount of the composition of any one of claims 1-42 to the subject.
  • the liver disease comprises NAFLD, NASH, alcoholic liver disease, liver fibrosis, liver cirrhosis, or hepatocellular carcinoma.
  • the cardiometabolic disease comprises hyperlipidemia, ischemic heart disease, or coronary heart disease.
  • DETAILED DESCRIPTION [005] Large-scale human genetic data can improve the success rate of pharmaceutical discovery and development.
  • a Genome Wide Association Study (GWAS) detects associations between genetic variants and traits in a population sample, and this improves understanding of the biology of disease and provides evidence of applicable treatments.
  • a GWAS generally utilizes genotyping and/or sequencing data, and often involves an evaluation of millions of genetic variants that are relatively evenly distributed across the genome.
  • the most common GWAS design is the case-control study, which involves comparing variant frequencies in cases versus controls. If a variant has a significantly different frequency in cases versus controls, that variant is considered associated with disease.
  • Association statistics used in a GWAS include p-values, as a measure of statistical significance; odds ratios (OR), as a measure of effect size; or beta coefficients (beta), as a measure of effect size.
  • OR odds ratios
  • beta coefficients beta coefficients
  • researchers often assume an additive genetic model and calculate an allelic odds ratio, which is the increased (or decreased) risk of disease conferred by each additional copy of an allele (compared to carrying no copies of that allele).
  • An additional concept in design and interpretation of GWAS is that of linkage disequilibrium, which is the non-random association of alleles.
  • the choice of therapeutic modality depends on factors such as the location of a target (for example, intracellular, extracellular, or secreted), a relevant tissue (for example, fat or liver) and a relevant indication.
  • the GPAM also known as GPAT or GPAT1
  • the GPAM may include 828 amino acids and have a mass of about 94 kDa.
  • GPAM may be expressed in liver, adipose, adrenal, thyroid, heart, gall bladder, brain, salivary gland, and testis cells.
  • GPAM may be intracellular.
  • GPAM may exist as two different enzymatic forms, in the mitochondria or in the endoplasmic reticulum. GPAM may catalyze the initial and committing step in glycerolipid biosynthesis and can play a significant role in the regulation of cellular triacylglycerol and phospholipid levels.
  • the mitochondrial enzyme GPAM may preferentially use saturated fatty acids as a substrate for the synthesis of glycerolipids.
  • GPAM may catalyze the first step in this metabolic pathway.
  • GPAM may interact with APP, SREBF1, AGPAT1, AGPAT2, AGPAT3, AGPAT4, AGPAT5, AGPAT6, AGPAT9, GPD1, or MBOAT2.
  • inhibition of GPAM may serve as a therapeutic for treatment of liver or cardiometabolic diseases or disorders such as non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH), alcoholic liver disease, liver fibrosis, liver cirrhosis, hepatocellular carcinoma, hyperlipidemia, ischemic heart disease, or coronary heart disease.
  • the GPAM inhibition may result in an improved liver function, cardiovascular function, and metabolic phenotypes including favorable liver fat percentage, liver fibrosis score, NAFLD activity score, liver enzyme function test, serum metabolite test, or serum lipid panel test.
  • compositions comprising an oligonucleotide that targets GPAM.
  • some embodiments may include inhibiting or targeting a GPAM protein or GPAM RNA.
  • a GPAM protein or GPAM RNA For example, by inhibiting or targeting an RNA (e.g. mRNA) encoded by the GPAM gene using an oligonucleotide described herein, the GPAM protein -5- Attorney Docket No.54462-742.601 may be inhibited or targeted as a result of there being less production of the GPAM protein by translation of the GPAM RNA; or a GPAM protein may be targeted or inhibited by an oligonucleotide that binds or interacts with a GPAM RNA and reduces production of the GPAM protein from the GPAM RNA.
  • an RNA e.g. mRNA
  • a GPAM protein may be targeted or inhibited by an oligonucleotide that binds or interacts with a GPAM RNA and reduces production of the GPAM protein from the GP
  • targeting GPAM may refer to binding a GPAM RNA and reducing GPAM RNA or protein levels.
  • the oligonucleotide may include a small interfering RNA (siRNA) or an antisense oligonucleotide (ASO). Administration of the oligonucleotide to a subject may improve (e.g.
  • compositions comprising an oligonucleotide.
  • the composition comprises an oligonucleotide that targets GPAM. In some embodiments, the composition consists of an oligonucleotide that targets GPAM. In some embodiments, the oligonucleotide reduces GPAM mRNA expression in the subject. In some embodiments, the oligonucleotide reduces GPAM protein expression in the subject.
  • the oligonucleotide may include a small interfering RNA (siRNA) described herein.
  • the oligonucleotide may include an antisense oligonucleotide (ASO) described herein.
  • a composition described herein is used in a method of treating a disorder in a subject in need thereof.
  • Some embodiments relate to a composition comprising an oligonucleotide for use in a method of treating a disorder as described herein. Some embodiments relate to use of a composition comprising an oligonucleotide, in a method of treating a disorder as described herein.
  • Some embodiments include a composition comprising an oligonucleotide that targets GPAM and when administered to a subject in an effective amount decreases GPAM mRNA or protein levels in a cell, fluid or tissue.
  • the composition comprises an oligonucleotide that targets GPAM and when administered to a subject in an effective amount decreases GPAM mRNA levels in a cell or tissue.
  • the tissue is liver tissue.
  • the tissue is fat tissue.
  • the cell is a hepatocyte.
  • the cell is an adipocyte.
  • the GPAM mRNA levels are decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, as compared to prior to administration.
  • the GPAM mRNA levels are decreased by about 10% or more, as compared to prior to administration.
  • the GPAM mRNA levels are decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100%, as compared to prior to administration.
  • the GPAM mRNA levels are decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, as compared to prior to administration. In some embodiments, the GPAM mRNA levels are decreased by no more than about 10%, as compared to prior to administration. In some embodiments, the GPAM mRNA levels are -6- Attorney Docket No.54462-742.601 decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, or no more than about 90%, as compared to prior to administration.
  • the GPAM mRNA levels are decreased by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the composition comprises an oligonucleotide that targets GPAM and when administered to a subject in an effective amount decreases GPAM protein levels in a cell, fluid or tissue.
  • the composition decreases GPAM protein levels in a cell or tissue.
  • the tissue is liver tissue.
  • the tissue is fat tissue.
  • the cell is a hepatocyte. In some embodiments, the cell is an adipocyte. In some embodiments, the GPAM protein levels are decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, as compared to prior to administration. In some embodiments, the GPAM protein levels are decreased by about 10% or more, as compared to prior to administration. In some embodiments, the GPAM protein levels are decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100%, as compared to prior to administration.
  • the GPAM protein levels are decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, as compared to prior to administration. In some embodiments, the GPAM protein levels are decreased by no more than about 10%, as compared to prior to administration. In some embodiments, the GPAM protein levels are decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, or no more than about 90%, as compared to prior to administration.
  • the GPAM protein levels are decreased by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the composition comprises an oligonucleotide that targets GPAM and when administered to a subject in an effective amount diminishes a liver disease phenotype.
  • the liver disease may include non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH), alcoholic liver disease, liver fibrosis, liver cirrhosis, or hepatocellular carcinoma.
  • NAFLD non-alcoholic fatty liver disease
  • NASH non-alcoholic steatohepatitis
  • alcoholic liver disease liver fibrosis, liver cirrhosis, or hepatocellular carcinoma.
  • the liver disease phenotype is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, as compared to prior to administration.
  • the liver disease phenotype is decreased by about 10% or more, as compared to prior to administration.
  • the liver disease phenotype is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100%, as compared to prior to administration. In some embodiments, the liver disease phenotype is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, as compared to prior to administration. In some embodiments, the liver disease phenotype is decreased by no more than about 10%, as compared to prior to administration.
  • the liver disease phenotype is -7- Attorney Docket No.54462-742.601 decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, or no more than about 90%, as compared to prior to administration. In some embodiments, the liver disease phenotype is decreased by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the composition comprises an oligonucleotide that targets GPAM and when administered to a subject in an effective amount diminishes a cardiometabolic disease phenotype.
  • the cardiometabolic disease may include hyperlipidemia, ischemic heart disease, or coronary heart disease.
  • the cardiometabolic disease phenotype is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, as compared to prior to administration. In some embodiments, the cardiometabolic disease phenotype is decreased by about 10% or more, as compared to prior to administration.
  • the cardiometabolic disease phenotype is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100%, as compared to prior to administration. In some embodiments, the cardiometabolic disease phenotype is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, as compared to prior to administration. In some embodiments, the cardiometabolic disease phenotype is decreased by no more than about 10%, as compared to prior to administration.
  • the cardiometabolic disease phenotype is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, or no more than about 90%, as compared to prior to administration. In some embodiments, the cardiometabolic disease phenotype is decreased by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the composition comprises an oligonucleotide that targets GPAM and when administered to a subject in an effective amount enhances a protective phenotype against a liver disease in the subject.
  • the liver disease may include non-alcoholic fatty liver disease (NAFLD), non- alcoholic steatohepatitis (NASH), alcoholic liver disease, liver fibrosis, liver cirrhosis, or hepatocellular carcinoma.
  • NAFLD non-alcoholic fatty liver disease
  • NASH non-alcoholic steatohepatitis
  • alcoholic liver disease liver fibrosis
  • liver cirrhosis liver cirrhosis
  • hepatocellular carcinoma hepatocellular carcinoma.
  • the protective phenotype is increased by about 2.5% or more, about 5% or more, or about 7.5% or more, as compared to prior to administration.
  • the protective phenotype is increased by about 10% or more, as compared to prior to administration.
  • the protective phenotype is increased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100% or more, as compared to prior to administration. In some embodiments, the protective phenotype is increased by about 200% or more, about 300% or more, about 400% or more, about 500% or more, about 600% or more, about 700% or more, about 800% or more, about 900% or more, or about 1000% or more, as compared to prior to administration.
  • the protective phenotype is increased by no more than about 2.5%, no more than about 5%, or no more than -8- Attorney Docket No.54462-742.601 about 7.5%, as compared to prior to administration. In some embodiments, the protective phenotype is increased by no more than about 10%, as compared to prior to administration. In some embodiments, the protective phenotype is increased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or no more than about 100%, as compared to prior to administration.
  • the protective phenotype is increased by no more than about 200%, no more than about 300%, no more than about 400%, no more than about 500%, no more than about 600%, no more than about 700%, no more than about 800%, no more than about 900%, or no more than about 1000%, as compared to prior to administration.
  • the protective phenotype is increased by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 150%, 200%, 250%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, or 1000%, or by a range defined by any of the two aforementioned percentages.
  • the composition comprises an oligonucleotide that targets GPAM and when administered to a subject in an effective amount enhances a protective phenotype against a cardiometabolic disease in the subject.
  • the cardiometabolic disease may include hyperlipidemia, ischemic heart disease, or coronary heart disease.
  • the protective phenotype is increased by about 2.5% or more, about 5% or more, or about 7.5% or more, as compared to prior to administration. In some embodiments, the protective phenotype is increased by about 10% or more, as compared to prior to administration. In some embodiments, the protective phenotype is increased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100% or more, as compared to prior to administration.
  • the protective phenotype is increased by about 200% or more, about 300% or more, about 400% or more, about 500% or more, about 600% or more, about 700% or more, about 800% or more, about 900% or more, or about 1000% or more, as compared to prior to administration. In some embodiments, the protective phenotype is increased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, as compared to prior to administration. In some embodiments, the protective phenotype is increased by no more than about 10%, as compared to prior to administration.
  • the protective phenotype is increased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or no more than about 100%, as compared to prior to administration. In some embodiments, the protective phenotype is increased by no more than about 200%, no more than about 300%, no more than about 400%, no more than about 500%, no more than about 600%, no more than about 700%, no more than about 800%, no more than about 900%, or no more than about 1000%, as compared to prior to administration.
  • the protective phenotype is increased by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 150%, 200%, 250%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, or 1000%, or by a range defined by any of the two aforementioned percentages.
  • the composition comprises an oligonucleotide that targets GPAM and when administered to a subject in an effective amount decreases circulating cholesterol in the subject.
  • the -9- Attorney Docket No.54462-742.601 circulating cholesterol may include total cholesterol or non-high density lipoprotein (HDL) cholesterol.
  • the circulating cholesterol may include total cholesterol.
  • the circulating cholesterol may include non- HDL cholesterol.
  • the circulating cholesterol is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, as compared to prior to administration.
  • the circulating cholesterol is decreased by about 10% or more, as compared to prior to administration.
  • the circulating cholesterol is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, or about 90% or more, or about 100%, as compared to prior to administration.
  • the circulating cholesterol is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, as compared to prior to administration. In some embodiments, the circulating cholesterol is decreased by no more than about 10%, as compared to prior to administration. In some embodiments, the circulating cholesterol is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, or no more than about 90%, as compared to prior to administration.
  • the circulating cholesterol is decreased by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the composition comprises an oligonucleotide that targets GPAM and when administered to a subject in an effective amount decreases circulating low density lipoproteins (LDL) in the subject.
  • the circulating LDL is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, as compared to prior to administration.
  • the circulating LDL is decreased by about 10% or more, as compared to prior to administration. In some embodiments, the circulating LDL is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, or about 90% or more, or about 100%, as compared to prior to administration. In some embodiments, the circulating LDL is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, as compared to prior to administration. In some embodiments, the circulating LDL is decreased by no more than about 10%, as compared to prior to administration.
  • the circulating LDL is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, or no more than about 90%, as compared to prior to administration. In some embodiments, the circulating LDL is decreased by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the composition comprises an oligonucleotide that targets GPAM and when administered to a subject in an effective amount decreases circulating apolipoprotein B (APOB) in the subject.
  • APOB circulating apolipoprotein B
  • the circulating APOB is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, as compared to prior to administration.
  • the circulating APOB is decreased by about 10% or more, as compared to prior to administration.
  • the circulating APOB is decreased by about 20% or more, about 30% or more, about 40% -10- Attorney Docket No.54462-742.601 or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, or about 90% or more, or about 100%, as compared to prior to administration. In some embodiments, the circulating APOB is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, as compared to prior to administration. In some embodiments, the circulating APOB is decreased by no more than about 10%, as compared to prior to administration.
  • the circulating APOB is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, or no more than about 90%, as compared to prior to administration. In some embodiments, the circulating APOB is decreased by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the composition comprises an oligonucleotide that targets GPAM and when administered to a subject in an effective amount decreases circulating alanine aminotransferase (ALT) in the subject.
  • ALT circulating alanine aminotransferase
  • the circulating ALT is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, as compared to prior to administration.
  • the circulating ALT is decreased by about 10% or more, as compared to prior to administration.
  • the circulating ALT is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, or about 90% or more, or about 100%, as compared to prior to administration.
  • the circulating ALT is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, as compared to prior to administration. In some embodiments, the circulating ALT is decreased by no more than about 10%, as compared to prior to administration. In some embodiments, the circulating ALT is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, or no more than about 90%, as compared to prior to administration.
  • the circulating ALT is decreased by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the composition comprises an oligonucleotide that targets GPAM and when administered to a subject in an effective amount decreases circulating aspartate aminotransferase (AST) in the subject.
  • the circulating AST is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, as compared to prior to administration.
  • the circulating AST is decreased by about 10% or more, as compared to prior to administration. In some embodiments, the circulating AST is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, or about 90% or more, or about 100%, as compared to prior to administration. In some embodiments, the circulating AST is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, as compared to prior to administration. In some embodiments, the circulating AST is decreased by no more than about 10%, as compared to prior to administration.
  • the circulating AST is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, or no more than -11- Attorney Docket No.54462-742.601 about 90%, as compared to prior to administration.
  • the circulating AST is decreased by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the composition comprises an oligonucleotide that targets GPAM and when administered to a subject in an effective amount decreases circulating aspartate aminotransferase (ASP) in the subject.
  • ASP circulating aspartate aminotransferase
  • the circulating ASP is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, as compared to prior to administration.
  • the circulating ASP is decreased by about 10% or more, as compared to prior to administration.
  • the circulating ASP is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, or about 90% or more, or about 100%, as compared to prior to administration.
  • the circulating ASP is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, as compared to prior to administration. In some embodiments, the circulating ASP is decreased by no more than about 10%, as compared to prior to administration. In some embodiments, the circulating ASP is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, or no more than about 90%, as compared to prior to administration.
  • the circulating ASP is decreased by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the composition comprises an oligonucleotide that targets GPAM and when administered to a subject in an effective amount decreases circulating bilirubin in the subject.
  • the circulating bilirubin is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, as compared to prior to administration.
  • the circulating bilirubin is decreased by about 10% or more, as compared to prior to administration. In some embodiments, the circulating bilirubin is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, or about 90% or more, or about 100%, as compared to prior to administration. In some embodiments, the circulating bilirubin is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, as compared to prior to administration. In some embodiments, the circulating bilirubin is decreased by no more than about 10%, as compared to prior to administration.
  • the circulating bilirubin is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, or no more than about 90%, as compared to prior to administration. In some embodiments, the circulating bilirubin is decreased by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the composition comprises an oligonucleotide that targets GPAM and when administered to a subject in an effective amount decreases a nonalcoholic fatty liver disease (NAFLD) activity score in the subject.
  • NAFLD activity score is decreased by -12- Attorney Docket No.54462-742.601 about 2.5% or more, about 5% or more, or about 7.5% or more, as compared to prior to administration.
  • the NAFLD activity score is decreased by about 10% or more, as compared to prior to administration.
  • the NAFLD activity score is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, or about 90% or more, or about 100%, as compared to prior to administration. In some embodiments, the NAFLD activity score is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, as compared to prior to administration. In some embodiments, the NAFLD activity score is decreased by no more than about 10%, as compared to prior to administration.
  • the NAFLD activity score is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, or no more than about 90%, as compared to prior to administration. In some embodiments, the NAFLD activity score is decreased by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the composition comprises an oligonucleotide that targets GPAM and when administered to a subject in an effective amount decreases a liver fat percentage in the subject.
  • the liver fat percentage is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, as compared to prior to administration.
  • the liver fat percentage is decreased by about 10% or more, as compared to prior to administration.
  • the liver fat percentage is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, or about 90% or more, or about 100%, as compared to prior to administration.
  • the liver fat percentage is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, as compared to prior to administration. In some embodiments, the liver fat percentage is decreased by no more than about 10%, as compared to prior to administration. In some embodiments, the liver fat percentage is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, or no more than about 90%, as compared to prior to administration.
  • the liver fat percentage is decreased by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the composition comprises an oligonucleotide that targets GPAM and when administered to a subject in an effective amount decreases a liver fibrosis score in the subject.
  • the liver fibrosis score is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, as compared to prior to administration.
  • the liver fibrosis score is decreased by about 10% or more, as compared to prior to administration. In some embodiments, the liver fibrosis score is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, or about 90% or more, or about 100%, as compared to prior to administration. In some embodiments, the liver fibrosis score is -13- Attorney Docket No.54462-742.601 decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, as compared to prior to administration. In some embodiments, the liver fibrosis score is decreased by no more than about 10%, as compared to prior to administration.
  • the liver fibrosis score is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, or no more than about 90%, as compared to prior to administration. In some embodiments, the liver fibrosis score is decreased by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the composition comprises an oligonucleotide that targets GPAM and when administered to a subject in an effective amount decreases a use of an HGM CoA reductase inhibitor (e.g. a statin) medication in the subject.
  • an HGM CoA reductase inhibitor e.g. a statin
  • the use of statin medications is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, as compared to prior to administration.
  • the use of statin medications is decreased by about 10% or more, as compared to prior to administration.
  • the use of statin medications is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, or about 90% or more, or about 100%, as compared to prior to administration. In some embodiments, the use of statin medications is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, as compared to prior to administration. In some embodiments, the use of statin medications is decreased by no more than about 10%, as compared to prior to administration.
  • the use of statin medications is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, or no more than about 90%, as compared to prior to administration. In some embodiments, the use of statin medications is decreased by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the composition comprises an oligonucleotide that targets GPAM and when administered to a subject in an effective amount increases circulating ketone bodies in the subject.
  • the circulating ketone body 3-hydroxybutyrate is increased by about 2.5% or more, about 5% or more, or about 7.5% or more, as compared to prior to administration.
  • the circulating ketone body 3-hydroxybutyrate is increased by about 10% or more, as compared to prior to administration.
  • the circulating ketone body 3-hydroxybutyrate is increased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100% or more, as compared to prior to administration. In some embodiments, the circulating ketone body 3-hydroxybutyrate is increased by about 200% or more, about 300% or more, about 400% or more, about 500% or more, about 600% or more, about 700% or more, about 800% or more, about 900% or more, or about 1000% or more, as compared to prior to administration.
  • the circulating ketone body 3-hydroxybutyrate is increased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, as compared to prior to administration. In some embodiments, the circulating ketone body 3-hydroxybutyrate -14- Attorney Docket No.54462-742.601 is increased by no more than about 10%, as compared to prior to administration. In some embodiments, the circulating ketone body 3-hydroxybutyrate is increased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or no more than about 100%, as compared to prior to administration.
  • the circulating ketone body 3-hydroxybutyrate is increased by no more than about 200%, no more than about 300%, no more than about 400%, no more than about 500%, no more than about 600%, no more than about 700%, no more than about 800%, no more than about 900%, or no more than about 1000%, as compared to prior to administration.
  • the circulating ketone body 3-hydroxybutyrate is increased by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 150%, 200%, 250%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, or 1000%, or by a range defined by any of the two aforementioned percentages.
  • the composition comprises an oligonucleotide that targets GPAM and when administered to a subject in an effective amount increases circulating ketone bodies in the subject.
  • the circulating ketone body acetoacetate is increased by about 2.5% or more, about 5% or more, or about 7.5% or more, as compared to prior to administration. In some embodiments, the circulating ketone body acetoacetate is increased by about 10% or more, as compared to prior to administration. In some embodiments, the circulating ketone body acetoacetate is increased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100% or more, as compared to prior to administration.
  • the circulating ketone body acetoacetate is increased by about 200% or more, about 300% or more, about 400% or more, about 500% or more, about 600% or more, about 700% or more, about 800% or more, about 900% or more, or about 1000% or more, as compared to prior to administration. In some embodiments, the circulating ketone body acetoacetate is increased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, as compared to prior to administration. In some embodiments, the circulating ketone body acetoacetate is increased by no more than about 10%, as compared to prior to administration.
  • the circulating ketone body acetoacetate is increased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or no more than about 100%, as compared to prior to administration. In some embodiments, the circulating ketone body acetoacetate is increased by no more than about 200%, no more than about 300%, no more than about 400%, no more than about 500%, no more than about 600%, no more than about 700%, no more than about 800%, no more than about 900%, or no more than about 1000%, as compared to prior to administration.
  • the circulating ketone body acetoacetate is increased by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 150%, 200%, 250%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, or 1000%, or by a range defined by any of the two aforementioned percentages.
  • the composition comprises an oligonucleotide that targets GPAM and when administered to a subject in an effective amount increases circulating ketone bodies in the subject.
  • the circulating ketone body acetone is increased by about 2.5% or more, about 5% or more, or about 7.5% or more, as compared to prior to administration. In some embodiments, the circulating ketone body acetone is increased by about 10% or more, as compared to prior to administration. In some embodiments, the circulating ketone body acetone is increased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100% or more, as compared to prior to administration.
  • the circulating ketone body acetone is increased by about 200% or more, about 300% or more, about 400% or more, about 500% or more, about 600% or more, about 700% or more, about 800% or more, about 900% or more, or about 1000% or more, as compared to prior to administration. In some embodiments, the circulating ketone body acetone is increased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, as compared to prior to administration. In some embodiments, the circulating ketone body acetone is increased by no more than about 10%, as compared to prior to administration.
  • the circulating ketone body acetone is increased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or no more than about 100%, as compared to prior to administration. In some embodiments, the circulating ketone body acetone is increased by no more than about 200%, no more than about 300%, no more than about 400%, no more than about 500%, no more than about 600%, no more than about 700%, no more than about 800%, no more than about 900%, or no more than about 1000%, as compared to prior to administration.
  • the circulating ketone body acetone is increased by 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 150%, 200%, 250%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, or 1000%, or by a range defined by any of the two aforementioned percentages.
  • the composition comprises an oligonucleotide that targets GPAM, wherein the oligonucleotide comprises a small interfering RNA (siRNA).
  • the composition comprises an oligonucleotide that targets GPAM, wherein the oligonucleotide comprises a small interfering RNA (siRNA) comprising a sense strand and an antisense strand.
  • siRNA small interfering RNA
  • the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises an siRNA comprising a sense strand and an antisense strand, wherein the sense strand is 12-30 nucleosides in length.
  • the composition comprises a sense strange that is 10, 11, 12, 13, 14, 15, 16, 17, 18,19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 nucleosides in length, or a range defined by any of the two aforementioned numbers.
  • the sense strand may be 14-30 nucleosides in length.
  • the composition comprises an antisense strand is 12-30 nucleosides in length.
  • the composition comprises an antisense strand that is 10, 11, 12, 13, 14, 15, 16, 17, 18,19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 nucleosides in length, or a range defined by any of the two aforementioned numbers.
  • the antisense strand may be 14-30 nucleosides in length.
  • the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises an siRNA comprising a sense strand and an antisense strand, each strand is independently about 12-30 nucleosides in length, and at least one of the sense strand and the antisense strand comprises a nucleoside sequence comprising about 12-30 contiguous nucleosides of a full-length human GPAM mRNA sequence such as SEQ ID NO: 12867.
  • the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises an siRNA comprising a sense strand and an antisense strand, wherein the sense strand and the antisense strand form a double-stranded RNA duplex.
  • the first base pair of the double-stranded RNA duplex is an AU base pair.
  • the sense strand further comprises a 3’ overhang.
  • the 3’ overhang comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 nucleosides, or a range of nucleotides defined by any two of the aforementioned numbers.
  • the 3’ overhang comprises 1, 2, or more nucleosides.
  • the 3’ overhang comprises 2 nucleosides.
  • the sense strand further comprises a 5’ overhang.
  • the 5’ overhang comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 nucleosides, or a range of nucleotides defined by any two of the aforementioned numbers.
  • the 5’ overhang comprises 1, 2, or more nucleosides. In some embodiments, the 5’ overhang comprises 2 nucleosides. [0037] In some embodiments, the antisense strand further comprises a 3’ overhang. In some embodiments, the 3’ overhang comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 nucleosides, or a range of nucleotides defined by any two of the aforementioned numbers. In some embodiments, the 3’ overhang comprises 1, 2, or more nucleosides. In some embodiments, the 3’ overhang comprises 2 nucleosides. In some embodiments, the antisense strand further comprises a 5’ overhang.
  • the 5’ overhang comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 nucleosides, or a range of nucleotides defined by any two of the aforementioned numbers. In some embodiments, the 5’ overhang comprises 1, 2, or more nucleosides. In some embodiments, the 5’ overhang comprises 2 nucleosides. [0038] In some embodiments, the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises an siRNA comprising a sense strand and an antisense strand, wherein the siRNA binds with a 19mer in a human GPAM mRNA.
  • the siRNA binds with a 12mer, a 13mer, a 14mer, a 15mer, a 16mer, a 17mer, a 18mer, a 19mer, a 20mer, a 21mer, a 22mer, a 23mer, a 24mer, or a 25mer in a human GPAM mRNA.
  • the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises an siRNA comprising a sense strand and an antisense strand, wherein the siRNA binds with a 17mer in a non-human primate GPAM mRNA.
  • the siRNA binds with a 12mer, a 13mer, a 14mer, a 15mer, a 16mer, a 17mer, a 18mer, a 19mer, a 20mer, a 21mer, a 22mer, a 23mer, a 24mer, or a 25mer in a non-human primate GPAM mRNA.
  • the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises an siRNA comprising a sense strand and an antisense strand, wherein the siRNA binds with a human GPAM mRNA and less than or equal to 20 human off-targets, with no more than 2 mismatches in the antisense strand. In some embodiments, the siRNA binds with a human GPAM mRNA and less than or equal to 10 human off-targets, with no more than 2 mismatches in the antisense strand.
  • the siRNA binds with a human GPAM mRNA and less than or equal to 30 human off-targets, with no more than 2 mismatches in the antisense strand. In some embodiments, the siRNA binds with a human GPAM mRNA and less than or equal to 40 human off-targets, with no more than 2 mismatches in the antisense strand. In some embodiments, the siRNA binds with a human GPAM mRNA and less than or equal to 50 human off-targets, with no more than 2 mismatches in the antisense strand.
  • the siRNA binds with a human GPAM mRNA and less than or equal to 10 human off-targets, with no more than 3 mismatches in the antisense strand. In some embodiments, the siRNA binds with a human GPAM mRNA and less than or equal to 20 human off-targets, with no more than 3 mismatches in the antisense strand. In some embodiments, the siRNA binds with a human GPAM mRNA and less than or equal to 30 human off-targets, with no more than 3 mismatches in the antisense strand.
  • the siRNA binds with a human GPAM mRNA and less than or equal to 40 human off-targets, with no more than 3 mismatches in the antisense strand. In some embodiments, the siRNA binds with a human GPAM mRNA and less than or equal to 50 human off-targets, with no more than 3 mismatches in the antisense strand.
  • the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises an siRNA comprising a sense strand and an antisense strand, siRNA binds with a human GPAM mRNA target site that does not harbor an SNP, with a minor allele frequency (MAF) greater or equal to 1% (pos.2-18).
  • siRNA binds with a human GPAM mRNA target site that does not harbor an SNP, with a minor allele frequency (MAF) greater or equal to 1% (pos.2-18).
  • the MAF is greater or equal to about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, or about 20%.
  • the siRNA comprises a sense strand having a sequence in accordance with any of SEQ ID NOs: 1-6354.
  • the sense strand sequence comprises or consists of sequence at least 75% identical to any one of SEQ ID NOs: 1-6354, at least 80% identical to any one of SEQ ID NOs: 1-6354, at least 85% identical to of any one of SEQ ID NOs: 1-6354, at least 90% identical to any one of SEQ ID NOs1-6354, or at least 95% identical to any one of SEQ ID NOs: 1-6354.
  • the sense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 1-6354, or a sense strand sequence thereof having 1, 2, 3, or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 1-6354, or a sense strand sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand sequence comprises or consists of a sequence 100% identical to SEQ ID NOs: 1-6354.
  • the sense strand sequence may include the first 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the sense strand sequence may include the last 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, -18- Attorney Docket No.54462-742.601 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the sense strand may comprise a modification pattern described herein.
  • the sense strand may comprise an overhang.
  • the sense strand may comprise a lipid moiety.
  • the sense strand may comprise a GalNAc moiety.
  • the siRNA comprises an antisense strand having a sequence in accordance with any of SEQ ID NOs: 6355-12708.
  • the antisense strand sequence comprises or consists of sequence at least 75% identical to any one of SEQ ID NOs: 6355-12708, at least 80% identical to any one of SEQ ID NOs: 6355-12708, at least 85% identical to of any one of SEQ ID NOs: 6355- 12708, at least 90% identical to any one of SEQ ID NOs: 6355-12708, or at least 95% identical to any one of SEQ ID NOs: 6355-12708.
  • the antisense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 6355-12708, or an antisense strand sequence thereof having 1, 2, 3, or 4 nucleoside substitutions, additions, or deletions. In some embodiments, the antisense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 6355-12708, or an antisense strand sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions. In some embodiments, the antisense strand sequence comprises or consists of a sequence 100% identical to SEQ ID NOs: 6355-12708.
  • the antisense strand sequence may include the first 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the antisense strand sequence may include the last 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the antisense strand may comprise an overhang.
  • the antisense strand may comprise a modification pattern described herein.
  • the antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand having a sequence in accordance with any of SEQ ID NOs: 13082-13402.
  • the sense strand sequence comprises or consists of sequence at least 75% identical to any one of SEQ ID NOs: 13082-13402, at least 80% identical to any one of SEQ ID NOs: 13082-13402, at least 85% identical to of any one of SEQ ID NOs: 13082-13402, at least 90% identical to any one of SEQ ID NOs13082-13402, or at least 95% identical to any one of SEQ ID NOs: 13082-13402.
  • the sense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 13082-13402, or a sense strand sequence thereof having 1, 2, 3, or 4 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 13082-13402, or a sense strand sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand sequence comprises or consists of a sequence 100% identical to SEQ ID NOs: 13082-13402.
  • the sense strand sequence may include the first 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the sense strand sequence may include the last 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the sense strand may comprise a modification pattern described herein.
  • the sense strand may comprise an overhang.
  • the sense strand may comprise a lipid moiety.
  • the sense strand may comprise a GalNAc moiety.
  • the siRNA comprises an antisense strand having a sequence in accordance with any of SEQ ID NOs: 13403- 13723.
  • the antisense strand sequence comprises or consists of sequence at least -19- Attorney Docket No.54462-742.601 75% identical to any one of SEQ ID NOs: 13403-13723, at least 80% identical to any one of SEQ ID NOs: 13403-13723, at least 85% identical to of any one of SEQ ID NOs: 13403-13723, at least 90% identical to any one of SEQ ID NOs: 13403-13723, or at least 95% identical to any one of SEQ ID NOs: 13403-13723.
  • the antisense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 13403-13723, or an antisense strand sequence thereof having 1, 2, 3, or 4 nucleoside substitutions, additions, or deletions. In some embodiments, the antisense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 13403-13723, or an antisense strand sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions. In some embodiments, the antisense strand sequence comprises or consists of a sequence 100% identical to SEQ ID NOs: 13403- 13723.
  • the antisense strand sequence may include the first 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the antisense strand sequence may include the last 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the antisense strand may comprise an overhang.
  • the antisense strand may comprise a modification pattern described herein.
  • the antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of subset A.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of subset A.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of subset A, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of subset B.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of subset B.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of subset B, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of subset B, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of subset B.
  • the sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of subset C.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of subset C.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of subset C, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of subset C, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of subset C.
  • the sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of subset D.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of subset D.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of subset D, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of subset D, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of subset D.
  • the sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of subset E, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of subset E.
  • the sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of subset F.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at -21- Attorney Docket No.54462-742.601 least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of subset F.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of subset F, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of subset F, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of subset F.
  • the sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of subset I.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of subset I.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of subset I, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of subset I, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of subset I.
  • the sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of subset J.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of subset J.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of subset J, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of subset J, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of subset J.
  • the sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA herein (such as an siRNA in a table herein).
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of an siRNA herein.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of an siRNA herein, -22- Attorney Docket No.54462-742.601 or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of an siRNA herein, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of an siRNA herein.
  • the sense strand or antisense strand may comprise an overhang.
  • the sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of Table 8.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of Table 8.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 8, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 8, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of Table 8. The sense strand or antisense strand may comprise an overhang. The sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of Table 9.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of Table 9.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 9, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 9, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of Table 9. The sense strand or antisense strand may comprise an overhang. The sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of Table 11A.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of Table 11A.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 11A, or a sequence thereof having 3 -23- Attorney Docket No.54462-742.601 or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 11A, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of Table 11A.
  • the sense strand or antisense strand may comprise an overhang.
  • the sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of Table 11B.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of Table 11B.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of Table 14.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of Table 14.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 14, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 14, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of Table 14. The sense strand or antisense strand may comprise an overhang. The sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of Table 18.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of Table 18.
  • the sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of Table 22.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of Table 22.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 22, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 22, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of Table 22. The sense strand or antisense strand may comprise an overhang. The sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 25, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 25, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of Table 25. The sense strand or antisense strand may comprise an overhang. The sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of Table 28.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of Table 28.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 28, or a sequence thereof having 3 or -25- Attorney Docket No.54462-742.601 4 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 28, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of Table 28.
  • the sense strand or antisense strand may comprise an overhang.
  • the sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of Table 31.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of Table 31.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 31, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 31, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of Table 31. The sense strand or antisense strand may comprise an overhang. The sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 35, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 35, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of Table 35. The sense strand or antisense strand may comprise an overhang. The sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of Table 49.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of Table 49.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 49, or a sequence thereof having 3 or -26- Attorney Docket No.54462-742.601 4 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 49, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of Table 49.
  • the sense strand or antisense strand may comprise an overhang.
  • the sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of Table 63.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of Table 63.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 63, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 63, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of Table 63. The sense strand or antisense strand may comprise an overhang. The sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of Table 66.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of Table 66.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 66, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 66, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of Table 66. The sense strand or antisense strand may comprise an overhang. The sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand having a sequence in accordance with any of SEQ ID NOs: 13329, 13334, 13335, 13091, 13259, 13260, 13261, 13262, 13263, 13264, 13265, 13266, 13336, 13311, 13312, 13341, 13099, 13165, 13166, 13167, 13168, 13169, 13170, 13171, 13172, 13173, 13100, 13157, 13158, 13159, 13160, 13161, 13162, 13163, 13164, 13344, 13103, 13216, 13217, 13218, 13219, 13220, 13221, 13222, 13223, 13224, 13307, 13308, 13309, 13310, 13345, 13346, 13106, 13267,
  • the sense -27- Attorney Docket No.54462-742.601 strand sequence comprises or consists of sequence at least 75% identical to any one of SEQ ID NOs: 13329, 13334, 13335, 13091, 13259, 13260, 13261, 13262, 13263, 13264, 13265, 13266, 13336, 13311, 13312, 13341, 13099, 13165, 13166, 13167, 13168, 13169, 13170, 13171, 13172, 13173, 13100, 13157, 13158, 13159, 13160, 13161, 13162, 13163, 13164, 13344, 13103, 13216, 13217, 13218, 13219, 13220, 13221, 13222, 13223, 13224, 13307, 13308, 13309, 13310, 13345, 13346, 13106, 13267, 13268, 13269, 13270, 13271
  • the sense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 13329, 13334, 13335, 13091, 13259, 13260, 13261, 13262, 13263, 13264, 13265, 13266, 13336, 13311, 13312, 13341, 13099, 13165, 13166, 13167, 13168, 13169, 13170, 13171, 13172, 13173, 13100, 13157, 13158, 13159, 13160, 13161, 13162, 13163, 13164, 13344, 13103, 13216, 13217, 13218, 13219, 13220, 13221, 13222, 13223, 13224, 13307, 13308, 13309, 13310, 13345, 13346, 13106, 13267, 13268, 13269, 13270, 13271, 13272, 13273, and 13274., or a sense
  • the sense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 13329, 13334, 13335, 13091, 13259, 13260, 13261, 13262, 13263, 13264, 13265, 13266, 13336, 13311, 13312, 13341, 13099, 13165, 13166, 13167, 13168, 13169, 13170, 13171, 13172, 13173, 13100, 13157, 13158, 13159, 13160, 13161, 13162, 13163, 13164, 13344, 13103, 13216, 13217, 13218, 13219, 13220, 13221, 13222, 13223, 13224, 13307, 13308, 13309, 13310, 13345, 13346, 13106, 13267, 13268, 13269, 13270, 13271, 13272, 13273, and 13274., or a sense
  • the sense strand sequence comprises or consists of a sequence 100% identical to -28- Attorney Docket No.54462-742.601 SEQ ID NOs: 13329, 13334, 13335, 13091, 13259, 13260, 13261, 13262, 13263, 13264, 13265, 13266, 13336, 13311, 13312, 13341, 13099, 13165, 13166, 13167, 13168, 13169, 13170, 13171, 13172, 13173, 13100, 13157, 13158, 13159, 13160, 13161, 13162, 13163, 13164, 13344, 13103, 13216, 13217, 13218, 13219, 13220, 13221, 13222, 13223, 13224, 13307, 13308, 13309, 13310, 13345, 13346, 13106, 13267, 13268, 13269, 13270, 13271, 13272
  • the siRNA comprises an antisense strand having a sequence in accordance with any of SEQ ID NOs: 13650, 13655, 13656, 13412, 13580, 13581, 13582, 13583, 13584, 13585, 13586, 13587, 13657, 13632, 13633, 13662, 13420, 13486, 13487, 13488, 13489, 13490, 13491, 13492, 13493, 13494, 13421, 13478, 13479, 13480, 13481, 13482, 13483, 13484, 13485, 13665, 13424, 13537, 13538, 13539, 13540, 13541, 13542, 13543, 13544, 13545, 13628, 13629, 13630, 13631, 13666, 13667, 13427, 13588, 13589, 13590, 13591, 13592, 13593, 13594, or
  • the antisense strand sequence comprises or consists of sequence at least 75% identical to any one of SEQ ID NOs: 13650, 13655, 13656, 13412, 13580, 13581, 13582, 13583, 13584, 13585, 13586, 13587, 13657, 13632, 13633, 13662, 13420, 13486, 13487, 13488, 13489, 13490, 13491, 13492, 13493, 13494, 13421, 13478, 13479, 13480, 13481, 13482, 13483, 13484, 13485, 13665, 13424, 13537, 13538, 13539, 13540, 13541, 13542, 13543, 13544, 13545, 13628, 13629, 13630, 13631, 13666, 13667, 13427, 13588, 13589, 13590, 13591, 13592, 13593, 13594, or 1359
  • the antisense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 13650, 13655, 13656, 13412, 13580, 13581, 13582, 13583, 13584, 13585, 13586, 13587, 13657, 13632, 13633, 13662, 13420, 13486, 13487, 13488, 13489, 13490, 13491, 13492, 13493, 13494, 13421, 13478, 13479, 13480, 13481, 13482, 13483, 13484, 13485, 13665, 13424, 13537, 13538, 13539, 13540, 13541, 13542, 13543, 13544, 13545, 13628, 13629, 13630, 13631, 13666, 13667, 13427, 13588, 13589, 13590, 13591, 13592, 13593, 13594, or 13595, or an anti
  • the antisense strand sequence comprises or consists of a sequence 100% identical to SEQ ID NOs: 13650, 13655, 13656, 13412, 13580, 13581, 13582, 13583, 13584, 13585, 13586, 13587, 13657, 13632, 13633, 13662, 13420, 13486, 13487, 13488, 13489, 13490, 13491, 13492, 13493, 13494, 13421, 13478, 13479, 13480, 13481, 13482, 13483, 13484, 13485, 13665, 13424, 13537, 13538, 13539, 13540, 13541, 13542, 13543, 13544, 13545, 13628, 13629, 13630, 13631, 13666, 13667, 13427, 13588, 13589, 13590, 13591, 13592, 13593, 13594, or 13595.
  • the antisense strand sequence may include the first 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the antisense strand sequence may include the last 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the antisense strand may comprise an overhang.
  • the antisense strand may comprise a modification pattern described herein.
  • the antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand having a sequence in accordance with any of SEQ ID NOs: 13329, 13334, 13335, 13260, 13264, 13336, 13311, 13341, 13168, 13173, 13161, 13344, 13224, 13345, 13346, 13267, 13268, 13353, 13358, 13359, 13276, 13280, 13360, 13317, 13365, 13185, 13190, 13178, 13368, 13241, 13369, 13370, 13283, 13284, 13377, 13382, 13383, 13292, 13296, 13384, 13323, 13389, 13202, 13207, 13195, 13392, 13258, 13393, 13394, 13299, or 13300.
  • the sense strand sequence comprises or consists of sequence at least 75% identical to any one of SEQ ID NOs: 13329, 13334, 13335, 13260, 13264, 13336, 13311, 13341, 13168, 13173, 13161, 13344, 13224, 13345, 13346, 13267, 13268, 13353, 13358, 13359, 13276, 13280, 13360, 13317, 13365, 13185, 13190, 13178, 13368, 13241, 13369, 13370, 13283, 13284, 13377, 13382, 13383, 13292, 13296, 13384, 13323, 13389, 13202, 13207, 13195, 13392, 13258, 13393, 13394, 13299, or 13300, at least 80% identical to any one of SEQ ID NOs: 13329, 13334, 13335, 13260, 13264, 13336,
  • the sense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 13329, 13334, 13335, 13260, 13264, 13336, 13311, 13341, 13168, 13173, 13161, 13344, 13224, 13345, 13346, 13267, 13268, 13353, 13358, 13359, 13276, 13280, 13360, 13317, 13365, 13185, 13190, 13178, 13368, 13241, 13369, 13370, 13283, 13284, 13377, 13382, 13383, 13292, 13296, 13384, 13323, 13389, 13202, 13207, 13195, 13392, 13258, 13393, 13394, 13299, or 13300, or a sense strand sequence thereof having 1, 2, 3, or 4 nucleoside substitutions, additions, or deletions.
  • the antisense strand sequence comprises or consists of sequence at least 75% identical to any one of SEQ ID NOs: 13650, 13655, 13656, 13581, 13585, 13657, 13632, 13662, 13489, 13494, 13482, 13665, 13545, 13666, 13667, 13588, 13589, 13674, 13679, 13680, 13597, 13601, 13681, 13638, 13686, 13506, 13511, 13499, 13689, 13562, 13690, 13691, 13604, 13605, 13698, 13703, 13704, 13613, 13617, 13705, 13644, 13710, 13523, 13528, 13516, 13713, 13579, 13714, 13715, 13620, or 13621, at least 80% identical to any one of SEQ ID NOs: 13650, 13655, 13656, 13581, 13
  • the antisense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 13650, 13655, 13656, 13581, 13585, 13657, 13632, 13662, 13489, 13494, 13482, 13665, 13545, 13666, 13667, 13588, 13589, 13674, 13679, 13680, 13597, 13601, 13681, 13638, 13686, 13506, 13511, 13499, 13689, 13562, 13690, 13691, 13604, 13605, 13698, 13703, 13704, 13613, 13617, 13705, 13644, 13710, 13523, 13528, 13516, 13713, 13579, 13714, 13715, 13620, or 13621, or an antisense strand sequence thereof having 1, 2, 3, or 4 nucleoside substitutions, additions, or deletions.
  • the antisense strand sequence -32- Attorney Docket No.54462-742.601 comprises or consists of a sequence 100% identical to SEQ ID NOs: 13650, 13655, 13656, 13581, 13585, 13657, 13632, 13662, 13489, 13494, 13482, 13665, 13545, 13666, 13667, 13588, 13589, 13674, 13679, 13680, 13597, 13601, 13681, 13638, 13686, 13506, 13511, 13499, 13689, 13562, 13690, 13691, 13604, 13605, 13698, 13703, 13704, 13613, 13617, 13705, 13644, 13710, 13523, 13528, 13516, 13713, 13579, 13714, 13715, 13620, or 13621.
  • the antisense strand sequence may include the first 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the antisense strand sequence may include the last 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the antisense strand may comprise an overhang.
  • the antisense strand may comprise a modification pattern described herein.
  • the antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises an antisense oligonucleotide (ASO).
  • ASO antisense oligonucleotide
  • the ASO is 12-30 nucleosides in length.
  • the ASO is 14-30 nucleosides in length.
  • the ASO is at least about 10, 11, 12, 13, 14, 15, 16, 17, 18,19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 nucleosides in length, or a range defined by any of the two aforementioned numbers.
  • the ASO is 15-25 nucleosides in length.
  • the ASO is 20 nucleosides in length.
  • the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises an ASO about 12-30 nucleosides in length and comprising a nucleoside sequence complementary to about 12-30 contiguous nucleosides of a full- length human GPAM mRNA sequence such as SEQ ID NO: 12867; wherein (i) the oligonucleotide comprises a modification comprising a modified nucleoside and/or a modified internucleoside linkage, and/or (ii) the composition comprises a pharmaceutically acceptable carrier.
  • the ASO comprise a nucleoside sequence complementary to at least about 10, 11, 12, 13, 14, 15, 16, 17, 18,19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, or more contiguous nucleosides of one of SEQ ID NO: 12867.
  • the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises a modification comprising a modified nucleoside and/or a modified internucleoside linkage, and/or (ii) the composition comprises a pharmaceutically acceptable carrier.
  • the oligonucleotide comprises a modification comprising a modified nucleoside and/or a modified internucleoside linkage. In some embodiments, the oligonucleotide comprises a modified internucleoside linkage. In some embodiments, the modified internucleoside linkage comprises alkylphosphonate, phosphorothioate, methylphosphonate, phosphorodithioate, alkylphosphonothioate, phosphoramidate, carbamate, carbonate, phosphate triester, acetamidate, or carboxymethyl ester, or a combination thereof. In some embodiments, the modified internucleoside linkage comprises one or more phosphorothioate linkages.
  • a phosphorothioate may include a nonbridging oxygen atom in a phosphate backbone of the oligonucleotide that is replaced by -33- Attorney Docket No.54462-742.601 sulfur.
  • Modified internucleoside linkages may be included in siRNAs or ASOs. Benefits of the modified internucleoside linkage may include decreased toxicity or improved pharmacokinetics.
  • the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises a modified internucleoside linkage, wherein the oligonucleotide comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 modified internucleoside linkages, or a range of modified internucleoside linkages defined by any two of the aforementioned numbers.
  • the oligonucleotide comprises no more than 18 modified internucleoside linkages.
  • the oligonucleotide comprises no more than 20 modified internucleoside linkages.
  • the oligonucleotide comprises 2 or more modified internucleoside linkages, 3 or more modified internucleoside linkages, 4 or more modified internucleoside linkages, 5 or more modified internucleoside linkages, 6 or more modified internucleoside linkages, 7 or more modified internucleoside linkages, 8 or more modified internucleoside linkages, 9 or more modified internucleoside linkages, 10 or more modified internucleoside linkages, 11 or more modified internucleoside linkages, 12 or more modified internucleoside linkages, 13 or more modified internucleoside linkages, 14 or more modified internucleoside linkages, 15 or more modified internucleoside linkages, 16 or more modified internucleoside linkages, 17 or more modified internucleoside linkages, 18 or more modified internucleoside linkages, 19 or more modified internucleoside linkages, or 20 or more modified internucleoside linkages.
  • the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises the modified nucleoside.
  • the modified nucleoside comprises a locked nucleic acid (LNA), hexitol nucleic acid (HLA), cyclohexene nucleic acid (CeNA), 2'-O-(2-methoxyethyl), 2'-O-alkyl, 2'-O-allyl, 2'-fluoro, or 2'- deoxy, or a combination thereof.
  • the modified nucleoside comprises a LNA.
  • the modified nucleoside comprises a 2’,4’ constrained ethyl nucleic acid. In some embodiments, the modified nucleoside comprises HLA. In some embodiments, the modified nucleoside comprises CeNA. In some embodiments, the modified nucleoside comprises a 2'-methoxyethyl group. In some embodiments, the modified nucleoside comprises a 2'-O-(2-methoxyethyl) group. In some embodiments, the modified nucleoside comprises a 2'-O-alkyl group. In some embodiments, the modified nucleoside comprises a 2'-O-allyl group. In some embodiments, the modified nucleoside comprises a 2'- fluoro group.
  • the modified nucleoside comprises a 2'-deoxy group. In some embodiments, the modified nucleoside comprises a 2'-O-(2-methoxyethyl). In some embodiments, the modified nucleoside comprises a 2'-O-methyl nucleoside, 2'-deoxyfluoro nucleoside, 2'-O-N- methylacetamido (2'-O-NMA) nucleoside, a 2'-O-dimethylaminoethoxyethyl (2'-O-DMAEOE) nucleoside, 2'-O-aminopropyl (2'-O-AP) nucleoside, or 2'-ara-F, or a combination thereof.
  • the modified nucleoside comprises a 2'-O-methyl nucleoside. In some embodiments, the modified nucleoside comprises a 2'-deoxyfluoro nucleoside. In some embodiments, the modified nucleoside comprises a 2'-O-NMA nucleoside. In some embodiments, the modified nucleoside comprises a 2'-O-DMAEOE nucleoside. In some embodiments, the modified nucleoside comprises a 2'-O- aminopropyl (2'-O-AP) nucleoside. In some embodiments, the modified nucleoside comprises 2'-ara-F.
  • the modified nucleoside comprises one or more 2’fluoro modified nucleosides. In some embodiments, the modified nucleoside comprises a 2' O-alkyl modified nucleoside. Benefits of the modified nucleoside may include decreased toxicity or improved pharmacokinetics. [0076] In some embodiments, the oligonucleotide comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or 21 modified nucleosides, or a range of nucleosides defined by any two of the aforementioned numbers. In some embodiments, the oligonucleotide comprises no more than 19 modified nucleosides.
  • the oligonucleotide comprises no more than 21 modified nucleosides. In some embodiments, the oligonucleotide comprises 2 or more modified nucleosides, 3 or more modified nucleosides, 4 or more modified nucleosides, 5 or more modified nucleosides, 6 or more modified nucleosides, 7 or more modified nucleosides, 8 or more modified nucleosides, 9 or more modified nucleosides, 10 or more modified nucleosides, 11 or more modified nucleosides, 12 or more modified nucleosides, 13 or more modified nucleosides, 14 or more modified nucleosides, 15 or more modified nucleosides, 16 or more modified nucleosides, 17 or more modified nucleosides, 18 or more modified nucleosides, 19 or more modified nucleosides, 20 or more modified nucleosides, or 21 or more modified nucleosides.
  • the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises a moiety attached at a 3’ or 5’ terminus of the oligonucleotide.
  • moieties include a hydrophobic moiety or a sugar moiety, or a combination thereof.
  • the oligonucleotide is an siRNA having a sense strand, and the moiety is attached to a 5’ end of the sense strand.
  • the oligonucleotide is an siRNA having a sense strand, and the moiety is attached to a 3’ end of the sense strand.
  • the oligonucleotide is an siRNA having an antisense strand, and the moiety is attached to a 5’ end of the antisense strand. In some embodiments, the oligonucleotide is an siRNA having an antisense strand, and the moiety is attached to a 3’ end of the antisense strand. In some embodiments, the oligonucleotide is an ASO, and the moiety is attached to a 5’ end of the ASO. In some embodiments, the oligonucleotide is an ASO, and the moiety is attached to a 3’ end of the ASO. [0078] The oligonucleotide may include purines.
  • all purines of the oligonucleotide comprise 2’ fluoro modified purines. In some embodiments, all purines of the oligonucleotide comprise 2’-O-methyl modified purines. In some embodiments, all purines of the oligonucleotide comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines. 2’-O-methyl may include 2’ O-methyl.
  • pyrimidines of the oligonucleotide comprise 2’ fluoro modified pyrimidines. In some embodiments, pyrimidines of the oligonucleotide comprise 2’-O-methyl modified pyrimidines.
  • pyrimidines of the oligonucleotide comprise 2’-O-methyl modified pyrimidines, and purines of the oligonucleotide comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines.
  • pyrimidines of the oligonucleotide comprise 2’ fluoro modified pyrimidines, and purines of the oligonucleotide comprise 2’- O-methyl modified purines.
  • pyrimidines of the oligonucleotide comprise 2’-O- methyl modified pyrimidines, and purines of the oligonucleotide comprise 2’ fluoro modified purines.
  • all purines of the oligonucleotide comprise 2’ fluoro modified purines, and all pyrimidines of the oligonucleotide comprise 2’-O-methyl modified pyrimidines. In some embodiments, all purines of the oligonucleotide comprise 2’-O-methyl modified purines, and all pyrimidines of the oligonucleotide comprise 2’ fluoro modified pyrimidines. In some embodiments, all pyrimidines of the oligonucleotide comprise 2’ fluoro modified pyrimidines, and all purines of the oligonucleotide comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines.
  • all pyrimidines of the oligonucleotide comprise 2’-O-methyl modified pyrimidines, and all purines of the oligonucleotide comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines. In some embodiments, all pyrimidines of the oligonucleotide comprise 2’ fluoro modified pyrimidines, and all purines of the oligonucleotide comprise 2’-O-methyl modified purines. In some embodiments, all pyrimidines of the oligonucleotide comprise 2’-O-methyl modified pyrimidines, and all purines of the oligonucleotide comprise 2’ fluoro modified purines.
  • the oligonucleotide comprises a particular modification pattern.
  • position 9 counting from the 5’ end of the of a strand of the oligonucleotide may have a 2’F modification.
  • position 9 of a strand of the oligonucleotide is a pyrimidine, then all purines in a strand of the oligonucleotide have a 2’OMe modification.
  • -36- Attorney Docket No.54462-742.601 when position 9 is the only pyrimidine between positions 5 and 11 of the sense stand, then position 9 is the only position with a 2’F modification in a strand of the oligonucleotide.
  • position 9 and only one other base between positions 5 and 11 of a strand of the oligonucleotide are pyrimidines, then both of these pyrimidines are the only two positions with a 2’F modification in a strand of the oligonucleotide.
  • any combination of 2’F modifications can be made that give three 2’F modifications in total.
  • all combinations of pyrimidines having the 2’F modification are allowed that have three to five 2’F modifications in total, provided that a strand of the oligonucleotide does not have three 2’F modifications in a row.
  • a strand of the oligonucleotide of any of the siRNAs comprises a modification pattern which conforms to any or all of these a strand of the oligonucleotide rules.
  • position 9 of a strand of the oligonucleotide is a purine
  • all purines in a strand of the oligonucleotide have a 2’OMe modification.
  • position 9 is the only purine between positions 5 and 11 of the sense stand, then position 9 is the only position with a 2’F modification in a strand of the oligonucleotide.
  • both of these purines are the only two positions with a 2’F modification in a strand of the oligonucleotide.
  • position 9 and only two other bases between positions 5 and 11 of a strand of the oligonucleotide are purines, and those two other purines are in adjacent positions so that there would be not three 2’F modifications in a row, then any combination of 2’F modifications can be made that give three 2’F modifications in total.
  • a strand of the oligonucleotide of any of the siRNAs comprises a modification pattern which conforms to any or all of these a strand of the oligonucleotide rules.
  • position 9 of a strand of the oligonucleotide can be a 2’deoxy.
  • a strand of the oligonucleotide of any of the siRNAs comprises a modification pattern which conforms to these a strand of the oligonucleotide rules.
  • position nine of the sense strand comprises a 2’ fluoro-modified pyrimidine.
  • all purines of the sense strand comprise 2’-O-methyl modified purines.
  • 1, 2, 3, 4, or 5 pyrimidines between positions 5 and 11 comprise a 2’flouro- modified pyrimidine, provided there are not three 2’ fluoro-modified pyrimidines in a row.
  • the odd-numbered positions of the antisense strand comprise 2’-O-methyl modified nucleotides.
  • the even-numbered positions of the antisense strand comprise 2’flouro-modified nucleotides and unmodified deoxyribonucleotide.
  • the even- -37- Attorney Docket No.54462-742.601 numbered positions of the antisense strand comprise 2’flouro-modified nucleotides, 2’-O-methyl modified nucleotides and unmodified deoxyribonucleotide.
  • position nine of the sense strand comprises a 2’ fluoro-modified pyrimidine; all purines of the sense strand comprises 2’-O-methyl modified purines; 1, 2, 3, 4, or 5 pyrimidines between positions 5 and 11 comprise a 2’flouro-modified pyrimidine, provided there are not three 2’ fluoro-modified pyrimidines in a row; the odd-numbered positions of the antisense strand comprise 2’-O-methyl modified nucleotides; and the even-numbered positions of the antisense strand comprise 2’flouro-modified nucleotides and unmodified deoxyribonucleotides.
  • position nine of the sense strand comprises a 2’ fluoro-modified purine.
  • all pyrimidines of the sense strand comprise 2’-O-methyl modified purines.
  • 1, 2, 3, 4, or 5 purines between positions 5 and 11 comprise a 2’flouro-modified purine, provided there are not three 2’ fluoro-modified purine in a row.
  • the odd- numbered positions of the antisense strand comprise 2’-O-methyl modified nucleotides.
  • the even-numbered positions of the antisense strand comprise 2’flouro-modified nucleotides and unmodified deoxyribonucleotide.
  • the even-numbered positions of the antisense strand comprise 2’flouro-modified nucleotides, 2’-O-methyl modified nucleotides and unmodified deoxyribonucleotide.
  • position nine of the sense strand comprises a 2’ fluoro-modified purine; all pyrimidine of the sense strand comprises 2’-O-methyl modified pyrimidines; 1, 2, 3, 4, or 5 purines between positions 5 and 11 comprise a 2’flouro-modified purines, provided there are not three 2’ fluoro-modified purines in a row; the odd-numbered positions of the antisense strand comprise 2’-O-methyl modified nucleotides; and the even-numbered positions of the antisense strand comprise 2’flouro-modified nucleotides and unmodified deoxyribonucleotides.
  • position nine of the sense strand comprises an unmodified deoxyribonucleotide.
  • positions 5, 7, and 8 of the sense strand comprise 2’fluoro- modifed nucleotides.
  • all pyrimidines in positions 10 to 21 of the sense strand comprise 2’-O-methyl modified pyrimidines and all purines in positions 10 to 21 of the comprise 2’-O- methyl modified purines or 2’fluoro-modified purines.
  • the odd-numbered positions of the antisense strand comprise 2’-O-methyl modified nucleotides. In some embodiments, the even- numbered positions of the antisense strand comprise 2’flouro-modified nucleotides and unmodified deoxyribonucleotides. In some embodiments, the even-numbered positions of the antisense strand comprise 2’flouro-modified nucleotides, 2’-O-methyl modified nucleotides and unmodified deoxyribonucleotides.
  • position nine of the sense strand comprises an unmodified deoxyribonucleotide; positions 5, 7, and 8 of the sense strand comprise 2’fluoro-modifed nucleotides; all pyrimidines in positions 10 to 21 of the sense strand comprise 2’-O-methyl modified pyrimidines and all purines in positions 10 to 21 of the comprise 2’-O-methyl modified purines or 2’fluoro-modified purines; the odd-numbered positions of the antisense strand comprise 2’-O-methyl modified nucleotides; and the -38- Attorney Docket No.54462-742.601 even-numbered positions of the antisense strand comprise 2’flouro-modified nucleotides and unmodified deoxyribonucleotides.
  • position nine of the sense strand comprises an unmodified deoxyribonucleotide.
  • positions 5, 7, and 8 of the sense strand comprise 2’fluoro- modifed nucleotides.
  • all purines in positions 10 to 21 of the sense strand comprise 2’-O-methyl modified purines and all pyrimidines in positions 10 to 21 of the comprise 2’-O-methyl modified pyrimidines or 2’fluoro-modified pyrimidines.
  • the odd-numbered positions of the antisense strand comprise 2’-O-methyl modified nucleotides.
  • the even-numbered positions of the antisense strand comprise 2’flouro-modified nucleotides and unmodified deoxyribonucleotides. In some embodiments, the even-numbered positions of the antisense strand comprise 2’flouro-modified nucleotides, 2’-O-methyl modified nucleotides and unmodified deoxyribonucleotides.
  • position nine of the sense strand comprises an unmodified deoxyribonucleotide; positions 5, 7, and 8 of the sense strand comprise 2’fluoro-modifed nucleotides; all purines in positions 10 to 21 of the sense strand comprise 2’-O-methyl modified purines and all pyrimidines in positions 10 to 21 of the comprise 2’-O-methyl modified pyrimidines or 2’fluoro-modified pyrimidines; the odd-numbered positions of the antisense strand comprise 2’-O-methyl modified nucleotides; and the even-numbered positions of the antisense strand comprise 2’flouro-modified nucleotides and unmodified deoxyribonucleotide.
  • the moiety includes a negatively charged group attached at a 5’ end of the oligonucleotide. This may be referred to as a 5’-end group.
  • the negatively charged group is attached at a 5’ end of an antisense strand of an siRNA disclosed herein.
  • the 5’-end group may be or include a 5’-end phosphorothioate, 5’-end phosphorodithioate, 5’-end vinylphosphonate (5’-VP), 5’-end methylphosphonate, 5’-end cyclopropyl phosphonate, or a 5’-deoxy-5’-C-malonyl.
  • the 5’-end group may comprise 5’-VP.
  • the 5’-VP comprises a trans-vinylphosphate or cis-vinylphosphate.
  • the 5’-end group may include an extra 5’ phosphate.
  • a combination of 5’-end groups may be used.
  • the oligonucleotide includes a negatively charged group.
  • the negatively charged group may aid in cell or tissue penetration.
  • the negatively charged group may be attached at a 5’ or 3’ end (e.g. a 5’ end) of the oligonucleotide. This may be referred to as an end group.
  • the end group may be or include a phosphorothioate, phosphorodithioate, vinylphosphonate, methylphosphonate, cyclopropyl phosphonate, or a deoxy-C-malonyl.
  • the end group may include an extra 5’ phosphate such as an extra 5’ phosphate.
  • a combination of end groups may be used.
  • the oligonucleotide includes a phosphate mimic.
  • the phosphate mimic comprises vinyl phosphonate.
  • the vinyl phosphonate comprises a trans-vinylphosphate.
  • the vinyl phosphonate comprises a cis-vinylphosphate.
  • the vinyl phosphonate increases the stability of the oligonucleotide. In some embodiments, the vinyl phosphonate increases the accumulation of the oligonucleotide in tissues. In some embodiments, the vinyl phosphonate protects the oligonucleotide from an exonuclease or a phosphatase. In some embodiments, the vinyl phosphonate improves the binding affinity of the oligonucleotide with the siRNA processing machinery.
  • the oligonucleotide includes 1 vinyl phosphonate. In some embodiments, the oligonucleotide includes 2 vinyl phosphonates. In some embodiments, the oligonucleotide includes 3 vinyl phosphonates. In some embodiments, the oligonucleotide includes 4 vinyl phosphonates. In some embodiments, the antisense strand of the oligonucleotide comprises a vinyl phosphonate at the 5’ end. In some embodiments, the antisense strand of the oligonucleotide comprises a vinyl phosphonate at the 3’ end.
  • the sense strand of the oligonucleotide comprises a vinyl phosphonate at the 5’ end. In some embodiments, the sense strand of the oligonucleotide comprises a vinyl phosphonate at the 3’ end.
  • Hydrophobic moieties [0095] In some embodiments, the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises a hydrophobic moiety.
  • the hydrophobic moiety may be attached at a 3’ or 5’ terminus of the oligonucleotide.
  • the hydrophobic moiety may include a lipid such as a fatty acid.
  • the hydrophobic moiety may include a hydrocarbon.
  • the hydrocarbon may be linear.
  • the hydrocarbon may be non-linear.
  • the hydrophobic moiety may include a lipid moiety or a cholesterol moiety, or a combination thereof.
  • the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises a lipid attached at a 3’ or 5’ terminus of the oligonucleotide.
  • the lipid comprises cholesterol, myristoyl, palmitoyl, stearoyl, lithocholoyl, docosanoyl, docosahexaenoyl, myristyl, palmityl, stearyl, or ⁇ -tocopherol, or a combination thereof.
  • the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises a hydrophobic ligand or moiety.
  • the hydrophobic ligand or moiety comprises cholesterol.
  • the hydrophobic ligand or moiety comprises a cholesterol derivative.
  • the hydrophobic -40- Attorney Docket No.54462-742.601 ligand or moiety is attached at a 3’ terminus of the oligonucleotide. In some embodiments, the hydrophobic ligand or moiety s attached at a 5’ terminus of the oligonucleotide. In some embodiments, the composition comprises a sense strand, and the hydrophobic ligand or moiety is attached to the sense strand (e.g. attached to a 5’ end of the sense strand, or attached to a 3’ end of the sense strand). In some embodiments, the composition comprises an antisense strand, and the hydrophobic ligand or moiety is attached to the antisense strand (e.g.
  • the composition comprises a hydrophobic ligand or moiety attached at a 3’ or 5’ terminus of the oligonucleotide.
  • a hydrophobic moiety is attached to the oligonucleotide (e.g. a sense strand and/or an antisense strand of a siRNA).
  • a hydrophobic moiety is attached at a 3’ terminus of the oligonucleotide.
  • a hydrophobic moiety is attached at a 5’ terminus of the oligonucleotide.
  • the hydrophobic moiety comprises cholesterol. In some embodiments, the hydrophobic moiety includes a cyclohexanyl. [0099] In some embodiments, the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises a lipid attached at a 3’ or 5’ terminus of the oligonucleotide. In some embodiments, a lipid is attached at a 3’ terminus of the oligonucleotide. In some embodiments, a lipid is attached at a 5’ terminus of the oligonucleotide.
  • the lipid comprises cholesterol, myristoyl, palmitoyl, stearoyl, lithocholoyl, docosanoyl, docosahexaenoyl, myristyl, palmityl, stearyl, or ⁇ -tocopherol, or a combination thereof.
  • the lipid comprises stearyl, lithocholyl, docosanyl, docosahexaenyl, or myristyl.
  • the lipid comprises cholesterol.
  • the lipid includes a sterol such as cholesterol.
  • the lipid comprises stearyl, t-butylphenol, n-butylphenol, octylphenol, dodecylphenol, phenyl n-dodecyl, octadecylbenzamide, hexadecylbenzamide, or octadecylcyclohexyl.
  • the lipid comprises phenyl para C12.
  • the oligonucleotide comprises any aspect of the following structure: de comprises any aspect of the following structure: .
  • the oligonucleotide comprises any aspect of the following structure: -41- Attorney Docket No.54462-742.601 n
  • the oligo The aspect included in the oligonucleotide may include the entire structure, or may include the lipid moiety, of any of the structures shown.
  • n is 1-3.
  • n is 1.
  • n is 2.
  • n is 3.
  • R is an alkyl group.
  • the alkyl group contains 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 carbons.
  • the alkyl group contains 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or 18 carbons, or a range defined by any two of the aforementioned numbers of carbons. In some embodiments, the alkyl group contains 4-18 carbons.
  • the lipid moiety comprises an alcohol or ether. [00101] In some embodiments, the lipid includes a fatty acid. In some embodiments, the lipid comprises a lipid depicted in Table 1. The example lipid moieties in Table 1 are shown attached at a 5’ end of an oligonucleotide, in which the 5’ terminal phosphate of the oligonucleotide is shown with the lipid moiety.
  • a lipid moiety in Table 1 may be attached at a different point of attachment than shown.
  • the point of attachment of any of the lipid moieties in the table may be at a 3’ oligonucleotide end.
  • the lipid is used for targeting the oligonucleotide to a non-hepatic cell or tissue.
  • Table 1 Hydrophobic moiety examples Moi t -42- Attorney Docket No.54462-742.601 n d ph octa hex -43- Attorney Docket No.54462-742.601 octa phe [00102]
  • the lipid or lipid moiety includes 16 to 18 carbons.
  • the hydrophobic moiety may include a linker that comprises a carbocycle.
  • the carbocycle may be six-membered. Some examples of a carbocycle include phenyl or cyclohexyl.
  • the linker may include a phenyl.
  • the linker may include a cyclohexyl.
  • the lipid may be attached to the carbocycle, which may in turn be attached at a phosphate (e.g.5’ or 3’ phosphate) of the oligonucleotide.
  • the lipid or hydrocarbon, and the end of the sense are connected to the phenyl or cyclohexyl linker in the 1,4; 1,3; or 1,2 substitution pattern (e.g. the para, meta, or ortho phenyl configuration).
  • the lipid or hydrocarbon, and the end of the sense are connected to the phenyl or cyclohexyl linker in the 1,4 substitution pattern (e.g. the para phenyl configuration).
  • the lipid may be attached to the carbocycle in the 1,4 substitution pattern relative to the oligonucleotide.
  • the lipid may be attached to the carbocycle in the 1,3 substitution pattern relative to the oligonucleotide.
  • the lipid may be attached to the carbocycle in the 1,2 substitution pattern relative to the oligonucleotide.
  • the lipid may be attached to the carbocycle in the ortho orientation relative to the oligonucleotide.
  • the lipid may be -44- Attorney Docket No.54462-742.601 attached to the carbocycle in the para orientation relative to the oligonucleotide.
  • the lipid may be attached to the carbocycle in the meta orientation relative to the oligonucleotide.
  • the lipid moiety may comprise or consist of the following structure .
  • the lipid moiety comprises or consists of the following structure: .
  • the lipid moiety comprises the following structure: .
  • the lipid moiety comprises or consist of the following structure: .
  • the dotted line indicates a covalent connection.
  • the covalent connection may between an end of the sense or antisense strand.
  • the connection may be to the 5’ end of the sense strand.
  • n is 0-3.
  • n is 1-3.
  • n is 0.
  • n is 1.
  • n is 2.
  • n is 3.
  • n is 4. In some embodiments, n is 5. In some embodiments, n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.
  • R is an alkyl group. In some embodiments, the alkyl group contains 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 carbons. In some embodiments, the alkyl group contains 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or 18 carbons, or a range defined by any two of the aforementioned numbers of carbons. In some embodiments, R comprises or consists of an alkyl group containing 4-18 carbons. [00105] The lipid moiety may be attached at a 5’ end of the oligonucleotide.
  • the 5’ end may have one phosphate linking the lipid moiety to a 5’ carbon of a sugar of the oligonucleotide.
  • the 5’ end may have two phosphates linking the lipid moiety to a 5’ carbon of a sugar of the oligonucleotide.
  • the 5’ end may have three phosphates linking the lipid moiety to a 5’ carbon of a sugar of the oligonucleotide.
  • the 5’ end may have one phosphate connected to the 5’ carbon of a sugar of the oligonucleotide, where the one phosphate is connected to the lipid moiety.
  • the 5’ end may have two phosphates connected to the 5’ carbon of a sugar of the oligonucleotide, where the one of the two phosphates is connected to the lipid -45- Attorney Docket No.54462-742.601 moiety.
  • the 5’ end may have three phosphates connected to the 5’ carbon of a sugar of the oligonucleotide, where the one of the three phosphates is connected to the lipid moiety.
  • the sugar may include a ribose.
  • the sugar may include a deoxyribose.
  • the sugar may be modified a such as a 2’ modified sugar (e.g. a 2’ O-methyl or 2’ fluoro ribose).
  • a phosphate of the 5’ end may include a modification such as a sulfur in place of an oxygen.
  • Two phosphates of the 5’ end may include a modification such as a sulfur in place of an oxygen.
  • Three phosphates of the 5’ end may include a modification such as a sulfur in place of an oxygen.
  • the oligonucleotide includes 1 lipid moiety. In some embodiments, the oligonucleotide includes 2 lipid moieties. In some embodiments, the oligonucleotide includes 3 lipid moieties. In some embodiments, the oligonucleotide includes 4 lipid moieties.
  • Some embodiments relate to a method of making an oligonucleotide comprising a hydrophobic conjugate.
  • a strategy for making hydrophobic conjugates may include use of a phosphoramidite reagent based upon a 6-membered ring alcohol such as a phenol or cyclohexanol. The phosphoramidite may be reacted to a nucleotide to connect the nucleotide to the hydrophobic moiety, and thereby produce the hydrophobic conjugate.
  • Some examples of phosphoramidite reagents that may be used to produce a hydrophobic conjugate are provided as follows: or .
  • n is 1-3.
  • n is 1.
  • n is 2.
  • n is 3.
  • R is an alkyl group.
  • the alkyl group contains 1, 2, 3, 4, 5, 6, 7, 8, 9, -46- Attorney Docket No.54462-742.601 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 carbons.
  • the alkyl group contains 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or 18 carbons, or a range defined by any two of the aforementioned numbers of carbons.
  • R comprises or consists of an alkyl group containing 4-18 carbons.
  • any one of the phosphoramidite reagents may be reacted to a 5’ end of an oligonucleotide to produce an oligonucleotide comprising a hydrophobic moiety.
  • the phosphoramidite reagents is reacted to a 5’ end of a sense strand of an siRNA.
  • the sense strand may then be hybridized to an antisense strand to form a duplex.
  • the hybridization may be performed by incubating the sense and antisense strands in solution at a given temperature.
  • the temperature may be gradually reduced.
  • the temperature may comprise or include a temperature comprising an annealing temperature for the sense and antisense strands.
  • the temperature may be below or include a temperature below the annealing temperature for the sense and antisense strands.
  • the temperature may be below a melting temperature of the sense and antisense strands.
  • the lipid may be attached to the oligonucleotide by a linker.
  • the linker may include a polyethyleneglycol (e.g. tetraethyleneglycol).
  • the modifications described herein may be useful for delivery to a cell or tissue, for example, extrahepatic delivery or targeting of an oligonucleotide composition.
  • the modifications described herein may be useful for targeting an oligonucleotide composition to a cell or tissue. 2.
  • the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises a sugar moiety.
  • the sugar moiety may include an N-acetyl galactose moiety (e.g. an N-acetylgalactosamine (GalNAc) moiety), an N-acetyl glucose moiety (e.g. an N-acetylglucosamine (GlcNAc) moiety), a fucose moiety, or a mannose moiety.
  • the sugar moiety may include 1, 2, 3, or more sugar molecules.
  • the sugar moiety may be attached at a 3’ or 5’ terminus of the oligonucleotide.
  • the sugar moiety may include an N-acetyl galactose moiety.
  • the sugar moiety may include an N-acetylgalactosamine (GalNAc) moiety.
  • the sugar moiety may include an N-acetyl glucose moiety.
  • the sugar moiety may include N-acetylglucosamine (GlcNAc) moiety.
  • the sugar moiety may include a fucose moiety.
  • the sugar moiety may include a mannose moiety.
  • N-acetyl glucose, GlcNAc, fucose, or mannose may be useful for targeting macrophages when they target or bind a mannose receptor such as CD206.
  • the sugar moiety may be useful for binding or targeting an asialoglycoprotein receptor such as an asialoglycoprotein receptor of a hepatocyte.
  • the GalNAc moiety may bind to an asialoglycoprotein receptor.
  • the GalNAc moiety may target a hepatocyte.
  • the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises an N-acetylgalactosamine (GalNAc) moiety.
  • GalNAc may be useful for hepatocyte targeting.
  • the GalNAc moiety may include a bivalent or trivalent branched linker.
  • the oligo may be attached to 1, 2 or 3 GalNAcs through a bivalent or trivalent branched linker.
  • the GalNAc moiety may include 1, 2, 3, or more GalNAc molecules.
  • the GalNAc moiety may be attached at a 3’ or 5’ terminus of the oligonucleotide.
  • the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises an N-acetylgalactosamine (GalNAc) ligand for hepatocyte targeting.
  • the composition comprises GalNAc.
  • the composition comprises a GalNAc derivative.
  • the GalNAc ligand is attached at a 3’ terminus of the oligonucleotide.
  • the GalNAc ligand is attached at a 5’ terminus of the oligonucleotide.
  • the composition comprises a sense strand, and the GalNAc ligand is attached to the sense strand (e.g. attached to a 5’ end of the sense strand, or attached to a 3’ end of the sense strand).
  • the composition comprises an antisense strand, and the GalNAc ligand is attached to the antisense strand (e.g. attached to a 5’ end of the antisense strand, or attached to a 3’ end of the antisense strand).
  • the composition comprises a GalNAc ligand attached at a 3’ or 5’ terminus of the oligonucleotide.
  • compositions comprising an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises a GalNAc moiety.
  • the GalNAc moiety may be included in any formula, structure, or GalNAc moiety shown below.
  • described herein is a compound (e.g.
  • oligonucleotide represented by Formula (I) or (II): (II); or a salt thereof, wherein J is an oligonucleotide; each w is independently selected from any value from 1 to 20; each v is independently selected from any value from 1 to 20; n is selected from any value from 1 to 20; m is selected from any value from 1 to 20; z is selected from any value from 1 to 3, wherein if z is 3, Y is C if z is 2, Y is CR 6 , or if z is 1, Y is C(R 6 ) 2 ; Q is selected from: C 3-10 carbocycle optionally substituted with one or more substituents independently selected from halogen, -CN, -NO 2 , -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -C(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7
  • each w is independently selected from any value from 1 to 10. In some embodiments, each w is independently selected from any value from 1 to 5. In some embodiments, each w is 1. In some embodiments, each v is independently selected from any value from 1 to 10. In some embodiments, each v is independently selected from any value from 1 to 5. In some embodiments, each v -49- Attorney Docket No.54462-742.601 is 1. In some embodiments, n is selected from any value from 1 to 10. In some embodiments, n is selected from any value from 1 to 5. In some embodiments, n is 2. In some embodiments, m is selected from any value from 1 to 10. In some embodiments, m is selected from any value from 1 to 5.
  • m is selected from 1 and 2.
  • z is 3 and Y is C.
  • Q is selected from C5-6 carbocycle optionally substituted with one or more substituents independently selected from halogen, -CN, -NO2, -OR 7 , -SR 7 , -N(R 7 )2, -C(O)R 7 , -C(O)N(R 7 )2, -N(R 7 )C(O)R 7 , - N(R 7 )C(O)N(R 7 )2, -OC(O)N(R 7 )2, -N(R 7 )C(O)OR 7 , -C(O)OR 7 , -OC(O)R 7 , and -S(O)R 7 .
  • Q is selected from C5-6 carbocycle optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2, and -NH2.
  • Q is selected from phenyl and cyclohexyl, each of which is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO2, and -NH2.
  • Q is selected from phenyl.
  • Q is selected from cyclohexyl.
  • R 1 is selected from -OP(O)(OR 7 )O-, -SP(O)(OR 7 )O-, -OP(S)(OR 7 )O-, -OP(O)(SR 7 )O-, - OP(O)(OR 7 )S-, -OP(O)(O-)O-, -SP(O)(O-)O-, -OP(S)(O-)O-, -OP(O)(S-)O-, -OP(O)(O-)S-, -OP(O)(OR 7 )NR 7 -, -OP(O)(N(R 7 )2)NR 7 -, -OP(OR 7 )O-, -OP(N(R 7 )2)O-, -OP(OR 7 )N(R 7 )-, and -OPN(R 7 )2- NR 7 .
  • R 1 is selected from -OP(O)(OR 7 )O-, -SP(O)(OR 7 )O-, -OP(S)(OR 7 )O-, - OP(O)(SR 7 )O-, -OP(O)(OR 7 )S-, -OP(O)(O-)O-, -SP(O)(O-)O-, -OP(S)(O-)O-, -OP(O)(S-)O-, -OP(O)(O- )S-, and -OP(OR 7 )O-.
  • R 1 is selected from -OP(O)(OR 7 )O-, -OP(S)(OR 7 )O-, - OP(O)(O-)O-, -OP(S)(O-)O-, -OP(O)(S-)O-, and -OP(OR 7 )O-. In some embodiments, R 1 is selected from - OP(O)(OR 7 )O- and -OP(OR 7 )O-.
  • R 2 is selected from C1-3 alkyl substituted with one or more substituents independently selected from halogen, -OR 7 , -OC(O)R 7 , -SR 7 , -N(R 7 )2, -C(O)R 7 , and -S(O)R 7 .
  • R 2 is selected from C1-3 alkyl substituted with one or more substituents independently selected from -OR 7 , -OC(O)R 7 , -SR 7 , and -N(R 7 )2.
  • R 2 is selected from C1-3 alkyl substituted with one or more substituents independently selected from -OR 7 and - OC(O)R 7 .
  • R 3 is selected from halogen, -OR 7 , -SR 7 , -N(R 7 )2, -C(O)R 7 , -OC(O)R 7 , and -S(O)R 7 . In some embodiments, R 3 is selected from -OR 7 -SR 7 , -OC(O)R 7 , and -N(R 7 )2. In some embodiments, R 3 is selected from -OR 7 - and -OC(O)R 7 .
  • R 4 is selected from halogen, -OR 7 , -SR 7 , -N(R 7 ) 2 , -C(O)R 7 , -OC(O)R 7 , and -S(O)R 7 . In some embodiments, R 4 is selected from -OR 7 -SR 7 , -OC(O)R 7 , and -N(R 7 ) 2. In some embodiments, R 4 is selected from -OR 7 - and -OC(O)R 7 .
  • R 5 is selected from -OC(O)R 7 , -OC(O)N(R 7 ) 2 , -N(R 7 )C(O)R 7 , -N(R 7 )C(O)N(R 7 ) 2 , and -N(R 7 )C(O)OR 7 . In some embodiments, R 5 is selected from -OC(O)R 7 and -N(R 7 )C(O)R 7 .
  • each R 7 is independently selected from C 1-6 alkyl optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, and -SH.
  • Q is phenyl or cyclohexyl, each of which is optionally substituted with one or more substituents independently selected from halogen, -CN, -OH, -SH, -NO 2 , -NH 2 , and C 1-3 alkyl;
  • R 1 is selected from -OP(O)(OR 7 )O-, -OP(S)(OR 7 )O-, -OP(O)(O-)O-, -OP(S)(O-)O-, -OP(O)(S-)O-, and - OP(OR 7 )O-;
  • R 2 is C 1 alkyl substituted with -OH or -OC(O)CH 3 ;
  • R 3 is -OH or -OC(O)CH3;
  • the oligonucleotide (J) is attached at a 5’ end or a 3’ end of the oligonucleotide.
  • the oligonucleotide comprises DNA. In some embodiments, the oligonucleotide comprises RNA. In some embodiments, the oligonucleotide comprises one or more modified internucleoside linkages. In some embodiments, the one or more modified internucleoside linkages comprise alkylphosphonate, phosphorothioate, methylphosphonate, phosphorodithioate, alkylphosphonothioate, phosphoramidate, carbamate, carbonate, phosphate triester, acetamidate, or carboxymethyl ester, or a combination thereof.
  • the oligonucleotide comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 modified internucleoside linkages.
  • the compound binds to an asialoglycoprotein receptor.
  • the compound targets a hepatocyte.
  • J is the oligonucleotide: -57- Attorney Docket No.54462-742.601 [00115] .
  • J may include one or more additional phosphates, or one or more phosphorothioates linking to the oligonucleotide.
  • J may include one or more additional phosphates linking to the oligonucleotide.
  • J may include one or more phosphorothioates linking to the oligonucleotide.
  • Some embodiments include the following, where J is the oligonucleotide: [00117] .
  • J may include one or more additional phosphates, or one or more phosphorothioates linking to the oligonucleotide.
  • J may include one or more additional phosphates linking to the oligonucleotide.
  • J may include one or more phosphorothioates linking to the oligonucleotide.
  • J is the oligonucleotide: -58- Attorney Docket No.54462-742.601 may include one or more phosphates or phosphorothioates linking to the oligonucleotide.
  • J may include one or more phosphates linking to the oligonucleotide.
  • J may include a phosphate linking to the oligonucleotide.
  • J may include one or more phosphorothioates linking to the oligonucleotide.
  • J may include a phosphorothioate linking to the oligonucleotide.
  • Some embodiments include the following, where J is the oligonucleotide: .
  • J The structure in this compound attached to the oligonucleotide (J) is an example of a GalNAc moiety.
  • J may include one or more phosphates or phosphorothioates linking to the oligonucleotide.
  • J may include one or more phosphates linking to the oligonucleotide.
  • J may include a phosphate linking to the oligonucleotide.
  • -59- Attorney Docket No.54462-742.601 J may include one or more phosphorothioates linking to the oligonucleotide.
  • J may include a phosphorothioate linking to the oligonucleotide.
  • J is the oligonucleotide: [001 .
  • J may include one or more additional phosphates, or one or more phosphorothioates linking to the oligonucleotide.
  • J may include one or more additional phosphates linking to the oligonucleotide.
  • J may include one or more phosphorothioates linking to the oligonucleotide.
  • Some embodiments include the following, where J is the oligonucleotide: -60- Attorney Docket No.54462-742.601 .
  • J may include one or more additional phosphates, or one or more phosphorothioates linking to the oligonucleotide.
  • J may include one or more additional phosphates linking to the oligonucleotide.
  • J may include one or more phosphorothioates linking to the oligonucleotide.
  • Some embodiments include the following, where J is the oligonucleotide: .
  • J may include one or more phosphates or phosphorothioates linking to the oligonucleotide.
  • J may include one or more phosphates linking to the oligonucleotide.
  • J may include a phosphate linking to the oligonucleotide.
  • J may include one or more phosphorothioates linking to the oligonucleotide.
  • J may include a phosphorothioate linking to the oligonucleotide.
  • J is the oligonucleotide: .
  • the structure in this compound attached to the oligonucleotide (J) may be referred to as “ETL17,” and is an example of a GalNAc moiety.
  • J may include one or more phosphates or phosphorothioates linking to the oligonucleotide.
  • J may include one or more phosphates linking to the oligonucleotide.
  • J may include a phosphate linking to the oligonucleotide.
  • J may include one or more phosphorothioates linking to the oligonucleotide.
  • J may include a phosphorothioate linking to the oligonucleotide.
  • Some embodiments include the following, where the phosphate or “5’” indicates a connection to the oligonucleotide: [00124] -62- Attorney Docket No.54462-742.601 [00125] Some embodiments include the following, where the phosphate or “5’” indicates a connection to the oligonucleotide: [001 [00127] Some embodiments include the following, where J is the oligonucleotide: [00128] include one or more phosphates or phosphorothioates linking to the oligonucleotide.
  • J may include one or more phosphates linking to the oligonucleotide.
  • J may include a phosphate linking to the oligonucleotide.
  • J may include one or more phosphorothioates linking to the oligonucleotide.
  • J may include a phosphorothioate linking to the oligonucleotide.
  • Some embodiments include the following, where J is the oligonucleotide: -63- Attorney Docket No.54462-742.601 [001 .
  • the structure in this compound attached to the oligonucleotide (J) may be referred to as ETL1, and is an example of a GalNAc moiety.
  • J may include one or more phosphates or phosphorothioates linking to the oligonucleotide.
  • J may include one or more phosphates linking to the oligonucleotide.
  • J may include a phosphate linking to the oligonucleotide.
  • J may include one or more phosphorothioates linking to the oligonucleotide.
  • J may include a phosphorothioate linking to the oligonucleotide 3.
  • the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises an siRNA comprising a sense strand and an antisense strand, wherein the sense strand comprises modification pattern 1S: 5’-NfsnsNfnNfnNfNfNfnNfnNfnNfnNfnNfnNfsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 2S: 5’-nsnsnnnNfnNfNfNfnnnnnnnnsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 3S: 5’-nsnsnnnNfnNfnNfnnnnnnnnsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 4S: 5’-NfsnsNfnNfnNfNfNfnNfnNfnNfnNfnNfnNfsnsnN-moiety-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, “s” is a phosphorothioate linkage, and N comprises one or more nucleosides.
  • the sense strand comprises modification pattern 5S: 5’-nsnsnnnNfnNfNfNfnnnnnnnnsnsnN-moiety-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, “s” is a phosphorothioate linkage, and N comprises one or more nucleosides.
  • the moiety in modification pattern 4S or 5S is a lipid moiety. In some embodiments, the moiety in modification pattern 4S or 5S is a sugar moiety.
  • the sense strand comprises modification pattern 6S: -64- Attorney Docket No.54462-742.601 5’-NfsnsNfnNfnNfnNfnNfnNfnNfnNfnNfnNfsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 7S: 5’-nsnsnnNfNfNfNfNfnnnnnnnsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 8S: 5’-nsnsnnnnNfNfNfNfnnnnnnnnsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 9S: 5’-nsnsnnnnnNfNfNfnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 10S: 5’-snnnnNfNfnnNfnnnnnnnsn -3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 11S: 5’-snnnnnNfnnNfnNfnnnnnnnsn -3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 12S: 5’-snnnnnNfNfNfdNnnnnnnnnnsnsn -3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “dN” is a 2’ deoxy-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 13S: 5’-snnnnnNfnnNfNfnnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 14S: 5’-snnnnNfnnNfNfnnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 15S: 5’-snnnnNfnnnNfnNfnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 16S: 5’-snnnnNfNfnNfNfnNfnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 17S: 5’-snnnnnnnNfNfnNfnnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 18S: 5’-snnnnnNfNfnNfNfnnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 19S: 5’-snnnnNfnnNfNfnNfnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 20S: 5’-snnnnnNfnNfNfnnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 21S: 5’-snnnnnNfNfNfNfnnnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- -65- Attorney Docket No.54462-742.601 methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 22S: 5’-nsnsnnnnnNfnNfnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 23S: 5’-nsnsnnnnNfnnNfnNfnnnnnnsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 24S: 5’-nsnsnnnnNfNfnNfnnnnnnnnsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 25S: 5’-nsnsnnnnNfNfnNfnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 26S: 5’-nsnsnnnNfnnnNfnNfnnnnnnsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 27S: 5’-nsnsnnnNfnNfnNfnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 28S: 5’-nsnsnnNfNfnnNfnnnnnnnnsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 29S: 5’-nsnsnnnnnnNfNfnnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 30S: 5’-nsnsnnnnnNfNfnnnnnnnnsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 31S: 5’-nsnsnnnnNfnnNfNfnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 32S: 5’-nsnsnnnNfnnnNfNfnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 33S: 5’-nsnsnnnNfNfnnNfNfnnnnnnnsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 34S: 5’-nsnsnnnnnnNfNfnNfnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 35S: 5’-nsnsnnnnNfnNfNfnnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 36S: 5’-nsnsnnnnNfnNfNfnNfnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 37S: -66- Attorney Docket No.54462-742.601 5’-nsnsnnnnNfNfNfNfnNfnnnnnnnsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 38S: 5’-nsnsnnnNfnnNfNfnnnnnnnnsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 39S: 5’-nsnsnnnNfnnNfNfnNfnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 40S: 5’-nsnsnnnNfNfnnNfnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 41S: 5’-nsnsnnNfNfnNfNfnnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 42S: 5’-nsnsnnnnNfnNfnnnnnnnnsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 43S: 5’-nsnsnnnnNfnnNfnnnnnnnnsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 44S: 5’-nsnsnnnnnnNfNfNfnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 45S: 5’-nsnsnnnnnNfnNfNfnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 46S: 5’-nsnsnnnnnNfNfNfNfnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 47S: 5’-nsnsnnnnNfnNfNfNfNfnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 48S: 5’-nsnsnnnnNfNfNfNfNfnnnnnnnsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 49S: 5’-nsnsnnnNfnnNfNfNfNfnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 50S: 5’-nsnsnnnnNfNfnNfNfnnnnnnnsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 51S: 5’-nsnsnnnNfnNfnNfNfnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 52S: 5’-nsnsnnnNfnNfNfNfNfnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate -67- Attorney Docket No.54462-742.601 linkage.
  • the sense strand comprises modification pattern 53S:5'- snnnnnNfNfNfdNnnnnnnnnnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, “Nm” is a 2’-O-(2-methoxyethyl) nucleoside, and “s” is a phosphorothioate linkage. In some embodiments, the Nm is a 2’-O-(2-methoxyethyl) thymine.
  • the sense strand comprises modification pattern 54S:5'-snnnnnNfNfNfNfnnnnnnNmnnnnnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, “Nm” is a 2’-O-(2- methoxyethyl) nucleoside, and “s” is a phosphorothioate linkage. In some embodiments, the Nm is a 2’- O-(2-methoxyethyl) thymine.
  • the sense strand comprises modification pattern 55S: 5'-snnnnTmNfNfNfNfnnnNmnnnnnnnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, “Nm” is a 2’-O-(2-methoxyethyl) nucleoside, and “s” is a phosphorothioate linkage. In some embodiments, the Nm is a 2’-O-(2-methoxyethyl) thymine.
  • the sense strand comprises modification pattern 56S:5'-snnnnnnnNfNfNfnnnnnnnsn- 3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 57S:5'- snnnnnnNfnNfNfnnnnnnnnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 58S:5'-snnnnNfndNnNfnNfnnnnnnnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 59S:5'- snnnNmnNfNfNfnnnNmnnnnnnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 60S:5'-snnnNmnNfNfNfnnNmnnnnnnnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 61S:5'- snnnNmnNfNfNfnnnNmnnnnnnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, “Nm” is a 2’-O-(2-methoxyethyl) nucleoside, and “s” is a phosphorothioate linkage. In some embodiments, the Nm is a 2’-O-(2-methoxyethyl) thymine.
  • the sense strand comprises modification pattern 62S:5'-snnnNmnNfNfNfNfnnnNmnnnnnnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, “Nm” is a 2’-O-(2- methoxyethyl) nucleoside, and “s” is a phosphorothioate linkage. In some embodiments, the Nm is a 2’- O-(2-methoxyethyl) thymine.
  • the sense strand comprises modification pattern 63S:5'-snnnNmnNfNfNfNfnnnNmnnnnnnnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, “Nm” is a 2’-O-(2-methoxyethyl) nucleoside, and “s” is a phosphorothioate linkage. In some embodiments, the Nm is a 2’-O-(2-methoxyethyl) thymine.
  • the sense strand comprises modification pattern 64S:5'-snnnnNfnNfnNfnnnnnnnnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 65AS:5'-snnnNmnnNfNfNfnnNmnnnnnnnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, “Nm” is a 2’-O-(2-methoxyethyl) nucleoside, and “s” is a -68- Attorney Docket No.54462-742.601 phosphorothioate linkage. In some embodiments, the Nm is a 2’-O-(2-methoxyethyl) thymine.
  • the sense strand comprises modification pattern 66AS:5'- snnnNmNfnNfNfNfNfnnNmnnnnnnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, “Nm” is a 2’-O-(2-methoxyethyl) nucleoside, and “s” is a phosphorothioate linkage. In some embodiments, the Nm is a 2’-O-(2-methoxyethyl) thymine.
  • the sense strand comprises modification pattern 67AS:5'- snnnNmnNfNfNfnnNmnnnnnnnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O- methyl modified nucleoside, “Nm” is a 2’-O-(2-methoxyethyl) nucleoside, and “s” is a phosphorothioate linkage. In some embodiments, the Nm is a 2’-O-(2-methoxyethyl) thymine.
  • the sense strand comprises modification pattern 68AS:5'-snnnNmnNfNfNfNfnnnnNmnnnnnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, “Nm” is a 2’-O-(2- methoxyethyl) nucleoside, and “s” is a phosphorothioate linkage. In some embodiments, the Nm is a 2’- O-(2-methoxyethyl) thymine.
  • the sense strand comprises modification pattern 69AS:5'-snnnNmNfnNfNfNfNfnNmnnnnnnnsn-3' wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, “Nm” is a 2’-O-(2-methoxyethyl) nucleoside, and “s” is a phosphorothioate linkage. In some embodiments, the Nm is a 2’-O-(2-methoxyethyl) thymine.
  • the sense strand comprises modification pattern 70AS:5'- snnnNmNfnNfNfNfNfnnnNmnnnnnsnsn-3' wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, “Nm” is a 2’-O-(2-methoxyethyl) nucleoside, and “s” is a phosphorothioate linkage. In some embodiments, the Nm is a 2’-O-(2-methoxyethyl) thymine.
  • the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises an siRNA comprising a sense strand and an antisense strand, wherein the antisense strand comprises modification pattern 1AS: 5’-nsNfsnNfnNfnNfnNfnnnNfnNfnsnsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the antisense strand comprises modification pattern 2AS: 5’-nsNfsnnnNfnNfNfnnnnNfnNfnnsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the antisense strand comprises modification pattern 3AS: 5’-nsNfsnnnNfnnnnnnnnNfnNfnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the antisense strand comprises modification pattern 4AS: 5’-nsNfsnNfnNfnnnnnnnNfnNfnnnsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the antisense strand comprises modification pattern 5AS: 5’-nsNfsnnnnnnnnnnnnNfnNfnnsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the antisense strand comprises modification pattern 6AS: 5’-nsNfsnnnNfnnNfnnnnNfnnnsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the antisense strand comprises modification pattern 7AS: 5’-nsNfsnNfnNfnNfnNfnNfnNfnNfnNfnsnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, -69- Attorney Docket No.54462-742.601 “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the antisense strand comprises modification pattern 8AS: 5’-nsNfsnnnnnnnnnnnnnnnsn-3’, wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 9AS:5'- nsNfsnnnNfnNfnNfnNfnNfnNfnNfnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 10AS:5'-nsNfsnNfnNfnnNfnnnnNfnNfnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 11AS:5'-nsNfsnNfnNfnnNfnnNfnNfnNfnNfnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 12AS:5'-nsNfsnNfnnNfnNfnNfnNfnNfnNfnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 13AS:5'-nsNfsnnNfnNfnNfnNfnNfnNfnNfnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 14AS:5'-nsNfsnNfnNfnNfnnnNfnNfnNfnNfnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 15AS:5'-nsNfsnnnNfnNfnnnNfnNfnNfnNfnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 16AS:5'-nsNfsnnnNfnNfnNfnnnNfnNfnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 17AS:5'-nsNfsnNfnnNfnnNfnNfnNfnNfnNfnNfnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 18AS:5'-nsNfsnNfnnNfnnNfnnnNfnNfnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 19AS:5'-nsNfsnnNfnNfnnNfnNfnNfnNfnNfnNfnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 20AS:5'-nsNfsnNfnnnNfnNfnNfnNfnNfnNfnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 21AS:5'-nsNfsnnnnNfnNfnnNfnNfnNfnNfnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 22AS:5'-nsNfsnNfnnNfnNfnnnnNfnNfnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern -70- Attorney Docket No.54462-742.601 23AS:5'-nsNfsnnNfnNfnNfnnNfnNfnNfnNfnNfnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 24AS:5'-nsNfsnnNfnNfnNfnnnnNfnNfnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 25AS:5'-nsNfsnNfnNfnNfnNfnNfnNfnNfnnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 26AS:5'-nsNfsnNfnnNfnnnnNfnNfnNfnNfnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 27AS:5'-nsNfsnNfnnNfnNfnnNfnNfnNfnNfnNfnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 28AS:5'-nsNfsnnNfnNfnnNfnnnNfnNfnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 29AS:5'-nsNfsnnnNfnnnNfnNfnNfnNfnNfnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 30AS:5'-nsNfsnnnNfNfnnNfnNfnNfnNfnNfnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 31AS:5'-nsNfsnnNfnNfNfnNfnNfnNfnNfnNfnNfnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the sense strand comprises modification pattern 32AS:5'-nsNfsnnNfnNfNfnnnNfnNfnNfnNfnsnsn-3', wherein “Nf” is a 2’ fluoro-modified nucleoside, “n” is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises an siRNA comprising a sense strand and an antisense strand, wherein the sense strand comprises pattern 1S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • siRNA comprising a sense strand and an antisense strand
  • the sense strand comprises pattern 1S
  • the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS,
  • the sense strand comprises pattern 2S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 3S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 4S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, -71- Attorney Docket No.54462-742.601 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 5S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 6S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 7S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 8S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 9S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 10S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 11S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 12S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 13S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 14S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 15S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 16S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 17S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, -72- Attorney Docket No.54462-742.601 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 18S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 19S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 20S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 21S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 22S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 23S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 24S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 26S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 27S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 28S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 29S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 30S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, -73- Attorney Docket No.54462-742.601 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 31S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 32S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 33S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 34S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 35S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 36S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 37S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 38S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 39S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 40S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 41S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 42S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 43S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, -74- Attorney Docket No.54462-742.601 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 44S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 46S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 47S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 48S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 49S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 50S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 51S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 52S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 53S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 54S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 55S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 56S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, -75- Attorney Docket No.54462-742.601 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 57S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 58S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 59S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 60S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 61S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 62S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 63S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 64S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 65S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 66S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 67S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 68S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 69S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, -76- Attorney Docket No.54462-742.601 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 70S and the antisense strand comprises pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 1AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70Sand the antisense strand comprises pattern 2AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 3AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 4AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 5AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 6AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 7AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 9AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 10AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 11AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 12AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 13AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 14AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 15AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 16AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, -78- Attorney Docket No.54462-742.601 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 18AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 19AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 20AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 21AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 22AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 23AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 24AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 25AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, -79- Attorney Docket No.54462-742.601 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 27AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 28AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 29AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 30AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 31AS.
  • the sense strand comprises pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S and the antisense strand comprises pattern 32AS.
  • the sense strand comprises modification pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S .
  • the sense strand comprises modification pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, or 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the antisense strand comprises modification pattern 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, or 8AS, 9AS, 10AS, 11AS, 12AS, 13AS, 14AS, 15AS, 16AS, 17AS, 18AS, 19AS, 20AS, 21AS, -80- Attorney Docket No.54462-742.601 22AS, 23AS, 24AS, 25AS, 26AS, 27AS, 28AS, 29AS, 30AS, 31AS, or 32AS.
  • the antisense strand comprises modification pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, or 70S .
  • the sense strand or the antisense strand comprises modification pattern ASO1.
  • purines of the sense strand comprise 2’ fluoro modified purines. In some embodiments, purines of the sense strand comprise 2’-O-methyl modified purines. In some embodiments, purines of the sense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines. In some embodiments, all purines of the sense strand comprise 2’ fluoro modified purines. In some embodiments, all purines of the sense strand comprise 2’-O-methyl modified purines. In some embodiments, all purines of the sense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines.
  • pyrimidines of the sense strand comprise 2’ fluoro modified pyrimidines. In some embodiments, pyrimidines of the sense strand comprise 2’-O-methyl modified pyrimidines. In some embodiments, pyrimidines of the sense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines. In some embodiments, all pyrimidines of the sense strand comprise 2’ fluoro modified pyrimidines. In some embodiments, all pyrimidines of the sense strand comprise 2’-O- methyl modified pyrimidines.
  • all pyrimidines of the sense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines.
  • purines of the sense strand comprise 2’ fluoro modified purines, and pyrimidines of the sense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines.
  • purines of the sense strand comprise 2’-O-methyl modified purines, and pyrimidines of the sense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines.
  • purines of the sense strand comprise 2’ fluoro modified purines, and pyrimidines of the sense strand comprise 2’-O-methyl modified pyrimidines. In some embodiments, purines of the sense strand comprise 2’-O-methyl modified purines, and pyrimidines of the sense strand comprise 2’ fluoro modified pyrimidines. In some embodiments, pyrimidines of the sense strand comprise 2’ fluoro modified pyrimidines, and purines of the sense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines.
  • pyrimidines of the sense strand comprise 2’-O-methyl modified pyrimidines, and purines of the sense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines. In some embodiments, pyrimidines of the sense strand comprise 2’ fluoro modified pyrimidines, and purines of the sense strand comprise 2’-O-methyl modified purines. In some embodiments, pyrimidines of the sense strand comprise 2’-O-methyl modified pyrimidines, and purines of the sense strand comprise 2’ fluoro modified purines.
  • all purines of the sense strand comprise 2’ fluoro modified purines, and all pyrimidines of the sense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines. In some embodiments, all purines of the sense strand comprise 2’-O-methyl modified purines, and all pyrimidines of the sense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified -81- Attorney Docket No.54462-742.601 pyrimidines. In some embodiments, all purines of the sense strand comprise 2’ fluoro modified purines, and all pyrimidines of the sense strand comprise 2’-O-methyl modified pyrimidines.
  • all purines of the sense strand comprise 2’-O-methyl modified purines, and all pyrimidines of the sense strand comprise 2’ fluoro modified pyrimidines. In some embodiments, all pyrimidines of the sense strand comprise 2’ fluoro modified pyrimidines, and all purines of the sense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines. In some embodiments, all pyrimidines of the sense strand comprise 2’-O-methyl modified pyrimidines, and all purines of the sense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines.
  • all pyrimidines of the sense strand comprise 2’ fluoro modified pyrimidines, and all purines of the sense strand comprise 2’-O- methyl modified purines. In some embodiments, all pyrimidines of the sense strand comprise 2’-O-methyl modified pyrimidines, and all purines of the sense strand comprise 2’ fluoro modified purines. [00140] In some embodiments, purines of the antisense strand comprise 2’ fluoro modified purines. In some embodiments, purines of the antisense strand comprise 2’-O-methyl modified purines. In some embodiments, purines of the antisense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines.
  • all purines of the antisense strand comprise 2’ fluoro modified purines. In some embodiments, all purines of the antisense strand comprise 2’-O-methyl modified purines. In some embodiments, all purines of the antisense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines. [00141] In some embodiments, pyrimidines of the antisense strand comprise 2’ fluoro modified pyrimidines. In some embodiments, pyrimidines of the antisense strand comprise 2’-O-methyl modified pyrimidines.
  • pyrimidines of the antisense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines. In some embodiments, all pyrimidines of the antisense strand comprise 2’ fluoro modified pyrimidines. In some embodiments, all pyrimidines of the antisense strand comprise 2’-O-methyl modified pyrimidines. In some embodiments, all pyrimidines of the antisense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines.
  • purines of the antisense strand comprise 2’ fluoro modified purines, and pyrimidines of the antisense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines. In some embodiments, purines of the antisense strand comprise 2’-O-methyl modified purines, and pyrimidines of the antisense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines. In some embodiments, purines of the antisense strand comprise 2’ fluoro modified purines, and pyrimidines of the antisense strand comprise 2’-O-methyl modified pyrimidines.
  • purines of the antisense strand comprise 2’-O-methyl modified purines
  • pyrimidines of the antisense strand comprise 2’ fluoro modified pyrimidines.
  • pyrimidines of the antisense strand comprise 2’ fluoro modified pyrimidines
  • purines of the antisense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines.
  • pyrimidines of the antisense strand comprise 2’-O-methyl modified pyrimidines
  • purines of the antisense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines.
  • pyrimidines of the antisense strand comprise 2’ fluoro modified pyrimidines
  • purines of the antisense strand comprise 2’- -82- Attorney Docket No.54462-742.601 O-methyl modified purines.
  • pyrimidines of the antisense strand comprise 2’-O- methyl modified pyrimidines
  • purines of the antisense strand comprise 2’ fluoro modified purines.
  • all purines of the antisense strand comprise 2’ fluoro modified purines
  • all pyrimidines of the antisense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines.
  • all purines of the antisense strand comprise 2’-O-methyl modified purines, and all pyrimidines of the antisense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines. In some embodiments, all purines of the antisense strand comprise 2’ fluoro modified purines, and all pyrimidines of the antisense strand comprise 2’-O-methyl modified pyrimidines. In some embodiments, all purines of the antisense strand comprise 2’-O-methyl modified purines, and all pyrimidines of the antisense strand comprise 2’ fluoro modified pyrimidines.
  • all pyrimidines of the antisense strand comprise 2’ fluoro modified pyrimidines, and all purines of the antisense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines. In some embodiments, all pyrimidines of the antisense strand comprise 2’-O-methyl modified pyrimidines, and all purines of the antisense strand comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines. In some embodiments, all pyrimidines of the antisense strand comprise 2’ fluoro modified pyrimidines, and all purines of the antisense strand comprise 2’-O-methyl modified purines.
  • all pyrimidines of the antisense strand comprise 2’-O-methyl modified pyrimidines
  • all purines of the antisense strand comprise 2’ fluoro modified purines.
  • the modified oligonucleotide may be an siRNA that includes modifications to the ribose rings, and phosphate linkages. The modifications may be in particular patterns that maximize cell delivery, stability, and efficiency.
  • the siRNA may also include a vinyl phosphonate and a hydrophobic group. These modifications may aid in delivery to a cell or tissue within a subject.
  • the modified oligonucleotide may be used in a method such as a treatment method or a method of reducing gene expression.
  • the oligonucleotide comprises a duplex consisting of 21 nucleotide single strands with base pairing between 19 of the base pairs.
  • the duplex comprises single-stranded 2 nucleotide overhangs are at the 3’ ends of each strand.
  • One strand (antisense strand) is complementary to a GPAM mRNA. Each end of the antisense strand has one to two phosphorothioate bonds. The 5’ end has an optional phosphate mimic such as a vinyl phosphonate.
  • the oligonucleotide is used to knock down a GPAM mRNA or a target protein.
  • the sense strand has the same sequence as the GPAM mRNA.
  • the sense strand of any of the siRNAs comprises siRNA with a particular modification pattern.
  • position 9 counting from the 5’ end of the sense strand may have a 2’F modification.
  • position 9 of the sense strand is a pyrimidine
  • all purines in the sense strand have a 2’OMe modification.
  • position 9 is the only pyrimidine between positions 5 and 11 of the sense stand, then -83- Attorney Docket No.54462-742.601
  • position 9 is the only position with a 2’F modification in the sense strand.
  • position 9 and only one other base between positions 5 and 11 of the sense strand are pyrimidines, then both of these pyrimidines are the only two positions with a 2’F modification in the sense strand.
  • any combination of 2’F modifications can be made that give three 2’F modifications in total.
  • all combinations of pyrimidines having the 2’F modification are allowed that have three to five 2’F modifications in total, provided that the sense strand does not have three 2’F modifications in a row.
  • the sense strand of any of the siRNAs comprises a modification pattern which conforms to any or all of these sense strand rules.
  • position 9 of the sense strand when position 9 of the sense strand is a purine, then all purines in the sense strand have a 2’OMe modification. In some embodiments, when position 9 is the only purine between positions 5 and 11 of the sense stand, then position 9 is the only position with a 2’F modification in the sense strand. In some embodiments, when position 9 and only one other base between positions 5 and 11 of the sense strand are purines, then both of these purines are the only two positions with a 2’F modification in the sense strand.
  • any combination of 2’F modifications can be made that give three 2’F modifications in total.
  • all combinations of purines having the 2’F modification are allowed that have three to five 2’F modifications in total, provided that the sense strand does not have three 2’F modifications in a row.
  • the sense strand of any of the siRNAs comprises a modification pattern which conforms to any or all of these sense strand rules.
  • the siRNA comprises a sense strand, an antisense strand, and a lipid moiety connected to an end of the sense or antisense strand; wherein the lipid moiety comprises a phenyl or cyclohexanyl linker, wherein the linker is connected to a lipid and to the end of the sense or antisense strand.
  • any one of the following is true with regard to the sense strand: all purines comprise 2’ fluoro modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines; all purines comprise 2’-O-methyl modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines; all purines comprise 2’ fluoro modified purines, and all pyrimidines comprise 2’-O-methyl modified pyrimidines; all pyrimidines comprise 2’ fluoro modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; all pyrimidines comprise 2’-O-methyl modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; all pyrimidines comprise 2’-O-methyl modified pyrimidines, and all purines comprise a
  • any one of the following is true with regard to the antisense strand: all purines comprise 2’ fluoro modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines; all purines comprise 2’-O-methyl modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’-O- -84- Attorney Docket No.54462-742.601 methyl modified pyrimidines; all purines comprise 2’-O-methyl modified purines, and all pyrimidines comprise 2’ fluoro modified pyrimidines; all pyrimidines comprise 2’ fluoro modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; all pyrimidines comprise 2’- O-methyl modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; or all pyr
  • the siRNA comprises comprising a sense strand and an antisense strand; wherein the antisense strand comprises a 5’ end comprising a vinyl phosphonate and 2 phosphorothioate linkages, and a 3’ end comprising 2 phosphorothioate linkages; wherein the sense strand comprises a 5’ end comprising a hydrophobic moiety, and a 3’ end comprising 2 phosphorothioate linkages; wherein any one of the following is true with regard to the sense strand: all purines comprise 2’ fluoro modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’- O-methyl modified pyrimidines, all purines comprise 2’-O-methyl modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines, all purines comprise 2’ fluoro modified purines, and all pyrimidines comprise 2’-O-methyl modified pyrimidines,
  • any one of the following is true with regard to the sense strand: all purines comprise 2’ fluoro modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’- O-methyl modified pyrimidines; all purines comprise 2’-O-methyl modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines; all purines comprise 2’ fluoro modified purines, and all pyrimidines comprise 2’-O-methyl modified pyrimidines; all pyrimidines comprise 2’ fluoro modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; all pyrimidines comprise 2’-O-methyl modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; all pyrimidines comprise 2’-O-methyl modified pyrimidines, and all purines comprise a
  • all purines comprise 2’ fluoro modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines; with the proviso that a deoxy nucleoside may be included in the sense strand.
  • all purines comprise 2’-O-methyl modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines; with the proviso that a deoxy nucleoside may be included in the sense strand.
  • all purines comprise 2’ fluoro modified purines, and all pyrimidines comprise 2’-O-methyl modified pyrimidines; with the proviso that a deoxy nucleoside may be included in the sense strand.
  • all pyrimidines comprise 2’ fluoro modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; with the proviso that a deoxy nucleoside may be included in the sense strand.
  • all pyrimidines comprise 2’-O-methyl modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; with the proviso that a deoxy nucleoside may be included in the sense strand.
  • all pyrimidines comprise 2’ fluoro modified pyrimidines, and all purines comprise 2’-O-methyl modified purines; with the proviso that a deoxy nucleoside may be included in the sense strand.
  • the sense strand includes the deoxy nucleoside. The deoxy nucleoside may be at nucleoside position 9 of the sense strand.
  • the sense strand does not include a deoxy nucleoside.
  • the deoxy nucleoside of the sense strand may be otherwise unmodified.
  • any one of the following is true with regard to the antisense strand: all purines comprise 2’ fluoro modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’- O-methyl modified pyrimidines; all purines comprise 2’-O-methyl modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines; all purines comprise 2’-O-methyl modified purines, and all pyrimidines comprise 2’ fluoro modified pyrimidines; all pyrimidines comprise 2’ fluoro modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; all pyrimidines comprise 2’-O-methyl modified pyrimidines, and all purines comprise 2’-O-methyl
  • all purines comprise 2’ fluoro modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines; with the proviso that a deoxy nucleoside may be included in the antisense strand.
  • all purines comprise 2’-O-methyl modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines; with the proviso that a deoxy nucleoside may be included in the antisense strand.
  • all purines comprise 2’-O-methyl modified purines, and all pyrimidines comprise 2’ fluoro modified pyrimidines; with the proviso that a deoxy nucleoside may be included in the antisense strand.
  • all pyrimidines comprise 2’ fluoro modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; with the proviso that a deoxy nucleoside may be included in the antisense strand.
  • all pyrimidines comprise 2’-O-methyl modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; with the proviso that a deoxy nucleoside may be included in the antisense strand.
  • all pyrimidines comprise 2’-O-methyl modified pyrimidines, and all purines comprise 2’ fluoro modified purines; with the proviso that a deoxy -86- Attorney Docket No.54462-742.601 nucleoside may be included in the antisense strand.
  • the antisense strand includes the deoxy nucleoside.
  • the deoxy nucleoside may be at nucleoside position 9 of the antisense strand.
  • the antisense strand does not include a deoxy nucleoside.
  • the deoxy nucleoside of the antisense strand may be otherwise unmodified.
  • any one of the following is true with regard to the sense strand: all purines comprise 2’ fluoro modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’- O-methyl modified pyrimidines; all purines comprise 2’-O-methyl modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines; all purines comprise 2’ fluoro modified purines, and all pyrimidines comprise 2’-O-methyl modified pyrimidines; all pyrimidines comprise 2’ fluoro modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; all pyrimidines comprise 2’-O-methyl modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; all pyrimidines comprise 2’-O-methyl modified pyrimidines, and all purines comprise a
  • all purines comprise 2’ fluoro modified purines
  • all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines; with the proviso that a deoxy nucleoside or a 2’-O-(2-methoxyethyl) nucleoside may be included in the sense strand.
  • all purines comprise 2’-O-methyl modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines; with the proviso that a deoxy nucleoside or a 2’-O-(2- methoxyethyl) nucleoside may be included in the sense strand.
  • all purines comprise 2’ fluoro modified purines, and all pyrimidines comprise 2’-O-methyl modified pyrimidines; with the proviso that a deoxy nucleoside or a 2’-O-(2-methoxyethyl) nucleoside may be included in the sense strand.
  • all pyrimidines comprise 2’ fluoro modified pyrimidines
  • all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; with the proviso that a deoxy nucleoside or a 2’-O-(2-methoxyethyl) nucleoside may be included in the sense strand.
  • all pyrimidines comprise 2’-O-methyl modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; with the proviso that a deoxy nucleoside or a 2’-O-(2-methoxyethyl) nucleoside may be included in the sense strand.
  • all pyrimidines comprise 2’ fluoro modified pyrimidines, and all purines comprise 2’-O-methyl modified purines; with the proviso that a deoxy nucleoside or a 2’-O-(2-methoxyethyl) nucleoside may be included in the sense strand.
  • the sense strand includes the deoxy nucleoside.
  • the deoxy nucleoside may be at nucleoside position 9 of the sense strand.
  • the sense strand does not include a deoxy nucleoside.
  • the deoxy nucleoside of the sense strand may be otherwise unmodified.
  • the sense strand includes the a 2’-O-(2-methoxyethyl) nucleoside.
  • the 2’-O-(2- methoxyethyl) nucleoside may be at nucleoside position 4 of the sense strand.
  • the 2’-O-(2-methoxyethyl) nucleoside may include a 2’-O-(2-methoxyethyl) thymine nucleoside.
  • the sense strand does not include the a 2’-O-(2-methoxyethyl) nucleoside.
  • the 2’-O-(2-methoxyethyl) nucleoside of the sense strand may be otherwise unmodified.
  • any one of the following is true with regard to the antisense strand: all purines comprise 2’ fluoro modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’- O-methyl modified pyrimidines; all purines comprise 2’-O-methyl modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines; all purines comprise 2’-O-methyl modified purines, and all pyrimidines comprise 2’ fluoro modified pyrimidines; all pyrimidines comprise 2’ fluoro modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; all pyrimidines comprise 2’-O-methyl modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; or
  • all purines comprise 2’ fluoro modified purines
  • all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines; with the proviso that a deoxy nucleoside or a 2’-O-(2-methoxyethyl) nucleoside may be included in the antisense strand.
  • all purines comprise 2’-O-methyl modified purines
  • all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines; with the proviso that a deoxy nucleoside or a 2’-O-(2-methoxyethyl) nucleoside may be included in the antisense strand.
  • all purines comprise 2’-O-methyl modified purines
  • all pyrimidines comprise 2’ fluoro modified pyrimidines; with the proviso that a deoxy nucleoside or a 2’-O-(2-methoxyethyl) nucleoside may be included in the antisense strand.
  • all pyrimidines comprise 2’ fluoro modified pyrimidines
  • all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; with the proviso that a deoxy nucleoside or a 2’-O-(2-methoxyethyl) nucleoside may be included in the antisense strand.
  • all pyrimidines comprise 2’-O-methyl modified pyrimidines
  • all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; with the proviso that a deoxy nucleoside or a 2’-O-(2-methoxyethyl) nucleoside may be included in the antisense strand.
  • all pyrimidines comprise 2’-O-methyl modified pyrimidines
  • all purines comprise 2’ fluoro modified purines; with the proviso that a deoxy nucleoside or a 2’-O-(2-methoxyethyl) nucleoside may be included in the antisense strand.
  • the antisense strand includes the deoxy nucleoside.
  • the deoxy nucleoside may be at nucleoside position 9 of the antisense strand.
  • the antisense strand does not include a deoxy nucleoside.
  • the deoxy nucleoside of the antisense strand may be otherwise unmodified.
  • the antisense strand includes the a 2’-O-(2- methoxyethyl) nucleoside.
  • the 2’-O-(2-methoxyethyl) nucleoside may be at nucleoside position 4 of the sense strand.
  • the 2’-O-(2-methoxyethyl) nucleoside may include a 2’-O-(2-methoxyethyl) thymine nucleoside.
  • the antisense strand does not include the a 2’-O-(2-methoxyethyl) nucleoside.
  • the 2’-O-(2-methoxyethyl) nucleoside of the antisense strand may be otherwise unmodified.
  • any one of the following is true with regard to the sense strand: all purines comprise 2’ fluoro modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’- O-methyl modified pyrimidines; all purines comprise 2’-O-methyl modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines; all purines comprise 2’ fluoro -88- Attorney Docket No.54462-742.601 modified purines, and all pyrimidines comprise 2’-O-methyl modified pyrimidines; all pyrimidines comprise 2’ fluoro modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; all pyrimidines comprise 2’-O-methyl modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; all pyrimidines comprise 2’-O
  • all purines comprise 2’ fluoro modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’- O-methyl modified pyrimidines; with the proviso that a 2’-O-(2-methoxyethyl) nucleoside may be included in the sense strand.
  • all purines comprise 2’-O-methyl modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines; with the proviso that a 2’-O-(2-methoxyethyl) nucleoside may be included in the sense strand.
  • all purines comprise 2’ fluoro modified purines, and all pyrimidines comprise 2’-O-methyl modified pyrimidines; with the proviso that a 2’-O-(2-methoxyethyl) nucleoside may be included in the sense strand.
  • all pyrimidines comprise 2’ fluoro modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; with the proviso that a 2’-O-(2-methoxyethyl) nucleoside may be included in the sense strand.
  • all pyrimidines comprise 2’-O-methyl modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; with the proviso that a 2’-O- (2-methoxyethyl) nucleoside may be included in the sense strand.
  • all pyrimidines comprise 2’ fluoro modified pyrimidines, and all purines comprise 2’-O-methyl modified purines; with the proviso that a 2’-O-(2-methoxyethyl) nucleoside may be included in the sense strand.
  • the sense strand includes the a 2’-O-(2-methoxyethyl) nucleoside.
  • the 2’- O-(2-methoxyethyl) nucleoside may be at nucleoside position 4 of the sense strand.
  • the 2’-O-(2- methoxyethyl) nucleoside may include a 2’-O-(2-methoxyethyl) thymine nucleoside.
  • the sense strand does not include the a 2’-O-(2-methoxyethyl) nucleoside.
  • the 2’-O-(2- methoxyethyl) nucleoside of the sense strand may be otherwise unmodified.
  • any one of the following is true with regard to the antisense strand: all purines comprise 2’ fluoro modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’- O-methyl modified pyrimidines; all purines comprise 2’-O-methyl modified purines, and all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines; all purines comprise 2’-O-methyl modified purines, and all pyrimidines comprise 2’ fluoro modified pyrimidines; all pyrimidines comprise 2’ fluoro modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; all pyrimidines comprise 2’-O-methyl modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; all pyrimidines comprise 2’-O-methyl modified pyrimidines, and all purines comprise
  • all purines comprise 2’ fluoro modified purines
  • all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines; with the proviso that a 2’-O-(2- methoxyethyl) nucleoside may be included in the antisense strand.
  • all purines comprise 2’-O-methyl modified purines
  • all pyrimidines comprise a mixture of 2’ fluoro and 2’-O-methyl modified pyrimidines; with the proviso that a 2’-O-(2-methoxyethyl) nucleoside may be included in the antisense strand.
  • all purines comprise 2’-O-methyl modified purines, and all pyrimidines comprise 2’ fluoro modified pyrimidines; with the proviso that a 2’-O-(2-methoxyethyl) nucleoside may be included in the antisense strand.
  • all pyrimidines comprise 2’ fluoro modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; with the proviso that a 2’- O-(2-methoxyethyl) nucleoside may be included in the antisense strand.
  • all pyrimidines comprise 2’-O-methyl modified pyrimidines, and all purines comprise a mixture of 2’ fluoro and 2’-O-methyl modified purines; with the proviso that a 2’-O-(2-methoxyethyl) nucleoside may be included in the antisense strand.
  • all pyrimidines comprise 2’-O-methyl modified pyrimidines, and all purines comprise 2’ fluoro modified purines; with the proviso that a 2’-O-(2-methoxyethyl) nucleoside may be included in the antisense strand.
  • the antisense strand includes the a 2’-O-(2-methoxyethyl) nucleoside.
  • the 2’-O-(2-methoxyethyl) nucleoside may be at nucleoside position 4 of the sense strand.
  • the 2’-O-(2- methoxyethyl) nucleoside may include a 2’-O-(2-methoxyethyl) thymine nucleoside.
  • the antisense strand does not include the a 2’-O-(2-methoxyethyl) nucleoside.
  • the 2’-O-(2- methoxyethyl) nucleoside of the antisense strand may be otherwise unmodified.
  • any one of the following is true with regard to the sense strand, with the proviso that the sense strand may include a 2’ deoxy nucleoside: all purine nucleosides comprise 2’ fluoro, and all pyrimidine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl, all purine nucleosides comprise 2’-O-methyl, and all pyrimidine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl, all purine nucleosides comprise 2’ fluoro, and all pyrimidine nucleosides comprise 2’-O-methyl, all pyrimidine nucleosides comprise 2’ fluoro, and all purine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl, all pyrimidine nucleosides comprise 2’-O-methyl, all pyrimidine nucleosides comprise 2’-O-methyl, all pyrimidine nucle
  • all purine nucleosides comprise 2’ fluoro, and all pyrimidine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl, with the proviso that the sense strand may include a 2’ deoxy nucleoside.
  • all purine nucleosides comprise 2’-O-methyl, and all pyrimidine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl, with the proviso that the sense strand may include a 2’ deoxy nucleoside.
  • all purine nucleosides comprise 2’ fluoro, and all pyrimidine nucleosides comprise 2’-O-methyl, with the proviso that the sense strand may include a 2’ deoxy nucleoside.
  • all pyrimidine nucleosides comprise 2’ fluoro, and all purine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl, with the proviso that the sense strand may include a 2’ deoxy nucleoside.
  • all pyrimidine nucleosides comprise 2’-O-methyl, and all purine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl, with the proviso that the sense strand may include a 2’ deoxy nucleoside.
  • all pyrimidine -90- Attorney Docket No.54462-742.601 nucleosides comprise 2’ fluoro, and all purine nucleosides comprise 2’-O-methyl, with the proviso that the sense strand may include a 2’ deoxy nucleoside.
  • the sense strand includes the 2’ deoxy nucleoside.
  • the sense strand does not include the 2’ deoxy nucleoside.
  • Some embodiments include a proviso that the sense strand may include a 2’-O-(2-methoxyethyl) nucleoside (e.g. at position 4, counting from 5’ to 3’). Some embodiments include the 2’-O-(2-methoxyethyl) nucleoside in the sense strand. Some embodiments do not include the 2’-O-(2-methoxyethyl) nucleoside in the sense strand.
  • any one of the following is true with regard to the sense strand: all purine nucleosides comprise 2’ fluoro, and all pyrimidine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl, all purine nucleosides comprise 2’-O-methyl, and all pyrimidine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl, all purine nucleosides comprise 2’ fluoro, and all pyrimidine nucleosides comprise 2’-O-methyl, all pyrimidine nucleosides comprise 2’ fluoro, and all purine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl, all pyrimidine nucleosides comprise 2’-O-methyl, and all purine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl, all pyrimidine nucleosides comprise 2’-O-methyl, and
  • all purine nucleosides comprise 2’ fluoro, and all pyrimidine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl. In some embodiments, in the sense strand, all purine nucleosides comprise 2’-O-methyl, and all pyrimidine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl. In some embodiments, in the sense strand, all purine nucleosides comprise 2’ fluoro, and all pyrimidine nucleosides comprise 2’-O-methyl.
  • all pyrimidine nucleosides comprise 2’ fluoro, and all purine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl. In some embodiments, in the sense strand, all pyrimidine nucleosides comprise 2’-O-methyl, and all purine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl. In some embodiments, in the sense strand, all pyrimidine nucleosides comprise 2’ fluoro, and all purine nucleosides comprise 2’-O-methyl.
  • the sense strand may include a 2’-O-methoxyethyl nucleoside (e.g. at position 4, counting from 5’ to 3’). Some embodiments include the 2’-O-methoxyethyl nucleoside in the sense strand. Some embodiments do not include the 2’-O-methoxyethyl nucleoside in the sense strand.
  • any one of the following is true with regard to the antisense strand, with the proviso that the antisense strand may include a 2’ deoxy nucleoside: all purine nucleosides comprise 2’ fluoro, and all pyrimidine nucleosides are modified with a mixture of 2’ fluoro and 2’-O- methyl, all purine nucleosides comprise 2’-O-methyl, and all pyrimidine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl, all purine nucleosides comprise 2’-O-methyl, and all pyrimidine nucleosides comprise 2’ fluoro, all pyrimidine nucleosides comprise 2’ fluoro, and all purine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl, all pyrimidine nucleosides comprise 2’-O- methyl, and all purine nucleosides are modified with a mixture of 2’ flu
  • all purine nucleosides comprise 2’ fluoro, and all pyrimidine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl, with the proviso that the antisense -91- Attorney Docket No.54462-742.601 strand may include a 2’ deoxy nucleoside.
  • all purine nucleosides comprise 2’-O-methyl, and all pyrimidine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl, with the proviso that the antisense strand may include a 2’ deoxy nucleoside.
  • all purine nucleosides comprise 2’ fluoro, and all pyrimidine nucleosides comprise 2’-O-methyl, with the proviso that the antisense strand may include a 2’ deoxy nucleoside.
  • all pyrimidine nucleosides comprise 2’ fluoro, and all purine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl, with the proviso that the antisense strand may include a 2’ deoxy nucleoside.
  • all pyrimidine nucleosides comprise 2’-O-methyl, and all purine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl, with the proviso that the antisense strand may include a 2’ deoxy nucleoside.
  • all pyrimidine nucleosides comprise 2’ fluoro, and all purine nucleosides comprise 2’-O-methyl, with the proviso that the antisense strand may include a 2’ deoxy nucleoside.
  • the antisense strand includes the 2’ deoxy nucleoside.
  • the antisense strand does not include the 2’ deoxy nucleoside.
  • Some embodiments include a proviso that the antisense strand may include a 2’-O-methoxyethyl nucleoside (e.g. at position 4, counting from 5’ to 3’).
  • Some embodiments include the 2’-O-methoxyethyl nucleoside in the antisense strand.
  • Some embodiments do not include the 2’-O-methoxyethyl nucleoside in the antisense strand.
  • any one of the following is true with regard to the antisense strand: all purine nucleosides comprise 2’ fluoro, and all pyrimidine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl, all purine nucleosides comprise 2’-O-methyl, and all pyrimidine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl, all purine nucleosides comprise 2’-O-methyl, and all pyrimidine nucleosides comprise 2’ fluoro, all pyrimidine nucleosides comprise 2’ fluoro, and all purine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl, all pyrimidine nucleosides comprise 2’-O-methyl, and all purine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl, or all pyrimidine nucle
  • all purine nucleosides comprise 2’ fluoro, and all pyrimidine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl. In some embodiments, in the antisense strand, all purine nucleosides comprise 2’-O-methyl, and all pyrimidine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl. In some embodiments, in the antisense strand, all purine nucleosides comprise 2’ fluoro, and all pyrimidine nucleosides comprise 2’-O-methyl.
  • all pyrimidine nucleosides comprise 2’ fluoro, and all purine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl. In some embodiments, in the antisense strand, all pyrimidine nucleosides comprise 2’-O-methyl, and all purine nucleosides are modified with a mixture of 2’ fluoro and 2’-O-methyl. In some embodiments, in the antisense strand, all pyrimidine nucleosides comprise 2’ fluoro, and all purine nucleosides comprise 2’-O-methyl.
  • the antisense strand may include a 2’-O-methoxyethyl nucleoside (e.g. at position 4, counting from 5’ to 3’). Some embodiments include the 2’-O-methoxyethyl nucleoside in the antisense strand. Some embodiments do not include the 2’-O-methoxyethyl nucleoside in the antisense strand. -92- Attorney Docket No.54462-742.601 [00159] In some embodiments, the antisense strand comprises one or two 3’ phosphorothioate linkages.
  • the sense strand comprises one or two 5’ phosphorothioate linkages.
  • the sense strand does not comprise one or two 5’ phosphorothioate linkages. For example, in some embodiments, there are no phosphorothioate linkages between the last 3 nucleotides at the 5’ end of the sense strand. In some embodiments, the sense strand comprises 5’ phosphate linkages. In some embodiments, the sense strand comprises one or two 3’ phosphorothioate linkages. For example, there may be a phosphorothioate linkage between the first and second nucleotides from the 3’ end of the sense strand, or there may be phosphorothioate linkages between the first, second and third nucleotides from the 3’ end of the sense strand.
  • the antisense strand comprises a 5’ end comprising 2 phosphorothioate linkages.
  • the 5’ end may comprise 3 nucleosides separated by the 2 phosphorothioate linkages.
  • the antisense strand comprises a 3’ end comprising 2 phosphorothioate linkages.
  • the 3’ end may comprise 3 nucleosides separated by the 2 phosphorothioate linkages.
  • position 9 of the sense strand can be a 2’deoxy. In these cases, 2’F and 2’OMe modifications may occur at the other positions of the sense strand.
  • the sense strand of any of the siRNAs comprises a modification pattern which conforms to these sense strand rules. [00162] In some cases, the sense strand of any of the siRNAs comprises a modification pattern which conforms to these sense strand rules.
  • compositions comprising an oligonucleotide that targets GPAM and when administered to a cell decreases expression of GPAM, wherein the oligonucleotide comprises a small interfering RNA (siRNA) comprising a sense strand and an antisense strand, wherein the sense strand comprises a sense strand sequence described herein in which at least one internucleoside linkage is modified and at least one nucleoside is modified, or an sense strand sequence comprising 1 or 2 nucleoside substitutions, additions, or deletions of the oligonucleotide sequence in which at least one internucleoside linkage is modified and at least one nucleoside is modified, and wherein the antisense strand comprises an antisense strand sequence described herein in which at least one internucleoside linkage is modified and at least one nucleoside is modified, or an oligonucleotide sequence comprising 1 or 2 nucleoside substitution
  • the siRNA comprises a sense strand having a sequence in accordance with any of SEQ ID NOs: 12709-12733.
  • the sense strand sequence comprises or consists of sequence at least 75% identical to any one of SEQ ID NOs: 12709-12733, at least 80% identical to any one of SEQ ID NOs: 12709-12733, at least 85% identical to of any one of SEQ ID NOs: -93- Attorney Docket No.54462-742.601 12709-12733, at least 90% identical to any one of SEQ ID NOs: 12709-12733, or at least 95% identical to any one of SEQ ID NOs: 12709-12733.
  • the sense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 12709-12733, or a sense strand sequence thereof having 1, 2, 3, or 4 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 12709-12733, or a sense strand sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand sequence comprises or consists of a sequence 100% identical to SEQ ID NOs: 12709-12733. In some embodiments, the siRNA comprises a sense strand having a sequence in accordance with any of SEQ ID NOs: 12759-12866.
  • the sense strand sequence comprises or consists of sequence at least 75% identical to any one of SEQ ID NOs: 12759-12866, at least 80% identical to any one of SEQ ID NOs: 12759-12866, at least 85% identical to of any one of SEQ ID NOs: 12759-12866, at least 90% identical to any one of SEQ ID NOs: 12759-12866, or at least 95% identical to any one of SEQ ID NOs: 12759-12866.
  • the sense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 12759-12866, or a sense strand sequence thereof having 1, 2, 3, or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 12759-12866, or a sense strand sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand sequence comprises or consists of a sequence 100% identical to SEQ ID NOs: 12759-12866.
  • the sense strand sequence may include the first 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the sense strand sequence may include the first 19 nucleotides of any of the aforementioned sequences.
  • the sense strand sequence may include the last 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the sense strand may comprise an overhang.
  • the sense strand may comprise a modification pattern described herein, such as a different set of modifications or modification pattern than the aforementioned sequences.
  • the sense strand may comprise a lipid moiety.
  • the sense strand may comprise a GalNAc moiety.
  • the siRNA comprises an antisense strand having a sequence in accordance with any of SEQ ID NOs: 12734-12758.
  • the antisense strand sequence comprises or consists of sequence at least 75% identical to any one of SEQ ID NOs: 12734-12758, at least 80% identical to any one of SEQ ID NOs: 12734-12758, at least 85% identical to of any one of SEQ ID NOs: 12734-12758, at least 90% identical to any one of SEQ ID NOs: 12734-12758, or at least 95% identical to any one of SEQ ID NOs: 12734-12758.
  • the antisense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 12734-12758, or an antisense strand sequence thereof having 1, 2, 3, or 4 nucleoside substitutions, additions, or deletions. In some embodiments, the antisense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 12734-12758, or an antisense strand sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions. In some embodiments, the antisense strand sequence comprises or consists of a sequence 100% identical to SEQ ID NOs: 12734- 12758.
  • the antisense strand sequence may include the first 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the antisense strand -94- Attorney Docket No.54462-742.601 sequence may include the first 19 nucleotides of any of the aforementioned sequences.
  • the antisense strand sequence may include the last 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the antisense strand may comprise an overhang.
  • the antisense strand may comprise a modification pattern described herein, such as a different set of modifications or modification pattern than the aforementioned sequences.
  • the antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand having a sequence in accordance with any of SEQ ID NOs: 12868-12974.
  • the sense strand sequence comprises or consists of sequence at least 75% identical to any one of SEQ ID NOs: 12868-12974, at least 80% identical to any one of SEQ ID NOs: 12868-12974, at least 85% identical to of any one of SEQ ID NOs: 12868-12974, at least 90% identical to any one of SEQ ID NOs: 12868-12974, or at least 95% identical to any one of SEQ ID NOs: 12868-12974.
  • the sense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 12868-12974, or a sense strand sequence thereof having 1, 2, 3, or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 12868-12974, or a sense strand sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand sequence comprises or consists of a sequence 100% identical to SEQ ID NOs: 12868-12974.
  • the sense strand sequence may include the first 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the sense strand sequence may include the first 19 nucleotides of any of the aforementioned sequences.
  • the sense strand sequence may include the last 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the sense strand may comprise an overhang.
  • the sense strand may comprise a modification pattern described herein, such as a different set of modifications or modification pattern than the aforementioned sequences.
  • the sense strand may comprise a lipid moiety.
  • the sense strand may comprise a GalNAc moiety.
  • the siRNA comprises an antisense strand having a sequence in accordance with any of SEQ ID NOs: 12975- 13081.
  • the antisense strand sequence comprises or consists of sequence at least 75% identical to any one of SEQ ID NOs: 12975-13081, at least 80% identical to any one of SEQ ID NOs: 12975-13081, at least 85% identical to of any one of SEQ ID NOs: 12975-13081, at least 90% identical to any one of SEQ ID NOs: 12975-13081, or at least 95% identical to any one of SEQ ID NOs: 12975-13081.
  • the antisense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 12975-13081, or an antisense strand sequence thereof having 1, 2, 3, or 4 nucleoside substitutions, additions, or deletions. In some embodiments, the antisense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 12975-13081, or an antisense strand sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions. In some embodiments, the antisense strand sequence comprises or consists of a sequence 100% identical to SEQ ID NOs: 12975- 13081.
  • the antisense strand sequence may include the first 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the antisense strand sequence may include the first 19 nucleotides of any of the aforementioned sequences.
  • the antisense -95- Attorney Docket No.54462-742.601 strand sequence may include the last 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the antisense strand may comprise an overhang.
  • the antisense strand may comprise a modification pattern described herein, such as a different set of modifications or modification pattern than the aforementioned sequences.
  • the antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand having a sequence in accordance with any of SEQ ID NOs: 13724-13751.
  • the sense strand sequence comprises or consists of sequence at least 75% identical to any one of SEQ ID NOs: 13724-13751, at least 80% identical to any one of SEQ ID NOs: 13724-13751, at least 85% identical to of any one of SEQ ID NOs: 13724-13751, at least 90% identical to any one of SEQ ID NOs: 13724-13751, or at least 95% identical to any one of SEQ ID NOs: 13724-13751.
  • the sense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 13724-13751, or a sense strand sequence thereof having 1, 2, 3, or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 13724-13751, or a sense strand sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions. In some embodiments, the sense strand sequence comprises or consists of a sequence 100% identical to SEQ ID NOs: 13724-13751.
  • the sense strand sequence may include the first 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the sense strand sequence may include the first 19 nucleotides of any of the aforementioned sequences.
  • the sense strand sequence may include the last 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the sense strand may comprise an overhang.
  • the sense strand may comprise a modification pattern described herein, such as a different set of modifications or modification pattern than the aforementioned sequences.
  • the sense strand may comprise a lipid moiety.
  • the sense strand may comprise a GalNAc moiety.
  • the siRNA comprises an antisense strand having a sequence in accordance with any of SEQ ID NOs: 13752- 13779.
  • the antisense strand sequence comprises or consists of sequence at least 75% identical to any one of SEQ ID NOs: 13752-13779, at least 80% identical to any one of SEQ ID NOs: 13752-13779, at least 85% identical to of any one of SEQ ID NOs: 13752-13779, at least 90% identical to any one of SEQ ID NOs: 13752-13779, or at least 95% identical to any one of SEQ ID NOs: 13752-13779.
  • the antisense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 13752-13779, or an antisense strand sequence thereof having 1, 2, 3, or 4 nucleoside substitutions, additions, or deletions.
  • the antisense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 13752-13779, or an antisense strand sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the antisense strand sequence comprises or consists of a sequence 100% identical to SEQ ID NOs: 13752- 13779.
  • the antisense strand sequence may include the first 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the antisense strand sequence may include the first 19 nucleotides of any of the aforementioned sequences.
  • the antisense strand sequence may include the last 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in -96- Attorney Docket No.54462-742.601 the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the antisense strand may comprise an overhang.
  • the antisense strand may comprise a modification pattern described herein, such as a different set of modifications or modification pattern than the aforementioned sequences.
  • the antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of subset G.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of subset G.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of subset G, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of subset G, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of subset G.
  • the sense strand or antisense strand sequence may include the first 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the sense strand or antisense strand sequence may include the first 19 nucleotides of any of the aforementioned sequences.
  • the sense strand or antisense strand sequence may include the last 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the sense strand or antisense strand may comprise an overhang.
  • the sense strand or antisense strand may comprise any modifications described herein (e.g. a different set of modifications or modification pattern than subset G).
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of subset H.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of subset H.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of subset H, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of subset H, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of subset H.
  • the sense strand or antisense strand sequence may include the first 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the sense strand or antisense strand sequence may include the first 19 nucleotides of any of the aforementioned sequences.
  • the sense strand or antisense strand sequence may include the last 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the sense strand or antisense strand may comprise an overhang.
  • the sense -97- Attorney Docket No.54462-742.601 strand or antisense strand may comprise any modifications described herein (e.g. a different set of modifications or modification pattern than subset H).
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of Table 10.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of Table 10.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 10, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 10, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence [00170]
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of Table 11.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of Table 11.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 11, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 11, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of Table 11.
  • the sense strand or antisense strand may comprise an overhang.
  • the sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of Table 15.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of Table 15.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 15, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 15, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence. 100% identical to a sense strand or antisense strand sequence of Table 15.
  • the sense strand or antisense strand may comprise an overhang.
  • the sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of Table 19.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of Table 19.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 19, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 19, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of Table 19.
  • the sense strand or antisense strand may comprise an overhang.
  • the sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of Table 21.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of Table 21.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 21, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 21, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of Table 21.
  • the sense strand or antisense strand may comprise an overhang.
  • the sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of Table 24.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of Table 24.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 24, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 24, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of Table 24.
  • the sense strand or antisense strand may comprise an overhang.
  • the sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of Table 27.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of Table 27.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 27, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 27, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of Table 27.
  • the sense strand or antisense strand may comprise an overhang.
  • the sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of Table 30.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of Table 30.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 30, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 30, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of Table 30.
  • the sense strand or antisense strand may comprise an overhang.
  • the sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of Table 34.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of Table 34.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 34, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 34, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of Table 34.
  • the sense strand or antisense strand may comprise an overhang.
  • the sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of Table 48.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of Table 48.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 48, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 48, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of Table 48.
  • the sense strand or antisense strand may comprise an overhang.
  • the sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand or antisense strand having a sequence in accordance with the sense strand or antisense strand sequence of an siRNA of Table 62.
  • the sense strand or antisense strand comprises a sequence at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to a sense strand or antisense strand sequence of Table 62.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 62, or a sequence thereof having 3 or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense strand comprises a sequence of a sense or antisense strand of Table 62, or a sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand or antisense sequence comprises or consists of a sequence 100% identical to a sense strand or antisense strand sequence of Table 62.
  • the sense strand or antisense strand may comprise an overhang.
  • the sense strand or antisense strand may comprise any modifications described herein.
  • the sense strand or antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand having a sequence in accordance with any of SEQ ID NOs: 12953, 12958, 13724, 13725, 13726, 13727, 13728, 13729, 13730, 13731, 13732, 13733, 13734, 13735, 12959, 13736, 13737, 13738, 13739, 13740, 13741, 13742, 13743, 13744, 13745, 13746, 13747, 12877, 12927, 12928, 12929, 12930, 12931, 12932, 12933, 12934, 12960, 13748, 13749, 13750, 13751, 12883, 12910, 12911, 12912, 12913, 12914, 12915, 12916, 12917, 12947, 12948, 12965, 12885, 12901, 12902, 12903
  • the sense strand sequence comprises or consists of sequence at least 75% identical to any one of SEQ ID NOs: 12953, 12958, 13724, 13725, 13726, 13727, 13728, 13729, 13730, 13731, 13732, 13733, 13734, 13735, 12959, 13736, 13737, 13738, 13739, 13740, 13741, 13742, 13743, 13744, 13745, 13746, 13747, 12877, 12927, 12928, 12929, 12930, 12931, 12932, 12933, 12934, 12960, 13748, 13749, 13750, 13751, 12883, 12910, 12911, 12912, 12913, 12914, 12915, -101- Attorney Docket No.54462-742.601 12916, 12917, 12947, 12948, 12965, 12
  • the sense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 12953, 12958, 13724, 13725, 13726, 13727, 13728, 13729, 13730, 13731, 13732, 13733, 13734, 13735, 12959, 13736, 13737, 13738, 13739, 13740, 13741, 13742, 13743, 13744, 13745, 13746, 13747, 12877, 12927, 12928, 12929, 12930, 12931, 12932, 12933, 12934, 12960, 13748, 13749, 13750, 13751, 12883, 12910, 12911, 12912, 12913, 12914, 12915, 12916, 12917, 12947, 12948, 12965, 12885, 12901, 12902, 12903, 12904, 129
  • the sense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 12953, 12958, 13724, 13725, 13726, 13727, 13728, 13729, 13730, 13731, 13732, 13733, 13734, 13735, 12959, 13736, 13737, 13738, 13739, 13740, 13741, 13742, 13743, 13744, 13745, -102- Attorney Docket No.54462-742.601 13746, 13747, 12877, 12927, 12928, 12929, 12930, 12931, 12932, 12933, 12934, 12960, 13748, 13749, 13750, 13751, 12883, 12910, 12911, 12912, 12913, 12914, 12915, 12916, 12917, 12947, 12948, 12965, 12885, 12901
  • the sense strand sequence comprises or consists of a sequence 100% identical to SEQ ID NOs: 12953, 12958, 13724, 13725, 13726, 13727, 13728, 13729, 13730, 13731, 13732, 13733, 13734, 13735, 12959, 13736, 13737, 13738, 13739, 13740, 13741, 13742, 13743, 13744, 13745, 13746, 13747, 12877, 12927, 12928, 12929, 12930, 12931, 12932, 12933, 12934, 12960, 13748, 13749, 13750, 13751, 12883, 12910, 12911, 12912, 12913, 12914, 12915, 12916, 12917, 12947, 12948, 12965, 12885, 12901, 12902, 12903, 12904, 129
  • the sense strand sequence may include the first 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the sense strand sequence may include the last 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the sense strand may comprise an overhang.
  • the sense strand may comprise a modification pattern described herein.
  • the sense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises an antisense strand having a sequence in accordance with any of SEQ ID NOs: 13060, 13065, 13752, 13753, 13754, 13755, 13756, 13757, 13758, 13759, 13760, 13761, 13762, 13763, 13066, 13764, 12984, 13034, 13035, 13036, 13037, 13038, 13039, 13040, 13041, 13067, 13765, 13766, 13767, 13768, 12990, 13017, 13018, 13019, 13020, 13021, 13022, 13023, 13024, 13054, 13055, 13072, 12992, 13008, 13009, 13010, 13011, 13012, 13013, 13014, 13015, 13016, 12993, 13000, 13001, 13002, 13003, 13004, 13005, 13006, 13007, 13075, 12996, 13025, 13026, 13027, 13028
  • the antisense strand sequence comprises or consists of sequence at least 75% identical to any one of SEQ ID NOs: 13060, 13065, 13752, 13753, 13754, 13755, 13756, 13757, 13758, 13759, 13760, 13761, 13762, 13763, 13066, 13764, 12984, 13034, 13035, 13036, 13037, 13038, 13039, 13040, 13041, 13067, 13765, 13766, 13767, 13768, 12990, 13017, 13018, 13019, 13020, 13021, 13022, 13023, 13024, 13054, 13055, 13072, 12992, 13008, 13009, 13010, 13011, 13012, 13013, 13014, 13015, 13016, 12993, 13000, 13001, 13002, 13003, 13004, 13005, 13006, 13007, 13075, 12996, 13025, 13026, 13027, 13028, 130
  • the antisense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 13060, 13065, 13752, 13753, 13754, 13755, 13756, 13757, 13758, 13759, 13760, 13761, 13762, 13763, 13066, 13764, 12984, 13034, 13035, 13036, 13037, 13038, 13039, 13040, 13041, 13067, 13765, 13766, 13767, 13768, 12990, 13017, 13018, 13019, 13020, 13021, 13022, 13023, 13024, 13054, 13055, 13072, 12992, 13008, 13009, 13010, 13011, 13012, 13013, 13014, 13015, 13016, 12993, 13000, 13001, 13002, 13003, 13004, 13005, 13006, 13007, 13075, 12996, 13025, 13026, 13027, 13028, 13029, 13030
  • the antisense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 13060, 13065, 13752, 13753, 13754, 13755, 13756, 13757, 13758, 13759, 13760, 13761, 13762, 13763, 13066, 13764, 12984, 13034, 13035, 13036, 13037, 13038, 13039, 13040, 13041, 13067, 13765, 13766, 13767, 13768, 12990, 13017, 13018, 13019, 13020, 13021, 13022, 13023, 13024, 13054, 13055, 13072, 12992, 13008, 13009, 13010, 13011, 13012, 13013, 13014, 13015, 13016, 12993, 13000, 13001, 13002, 13003, 13004, 13005, 13006, 13007, 13075, 12996, 13025, 13026, 13027, 13028, 13029, 13030
  • the antisense strand sequence comprises or consists of a sequence 100% identical to SEQ ID NOs: 13060, 13065, 13752, 13753, 13754, 13755, 13756, 13757, 13758, 13759, 13760, 13761, 13762, 13763, 13066, 13764, 12984, 13034, 13035, 13036, 13037, 13038, 13039, 13040, 13041, 13067, 13765, 13766, 13767, 13768, 12990, 13017, 13018, 13019, 13020, 13021, 13022, 13023, 13024, 13054, 13055, 13072, 12992, 13008, 13009, 13010, 13011, 13012, 13013, 13014, 13015, 13016, 12993, 13000, 13001, 13002, 13003, 13004, 13005, 13006, 13007, 13075, 12996, 13025, 13026, 13027, 13028, 13029, 13030
  • the antisense strand sequence may include the first 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the antisense strand sequence may include the last 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the antisense strand may comprise an overhang.
  • the antisense strand may comprise a modification pattern described herein.
  • the antisense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises a sense strand having a sequence in accordance with any of SEQ ID NOs: 12959, 12928, 12932, 12960, 12947, 12965, 12904, 12909, 12897, 12968, 12926, 12969, 12970, 12935, or 12936.
  • the sense strand sequence comprises or consists of sequence at least 75% identical to any one of SEQ ID NOs: 12959, 12928, 12932, 12960, 12947, 12965, 12904, 12909, 12897, 12968, 12926, 12969, 12970, 12935, or 12936., at least 80% identical to any one of SEQ ID NOs: 12959, 12928, 12932, 12960, 12947, 12965, 12904, 12909, 12897, 12968, 12926, 12969, 12970, 12935, or 12936., at least 85% identical to of any one of SEQ ID NOs: 12959, 12928, 12932, 12960, 12947, 12965, 12904, 12909, 12897, 12968, 12926, 12969, 12970, 12935, or 12936., at least 90% identical to any one of SEQ ID NOs:
  • the sense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 12959, 12928, 12932, 12960, 12947, 12965, 12904, 12909, 12897, 12968, 12926, 12969, 12970, 12935, or 12936., or a sense strand sequence thereof having 1, 2, 3, or 4 nucleoside substitutions, additions, or deletions.
  • the sense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 12959, 12928, 12932, 12960, 12947, 12965, 12904, 12909, 12897, 12968, 12926, 12969, 12970, 12935, or 12936., or a sense strand sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the sense strand sequence comprises or consists of a sequence 100% identical to SEQ ID NOs: 12959, 12928, 12932, 12960, 12947, 12965, 12904, 12909, 12897, 12968, 12926, 12969, 12970, 12935, or 12936.
  • the sense strand sequence may include the first 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the sense strand sequence may include the last 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the sense strand may comprise an overhang.
  • the sense strand may comprise a modification pattern described herein.
  • the sense strand may comprise a lipid moiety or a GalNAc moiety.
  • the siRNA comprises an antisense strand having a sequence in accordance with any of SEQ ID NOs: 13060, 13065, 13066, 13035, 13039, 13067, 13054, 13072, 13011, 13016, 13004, 13075, 13033, 13076, 13077, 13042, or 13043.
  • the antisense strand sequence comprises or consists of sequence at least 75% identical to any one of SEQ ID NOs: 13060, 13065, 13066, 13035, 13039, 13067, 13054, 13072, 13011, 13016, 13004, 13075, 13033, 13076, 13077, 13042, or 13043, at least 80% identical to any one of SEQ ID NOs: 13060, 13065, 13066, 13035, 13039, 13067, 13054, 13072, 13011, 13016, 13004, 13075, 13033, 13076, 13077, 13042, or 13043, at least 85% identical to of any one of SEQ ID NOs: 13060, 13065, 13066, 13035, 13039, 13067, 13054, 13072, 13011, 13016, 13004, 13075, 13033, 13076, 13077, 13042, or 13043, at least 90% identical to any one of SEQ ID NOs: 13060, 13065, 13066, 13035, 13039, 13067, 13054, 13072, 130
  • the antisense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 13060, 13065, 13066, 13035, 13039, 13067, 13054, 13072, 13011, 13016, 13004, 13075, 13033, 13076, 13077, 13042, or 13043, or an antisense strand sequence thereof having 1, 2, 3, or 4 nucleoside substitutions, additions, or deletions.
  • the antisense strand sequence comprises or consists of the sequence of any one of SEQ ID NOs: 13060, 13065, 13066, 13035, 13039, 13067, 13054, 13072, 13011, 13016, 13004, 13075, 13033, 13076, 13077, 13042, or 13043, or an antisense strand sequence thereof having 1 or 2 nucleoside substitutions, additions, or deletions.
  • the antisense strand sequence comprises or consists of a sequence 100% identical to SEQ ID NOs: 13060, 13065, 13066, 13035, 13039, 13067, 13054, 13072, 13011, 13016, 13004, 13075, 13033, 13076, 13077, 13042, or 13043.
  • the antisense strand sequence may include the first 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the antisense strand sequence may include the last 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides (in the 5’ to 3’ direction) of any of the aforementioned sequences.
  • the antisense strand may comprise an overhang.
  • the antisense strand may comprise a modification pattern described herein.
  • the antisense strand may comprise a lipid moiety or a GalNAc moiety. 4.
  • the composition comprises an oligonucleotide that inhibits the expression of GPAM, wherein the oligonucleotide comprises an antisense oligonucleotide (ASO).
  • ASO antisense oligonucleotide
  • the ASO comprises modification pattern ASO1: 5’-nsnsnsnsnsnsdNsdNsdNsdNsdNsdNsdNsdNsdNsnsnsnsn-3’ (SEQ ID NO: 12928), wherein “dN” is any deoxynucleotide, “n” is a 2’O-methyl or 2’-O-(2-methoxyethyl)-modified nucleoside, and “s” is a phosphorothioate linkage.
  • the ASO comprises modification pattern 1S, 2S, 3S, 4S, 5S, 6S, 7S, 8S, 9S, 10S, 11S, 12S, 13S, 14S, 15S, 16S, 17S, 18S, 19S, 20S, 21S, 22S, 23S, 24S, 25S, 26S, 27S, 28S, 29S, 30S, 31S, 32S, 33S, 34S, 35S, 36S, 37S, 38S, 39S, 40S, 41S, 42S, 43S, 44S, 45S, 46S, 47S, 48S, 49S, 50S, 51S, 52S, 52S, 54S, 55S, 56S, 57S, 58S, 59S, 60S, 61S, 62S, 63S, 64S, 65S, 66S, 67S, 68S, 69S, 70S, 1AS, 2AS, 3AS, 4AS, 5AS, 6AS, 7AS, or 8AS, 9S,
  • the composition is a pharmaceutical composition. In some embodiments, the composition is sterile. In some embodiments, the composition further comprises a pharmaceutically acceptable carrier. [00186] In some embodiments, the pharmaceutically acceptable carrier comprises water. In some embodiments, the pharmaceutically acceptable carrier comprises a buffer. In some embodiments, the pharmaceutically acceptable carrier comprises a saline solution. In some embodiments, the pharmaceutically acceptable carrier comprises water, a buffer, or a saline solution. In some embodiments, the composition comprises a liposome. In some embodiments, the pharmaceutically acceptable carrier comprises liposomes, lipids, nanoparticles, proteins, protein-antibody complexes, peptides, cellulose, nanogel, or a combination thereof.
  • METHODS AND USES Disclosed herein, in some embodiments, are methods of administering a composition described herein to a subject. Some embodiments relate to use a composition described herein, such as administering the composition to a subject. [00188] Some embodiments relate to a method of treating a disorder in a subject in need thereof. Some embodiments relate to use of a composition described herein in the method of treatment. Some embodiments include administering a composition described herein to a subject with the disorder. In some embodiments, the administration treats the disorder in the subject. In some embodiments, the composition treats the disorder in the subject. The disorder may comprise a disease. [00189] In some embodiments, the treatment comprises prevention, inhibition, or reversion of the disorder in the subject.
  • Some embodiments relate to use of a composition described herein in the method of preventing, inhibiting, or reversing the disorder. Some embodiments relate to a method of preventing, inhibiting, or reversing a disorder a disorder in a subject in need thereof. Some embodiments include administering a composition described herein to a subject with the disorder. In some embodiments, the administration prevents, inhibits, or reverses the disorder in the subject. In some embodiments, the composition prevents, inhibits, or reverses the disorder in the subject. [00190] Some embodiments relate to a method of preventing a disorder a disorder in a subject in need thereof. Some embodiments relate to use of a composition described herein in the method of preventing the disorder.
  • Some embodiments include administering a composition described herein to a subject with the disorder. In some embodiments, the administration prevents the disorder in the subject. In some embodiments, the composition prevents the disorder in the subject. [00191] Some embodiments relate to a method of inhibiting a disorder a disorder in a subject in need thereof. Some embodiments relate to use of a composition described herein in the method of inhibiting the disorder. Some embodiments include administering a composition described herein to a subject with the -107- Attorney Docket No.54462-742.601 disorder. In some embodiments, the administration inhibits the disorder in the subject. In some embodiments, the composition inhibits the disorder in the subject.
  • Some embodiments relate to a method of reversing a disorder a disorder in a subject in need thereof. Some embodiments relate to use of a composition described herein in the method of reversing the disorder. Some embodiments include administering a composition described herein to a subject with the disorder. In some embodiments, the administration reverses the disorder in the subject. In some embodiments, the composition reverses the disorder in the subject. [00193] In some embodiments, the administration is systemic. In some embodiments, the administration is intravenous. In some embodiments, the administration is by injection. A. Disorders [00194] Some embodiments of the methods described herein include treating a disorder in a subject in need thereof. In some embodiments, the disorder is a liver disease.
  • Non-limiting examples of liver diseases may include non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH), alcoholic liver disease, liver fibrosis, liver cirrhosis, or hepatocellular carcinoma.
  • the disorder is a cardiometabolic disease.
  • cardiometabolic diseases may include hyperlipidemia, ischemic heart disease, or coronary heart disease.
  • the disorder comprises NAFLD.
  • the disorder comprises NASH.
  • the disorder comprises alcoholic liver disease.
  • the disorder comprises liver fibrosis.
  • the disorder comprises liver cirrhosis.
  • the disorder comprises hepatocellular carcinoma.
  • the disorder comprises hyperlipidemia. In some embodiments, the disorder comprises a heart disease. In some embodiments, the heart disease comprises ischemic heart disease. In some embodiments, the heart disease comprises coronary heart disease.
  • subjects include vertebrates, animals, mammals, dogs, cats, cattle, rodents, mice, rats, primates, monkeys, and humans. In some embodiments, the subject is a vertebrate. In some embodiments, the subject is an animal. In some embodiments, the subject is a mammal. In some embodiments, the subject is a dog. In some embodiments, the subject is a cat. In some embodiments, the subject is a cattle.
  • the subject is a mouse. In some embodiments, the subject is a rat. In some embodiments, the subject is a primate. In some embodiments, the subject is a monkey. In some embodiments, the subject is an animal, a mammal, a dog, a cat, cattle, a rodent, a mouse, a rat, a primate, or a monkey. In some embodiments, the subject is a human. [00196] In some embodiments, the subject is male. In some embodiments, the subject is female. In some embodiments, the subject is an adult (e.g. at least 18 years old).
  • the subject has a body mass index (BMI) of 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, or more, or a range defined by any two of the aforementioned integers.
  • BMI body mass index
  • the subject is overweight.
  • the subject has a BMI of 25 or more.
  • the subject has a BMI of 25- 29.
  • the subject is obese.
  • the subject has a BMI of 30 or -108- Attorney Docket No.54462-742.601 more.
  • the subject has a BMI of 30-39.
  • the subject has a BMI of 40-50. In some embodiments, the subject has a BMI of 25-50.
  • C. Baseline measurements [00198] Some embodiments of the methods described herein include obtaining a baseline measurement from a subject. For example, in some embodiments, a baseline measurement is obtained from the subject prior to treating the subject.
  • baseline measurements include a baseline liver fat percentage measurement, a baseline liver fibrosis score measurement, a baseline NAFLD activity score measurement, a baseline blood alanine aminotransferase (ALT) measurement, a baseline blood aspartate aminotransferase (AST) measurement, a baseline blood aspartate aminotransferase (ASP) measurement, a baseline blood bilirubin measurement, a baseline low-density lipoprotein (LDL) measurement, a baseline total cholesterol measurement, a baseline non-HDL cholesterol measurement, a baseline apolipoprotein B (APOB), a baseline GPAM protein measurement, or a baseline GPAM mRNA measurement.
  • the baseline measurement is obtained directly from the subject.
  • the baseline measurement is obtained by observation, for example by observation of the subject or of the subject’s tissue. In some embodiments, the baseline measurement is obtained noninvasively using an imaging device. [00200] In some embodiments, the baseline measurement is obtained in a sample from the subject. In some embodiments, the baseline measurement is obtained in one or more histological tissue sections. In some embodiments, the baseline measurement is obtained by performing an assay such as an immunoassay, a colorimetric assay, or a fluorescence assay, on the sample obtained from the subject. In some embodiments, the baseline measurement is obtained by an immunoassay, a colorimetric assay, a fluorescence assay, or a chromatography (e.g. HPLC) assay.
  • an immunoassay such as an immunoassay, a colorimetric assay, a fluorescence assay, or a chromatography (e.g. HPLC) assay.
  • the baseline measurement is obtained by PCR.
  • the baseline measurement is a baseline liver fat percentage measurement.
  • the baseline liver fat percentage measurement is a baseline percentage as measured by MRI.
  • the baseline liver fat percentage measurement is a baseline percentage as measured by CT scan.
  • the baseline liver fat percentage is measured in a liver biopsy.
  • the baseline measurement is a baseline NAFLD activity score measurement.
  • the baseline NAFLD activity score measurement is measured by histological analysis.
  • the baseline NAFLD activity score measurement includes a numerical score for steatosis (0–3), hepatocyte ballooning (1–2), and lobular inflammation (0–3).
  • the score thresholds of ⁇ 3 correlates with a diagnosis of not-NASH. In some embodiments, the score thresholds of > 5 correlates with a diagnosis of NASH. In some embodiments, a baseline NAFLD activity score measurement is assayed by imaging. [00203] In some embodiments, the baseline measurement is a baseline alanine aminotransferase (ALT) measurement. In some embodiments, the baseline ALT measurement includes a baseline blood ALT measurement. In some embodiments, the baseline ALT measurement is measured by enzymatic -109- Attorney Docket No.54462-742.601 activity. In some embodiments, the baseline ALT measurement is measured by presence of an ALT epitope.
  • ALT a baseline alanine aminotransferase
  • the baseline ALT measurement is a baseline circulating ALT measurement.
  • the baseline ALT measurement is obtained by an assay such as an immunoassay, a colorimetric assay, or a fluorescence assay.
  • the baseline measurement is a baseline aspartate aminotransferase (AST) measurement.
  • the baseline AST measurement includes a baseline blood AST measurement.
  • the baseline AST measurement is measured by enzymatic activity.
  • the baseline AST measurement is measured by presence of an AST epitope.
  • the baseline AST measurement is a baseline circulating AST measurement.
  • the baseline AST measurement is obtained by an assay such as an immunoassay, a colorimetric assay, or a fluorescence assay.
  • the baseline measurement is a baseline aspartate aminotransferase (ASP) measurement.
  • the baseline ASP measurement includes a baseline blood ASP measurement.
  • the baseline ASP measurement is measured by enzymatic activity.
  • the baseline ASP measurement is measured by presence of an ASP epitope.
  • the baseline ASP measurement is a baseline circulating ASP measurement.
  • the baseline ASP measurement is obtained by an assay such as an immunoassay, a colorimetric assay, or a fluorescence assay.
  • the baseline measurement is a baseline bilirubin measurement.
  • the baseline bilirubin measurement includes a baseline blood bilirubin measurement.
  • the baseline bilirubin measurement is measured by presence of a bilirubin epitope.
  • the baseline bilirubin measurement is a baseline circulating bilirubin measurement.
  • the baseline bilirubin measurement is obtained by an assay such as an immunoassay, a colorimetric assay, or a fluorescence assay.
  • the baseline measurement is a baseline low-density lipoprotein (LDL) measurement.
  • LDL low-density lipoprotein
  • the baseline LDL measurement includes a baseline blood LDL measurement. In some embodiments, the baseline LDL measurement is measured by presence of an LDL epitope. In some embodiments, the baseline LDL measurement is a baseline circulating LDL measurement. In some embodiments, the baseline LDL measurement is obtained by an assay such as an immunoassay, a colorimetric assay, or a fluorescence assay. [00208] In some embodiments, the baseline measurement is a baseline total cholesterol measurement. In some embodiments, the baseline total cholesterol measurement includes a baseline blood total cholesterol measurement. In some embodiments, the baseline total cholesterol is measured by presence of a total cholesterol epitope. In some embodiments, the baseline total cholesterol measurement is a baseline circulating total cholesterol measurement.
  • the baseline total cholesterol measurement is obtained by an assay such as an immunoassay, a colorimetric assay, or a fluorescence assay.
  • the baseline measurement is a baseline non-HDL cholesterol measurement.
  • the baseline non-HDL cholesterol measurement includes a baseline -110- Attorney Docket No.54462-742.601 blood non-HDL cholesterol measurement.
  • the baseline non-HDL cholesterol measurement is a baseline circulating non-HDL cholesterol measurement.
  • the baseline non-HDL cholesterol measurement is obtained by an assay such as an immunoassay, a colorimetric assay, or a fluorescence assay.
  • the baseline measurement is a baseline apolipoprotein B (APOB) measurement.
  • the baseline APOB includes a baseline blood APOB measurement.
  • the baseline APOB is measured by presence of an APOB epitope.
  • the baseline APOB measurement is a baseline circulating APOB measurement.
  • the baseline APOB measurement is obtained by an assay such as an immunoassay, a colorimetric assay, or a fluorescence assay.
  • the baseline measurement is a baseline liver fibrosis measurement.
  • the baseline liver fibrosis measurement is a baseline liver fibrosis score (LFS).
  • the baseline LFS comprises a score of 0, 1, 2, 3, or 4, or a range of scores defined by any two of the aforementioned numbers. In some embodiments, the baseline LFS comprises a score of 0- 4. In some embodiments, the baseline LFS is obtained using a scoring system exemplified in Table 2. In some embodiments, the baseline LFS measurement is obtained noninvasively. In some embodiments, the baseline LFS measurement is obtained by a medical imaging device such as a vibration-controlled transient elastography (VCTE) device, a shear wave elastography device, a medical resonance imaging (MRI) device, a magnetic resonance spectroscopy device, a computed tomography device, or an ultrasound device.
  • VCTE vibration-controlled transient elastography
  • MRI medical resonance imaging
  • spectroscopy device a computed tomography device
  • computed tomography device or an ultrasound device.
  • the baseline LFS measurement is obtained in a liver sample.
  • the baseline LFS is obtained by an assay such as an immunoassay, a colorimetric assay, or a fluorescence assay.
  • the baseline LFS is obtained using one or more indirect markers or measures of liver fibrosis such as an aspartate aminotransferase-to-platelet ratio index (APRI), a Fibrosis-4 (FIB-4) index, a FibroIndex, a Forns Index, a Hepascore, or a FibroTest.
  • APRI aspartate aminotransferase-to-platelet ratio index
  • FIB-4 Fibrosis-4
  • FibroIndex a FibroIndex
  • Forns Index a Hepascore
  • Hepascore or a FibroTest.
  • the baseline LFS is obtained using one or more indirect markers or measures of liver fibrosis such as a FIBROSpect test or a FIBROSpect II test.
  • the baseline LFS is obtained by RT-qPCR or RNA sequencing of one or more fibrosis-related genes such as a collagen gene.
  • the baseline LFS or the baseline LFS is obtained using a scoring system upon a visual inspection of a sample such as a histological sample.
  • the baseline LFS or the baseline LFS is obtained using a stain with an affinity to collagen.
  • the baseline LFS measurement is measured by FibroScan® in kilopascals (kPa).
  • the baseline LFS is measured by histological analysis.
  • the baseline LFS measurement is a baseline LFS measurement assayed by imaging.
  • the imaging is by MRI or CT scan. Sc 1 Mild fibrosis Fibrous portal expansion Periportal fibrotic expansion 2 Moderate fibrosis Rare bridges or septae Periportal septae (> 1 septum) 3 Severe fibrosis Numerous bridges or septae Portal-central septae -111- Attorney Docket No.54462-742.601 4 Cirrhosis Cirrhosis Cirrhosis Cirrhosis [00212]
  • the baseline measurement is a baseline GPAM mRNA measurement.
  • the baseline GPAM mRNA measurement comprises a baseline GPAM mRNA level. In some embodiments, the baseline GPAM mRNA level is indicated as an amount or percentage of GPAM mRNA per sample weight. In some embodiments, the baseline GPAM mRNA level is indicated as an amount or percentage of GPAM mRNA per sample volume. In some embodiments, the baseline GPAM mRNA level is indicated as an amount or percentage of GPAM mRNA per total mRNA within the sample. In some embodiments, the baseline GPAM mRNA level is indicated as an amount or percentage of GPAM mRNA per total nucleic acids within the sample.
  • the baseline GPAM mRNA level is indicated relative to another mRNA level, such as an mRNA level of a housekeeping gene, within the sample.
  • the baseline GPAM mRNA measurement is a baseline tissue GPAM mRNA measurement.
  • the baseline GPAM mRNA measurement is obtained by an assay such as a polymerase chain reaction (PCR) assay.
  • the PCR comprises quantitative PCR (qPCR).
  • the PCR comprises reverse transcription of the GPAM mRNA.
  • the baseline measurement is a baseline GPAM protein measurement.
  • the baseline GPAM protein measurement comprises a baseline GPAM protein level.
  • the baseline GPAM protein level is indicated as a mass or percentage of GPAM protein per sample weight. In some embodiments, the baseline GPAM protein level is indicated as a mass or percentage of GPAM protein per sample volume. In some embodiments, the baseline GPAM protein level is indicated as a mass or percentage of GPAM protein per total protein within the sample. In some embodiments, the baseline GPAM protein measurement is a baseline tissue GPAM protein measurement. In some embodiments, the baseline GPAM protein measurement is obtained by an assay such as an immunoassay, a colorimetric assay, or a fluorescence assay. [00214] Some embodiments of the methods described herein include obtaining a sample from a subject.
  • the baseline measurement is obtained in a sample obtained from the subject.
  • the sample is obtained from the subject prior to administration or treatment of the subject with a composition described herein.
  • a baseline measurement is obtained in a sample obtained from the subject prior to administering the composition to the subject.
  • the sample is obtained from the subject in a fasted state.
  • the sample is obtained from the subject after an overnight fasting period.
  • the sample is obtained from the subject in a fed state.
  • the sample comprises a fluid.
  • the sample is a fluid sample.
  • the sample is a blood, plasma, or serum sample.
  • the sample comprises blood.
  • the sample is a blood sample. In some embodiments, the sample is a whole-blood sample. In some embodiments, the blood is fractionated or centrifuged. In some embodiments, the sample comprises plasma. In some embodiments, the sample is a -112- Attorney Docket No.54462-742.601 plasma sample.
  • a blood sample may be a plasma sample. In some embodiments, the sample comprises serum. In some embodiments, the sample is a serum sample. A blood sample may be a serum sample.
  • the sample comprises a tissue. In some embodiments, the sample is a tissue sample. In some embodiments, the tissue comprises liver or adipose tissue.
  • the baseline GPAM mRNA measurement, or the baseline GPAM protein measurement may be obtained in a liver or adipose sample obtained from the patient.
  • the tissue comprises adipose tissue.
  • the adipose tissue comprises white adipose tissue.
  • the adipose tissue may include adipocytes.
  • the adipose tissue may include preadipocytes.
  • the tissue comprises liver tissue.
  • the liver may include hepatocytes.
  • the tissue comprises vasculature tissue.
  • the vasculature tissue comprises endothelial cells. [00217]
  • the sample includes cells.
  • the sample comprises a cell.
  • the cell comprises an adipose cell or a liver cell.
  • the cell is an adipose cell.
  • the adipose cell is an adipocyte.
  • the cell is a liver cell.
  • the liver cell is a hepatocyte.
  • the cell is a vasculature cell.
  • the vasculature cell is an endothelial cell. D.
  • the composition or administration of the composition affects a measurement such as a liver fat percentage measurement, a liver fibrosis score measurement, a NAFLD activity score measurement, a blood alanine aminotransferase (ALT) measurement, a blood aspartate aminotransferase (AST) measurement, a blood aspartate aminotransferase (ASP) measurement, a blood bilirubin measurement, a low-density lipoprotein (LDL) measurement, a total cholesterol measurement, a non-HDL cholesterol measurement, an apolipoprotein B (APOB), a GPAM protein measurement, or a GPAM mRNA measurement, relative to the baseline measurement.
  • a measurement such as a liver fat percentage measurement, a liver fibrosis score measurement, a NAFLD activity score measurement, a blood alanine aminotransferase (ALT) measurement, a blood aspartate aminotransferase (AST) measurement, a blood aspartate aminotransferase (ASP) measurement
  • Some embodiments of the methods described herein include obtaining the measurement from a subject.
  • the measurement may be obtained from the subject after treating the subject.
  • the measurement is obtained in a second sample (such as a fluid or tissue sample described herein) obtained from the subject after the composition is administered to the subject.
  • the measurement is an indication that the disorder has been treated.
  • the measurement is obtained directly from the subject.
  • the measurement is obtained noninvasively using an imaging device.
  • the measurement is obtained in a second sample from the subject.
  • the measurement is obtained in one or more histological tissue sections.
  • the measurement is obtained by performing an assay on the second sample obtained from the subject.
  • the measurement is obtained by an assay, such as an assay described herein.
  • the assay is an immunoassay, a colorimetric assay, a fluorescence assay, a chromatography (e.g. HPLC) assay, or a PCR assay.
  • the measurement is obtained by an assay such as an immunoassay, a colorimetric assay, a fluorescence assay, or a chromatography (e.g. HPLC) assay.
  • the measurement is obtained by PCR.
  • the measurement is -113- Attorney Docket No.54462-742.601 obtained by histology. In some embodiments, the measurement is obtained by observation. In some embodiments, additional measurements are made, such as in a 3rd sample, a 4th sample, or a fifth sample. [00221] In some embodiments, the measurement is obtained within 1 hour, within 2 hours, within 3 hours, within 4 hours, within 5 hours, within 6 hours, within 12 hours, within 18 hours, or within 24 hours after the administration of the composition. In some embodiments, the measurement is obtained within 1 day, within 2 days, within 3 days, within 4 days, within 5 days, within 6 days, or within 7 days after the administration of the composition.
  • the measurement is obtained within 1 week, within 2 weeks, within 3 weeks, within 1 month, within 2 months, within 3 months, within 6 months, within 1 year, within 2 years, within 3 years, within 4 years, or within 5 years after the administration of the composition. In some embodiments, the measurement is obtained after 1 hour, after 2 hours, after 3 hours, after 4 hours, after 5 hours, after 6 hours, after 12 hours, after 18 hours, or after 24 hours after the administration of the composition. In some embodiments, the measurement is obtained after 1 day, after 2 days, after 3 days, after 4 days, after 5 days, after 6 days, or after 7 days after the administration of the composition.
  • the measurement is obtained after 1 week, after 2 weeks, after 3 weeks, after 1 month, after 2 months, after 3 months, after 6 months, after 1 year, after 2 years, after 3 years, after 4 years, or after 5 years, following the administration of the composition.
  • the composition reduces the measurement relative to the baseline measurement. For example, an adverse phenotype of NAFLD, NASH, alcoholic liver disease, liver fibrosis, liver cirrhosis, hepatocellular carcinoma, hyperlipidemia, ischemic heart disease, or coronary heart disease may be reduced upon administration of the composition.
  • the reduction is measured in a second sample obtained from the subject after administering the composition to the subject.
  • the reduction is measured directly in the subject after administering the composition to the subject.
  • the measurement is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, relative to the baseline measurement.
  • the measurement is decreased by about 10% or more, relative to the baseline measurement.
  • the measurement is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, relative to the baseline measurement.
  • the measurement is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, relative to the baseline measurement.
  • the measurement is decreased by no more than about 10%, relative to the baseline measurement. In some embodiments, the measurement is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or no more than about 100% relative to the baseline measurement. In some embodiments, the measurement is decreased by 2.5%, 5%, 7.5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages. [00223] In some embodiments, the composition increases the measurement relative to the baseline measurement.
  • a protective liver function phenotype, metabolic phenotype, or level of circulating ketone bodies may be increased upon administration of the composition.
  • the increase is measured in a second sample obtained from the subject after administering the composition to the subject. In some embodiments, the increase is measured directly in the subject after administering the composition to the subject. In some embodiments, the measurement is increased by about 2.5% or more, about 5% or more, or about 7.5% or more, relative to the baseline measurement. In some embodiments, the measurement is increased by about 10% or more, relative to the baseline measurement.
  • the measurement is increased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, relative to the baseline measurement. In some embodiments, the measurement is increased by about 100% or more, increased by about 250% or more, increased by about 500% or more, increased by about 750% or more, or increased by about 1000% or more, relative to the baseline measurement. In some embodiments, the measurement is increased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, relative to the baseline measurement. In some embodiments, the measurement is increased by no more than about 10%, relative to the baseline measurement.
  • the measurement is increased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or no more than about 100% relative to the baseline measurement. In some embodiments, the measurement is increased by no more than about 100%, increased by no more than about 250%, increased by no more than about 500%, increased by no more than about 750%, or increased by no more than about 1000%, relative to the baseline measurement.
  • the measurement is increased by 2.5%, 5%, 7.5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 250%, 500%, 750%, or 1000%, or by a range defined by any of the two aforementioned percentages.
  • the measurement is a liver fat percentage (LFP) measurement.
  • the LFP measurement is a percentage as measured by MRI.
  • the LFP measurement is a percentage as measured by CT scan.
  • the LFP is measured in a liver biopsy.
  • the composition reduces the LFP measurement relative to the baseline LFP measurement.
  • the LFP measurement is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, relative to the baseline LFP measurement. In some embodiments, the LFP measurement is decreased by about 10% or more, relative to the baseline LFP measurement. In some embodiments, the LFP measurement is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100% or more relative to the baseline LFP measurement. In some embodiments, the LFP measurement is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, relative to the baseline LFP measurement.
  • the LFP measurement is decreased by no more than about 10%, relative to the baseline LFP measurement. In some embodiments, the LFP measurement is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or about 100% relative to the baseline LFP -115- Attorney Docket No.54462-742.601 measurement. In some embodiments, the LFP measurement is decreased by 2.5%, 5%, 7.5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the measurement is a liver fibrosis measurement.
  • the liver fibrosis measurement is a liver fibrosis score (LFS).
  • the LFS comprises a score of 0, 1, 2, 3, or 4, or a range of scores defined by any two of the aforementioned numbers.
  • the LFS comprises a score of 0-4.
  • the LFS is obtained using a scoring system exemplified in Table 2.
  • the LFS measurement is obtained noninvasively.
  • the LFS measurement is obtained by a medical imaging device such as a vibration-controlled transient elastography (VCTE) device, a shear wave elastography device, a medical resonance imaging (MRI) device, a magnetic resonance spectroscopy device, a computed tomography device, or an ultrasound device.
  • a medical imaging device such as a vibration-controlled transient elastography (VCTE) device, a shear wave elastography device, a medical resonance imaging (MRI) device, a magnetic resonance spectroscopy device, a computed tomography device, or an ultrasound device.
  • the LFS measurement is obtained in a liver sample.
  • the LFS is obtained by an assay such as an immunoassay, a colorimetric assay, or a fluorescence assay.
  • the LFS is obtained using one or more indirect markers or measures of liver fibrosis such as an aspartate aminotransferase-to- platelet ratio index (APRI), a Fibrosis-4 (FIB-4) index, a FibroIndex, a Forns Index, a Hepascore, or a FibroTest.
  • the LFS is obtained using one or more indirect markers or measures of liver fibrosis such as a FIBROSpect test or a FIBROSpect II test.
  • the LFS is obtained by RT-qPCR or RNA sequencing of one or more fibrosis-related genes such as a collagen gene.
  • the LFS or the LFS is obtained using a scoring system upon a visual inspection of a sample such as a histological sample. In some embodiments, the LFS or the LFS is obtained using a stain with an affinity to collagen. In some embodiments, the LFS measurement is measured by FibroScan® in kilopascals (kPa). In some embodiments, the LFS is measured by histological analysis. In some embodiments, the LFS measurement is a LFS measurement assayed by imaging. In some embodiments, the imaging is by MRI or CT scan. [00227] In some embodiments, the composition reduces the LFS measurement relative to the baseline LFS measurement.
  • the LFS measurement is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, relative to the baseline LFS measurement. In some embodiments, the LFS measurement is decreased by about 10% or more, relative to the baseline LFS measurement. In some embodiments, the LFS measurement is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100% or more relative to the baseline LFS measurement. In some embodiments, the LFS measurement is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, relative to the baseline LFS measurement.
  • the LFS measurement is decreased by no more than about 10%, relative to the baseline LFS measurement. In some embodiments, the LFS measurement is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or about 100% relative to the baseline LFS measurement. In some embodiments, the LFS measurement is decreased by 2.5%, 5%, 7.5%, 10%, 20%, -116- Attorney Docket No.54462-742.601 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the measurement is a NAFLD activity score measurement.
  • the NAFLD activity score measurement is measured by histological analysis.
  • the NAFLD activity score measurement includes a numerical score for steatosis (0–3), hepatocyte ballooning (1–2), and lobular inflammation (0–3).
  • the score thresholds of ⁇ 3 correlates with a diagnosis of not-NASH.
  • the score thresholds of > 5 correlates with a diagnosis of NASH.
  • a NAFLD activity score measurement is assayed by imaging.
  • the composition reduces the NAFLD activity score measurement relative to the baseline NAFLD activity score measurement.
  • the NAFLD activity score measurement is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, relative to the baseline NAFLD activity score measurement. In some embodiments, the NAFLD activity score measurement is decreased by about 10% or more, relative to the baseline NAFLD activity score measurement. In some embodiments, the NAFLD activity score measurement is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100% or more relative to the baseline NAFLD activity score measurement.
  • the NAFLD ACTIVITY SCORE measurement is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, relative to the baseline NAFLD ACTIVITY SCORE measurement. In some embodiments, the NAFLD ACTIVITY SCORE measurement is decreased by no more than about 10%, relative to the baseline NAFLD ACTIVITY SCORE measurement. In some embodiments, the NAFLD ACTIVITY SCORE measurement is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or about 100% relative to the baseline NAFLD ACTIVITY SCORE measurement.
  • the NAFLD ACTIVITY SCORE measurement is decreased by 2.5%, 5%, 7.5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the measurement is an alanine aminotransferase (ALT) measurement.
  • the ALT measurement includes a blood ALT measurement.
  • the ALT measurement is measured by enzymatic activity.
  • the ALT measurement is measured by presence of an ALT epitope.
  • the ALT measurement is a circulating ALT measurement.
  • the ALT measurement is obtained by an assay such as an immunoassay, a colorimetric assay, or a fluorescence assay.
  • the ALT measurement is a blood ALT concentration (for example, units per liter (U/L)).
  • the ALT measurement is a circulating ALT measurement.
  • the ALT measurement is obtained by an assay such as an immunoassay, a colorimetric assay, or a fluorescence assay.
  • the composition reduces the ALT measurement relative to the baseline ALT measurement. In some embodiments, the composition reduces circulating ALTs relative to the -117- Attorney Docket No.54462-742.601 baseline ALT measurement.
  • the reduced ALTs are measured in a second sample obtained from the subject after administering the composition to the subject.
  • the ALT measurement is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, relative to the baseline ALT measurement.
  • the ALT measurement is decreased by about 10% or more, relative to the baseline ALT measurement.
  • the ALT measurement is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100% or more relative to the baseline ALT measurement.
  • the ALT measurement is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, relative to the baseline ALT measurement. In some embodiments, the ALT measurement is decreased by no more than about 10%, relative to the baseline ALT measurement. In some embodiments, the ALT measurement is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or about 100% relative to the baseline ALT measurement.
  • the ALT measurement is decreased by 2.5%, 5%, 7.5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the measurement is an aspartate aminotransferase (AST) measurement.
  • the AST measurement includes a blood AST measurement.
  • the AST measurement is measured by enzymatic activity.
  • the AST measurement is measured by presence of an AST epitope.
  • the AST measurement is a blood AST concentration (for example, units per liter (U/L)).
  • the AST measurement is a circulating AST measurement.
  • the AST measurement is obtained by an assay such as an immunoassay, a colorimetric assay, or a fluorescence assay.
  • the composition reduces the AST measurement relative to the baseline AST measurement. In some embodiments, the composition reduces circulating ASTs relative to the baseline AST measurement. In some embodiments, the reduced ASTs are measured in a second sample obtained from the subject after administering the composition to the subject. In some embodiments, the AST measurement is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, relative to the baseline AST measurement. In some embodiments, the AST measurement is decreased by about 10% or more, relative to the baseline AST measurement.
  • the AST measurement is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100% or more relative to the baseline AST measurement. In some embodiments, the AST measurement is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, relative to the baseline AST measurement. In some embodiments, the AST measurement is decreased by no more than about 10%, relative to the baseline AST measurement.
  • the AST measurement is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or about 100% relative to the baseline AST measurement.
  • the AST -118- Attorney Docket No.54462-742.601 measurement is decreased by 2.5%, 5%, 7.5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the measurement is an aspartate aminotransferase (ASP) measurement.
  • the ASP measurement includes a blood ASP measurement.
  • the ASP measurement is a blood ASP concentration (for example, units per liter (U/L)).
  • the ASP measurement is measured by enzymatic activity.
  • the ASP measurement is measured by presence of an ASP epitope.
  • the ASP measurement is a circulating ASP measurement.
  • the ASP measurement is obtained by an assay such as an immunoassay, a colorimetric assay, or a fluorescence assay.
  • the composition reduces the ASP measurement relative to the baseline ASP measurement. In some embodiments, the composition reduces circulating ASPs relative to the baseline ASP measurement.
  • the reduced ASPs are measured in a second sample obtained from the subject after administering the composition to the subject.
  • the ASP measurement is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, relative to the baseline ASP measurement.
  • the ASP measurement is decreased by about 10% or more, relative to the baseline ASP measurement.
  • the ASP measurement is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100% or more relative to the baseline ASP measurement.
  • the ASP measurement is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, relative to the baseline ASP measurement. In some embodiments, the ASP measurement is decreased by no more than about 10%, relative to the baseline ASP measurement. In some embodiments, the ASP measurement is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or about 100% relative to the baseline ASP measurement.
  • the ASP measurement is decreased by 2.5%, 5%, 7.5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the measurement is a bilirubin measurement.
  • the bilirubin measurement includes a blood bilirubin measurement.
  • the bilirubin measurement is a blood bilirubin concentration (for example, units per liter ( ⁇ mol/L)).
  • the bilirubin measurement is measured by presence of a bilirubin epitope.
  • the bilirubin measurement is a circulating bilirubin measurement.
  • the bilirubin measurement is obtained by an assay such as an immunoassay, a colorimetric assay, or a fluorescence assay.
  • the composition reduces the bilirubin measurement relative to the baseline bilirubin measurement.
  • the composition reduces circulating bilirubin relative to the baseline bilirubin measurement.
  • the reduced bilirubin is measured in a second sample obtained from the subject after administering the composition to the subject.
  • the bilirubin measurement is decreased by about 2.5% or more, about 5% or more, or about -119- Attorney Docket No.54462-742.601 7.5% or more, relative to the baseline bilirubin measurement. In some embodiments, the bilirubin measurement is decreased by about 10% or more, relative to the baseline bilirubin measurement. In some embodiments, the bilirubin measurement is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100% or more relative to the baseline bilirubin measurement.
  • the bilirubin measurement is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, relative to the baseline bilirubin measurement. In some embodiments, the bilirubin measurement is decreased by no more than about 10%, relative to the baseline bilirubin measurement. In some embodiments, the bilirubin measurement is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or about 100% relative to the baseline bilirubin measurement.
  • the bilirubin measurement is decreased by 2.5%, 5%, 7.5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the measurement is a low-density lipoprotein (LDL) measurement.
  • the LDL measurement includes a blood LDL measurement.
  • the LDL measurement is a blood LDL concentration (for example, units per liter (mg/dL)).
  • the LDL measurement is measured by presence of an LDL epitope.
  • the LDL measurement is a circulating LDL measurement.
  • the LDL measurement is obtained by an assay such as an immunoassay, a colorimetric assay, or a fluorescence assay.
  • the composition reduces the LDL measurement relative to the baseline LDL measurement. In some embodiments, the composition reduces circulating LDLs relative to the baseline LDL measurement. In some embodiments, the reduced LDLs are measured in a second sample obtained from the subject after administering the composition to the subject. In some embodiments, the LDL measurement is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, relative to the baseline LDL measurement. In some embodiments, the LDL measurement is decreased by about 10% or more, relative to the baseline LDL measurement.
  • the LDL measurement is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100% or more relative to the baseline LDL measurement. In some embodiments, the LDL measurement is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, relative to the baseline LDL measurement. In some embodiments, the LDL measurement is decreased by no more than about 10%, relative to the baseline LDL measurement.
  • the LDL measurement is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or about 100% relative to the baseline LDL measurement. In some embodiments, the LDL measurement is decreased by 2.5%, 5%, 7.5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages. -120- Attorney Docket No.54462-742.601 [00240] In some embodiments, the measurement is an APOB measurement.
  • the APOB measurement is a blood APOB concentration (for example, units per liter (mg/dL)). In some embodiments, the APOB measurement is a circulating APOB measurement. In some embodiments, the APOB measurement is obtained by an assay such as an immunoassay, a colorimetric assay, or a fluorescence assay. [00241] In some embodiments, the composition reduces the APOB measurement relative to the baseline APOB measurement. In some embodiments, the composition reduces circulating APOBs relative to the baseline APOB measurement. In some embodiments, the reduced APOBs are measured in a second sample obtained from the subject after administering the composition to the subject.
  • the APOB measurement is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, relative to the baseline APOB measurement. In some embodiments, the APOB measurement is decreased by about 10% or more, relative to the baseline APOB measurement. In some embodiments, the APOB measurement is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100% or more relative to the baseline APOB measurement. In some embodiments, the APOB measurement is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, relative to the baseline APOB measurement.
  • the APOB measurement is decreased by no more than about 10%, relative to the baseline APOB measurement. In some embodiments, the APOB measurement is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or about 100% relative to the baseline APOB measurement. In some embodiments, the APOB measurement is decreased by 2.5%, 5%, 7.5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages. [00242] In some embodiments, the measurement is a total cholesterol measurement.
  • the total cholesterol measurement is a blood total cholesterol concentration (for example, units per liter (mg/dL)). In some embodiments, the total cholesterol measurement is a circulating total cholesterol measurement. In some embodiments, the total cholesterol measurement is obtained by an assay such as an immunoassay, a colorimetric assay, or a fluorescence assay. [00243] In some embodiments, the composition reduces the total cholesterol measurement relative to the baseline total cholesterol measurement. In some embodiments, the composition reduces circulating total cholesterol relative to the baseline total cholesterol measurement. In some embodiments, the reduced total cholesterol is measured in a second sample obtained from the subject after administering the composition to the subject.
  • an assay such as an immunoassay, a colorimetric assay, or a fluorescence assay.
  • the total cholesterol measurement is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, relative to the baseline total cholesterol measurement. In some embodiments, the total cholesterol measurement is decreased by about 10% or more, relative to the baseline total cholesterol measurement. In some embodiments, the total cholesterol measurement is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100% or -121- Attorney Docket No.54462-742.601 more relative to the baseline total cholesterol measurement. In some embodiments, the total cholesterol measurement is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, relative to the baseline total cholesterol measurement.
  • the total cholesterol measurement is decreased by no more than about 10%, relative to the baseline total cholesterol measurement. In some embodiments, the total cholesterol measurement is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or about 100% relative to the baseline total cholesterol measurement. In some embodiments, the total cholesterol measurement is decreased by 2.5%, 5%, 7.5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages. [00244] In some embodiments, the measurement is a non-HDL cholesterol measurement.
  • the non-HDL cholesterol measurement is a blood non-HDL cholesterol concentration (for example, units per liter (mg/dL)). In some embodiments, the non-HDL cholesterol measurement is a circulating non-HDL cholesterol measurement. In some embodiments, the non-HDL cholesterol measurement is obtained by an assay such as an immunoassay, a colorimetric assay, or a fluorescence assay. [00245] In some embodiments, the composition reduces the non-HDL cholesterol measurement relative to the baseline non-HDL cholesterol measurement. In some embodiments, the composition reduces circulating non-HDL cholesterol relative to the baseline non-HDL cholesterol measurement.
  • the reduced non-HDL cholesterol is measured in a second sample obtained from the subject after administering the composition to the subject.
  • the non-HDL cholesterol measurement is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, relative to the baseline non-HDL cholesterol measurement.
  • the non-HDL cholesterol measurement is decreased by about 10% or more, relative to the baseline non-HDL cholesterol measurement.
  • the non-HDL cholesterol measurement is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100% or more relative to the baseline non-HDL cholesterol measurement.
  • the non-HDL cholesterol measurement is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, relative to the baseline non-HDL cholesterol measurement. In some embodiments, the non-HDL cholesterol measurement is decreased by no more than about 10%, relative to the baseline non-HDL cholesterol measurement. In some embodiments, the non-HDL cholesterol measurement is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or about 100% relative to the baseline non-HDL cholesterol measurement.
  • the non-HDL cholesterol measurement is decreased by 2.5%, 5%, 7.5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the measurement is an GPAM protein measurement.
  • the GPAM protein measurement comprises an GPAM protein level.
  • -122- Attorney Docket No.54462-742.601 the GPAM protein level is indicated as a mass or percentage of GPAM protein per sample weight.
  • the GPAM protein level is indicated as a mass or percentage of GPAM protein per sample volume.
  • the GPAM protein level is indicated as a mass or percentage of GPAM protein per total protein within the sample.
  • the GPAM protein measurement is a tissue GPAM protein measurement.
  • the GPAM protein measurement is obtained by an assay such as an immunoassay, a colorimetric assay, or a fluorescence assay.
  • the composition reduces the GPAM protein measurement relative to the baseline GPAM protein measurement..
  • the composition reduces tissue GPAM protein levels relative to the baseline GPAM protein measurement.
  • the reduced GPAM protein levels are measured in a second sample obtained from the subject after administering the composition to the subject.
  • the GPAM protein measurement is decreased by about 2.5% or more, about 5% or more, or about 7.5% or more, relative to the baseline GPAM protein measurement. In some embodiments, the GPAM protein measurement is decreased by about 10% or more, relative to the baseline GPAM protein measurement. In some embodiments, the GPAM protein measurement is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100%, relative to the baseline GPAM protein measurement. In some embodiments, the GPAM protein measurement is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, relative to the baseline GPAM protein measurement.
  • the GPAM protein measurement is decreased by no more than about 10%, relative to the baseline GPAM protein measurement. In some embodiments, the GPAM protein measurement is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or no more than about 100% relative to the baseline GPAM protein measurement. In some embodiments, the GPAM protein measurement is decreased by 2.5%, 5%, 7.5%, 19%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%, or by a range defined by any of the two aforementioned percentages.
  • the measurement is an GPAM mRNA measurement.
  • the GPAM mRNA measurement comprises an GPAM mRNA level.
  • the GPAM mRNA level is indicated as an amount or percentage of GPAM mRNA per sample weight.
  • the GPAM mRNA level is indicated as an amount or percentage of GPAM mRNA per sample volume.
  • the GPAM mRNA level is indicated as an amount or percentage of GPAM mRNA per total mRNA within the sample.
  • the GPAM mRNA level is indicated as an amount or percentage of GPAM mRNA per total nucleic acids within the sample.
  • the GPAM mRNA level is indicated relative to another mRNA level, such as an mRNA level of a housekeeping gene, within the sample.
  • the GPAM mRNA measurement is a tissue GPAM mRNA measurement.
  • the GPAM mRNA measurement is obtained by an assay such as a PCR assay.
  • the PCR comprises qPCR.
  • the PCR comprises reverse transcription of the GPAM mRNA. -123- Attorney Docket No.54462-742.601 [00249]
  • the composition reduces the GPAM mRNA measurement relative to the baseline GPAM mRNA measurement.
  • the GPAM mRNA measurement is obtained in a second sample obtained from the subject after administering the composition to the subject.
  • the composition reduces GPAM mRNA levels relative to the baseline GPAM mRNA levels.
  • the reduced GPAM mRNA levels are measured in a second sample obtained from the subject after administering the composition to the subject.
  • the second sample is a liver sample.
  • the second sample is an adipose sample.
  • the GPAM mRNA measurement is reduced by about 2.5% or more, about 5% or more, or about 7.5% or more, relative to the baseline GPAM mRNA measurement.
  • the GPAM mRNA measurement is decreased by about 10% or more, relative to the baseline GPAM mRNA measurement. In some embodiments, the GPAM mRNA measurement is decreased by about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, or about 100%, relative to the baseline GPAM mRNA measurement. In some embodiments, the GPAM mRNA measurement is decreased by no more than about 2.5%, no more than about 5%, or no more than about 7.5%, relative to the baseline GPAM mRNA measurement. In some embodiments, the GPAM mRNA measurement is decreased by no more than about 10%, relative to the baseline GPAM mRNA measurement.
  • the GPAM mRNA measurement is decreased by no more than about 20%, no more than about 30%, no more than about 40%, no more than about 50%, no more than about 60%, no more than about 70%, no more than about 80%, no more than about 90%, or no more than about 100%, relative to the baseline GPAM mRNA measurement. In some embodiments, the GPAM mRNA measurement is decreased by 2.5%, 5%, 7.5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% or by a range defined by any of the two aforementioned percentages. III.
  • a range should be considered to have specifically disclosed all the possible subranges as well as individual numerical values within that range.
  • description of a range such as from 1 to 6 should be considered to have specifically disclosed subranges such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6 etc., as well as individual numbers within that range, for example, 1, 2, 3, 4, 5, and 6.
  • the singular forms “a”, “an” and “the” include plural references unless the context clearly dictates otherwise.
  • the term “a sample” includes a plurality of samples, including mixtures thereof.
  • determining means determining if an element is present or not (for example, detection). These terms can include quantitative, qualitative or quantitative and qualitative determinations. Assessing can be relative or absolute. “Detecting the presence of” can include determining the amount of something present in addition to determining whether it is present or absent depending on the context.
  • subject and “patient” may be used interchangeably herein. A “subject” can be a biological entity containing expressed genetic materials.
  • the biological entity can be a plant, animal, or microorganism, including, for example, bacteria, viruses, fungi, and protozoa.
  • the subject can be a mammal.
  • the mammal can be a human.
  • the subject may be diagnosed or suspected of being at high risk for a disease. In some cases, the subject is not necessarily diagnosed or suspected of being at high risk for the disease.
  • the term “about” a number refers to that number plus or minus 10% of that number.
  • the term “about” a range refers to that range minus 10% of its lowest value and plus 10% of its greatest value.
  • treatment or “treating” are used in reference to a pharmaceutical or other intervention regimen for obtaining beneficial or desired results in the recipient.
  • beneficial or desired results include but are not limited to a therapeutic benefit and/or a prophylactic benefit.
  • a therapeutic benefit may refer to eradication or amelioration of symptoms or of an underlying disorder being treated.
  • a therapeutic benefit can be achieved with the eradication or amelioration of one or more of the physiological symptoms associated with the underlying disorder such that an improvement is observed in the subject, notwithstanding that the subject may still be afflicted with the underlying disorder.
  • a prophylactic effect includes delaying, preventing, or eliminating the appearance of a disease or condition, delaying or eliminating the onset of symptoms of a disease or condition, slowing, halting, or reversing the progression of a disease or condition, or any combination thereof.
  • a subject at risk of developing a particular disease, or to a subject reporting one or more of the physiological symptoms of a disease may undergo treatment, even though a diagnosis of this disease may not have been made.
  • the term “Cx-y” or “Cx-Cy” when used in conjunction with a chemical moiety, such as alkyl, alkenyl, or alkynyl is meant to include groups that contain from x to y carbons in the chain.
  • C1-6alkyl refers to substituted or unsubstituted saturated hydrocarbon groups, including straight-chain alkyl and branched-chain alkyl groups that contain from 1 to 6 carbons.
  • Cx-yalkenyl and Cx-yalkynyl refer to substituted or unsubstituted unsaturated aliphatic groups analogous in length and possible substitution to the alkyls described above, but that contain at least one double or triple bond, respectively.
  • -125- Attorney Docket No.54462-742.601 refers to a saturated, unsaturated or aromatic ring in which each atom of the ring is carbon.
  • Carbocycle includes 3- to 10-membered monocyclic rings, 5- to 12-membered bicyclic rings, 5- to 12-membered spiro bicycles, and 5- to 12-membered bridged rings.
  • Each ring of a bicyclic carbocycle may be selected from saturated, unsaturated, and aromatic rings.
  • an aromatic ring e.g., phenyl
  • a bicyclic carbocycle includes any combination of saturated, unsaturated and aromatic bicyclic rings, as valence permits.
  • a bicyclic carbocycle further includes spiro bicyclic rings such as spiropentane.
  • a bicyclic carbocycle includes any combination of ring sizes such as 3-3 spiro ring systems, 4-4 spiro ring systems, 4-5 fused ring systems, 5-5 fused ring systems, 5-6 fused ring systems, 6-6 fused ring systems, 5-7 fused ring systems, 6-7 fused ring systems, 5- 8 fused ring systems, and 6-8 fused ring systems.
  • Exemplary carbocycles include cyclopentyl, cyclohexyl, cyclohexenyl, adamantyl, phenyl, indanyl, naphthyl, and bicyclo[1.1.1]pentanyl.
  • aryl refers to an aromatic monocyclic or aromatic multicyclic hydrocarbon ring system.
  • the aromatic monocyclic or aromatic multicyclic hydrocarbon ring system contains only hydrogen and carbon and from five to eighteen carbon atoms, where at least one of the rings in the ring system is aromatic, i.e., it contains a cyclic, delocalized (4n+2) ⁇ -electron system in accordance with the Hückel theory.
  • aryl groups include, but are not limited to, groups such as benzene, fluorene, indane, indene, tetralin and naphthalene.
  • cycloalkyl refers to a saturated ring in which each atom of the ring is carbon. Cycloalkyl may include monocyclic and polycyclic rings such as 3- to 10-membered monocyclic rings, 5- to 12-membered bicyclic rings, 5- to 12-membered spiro bicycles, and 5- to 12-membered bridged rings. In certain embodiments, a cycloalkyl comprises three to ten carbon atoms.
  • a cycloalkyl comprises five to seven carbon atoms.
  • the cycloalkyl may be attached to the rest of the molecule by a single bond.
  • monocyclic cycloalkyls include, e.g., cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl.
  • Polycyclic cycloalkyl radicals include, for example, adamantyl, spiropentane, norbornyl (i.e., bicyclo[2.2.1]heptanyl), decalinyl, 7,7 dimethyl bicyclo[2.2.1]heptanyl, bicyclo[1.1.1]pentanyl, and the like.
  • cycloalkenyl refers to a saturated ring in which each atom of the ring is carbon and there is at least one double bond between two ring carbons. Cycloalkenyl may include monocyclic and polycyclic rings such as 3- to 10-membered monocyclic rings, 6- to 12-membered bicyclic rings, and 5- to 12-membered bridged rings.
  • a cycloalkenyl comprises five to seven carbon atoms.
  • the cycloalkenyl may be attached to the rest of the molecule by a single bond.
  • monocyclic cycloalkenyls include, e.g., cyclopentenyl, cyclohexenyl, cycloheptenyl, and cyclooctenyl.
  • halo or, alternatively, “halogen” or “halide,” means fluoro, chloro, bromo or iodo. In some embodiments, halo is fluoro, chloro, or bromo.
  • haloalkyl refers to an alkyl radical, as defined above, that is substituted by one or more halo radicals, for example, trifluoromethyl, dichloromethyl, bromomethyl, 2,2,2 trifluoroethyl, 1 chloromethyl 2 fluoroethyl, and the like.
  • the alkyl part of the haloalkyl radical is optionally further substituted as described herein.
  • heterocycle as used herein refers to a saturated, unsaturated or aromatic ring comprising one or more heteroatoms.
  • heteroatoms include N, O, Si, P, B, and S atoms.
  • Heterocycles include 3- to 10-membered monocyclic rings, 6- to 12-membered bicyclic rings, 5- to 12- membered spiro bicycles, and 5- to 12-membered bridged rings.
  • a bicyclic heterocycle includes any combination of saturated, unsaturated and aromatic bicyclic rings, as valence permits.
  • an aromatic ring e.g., pyridyl
  • a bicyclic heterocycle includes any combination of ring sizes such as 4-5 fused ring systems, 5-5 fused ring systems, 5-6 fused ring systems, 6-6 fused ring systems, 5-7 fused ring systems, 6-7 fused ring systems, 5-8 fused ring systems, and 6-8 fused ring systems.
  • a bicyclic heterocycle further includes spiro bicyclic rings, e.g., 5 to 12-membered spiro bicycles, such as 2-oxa-6-azaspiro[3.3]heptane.
  • heteroaryl refers to a radical derived from a 5 to 18 membered aromatic ring radical that comprises two to seventeen carbon atoms and from one to six heteroatoms selected from nitrogen, oxygen and sulfur.
  • the heteroaryl radical is a monocyclic, bicyclic, tricyclic or tetracyclic ring system, wherein at least one of the rings in the ring system is aromatic, i.e., it contains a cyclic, delocalized (4n+2) ⁇ -electron system in accordance with the Hückel theory.
  • Heteroaryl includes fused or bridged ring systems.
  • the heteroatom(s) in the heteroaryl radical is optionally oxidized.
  • heteroaryl is attached to the rest of the molecule through any atom of the ring(s).
  • heteroaryls include, but are not limited to, azepinyl, acridinyl, benzimidazolyl, benzindolyl, 1,3 benzodioxolyl, benzofuranyl, benzoxazolyl, benzo[d]thiazolyl, benzothiadiazolyl, benzo[b][1,4]dioxepinyl, benzo[b][1,4]oxazinyl, 1,4 benzodioxanyl, benzonaphthofuranyl, benzoxazolyl, benzodioxolyl, benzodioxinyl, benzopyranyl, benzopyranonyl, benzofuranyl, benzofuranonyl, benzothienyl (benzothiophenyl (benzothiopheny
  • heterocycloalkyl refers to a saturated ring with carbon atoms and at least one heteroatom.
  • exemplary heteroatoms include N, O, Si, P, B, and S atoms.
  • Heterocycloalkyl may include monocyclic and polycyclic rings such as 3- to 10-membered monocyclic rings, 6- to 12-membered bicyclic rings, 5- to 12-membered spiro bicycles, and 5- to 12-membered bridged rings.
  • the heteroatoms in the heterocycloalkyl radical are optionally oxidized.
  • One or more nitrogen atoms, if present, are optionally quaternized.
  • heterocycloalkyl is attached to the rest of the molecule through any atom of the heterocycloalkyl, valence permitting, such as any carbon or nitrogen atoms of the heterocycloalkyl.
  • heterocycloalkyl radicals include, but are not limited to, dioxolanyl, thienyl[1,3]dithianyl, decahydroisoquinolyl, imidazolinyl, imidazolidinyl, isothiazolidinyl, isoxazolidinyl, morpholinyl, octahydroindolyl, octahydroisoindolyl, 2 oxopiperazinyl, 2 oxopiperidinyl, 2 oxopyrrolidinyl, oxazolidinyl, piperidinyl, piperazinyl, 4 piperidonyl, pyrrolidinyl, pyrazolidinyl, quinuclidinyl
  • heterocycloalkenyl refers to an unsaturated ring with carbon atoms and at least one heteroatom and there is at least one double bond between two ring carbons. Heterocycloalkenyl does not include heteroaryl rings. Exemplary heteroatoms include N, O, Si, P, B, and S atoms. Heterocycloalkenyl may include monocyclic and polycyclic rings such as 3- to 10-membered monocyclic rings, 6- to 12-membered bicyclic rings, and 5- to 12-membered bridged rings. In other embodiments, a heterocycloalkenyl comprises five to seven ring atoms.
  • the heterocycloalkenyl may be attached to the rest of the molecule by a single bond.
  • monocyclic cycloalkenyls include, e.g., pyrroline (dihydropyrrole), pyrazoline (dihydropyrazole), imidazoline (dihydroimidazole), triazoline (dihydrotriazole), dihydrofuran, dihydrothiophene, oxazoline (dihydrooxazole), isoxazoline (dihydroisoxazole), thiazoline (dihydrothiazole), isothiazoline (dihydroisothiazole), oxadiazoline (dihydrooxadiazole), thiadiazoline (dihydrothiadiazole), dihydropyridine, tetrahydropyridine, dihydropyridazine, tetrahydropyridazine, dihydropyrimidine, tetrahydro
  • substituted refers to moieties having substituents replacing a hydrogen on one or more carbons or substitutable heteroatoms, e.g., an NH or NH2 of a compound. It will be understood that “substitution” or “substituted with” includes the implicit proviso that such substitution is in accordance with permitted valence of the substituted atom and the substituent, and that the substitution results in a stable compound, i.e., a compound which does not spontaneously undergo transformation such as by rearrangement, cyclization, elimination, etc.
  • substituted refers to moieties having substituents replacing two hydrogen atoms on the same carbon atom, such as substituting the two hydrogen atoms on a single carbon with an oxo, imino or thioxo group.
  • substituted is contemplated to include all permissible substituents of organic compounds.
  • the permissible substituents include acyclic and cyclic, branched and unbranched, carbocyclic and heterocyclic, aromatic and non-aromatic substituents of organic compounds.
  • the permissible substituents can be one or more and the same or different for appropriate organic compounds.
  • a "derivative" polypeptide or peptide is one that is modified, for example, by glycosylation, pegylation, phosphorylation, sulfation, reduction/alkylation, acylation, chemical coupling, or mild formalin treatment.
  • a derivative may also be modified to contain a detectable label, either directly or indirectly, including, but not limited to, a radioisotope, fluorescent, and enzyme label.
  • a detectable label either directly or indirectly, including, but not limited to, a radioisotope, fluorescent, and enzyme label.
  • Some embodiments refer to nucleic acid sequence information. It is contemplated that in some embodiments, thymine (T) may be interchanged with uracil (U), or vice versa. For example, some sequences in the sequence listing may recite Ts, but these may be replaced with Us in some embodiments. In some oligonucleotides with nucleic acid sequences that include uracil, the uracil may be replaced with thymine.
  • oligonucleotides with nucleic acid sequences that include thymine may be replaced with uracil.
  • an oligonucleotide such as an siRNA comprises or consists of RNA.
  • the oligonucleotide may include DNA.
  • the oligonucleotide may include 2’ deoxyribonucleotides.
  • An ASO may comprise or consist of -129- Attorney Docket No.54462-742.601 DNA. To any extent that the sequence listing contradicts the disclosure in the specification, the specification takes precedence. [00274] Some aspects include sequences with nucleotide modifications or modified internucleoside linkages.
  • Nf refers to a 2’ fluoro- modified nucleoside
  • dN e.g. dA, dC, dG, dT, or dU
  • n e.g. a, c, g, t, or u
  • s refers to a phosphorothioate linkage.
  • a pyrimidine may include cytosine (C), thymine (T), or uracil (U).
  • a pyrimidine may include C or U.
  • a pyrimidine may include C or T.
  • a reference to a pyrimidine may include a nucleoside or nucleotide comprising the pyrimidine.
  • a purine may include guanine (G) or adenine (A).
  • a reference to a purine may include a nucleoside or nucleotide comprising a purine.
  • Example 1 Functional Variants in GPAM Demonstrate Protective Associations for Non-Alcoholic Fatty Liver Disease, Alcoholic Liver Disease, Decreased Liver Fat Percentage and Decreased Blood Lipids
  • Variants in GPAM were evaluated for associations with liver disease, liver fat percentage, liver function parameters, and blood lipid levels in approximately 452,000 individuals from the UK Biobank cohort.
  • eQTL quantitative trait locus
  • NAFLD non-alcoholic fatty liver disease
  • alcoholic liver disease liver fibrosis and cirrhosis
  • sequelae of chronic liver disease such as esophageal varices and portal hypertension
  • MRI-derived liver fat percentage Table 5
  • GPAM variants are also associated with decreased blood levels of liver enzymes, including alanine aminotransferase (ALT), aspartate aminotransferase (AST) and alkaline phosphatase (ALP), and with decreased blood bilirubin (Table 6).
  • Table 5 GPAM variant liver disease-related associations ase V rs2 8 rs5 3 n V rs2 9 rs57 . ⁇ . . - ⁇ . . ⁇ .02 Tbl 6 GPAM rint li r f ntin itin V ) eta rs2 . . . . .
  • GPAM variants are also associated with decreased LDL-cholesterol, total cholesterol and APOB, and with decreased use of ‘statin’ (HMG CoA reductase inhibitor) medications (Table 7). These protective associations with putative loss of function variants in GPAM across several related and distinct diseases and traits su t tht ihibiti f GPAM ld b th ti i th d related diseases.
  • siRNAs with high specificity and a low number of predicted off-targets provided a benefit of increased targeting specificity.
  • siRNA sequences within the seed region were analyzed for similarity to seed regions of known miRNAs.
  • siRNAs can function in a miRNA like manner via base-pairing with complementary sequences within the 3’-UTR of mRNA molecules. The complementarity typically encompasses the 5‘-bases at positions 2-7 of the miRNA (seed region).
  • siRNA strands containing natural miRNA seed regions can be avoided. Seed regions identified in miRNAs from human, mouse, rat, rhesus monkey, dog, rabbit, and pig are referred to as “conserved”.
  • a “specificity category” This is divided into categories 1-4, with 1 having the highest specificity and 4 having the lowest specificity. Each strand of the siRNA is assigned to a specificity category.
  • gnomAD Genome Aggregation Database
  • siRNA sequences derived from human GPAM mRNA (ENST00000348367.9, SEQ ID NO: 12867) without consideration of specificity or species cross-reactivity was 6354 (sense and antisense strand sequences included in SEQ ID NOS: 1-6354 and 6355-12708, respectively).
  • 6354 sense and antisense strand sequences included in SEQ ID NOS: 1-6354 and 6355-12708, respectively.
  • Prioritizing sequences for target specificity, miRNA seed region sequences and SNPs as described above yields subset A.
  • Subset A contains 1348 siRNAs whose base sequences are shown in Table 8.
  • the above methods can be used to identify therapeutic siRNAs to downmodulate expression of the GPAM mRNA. Table 8.
  • Subset B included the following 1344 siRNAs: 13, 18, 23, 26, 28, 54, 62, 79, 81, 82, 83, 84, 88, 90, 91, 94, 95, 96, 98, 100, 101, 106, 107, 108, 109, 110, 111, 113, 114, 118, 119, 127, 161, 162, 170, 173, 175, 176, 177, 179, 181, 190, 191, 194, 202, 206, 207, 211, 216, 218, 221, 232, 241, 246, 247, 257, 260, 261, 262, 263, 264, 265, 267, 275, 278, 279, 280, 284, 303, 331, 354, 356, 357, 358, 359, 360, 361, 362, 363, 364, 365, 370, 386, 397, 401, 403, 417, 425, 427, 432, 435, 436, 444, 450, 454,
  • the siRNAs in subset B had the following characteristics: Cross-reactivity: With 19mer in human GPAM mRNA; Specificity category: For human: AS2 or better, SS3 or better; miRNA seeds: AS+SS strand: seed region not conserved in human, mouse, and rat and not present in >4 species; Off- target frequency: ⁇ 20 human off-targets matched with 2 mismatches in antisense strand; and SNPs: siRNA target sites do not harbor SNPs with a MAF ⁇ 1% (pos.2-18). [00292] The siRNA sequences in subset B were further selected for absence of seed regions in the AS strand that are identical to a seed region of known human miRNA to yield subset C.
  • Subset C included the following 931 siRNAs: 18, 23, 62, 79, 81, 82, 83, 84, 88, 90, 94, 95, 96, 98, 101, 106, 107, 108, 110, 113, 114, 118, 119, 127, 173, 175, 190, 191, 194, 207, 211, 218, 221, 232, 241, 257, 260, 261, 262, 263, 264, 265, 278, 280, 284, 331, 354, 357, 358, 359, 360, 361, 363, 364, 365, 386, 432, 435, 444, 450, 454, 458, 487, 493, 494, 495, 496, 497, 498, 499, 501, 506, 507, 515, 517, 518, 519, 520, 521, 537, 542, 543, 547, 549, 550, 551, 554, 562, 564, 565, 596, 601, 60
  • siRNAs in subset C had the following characteristics: Cross-reactivity: With 19mer in human GPAM mRNA; Specificity category: For human: AS2 or better, SS3 or better; miRNA seeds: AS+SS strand: seed region not conserved in human, mouse, and rat and not present in >4 species. AS strand: seed region not identical to seed region of known human miRNA; Off-target frequency: ⁇ 30 human off-targets matched with 2 mismatches by antisense strand; and SNPs: siRNA target sites do not harbor SNPs with a MAF ⁇ 1% (pos.2-18).
  • siRNA sequences in subset C were also selected for absence of seed regions in the AS or S strands that are identical to a seed region of known human miRNA in addition to having an off-target frequency of ⁇ 30 human off-targets matched with 2 mismatches by antisense strand to yield subset D.
  • Subset D included the following 595 siRNAs: 18, 23, 79, 83, 94, 95, 98, 106, 107, 108, 110, 114, 118, 127, 173, 175, 190, 191, 207, 218, 221, 241, 257, 260, 261, 262, 264, 278, 284, 354, 357, 359, 360, 361, 363, 364, 365, 386, 435, 450, 454, 494, 495, 497, 498, 499, 501, 507, 517, 518, 520, 537, 547, 549, 550, 551, 554, 596, 601, 613, 646, 647, 664, 673, 675, 681, 683, 703, 737, 745, 746, 748, 753, 756, 819, 826, 839, 845, 850, 854, 855, 857, 859, 874, 878, 879, 887, 889, 891, 893, 894
  • subset D The siRNA sequences in subset D were also selected to have an off-target frequency of ⁇ 20 human off-targets matched with 2 mismatches by antisense strand to yield subset E.
  • Subset E included the following 593 siRNAs: 18, 23, 79, 83, 94, 95, 98, 106, 107, 108, 110, 114, 118, 127, 173, 175, 190, 191, 207, 218, 221, 241, 257, 260, 261, 262, 264, 278, 284, 354, 357, 359, 360, 361, 363, 364, 365, 386, 435, 450, 454, 494, 495, 497, 498, 499, 501, 507, 517, 518, 520, 537, 547, 549, 550, 551, 554, 596, 601, 613, 646, 647, 664, 673, 675, 681, 683, 703, 737, 745, 746, 748,
  • any siRNA among any of subsets A-E may comprise any modification pattern described herein. If a sequence has a different number of nucleotides in length than a modification pattern, the modification pattern may still be used with the appropriate number of additional nucleotides added 5’ or 3’ to match the number of nucleotides in the modification pattern. For example, if a sense or antisense strand of the siRNA among any of subsets A-E comprises 19 nucleotides, and a modification pattern comprises 21 nucleotides, UU may be added onto the 5’ end of the sense or antisense strand.
  • siRNAs were designed to target human GPAM as described above and, in some cases, the GPAM sequence of at least one toxicology-relevant species, in this case, the non-human primate (NHP) cynomolgus monkey.
  • the siRNAs included in subset F had the following characteristics: Cross-reactivity: With 19mer in human GPAM mRNA, with 17mer/19mer in NHP GPAM; Specificity category: For human and NHP: AS2 or better, SS3 or better.
  • Subset F included 25 siRNAs whose base sequences are shown in Table 9. Table 9.
  • subset F Sequences and siRNA names in subset F (Screening Set) siR ⁇ -3 ⁇ ) siR AU siR UC siR AG siR AU siR CU siR GA siR AG siR AG siR GU siR GA siR UA siR AU siR CU siR AA siR CU siR CA siR GA siR CA siR UU siR UU siR UA siR AU siR AA siRNA 2486 2486 AACAGAAAGAAAUGUUGCA 8840 UGCAACAUUUCUUUCUGUU siRNA 2660 2660 AUAUAUUCUGAGUUUUGUG 9014 CACAAAACUCAGAAUAUAU -167- Attorney Docket No.54462-742.601 [00298]
  • the sense strand of any of the siRNAs of subset F comprises siRNA with a particular modification pattern.
  • position 9 counting from the 5’ end of the of the sense strand is has the 2’F modification. If position 9 of the sense strand is a pyrimidine, then all purines in the sense strand have the 2’OMe modification. If position 9 is the only pyrimidine between positions 5 and 11 of the sense stand, then position 9 is the only position with the 2’F modification in the sense strand. If position 9 and only one other base between positions 5 and 11 of the sense strand are pyrimidines, then both of these pyrimidines are the only two positions with the 2’F modification in the sense strand.
  • position 9 and only two other bases between positions 5 and 11 of the sense strand are pyrimidines, and those two other pyrimidines are in adjacent positions so that there would be not three 2’F modifications in a row, then any combination of 2’F modifications can be made that give three 2’F modifications in total. If there are >2 pyrimidines between positions 5 and 11 of the sense strand, then all combinations of pyrimidines having the 2’F modification are allowed that have three to five 2’F modifications in total, provided that the sense strand does not have three 2’F modifications in a row. [00299] If position 9 of the sense strand is a purine, then all purines in the sense strand have the 2’OMe modification.
  • position 9 is the only purine between positions 5 and 11 of the sense stand, then position 9 is the only position with the 2’F modification in the sense strand. If position 9 and only one other base between positions 5 and 11 of the sense strand are purines, then both of these purines are the only two positions with the 2’F modification in the sense strand. If position 9 and only two other bases between positions 5 and 11 of the sense strand are purines, and those two other purines are in adjacent positions so that there would be not three 2’F modifications in a row, then any combination of 2’F modifications can be made that give three 2’F modifications in total.
  • the sense strand of any of the siRNAs of subset F comprises a modification pattern which conforms to these sense strand rules (Table 10).
  • the antisense strand of any of the siRNAs of subset F comprise a modification or modification pattern.
  • the siRNAs in Table 10 include a GalNAc moiety.
  • Table 11 includes some additional sense strand modifications of the siRNAs in subset F.
  • the siRNAs in subset F may comprise any other modification pattern(s).
  • Nf e.g. Af, Cf, Gf, Tf, or Uf
  • dN e.g. dA, dC, dG, dT, or dU
  • n e.g.
  • a, c, g, t, or u is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • Table 10 Modified Screening Set (Subset G) siRNA SEQ ID Sense strand sequence (5 ⁇ -3 ⁇ ), with SEQ ID Name NO: GalNAc moiety NO: Antisense strand sequence (5 ⁇ -3 ⁇ ) ETD01994 12709 [ETL17]saucaGfAfauAfcAfguuggasusu 12734 usCfscAfaCfaCfuGfuAfuUfcUfgAfususu ETD susu ETD usu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu
  • Subset I includes 25 siRNAs whose base sequences are shown in Table 11A.
  • Tbl 11A S b t I iRNA siR ⁇ -3 ⁇ ) siR AU siR UC siR AG siR AU siR CU siR GA siR AG siR AG siR GU siR GA siR UA siRNA 1098 1098 AUAGUUGAAUUACUUCGAC 7452 GUCGAAGUAAUUCAACUAU siRNA 1100 1100 AGUUGAAUUACUUCGACAG 7454 CUGUCGAAGUAAUUCAACU -171- Attorney Docket No.54462-742.601 siRNA 1111 1111 UUCGACAGCAGCAAUUCUU 7465 AAGAAUUGCUGCUGUCGAA siRNA 1199 1199 AGUUGUGGUAGAUACUCUG 7553 CAGAGUAUCUACCACAACU siR CA siR GA siR CA siR UU siR UU siR UA siR AU siR
  • NHP and mouse cross-reactive siRNAs For human and NHP only cross-reactive siRNAs: human AS1 or better, human SS3 or better. For human, NHP and mouse cross-reactive siRNAs, human AS2 or better, human AS3 or better. • miRNA seeds: For human and NHP only cross-reactive siRNAs: AS and SS: seed region not in any human siRNA, seed region not conserved in human, mouse, and rat and not present in >3 species. For human, NHP and mouse cross-reactive siRNAs: seed region not in any human siRNA, seed region not conserved in human, mouse, and rat and not present in >1 species.
  • siRNA target sites do not harbor SNPs with a MAF ⁇ 1% (pos.2-18)
  • the siRNAs targeting human GPAM can also be selected using at least Selection Set C criteria, and including cross-reactivity with the NHP cynomolgus GPAM. Selection of these siRNAs yields Subset J.
  • Subset J includes 24 siRNAs whose base sequences are shown in Table 11B.
  • the siRNAs targeting GPAM can be synthesized with chemical modifications with the sense strand having modification pattern 1S and antisense strand having modification pattern 1AS.
  • the siRNAs targeting GPAM can also be synthesized with chemical modifications with the sense strand having modification pattern 2S and antisense strand having modification pattern 3AS.
  • the modifications included in Table 10 or Table 11 are used.
  • adenosine can be placed at position 19 in the sense strand and uridine at position 1 in the antisense strand.
  • Example 4 siRNA-mediated knockdown of GPAM in human hepatocyte cells
  • siRNAs targeted to the GPAM mRNA that downregulate levels of GPAM mRNA may lead to subsequent decrease of lysophosphatidic acid (LPA), a metabolite produced by the GPAM gene product, when administered to cultured hepatocyte cells.
  • LPA lysophosphatidic acid
  • the hepatocyte cells are to be seeded at 150,000 cells/mL into a Falcon 24-well tissue culture plate (ThermoFisher Cat. No. 353047) at 0.5 mL per well .
  • the GPAM siRNA and negative control siRNA master mixes are prepared.
  • the GPAM siRNA master mix contains 350 uL of Opti-MEM (ThermoFisher Cat. No.4427037 - s1288 Lot No. AS02B02D) and 3.5 ul of a mixture of the two GPAM siRNAs (10 uM stock).
  • the negative control siRNA master mix contains 350 uL of Opti-MEM and 3.5 ul of negative control siRNA (ThermoFisher -173- Attorney Docket No.54462-742.601 Cat. No.4390843, 10 uM stock).
  • 3 uL of TransIT-X2 (Mirus Cat. No. MIR6000) is added to each master mix.
  • the Stop Solution (5 ul/well) is added to each well and mixed by pipetting up and down five times and incubating at room temperature for 2 minutes.
  • the reverse transcriptase reaction is performed using 22.5 ul of the lysate according to the manufacturer’s protocol. Samples are stored at -80 °C until real-time qPCR was performed in triplicate using TaqMan Gene Expression Assays (Applied Biosystems FAM/GPAM using a BioRad iCycler).
  • a decrease in GPAM mRNA expression in the hepatocyte cells is expected after transfection with the GPAM siRNAs compared to GPAM mRNA levels in hepatocyte cells transfected with the non-specific control siRNA 48 hours after transfection.
  • Example 5 siRNA-mediated knockdown of GPAM in human adipocyte cells
  • Adipose tissue is a major site of endogenous triglyceride synthesis.
  • siRNAs targeted to the GPAM mRNA that downregulate levels of GPAM mRNA may lead to subsequent decrease of lysophosphatidic acid (LPA), a metabolite produced by the GPAM gene product, when administered to cultured adipocyte cells.
  • LPA lysophosphatidic acid
  • a decrease in LPA production may lead to a subsequent decrease in the levels of other components of the glycerol phosphate pathway for de novo triacylglycerol synthesis.
  • phosphatidic acid (PA), phosphatidylserine (DAG), and triacylglycerol (TAG) may be decreased after siRNA-mediated knockdown of GPAM in human adipocyte cells.
  • PA phosphatidic acid
  • DAG phosphatidylserine
  • TAG triacylglycerol
  • the GPAM siRNA master mix contains 350 uL of Opti-MEM (ThermoFisher Cat. No.4427037 - s1288 Lot No. AS02B02D) and 3.5 ul of a mixture of the two GPAM siRNAs (10 uM stock).
  • the negative control siRNA master mix contains 350 uL of Opti-MEM and 3.5 ul of negative control siRNA (ThermoFisher -174- Attorney Docket No.54462-742.601 Cat. No.4390843, 10 uM stock).
  • 3 uL of TransIT-X2 (Mirus Cat. No. MIR6000) is added to each master mix.
  • the Stop Solution (5 ul/well) is added to each well and mixed by pipetting up and down five times and incubating at room temperature for 2 minutes.
  • the reverse transcriptase reaction is performed using 22.5 ul of the lysate according to the manufacturer’s protocol. Samples are stored at -80 °C until real-time qPCR was performed in triplicate using TaqMan Gene Expression Assays (Applied Biosystems FAM/GPAM using a BioRad iCycler).
  • a decrease in GPAM mRNA expression in the adipocyte cells is expected after transfection with the GPAM siRNAs compared to GPAM mRNA levels in adipocyte cells transfected with the non-specific control siRNA 48 hours after transfection.
  • Example 6 ASO-mediated knockdown of GPAM in human hepatocyte cells
  • ASOs targeted to the GPAM mRNA that downregulate levels of GPAM mRNA leading to subsequent decrease of LPA, a metabolite produced by the GPAM gene product, when administered to cultured hepatocyte cells.
  • the hepatocyte cells are seeded at 150,000 cells/mL into a Falcon 24-well tissue culture plate (Cat. No.353047) at 0.5 mL per well.
  • the GPAM ASO and negative control ASO master mixes are prepared.
  • the GPAM ASO master mix contains 350 uL of Opti-MEM (ThermoFisher Cat.
  • the negative control ASO master mix contains 350 uL of Opti-MEM and 3.5 ul of negative control ASO (ThermoFisher Cat. No. 4390843, 10 uM stock).
  • 3 uL of TransIT-X2 (Mirus Cat. No. MIR6000) is added to each master -175- Attorney Docket No.54462-742.601 mix.
  • the Stop Solution (5 ul/well) is added to each well and mixed by pipetting up and down five times and incubating at room temperature for 2 minutes.
  • the reverse transcriptase reaction is performed using 22.5 ul of the lysate according to the manufacturer’s protocol. Samples are stored at -80 °C until real-time qPCR was performed in triplicate using TaqMan Gene Expression Assays (Applied Biosystems FAM/GPAM using a BioRad iCycler).
  • a decrease in GPAM mRNA expression in the hepatocyte cells is expected after transfection with the GPAM ASOs compared to GPAM mRNA levels in hepatocyte cells transfected with the non-specific control ASO 48 hours after transfection.
  • fatty liver disease is induced by feeding mice a Western Diet (WD) containing 21.1% fat, 41% Sucrose, and 1.25% Cholesterol by weight (Teklad diets, TD.120528) and a high sugar solution (23.1g/L d-fructose (Sigma-Aldrich, G8270) and 18.9 g/L d-glucose (Sigma- Aldrich, F0127)) for 12 weeks.
  • WD Western Diet
  • Teklad diets TD.1205208
  • a high sugar solution (23.1g/L d-fructose (Sigma-Aldrich, G8270) and 18.9 g/L d-glucose (Sigma- Aldrich, F0127)
  • mice are divided into five groups: Group 1 - a fatty liver disease group treated with non-targeting control siRNA, Group 2 - a fatty liver disease group treated with non-targeting control ASO, Group 3 - a fatty liver disease group treated with GPAM siRNA1, Group 4 – a fatty liver disease group treated with GPAM ASO1, Group 5 - control mice on a normal chow diet. Each group contains eight mice (4 males, 4 females). [00327] At weeks 12 weeks of Western Diet, blood samples are collected from each group prior to the first treatment.
  • siRNA or ASO administration of siRNA or ASO is achieved with a 200ul subcutaneous injection of siRNA or ASO resuspended in PBS at concentration of 10uM.
  • Group 1 mice are injected subcutaneously with non-targeting control siRNA
  • Group 2 mice are injected subcutaneously with non- targeting control ASO
  • Group 3 mice are injected subcutaneously with siRNA1 targeting mouse GPAM
  • Group 4 mice are injected subcutaneously with ASO1 targeting mouse GPAM
  • Group 5 mice are -176- Attorney Docket No.54462-742.601 injected subcutaneously with vehicle. Every other week thereafter starting on Day 14 the animals from each group are dosed as on Day 0 for a total of 5 injections.
  • ALT, AST, ALP bilirubin, total cholesterol, HDL cholesterol and triglyceride levels are measured using VITROS 5,1 FS (Ortho Clinical Diagnostics).
  • Non-fasting plasma insulin is measured with the Ultrasensitive Mouse Insulin ELISA kit (Crystal Chem, 90080) according to the manufacturer’s instructions.
  • Non-fasting blood glucose is assayed with the One Touch Ultra (Life Scan).
  • HOMA IR and QUICKI are calculated. Blood ketones, including 3-hydroxybutyrate, acetoacetate and acetone are measured using EnzyChrom Ketone Body Assay Kit (BioAssay Systems, EKBD-100).
  • mice are sacrificed by cervical dislocation following an intraperitoneal injection of 0.3 ml Nembutal (5 mg/ml).
  • Terminal blood draw is collected via cardiac puncture and final ALT, AST, ALP bilirubin, total cholesterol, HDL cholesterol and triglyceride levels are measured along with non-fasting plasma insulin and glucose and blood ketones.
  • Livers and adipose tissue are removed and divided into three sections each; one section placed in RNAlater for mRNA isolation, one section flash-frozen for protein isolation, one section fixed in formalin and then paraffin-embedded.
  • mRNA is isolated from tissue placed in RNAlater solution using the PureLink kit according to the manufacturer’s protocol (ThermoFisher Cat. No.12183020). The reverse transcriptase reaction is performed according to the manufacturer’s protocol. Samples are stored at -80 °C until real-time qPCR was performed in triplicate using TaqMan Gene Expression Assays (Applied Biosystems FAM/GPAM using a BioRad iCycler).
  • a decrease in GPAM mRNA expression in the liver and adipose tissue from mice is dosed with the GPAM siRNAs and ASOs compared to GPAM mRNA levels in the liver and adipose tissue from mice is dosed with the non-specific control siRNA and ASO.
  • NAFLD Activity Score (NAS) and fibrosis stage are evaluated by an expert pathologist according to the NASH CRN scoring system. The histological scoring is performed blinded, with no knowledge by the pathologist of the treatment(s) received.
  • GPAM siRNAs and ASOs elicit -177- Attorney Docket No.54462-742.601 knockdown of GPAM mRNA in liver tissue and that the decrease in GPAM expression is correlated with a decrease in NAS and NASH CRN.
  • Oligonucleotide Synthesis [00333] Oligonucleotides such as siRNAs may be synthesized according to phosphoramidite technology on a solid phase.
  • a K&A oligonucleotide synthesizer may be used. Syntheses may be performed on a solid support made of controlled pore glass (CPG, 500 ⁇ or 600 ⁇ , obtained from AM Chemicals, Oceanside, CA, USA). All 2′-OMe and 2’-F phosphoramidites may be purchased from Hongene Biotech (Union City, CA, USA).
  • CPG controlled pore glass
  • All phosphoramidites may be dissolved in anhydrous acetonitrile (100 mM) and molecular sieves (3 ⁇ ) may be added.5-Benzylthio-1H-tetrazole (BTT, 250 mM in acetonitrile) or 5-Ethylthio-1H-tetrazole (ETT, 250 mM in acetonitrile) may be used as activator solution. Coupling times may be 9-18 min (e.g. with a GalNAc such as ETL17), 6 min (e.g. with 2′OMe and 2′F).
  • a 100 mM solution of 3-phenyl 1,2,4- dithiazoline-5-one (POS, obtained from PolyOrg, Inc., Leominster, Mass., USA) in anhydrous acetonitrile may be employed.
  • POS 3-phenyl 1,2,4- dithiazoline-5-one
  • the dried solid support may be treated with a 1:1 volume solution of 40 wt. % methylamine in water and 28% ammonium hydroxide solution (Aldrich) for two hours at 30° C.
  • the solution may be evaporated and the solid residue may be reconstituted in water and purified by anionic exchange HPLC using a TKSgel SuperQ-5PW 13u column.
  • Buffer A may be 20 mM Tris, 5 mM EDTA, pH 9.0 and contained 20% Acetonitrile and buffer B may be the same as buffer A with the addition of 1 M sodium chloride. UV traces at 260 nm may be recorded. Appropriate fractions may be pooled then desalted using Sephadex G-25 medium. [00335] Equimolar amounts of sense and antisense strand may be combined to prepare a duplex.
  • the duplex solution may be prepared in 0.1 ⁇ PBS (Phosphate-Buffered Saline, 1 ⁇ , Gibco). The duplex solution may be annealed at 95° C. for 5 min, and cooled to room temperature slowly.
  • PBS Phosphate-Buffered Saline
  • Duplex concentration may be determined by measuring the solution absorbance on a UV-Vis spectrometer at 260 nm in 0.1 ⁇ PBS. For some experiments, a conversion factor may be calculated from an experimentally determined extinction coefficient.
  • Example 9 GalNAc ligand for hepatocyte targeting of oligonucleotides [00336]
  • GalNAc ligands may be attached to solid phase resin for 3’ conjugation or at the 5’ terminus using GalNAc phosphoramidite reagents.
  • GalNAc phosphoramidites may be coupled on solid phase as for other nucleosides in the oligonucleotide sequence at any position in the sequence.
  • Reagents for GalNAc conjugation to oligonucleotides are shown in Table 12. -178- Attorney Docket No.54462-742.601 Table 12.
  • GalNAc Conjugation Reagents Type of Structure conjugation Sol atta wh is r olig cha OH atta pha Sol atta pho -179- Attorney Docket No.54462-742.601 Solid phase 5’ atta Pho Sol Car ami any olig y g p y y p g g p oved under basic, acid or reducing conditions.
  • the oligonucleotide is then removed from the resin and GalNAc is conjugated to the reactive site.
  • the carboxy GalNAc derivatives may be coupled to amino-modified oligonucleotides.
  • peptide coupling conditions are known to the skilled in the art using a carbodiimide coupling agent like DCC (N,N′-Dicyclohexylcarbodiimide), EDC (N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide) or EDC.HCl (N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride and an additive like HOBt (1- hydroxybenztriazole), HOSu (N-hydroxysuccinimide), TBTU (N,N,N′,N′-Tetramethyl-O-(benzotriazol-1- yl)uronium tetrafluoroborate, HBTU (2-(1H-benzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate) or HOAt (1-Hydroxy-7-azabenzotriazole
  • Amine groups may be incorporated into oligonucleotides using a number of known, commercially available reagents at the 5’ terminus, 3’ terminus or anywhere in between.
  • Non-limiting examples of reagents for oligonucleotide synthesis to incorporate an amino group include: • 5’ attachment: • 6-(4-Monomethoxytritylamino)hexyl-(2-cyanoethyl)-(N,N-diisopropyl)-phosphoramidite CAS Number: 114616-27-2 • 5'-Amino-Modifier TEG CE-Phosphoramidite • 10-(O-trifluoroacetamido-N-ethyl)-triethyleneglycol-1-[(2-cyanoethyl)-(N,N-diisopropyl)]- phosphoramidite • 3’ attachment: • 3'-Amino-Modifier Serinol CPG • 3-Dimethoxytr
  • Solution phase conjugations may occur after oligonucleotide synthesis via reactions between non-nucleosidic nucleophilic functional groups that are attached to the oligonucleotide and electrophilic GalNAc reagents.
  • nucleophilic groups include amines and thiols
  • electrophilic reagents include activated esters (e.g. N-hydroxysuccinimide, pentafluorophenyl) and maleimides.
  • GalNAc ligands for hepatocyte targeting of oligonucleotides
  • GalNAc ligands may be attached to solid phase resin for 3’ conjugation or at the 5’ terminus using GalNAc phosphoramidite reagents.
  • GalNAc phosphoramidites may be coupled on solid phase as for other nucleosides in the oligonucleotide sequence at any position in the sequence.
  • a non-limiting example of a phosphoramidite reagent for GalNAc conjugation to a 5’ end oligonucleotide is shown in Table 13.
  • Table 13 GalNAc Conjugation Reagent Type of c Structure S ph
  • the following includes examples of synthesis reactions used to create a GalNAc moiety: Scheme for the preparation of NAcegal-Linker-TMSOTf -182- Attorney Docket No.54462-742.601 General procedure for preparation of Compound 2A [00345] To a solution of Compound 1A (500 g, 4.76 mol, 476 mL) in 2-Methly-THF (2.00 L) is added CbzCl (406 g, 2.38 mol, 338 mL) in 2-Methyl-THF (750 mL) dropwise at 0 °C.
  • reaction mixture is added to cold water (30.0 L) -183- Attorney Docket No.54462-742.601 and stirred at 0 °C for 0.5 hr, white solid formed, filtered and dried to give Compound 4A (1.55 kg, 3.98 mol, 85.8% yield) as a white solid and used in the next step without further purification.
  • the reaction mixture is diluted with DCM (100 mL) then washed with aq.NaHCO 3 (250 mL * 1) and brine (250 mL), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue.
  • Example 12 Modification motif 2 [00359]
  • An example GPAM siRNA includes a combination of the following modifications: • Position 9 (from 5’ to 3’) of the sense strand is 2’ deoxy. • Sense strand positions 5, 7 and 8 are 2’ F. • All pyrimidines in positions 10-21 are 2’ OMe, and purines are a mixture of 2’ OMe and 2’ F. Alternatively, all purines in positions 10-21 are 2’ OMe and all pyrimidines in positions 10-21 are a mixture of 2’ OMe and 2’ F. • Antisense strand odd-numbered positions are 2'OMe and even-numbered positions are a mixture of 2’ F, 2’OMe and 2’ deoxy. Example 12.
  • siRNAs ETD01994-ETD02018 targeting human GPAM mRNA in mice transfected with AAV8-TBG-h-GPAM [00360]
  • the siRNAs contain the GalNAc ligand ETL17 followed by a phosphorothioate linkage at the 5’ end of the sense strand.
  • the siRNA sequences that were used are shown in Table 14, where Nf (e.g. Af, Cf, Gf, Tf, or Uf) is a 2’ fluoro-modified nucleoside, n (e.g.
  • a, c, g, t, or u is a 2’ O-methyl modified nucleoside
  • dN e.g. dA, dC, dG, dT, or dU
  • s is a phosphorothioate linkage.
  • the base sequences for each siRNA, with and without the 3’ UU extension, are shown in Table 15. ETD01994-ETD02004 were tested in Part 1 of the study and ETD02005-ETD02018 were tested in Part 2.
  • AAV8 adeno- associated virus 8 vector
  • the recombinant AAV8 contains the open reading frame, the 5’ UTR, and a portion of the 3’UTR of the human GPAM sequence (ENST00000348367) under the control of the human thyroxine binding globulin promoter in an AAV2 backbone packaged in AAV8 capsid (AAV8-TBG-h-GPAM).
  • mice were euthanized on Day 10 after subcutaneous injection and a liver sample from each was collected and placed in RNAlater (ThermoFisher Catalog# AM7020) until processing.
  • Total liver RNA was prepared by homogenizing the liver tissue in homogenization buffer (Maxwell RSC simplyRNA Tissue Kit) using a Percellys 24 tissue homogenizer (Bertin Instruments) set at 5000 rpm for two 10 second cycles.
  • RNA from the lysate was purified on a Maxwell RSC 48 platform (Promega Corporation) according to the manufacturer’s recommendations.
  • Preparation of cDNA was performed using Quanta qScript cDNA SuperMix (VWR, Catalog# 95048-500) according to the manufacturer’s instructions.
  • liver GPAM mRNA The relative levels of liver GPAM mRNA were assessed by RT-qPCR in triplicate on a QuantStudioTM 6 Pro Real-Time PCR System using TaqMan assays for human GPAM (ThermoFisher, assay# Hs01573684_m1), or mouse GPAM (ThermoFisher, assay# Mm01261106_m1), and the mouse housekeeping gene PPIA (ThermoFisher, assay# Mm02342430_g1), and PerfeCTa® qPCR FastMix®, Low ROXTM (VWR, Catalog# 101419-222).
  • mice injected with ETD02009, ETD02011, ETD02012 or ETD02015 had the highest level of human GPAM mRNA knockdown in the liver.
  • ETD usu ETD usu ETD02010 12884 [ETL17]sucgaaggUfCfaCfuacaaugasusu 12991 usCfsaUfuGfuAfgUfgAfcCfuUfcGfasusu ETD02011 12885 [ETL17]suguuAfuuAfGfaAfuguuacgasusu 12992 usCfsgUfaAfcAfuUfcUfaAfuAfaCfasusu ETD02012 12886 [ETL17]saaacuAfuGfGfuuguguccgasusu 129
  • siRNAs with alternative modification patterns of ETD02012 and ETD02011 in mice transfected with AAV8-TBG-h-GPAM [00363]
  • the base sequences of ETD02012 and ETD02011 were synthesized to generate siRNAs (ETD02012, ETD02285-ETD02292 and ETD02011, ETD02301-ETD02309, respectively) with alternative modification patterns.
  • the activities of the siRNAs were assessed using mice transiently expressing human GPAM.
  • the siRNAs contain the GalNAc ligand ETL17 followed by a phosphorothioate linkage at the 5’ end of the sense strand.
  • siRNA sequences that were used are -197- Attorney Docket No.54462-742.601 shown in Table 18, where Nf (e.g. Af, Cf, Gf, Tf, or Uf) is a 2’ fluoro-modified nucleoside, n (e.g. a, c, g, t, or u) is a 2’ O-methyl modified nucleoside, Nm (e.g. Am, Cm, Gm, Tm, or Um) is a 2’-O-(2- methoxyethyl) modified nucleoside, dN (e.g.
  • Nf e.g. Af, Cf, Gf, Tf, or Uf
  • n e.g. a, c, g, t, or u
  • Nm e.g. Am, Cm, Gm, Tm, or Um
  • dN e.g.
  • dA, dC, dG, dT, or dU is a 2’ deoxynucleoside, and “s” is a phosphorothioate linkage.
  • the base sequences for each siRNA, with and without the 3’ UU extension, are shown in Table 19. [00364] Six to eight week old female mice (C57Bl/6) were injected with 5 uL of a recombinant adeno- associated virus 8 (AAV8) vector (1.2 x 10E13 genome copies/mL) by the retroorbital route on Day -14.
  • AAV8 recombinant adeno- associated virus 8
  • the recombinant AAV8 contains the open reading frame, the 5’ UTR, and a portion of the 3’UTR of the human GPAM sequence (ENST00000348367) under the control of the human thyroxine binding globulin promoter in an AAV2 backbone packaged in AAV8 capsid (AAV8-TBG-h-GPAM).
  • AAV8-TBG-h-GPAM AAV8 capsid
  • mice were euthanized on Day 14 after subcutaneous injection and a liver sample from each was collected and placed in RNAlater (ThermoFisher Catalog# AM7020) until processing.
  • Total liver RNA was prepared by homogenizing the liver tissue in homogenization buffer (Maxwell RSC simplyRNA Tissue Kit) using a Percellys 24 tissue homogenizer (Bertin Instruments) set at 5000 rpm for two 10 second cycles. Total RNA from the lysate was purified on a Maxwell RSC 48 platform (Promega Corporation) according to the manufacturer’s recommendations. Preparation of cDNA was performed using Quanta qScript cDNA SuperMix (VWR, Catalog# 95048-500) according to the manufacturer’s instructions.
  • liver GPAM mRNA The relative levels of liver GPAM mRNA were assessed by RT-qPCR in triplicate on a QuantStudioTM 6 Pro Real-Time PCR System using TaqMan assays for human GPAM (ThermoFisher, assay# Hs01573684_m1) and the mouse housekeeping gene PPIA (ThermoFisher, assay# Mm02342430_g1), and PerfeCTa® qPCR FastMix®, Low ROXTM (VWR, Catalog# 101419-222). Mice that gave low liver expression of human GPAM, as defined by having a Ct value of >30, were omitted from further analysis.
  • Example 14 Screening siRNAs with alternative modification patterns of ETD02009 and ETD02015 in mice transfected with AAV8-TBG-h-GPAM [00366]
  • the base sequences of ETD02009 and ETD02015 were synthesized to generate siRNAs (ETD02293-ETD02300, and ETD02015-ETD02318, respectively) with alternative modification patterns.
  • the activities of the siRNAs were assessed using mice transiently expressing human GPAM.
  • the siRNAs contain the GalNAc ligand ETL17 followed by a phosphorothioate linkage at the 5’ end of the sense strand.
  • siRNA sequences that were used are shown in Table 21, where Nf (e.g. Af, Cf, Gf, Tf, or Uf) is a 2’ fluoro-modified nucleoside, n (e.g. a, c, g, t, or u) is a 2’ O-methyl modified nucleoside, Nm (e.g. Am, Cm, Gm, Tm, or Um) is a 2’-O-(2-methoxyethyl) modified nucleoside, dN (e.g.
  • Nf e.g. Af, Cf, Gf, Tf, or Uf
  • n e.g. a, c, g, t, or u
  • Nm e.g. Am, Cm, Gm, Tm, or Um
  • dN e.g.
  • dA, dC, dG, dT, or -200- Attorney Docket No.54462-742.601 dU) is a 2’ deoxynucleoside, and “s” is a phosphorothioate linkage.
  • the base sequences for each siRNA, with and without the 3’ UU extension, are shown in Table 22. [00367] Six to eight week old female mice (C57Bl/6) were injected with 5 uL of a recombinant adeno- associated virus 8 (AAV8) vector (1.2 x 10E13 genome copies/mL) by the retroorbital route on Day -14.
  • AAV8 vector 1.2 x 10E13 genome copies/mL
  • the recombinant AAV8 contains the open reading frame, the 5’ UTR, and a portion of the 3’UTR of the human GPAM sequence (ENST00000348367) under the control of the human thyroxine binding globulin promoter in an AAV2 backbone packaged in AAV8 capsid (AAV8-TBG-h-GPAM).
  • AAV8-TBG-h-GPAM AAV8 capsid
  • mice were euthanized on Day 14 after subcutaneous injection and a liver sample from each was collected and placed in RNAlater (ThermoFisher Catalog# AM7020) until processing.
  • Total liver RNA was prepared by homogenizing the liver tissue in homogenization buffer (Maxwell RSC simplyRNA Tissue Kit) using a Percellys 24 tissue homogenizer (Bertin Instruments) set at 5000 rpm for two 10 second cycles. Total RNA from the lysate was purified on a Maxwell RSC 48 platform (Promega Corporation) according to the manufacturer’s recommendations. Preparation of cDNA was performed using Quanta qScript cDNA SuperMix (VWR, Catalog# 95048-500) according to the manufacturer’s instructions.
  • liver GPAM mRNA The relative levels of liver GPAM mRNA were assessed by RT-qPCR in triplicate on a QuantStudioTM 6 Pro Real-Time PCR System using TaqMan assays for human GPAM (ThermoFisher, assay# Hs01573684_m1) and the mouse housekeeping gene PPIA (ThermoFisher, assay# Mm02342430_g1), and PerfeCTa® qPCR FastMix®, Low ROXTM (VWR, Catalog# 101419-222). Mice that gave low liver expression of human GPAM, as defined by having a Ct value of >30, were omitted from further analysis.
  • siRNAs with alternative modification patterns of ETD02003 and ETD02018 in mice transfected with AAV8-TBG-h-GPAM [00369]
  • the base sequences of ETD02003 and ETD02018 were synthesized to generate siRNAs (ETD02223-ETD02230, and ETD02201-ETD02208, respectively) with alternative modification patterns.
  • the activities of the siRNAs were assessed using mice transiently expressing human GPAM.
  • the siRNAs contain the GalNAc ligand ETL17 followed by a phosphorothioate linkage at the 5’ end of the sense strand.
  • the siRNA sequences that were used are shown in Table 24, where Nf (e.g.
  • Af, Cf, Gf, Tf, or Uf is a 2’ fluoro-modified nucleoside
  • n e.g. a, c, g, t, or u
  • dN e.g. dA, dC, dG, dT, or dU
  • s is a phosphorothioate linkage.
  • Table 25 The base sequences for each siRNA, with and without the 3’ UU extension, are shown in Table 25.
  • AAV8 adeno- associated virus 8 vector
  • the recombinant AAV8 contains the open reading frame, the 5’ UTR, and a portion of the 3’UTR of the human GPAM sequence (ENST00000348367) under the control of the human thyroxine binding globulin -203- Attorney Docket No.54462-742.601 promoter in an AAV2 backbone packaged in AAV8 capsid (AAV8-TBG-h-GPAM).
  • mice were euthanized on Day 14 after subcutaneous injection and a liver sample from each was collected and placed in RNAlater (ThermoFisher Catalog# AM7020) until processing.
  • Total liver RNA was prepared by homogenizing the liver tissue in homogenization buffer (Maxwell RSC simplyRNA Tissue Kit) using a Percellys 24 tissue homogenizer (Bertin Instruments) set at 5000 rpm for two 10 second cycles.
  • RNA from the lysate was purified on a Maxwell RSC 48 platform (Promega Corporation) according to the manufacturer’s recommendations.
  • Preparation of cDNA was performed using Quanta qScript cDNA SuperMix (VWR, Catalog# 95048-500) according to the manufacturer’s instructions.
  • liver GPAM mRNA The relative levels of liver GPAM mRNA were assessed by RT-qPCR in triplicate on a QuantStudioTM 6 Pro Real-Time PCR System using TaqMan assays for human GPAM (ThermoFisher, assay# Hs01573684_m1) or mouse GPAM (ThermoFisher, assay# Mm01261106_m1), and the mouse housekeeping gene PPIA (ThermoFisher, assay# Mm02342430_g1), and PerfeCTa® qPCR FastMix®, Low ROXTM (VWR, Catalog# 101419-222). Data were normalized to the mean GPAM mRNA level in animals receiving a subcutaneous injection of PBS.
  • Results are shown in Table 26.
  • ETD02224 and ETD02228 had the greatest activity in terms of knockdown of the human GPAM mRNA.
  • ETD02202 had the greatest activity in terms of knockdown of the human GPAM mRNA. Table 24.
  • ETD02009 and ETD02015 were synthesized to generate siRNAs (ETD02543-ETD02544, and ETD02539-ETD02542, respectively) with alternative modification patterns including 2’-O-(2-methoxyethyl).
  • the activities of the siRNAs were assessed using mice transiently expressing human GPAM.
  • the siRNAs contain the GalNAc ligand ETL17 followed by a phosphorothioate linkage at the 5’ end of the sense strand.
  • siRNA sequences that were used are shown in Table 27, where Nf (e.g. Af, Cf, Gf, Tf, or Uf) is a 2’ fluoro-modified nucleoside, n (e.g. a, c, g, t, or u) is a 2’ O-methyl modified nucleoside, Nm (e.g. Am, Cm, Gm, Tm, or Um) is a 2’-O-(2- methoxyethyl) modified nucleoside, and “s” is a phosphorothioate linkage.
  • Nf e.g. Af, Cf, Gf, Tf, or Uf
  • n e.g. a, c, g, t, or u
  • Nm e.g. Am, Cm, Gm, Tm, or Um
  • s is a phosphorothioate linkage.
  • Table 28 The base sequences for each siRNA, with and without
  • AAV8 adeno- associated virus 8 vector
  • the recombinant AAV8 contains the open reading frame, the 5’ UTR, and a portion of the 3’UTR of the human GPAM sequence (ENST00000348367) under the control of the human thyroxine binding globulin promoter in an AAV2 backbone packaged in AAV8 capsid (AAV8-TBG-h-GPAM).
  • mice were euthanized on Day 14 after subcutaneous injection and a liver sample from each was collected and placed in RNAlater (ThermoFisher Catalog# AM7020) until processing.
  • Total liver RNA was prepared by homogenizing the liver tissue in homogenization buffer (Maxwell RSC simplyRNA Tissue Kit) using a Percellys 24 tissue homogenizer (Bertin Instruments) set at 5000 rpm for two 10 second cycles.
  • RNA from the lysate was purified on a Maxwell RSC 48 platform (Promega Corporation) according to the manufacturer’s recommendations.
  • Preparation of cDNA was performed using Quanta qScript cDNA SuperMix (VWR, Catalog# 95048-500) according to the manufacturer’s instructions.
  • liver GPAM mRNA The relative levels of liver GPAM mRNA were assessed by RT-qPCR in triplicate on a QuantStudioTM 6 Pro Real-Time PCR System using TaqMan assays for human GPAM (ThermoFisher, assay# Hs00326039_m1) and the mouse housekeeping gene PPIA (ThermoFisher, assay# Mm02342430_g1), and PerfeCTa® qPCR FastMix®, Low ROXTM (VWR, Catalog# 101419-222). Data -206- Attorney Docket No.54462-742.601 were normalized to the mean GPAM mRNA level in animals receiving a subcutaneous injection of PBS. Results are shown in Table 29.
  • ETD02541 had the greatest activity.
  • ETD02009 ETD02543 and ETD2544 had the greatest activity.
  • Table 27 Example siRNA Sequences ETD# SEQ Sense Strand Sequence (5 ⁇ -3 ⁇ ) with GalNAc SEQ Antisense Strand Sequence (5 ⁇ -3 ⁇ ) ID moiety ID ETD susu ETD susu ETD usu ETD usu ETD su ETD su ETD su ETD su Table 28.
  • siRNA BASE Sequences siRNA SEQ ID Sense Strand Base Sequence SEQ ID Antisense Strand Base Sequence N ET UU ET UU ET UU ET UU ET UU ET UU ET UU ET UU ET UU ET UU ET UU ET UU si ce N gs ET U ET A ET U ET U ET U ET U ET U ET U ET A ET A si ce N gs ET U ET A ET U ET U ET U ET U ET U ET U ET U ET U ETD02543 13323 UGGAAUCUCCUAUGAUCG 13644 CGAUCAUAGGAGAUUCCA ETD02544 13324 UGGAAUCUCCUAUGAUCG 13645 CGAUCAUAGGAGAUUCCA Table 29 h-GPAM .
  • mice transfected with AAV8-TBG-h-GPAM [00375] The activities of siRNAs, namely ETD02553-ETD02576, were assessed in mice transiently expressing human GPAM.
  • the siRNAs contain the GalNAc ligand ETL17 followed by a phosphorothioate linkage at the 5’ end of the sense strand.
  • the siRNA sequences that were used are shown in Table 30, where Nf (e.g.
  • Af, Cf, Gf, Tf, or Uf is a 2’ fluoro-modified nucleoside
  • n e.g. a, c, g, t, or u
  • Nm e.g. Am, Cm, Gm, Tm, or Um
  • s is a phosphorothioate linkage.
  • the base sequences for each siRNA, with and without the 3’ UU extension, are shown in Table 31. ETD02553-ETD02664 were tested in Part 1 of the study and ETD02565-ETD02576 were tested in Part 2.
  • AAV8 adeno-associated virus 8 vector
  • the recombinant AAV8 contains the open reading frame, the 5’ UTR, and a portion of the 3’UTR of the human GPAM sequence (ENST00000348367) under the control of the human thyroxine binding globulin promoter in an AAV2 backbone packaged in AAV8 capsid (AAV8-TBG-h-GPAM).
  • mice were euthanized on Day 14 after subcutaneous injection and a liver sample from each was collected and placed in RNAlater (ThermoFisher Catalog# AM7020) until processing.
  • Total liver RNA was prepared by homogenizing the liver tissue in homogenization buffer (Maxwell RSC simplyRNA Tissue Kit) using a Percellys 24 tissue homogenizer (Bertin Instruments) set at 5000 rpm for two 10 second cycles.
  • RNA from the lysate was purified on a Maxwell RSC 48 platform (Promega Corporation) according to the manufacturer’s recommendations.
  • Preparation of cDNA was performed using Quanta qScript cDNA SuperMix (VWR, Catalog# 95048-500) according to the manufacturer’s instructions.
  • liver GPAM mRNA The relative levels of liver GPAM mRNA were assessed by RT-qPCR in triplicate on a QuantStudioTM 6 Pro Real-Time PCR System using TaqMan assays for human GPAM (ThermoFisher, assay# Hs01573684_m1) and the mouse housekeeping gene PPIA (ThermoFisher, assay# Mm02342430_g1), and PerfeCTa® qPCR FastMix®, Low ROXTM (VWR, Catalog# 101419-222). Mice that gave low liver expression of human GPAM in Part 2, as defined by having a Ct value of >32, were omitted from further analysis.
  • ETD# SEQ Sense Strand Sequence (5 ⁇ -3 ⁇ ) with GalNAc SEQ ID Antisense Strand Sequence (5 ⁇ -3 ⁇ ) ID NO: moiety NO: ETD2 124 1 fAf fAf 1 f Af f Af f f f susu ETD susu ETD susu ETD susu ETD susu ETD usu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu ETD susu Table 31.
  • Example 18 Therapeutic siRNA-mediated knockdown of GPAM in a mouse model of non-alcoholic fatty liver disease (NAFLD) [00378] The effects of siRNA-mediated knockdown of GPAM in the liver was investigated in a high fat, high fructose diet-based mouse model of non-alcoholic fatty liver disease (NAFLD).
  • PBS phosphate buffered saline
  • the siRNAs contain the GalNAc ligand ETL17 followed by a phosphorothioate linkage at the 5’ end of the sense strand.
  • the siRNA sequences that were used are shown in Table 34, where Nf (e.g. Af, Cf, Gf, Tf, or Uf) is a 2’ fluoro-modified nucleoside, n (e.g.
  • a, c, g, t, or u is a 2’ O- methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the base sequences for each siRNA, with and without the 3’ UU extension, are shown in Table 35.
  • PBS phosphate buffered saline
  • the [00381] Body weights were recorded biweekly at the start of the Western diet until the end of the study. Mice were fasted 4-6 hours for serum collection prior to the first siRNA treatment and then 1 day prior to each biweekly siRNA injection.
  • Serum was sent for the following clinical chemistry assays (IDEXX Laboratories, Incorporated): ALP, ALT, BUN, cholesterol, glucose, total bilirubin, total protein, triglycerides, and beta-hydroxybutarate. Ketones were measured in a drop of whole blood using Precision Xtra Ketone Test Strips (Abbott). [00382] All mice in Groups 1-4 were euthanized via cervical dislocation following isoflurane exposure with a final serum bleed at 16 weeks and all mice in Groups 5-8 were euthanized via cervical dislocation following isoflurane exposure with a final serum bleed at 24 weeks and the liver was separated -212- Attorney Docket No.54462-742.601 into 4 sections.
  • RNAlater for qRT-PCR (Thermo Fisher #AM7020)
  • one section was snap frozen in liquid nitrogen for liver triglyceride measurement
  • one section was placed into 10% formalin for fixation and paraffin embedding for H&E and Picrosirius Red staining
  • one section was fixed with 10% formalin at 4C for 24 hours and then embedded in OCT for Oil Red O staining.
  • Total liver RNA was prepared by homogenizing the RNAlater liver tissue in homogenization buffer (Maxwell RSC simplyRNA Tissue Kit) using a Percellys 24 tissue homogenizer (Bertin Instruments) set at 5000 rpm for two 10 second cycles.
  • RNA from the lysate was purified on a Maxwell RSC 48 platform (Promega Corporation) according to the manufacturer’s recommendations.
  • Preparation of cDNA was performed using Quanta qScript cDNA SuperMix (VWR, Catalog# 95048-500) according to the manufacturer’s instructions.
  • liver GPAM mRNA were assessed by RT- qPCR in triplicate on a QuantStudioTM 6 Pro Real-Time PCR System using TaqMan assays for mouse GPAM (ThermoFisher, assay# Mm01261106_m1), mouse COL1A1 (ThermoFisher, assay # Mm00801666_g1), mouse TIMP1 (ThermoFisher, assay # Mm00801666_g1), mouse TGFB1 (ThermoFisher, assay # Mm01341361_m1), mouse Acta2/SMA (ThermoFisher, assay # Mm00725412_s1), mouse Tnf- ⁇ (ThermoFisher, assay # Mm00443258_m1), mouse Il1B (ThermoFisher, assay # Mm00434228_m1), mouse CCL2 (ThermoFisher, assay # Mm0000
  • the 16 week GPAM mRNA levels are shown in Table . Data were normalized to the level in animals receiving PBS and on the Western Diet (Group 2). Mice on the Western Diet treated with ETD02282 (Group 3) or ETD02284 (Group 4) had reduced liver GPAM mRNA levels compared to mice on the Western Diet receiving PBS (Group 2). The mice on the Western Diet had elevated levels of GPAM compared to the mice on normal chow.
  • Table 37.24-week GPAM mRNA Levels in Mice Treated with ETD02282 and ETD02284 Relative Liver Mean Relative 40 0.16 [00386] The starting (Day 0) and ending (Day 112) body weights are shown in Table 38 for the 16- week study. There was reduced weight gain in the mice fed the Western Diet and treated with ETD02284 (Group 4) compared to mice on the Western Diet and treated with PBS (Group 2) at 16 weeks.
  • mice Treated with ETD02282 and ETD02284 ody G 39 19.60 39.69 40 18.80 23.56 [00388]
  • the 16-week serum levels of ALT, cholesterol, and ketones are shown in Table 40.
  • the ALT and cholesterol levels at 16 weeks in mice treated with ETD02282 (Group 3) and ETD02284 (Group 4) were lower than mice treated with PBS (Group 2) on Western Diet.
  • the ketone levels in mice at 16 weeks treated with ETD02284 (Group 4) were higher than mice treated with PBS (Group 2) on Western Diet.
  • the % steatotic area was calculated by measuring the white areas left behind by lipid droplets in the tissue divided by the total area of the tissue section. Five different liver serial sections were used per animal for quantification. The 16-week % steatotic area of the H&E stained formalin fixed liver sections is in Table 42. The % steatotic area at 16 weeks was reduced in mice treated with ETD02282 (Group 3) and ETD02284 (Group 4) compared to mice treated with PBS (Group 2) on Western Diet.
  • Table 43.24-week % Steatotic Area of Livers in Mice Treated with ETD02282 and ETD02284 Individual % Mean % Group # Animal # Steatotic Area Steatotic Area [00392] Liver triglycerides were measured in a piece of flash frozen liver using a Triglyceride Colorimetric Kit (Elabscience #E-BC-K238) following the manufacturer’s instructions.
  • mice on normal diet treated with PBS The level of triglycerides in the liver at 16 weeks was reduced in mice treated with ETD02282 (Group 3) and ETD02284 (Group 4) compared to mice treated with PBS (Group 2) on Western Diet.
  • Table 44 16-week Liver Triglyceride Levels in Mice Treated with ETD02282 and ETD02284 l 31 0.61 4 32 0.27 0.79 33 0.56 -218- Attorney Docket No.54462-742.601 34 1.55 35 0.94 [00393] ide Colorimetric Kit (Elabscience #E-BC-K238) following the manufacturer’s instructions.
  • the levels of triglycerides in the flash frozen liver sections are shown in Table 45 with the data normalized to mice on normal diet treated with PBS (Group 5).
  • the level of triglycerides in the liver at 24 weeks was not reduced in mice treated with ETD02282 (Group 7) and ETD02284 (Group 8) compared to mice treated with PBS (Group 6) on Western Diet.
  • Expression of fibrotic and pro-inflammatory genes in the liver were measured by qRT-PCR and normalized to the mice on Western Diet receiving PBS (Group 2).
  • pro-fibrotic genes COL1A1 and TIMP1 and pro-inflammatory genes TNF- ⁇ , IL1b, and CCL2 are shown in Table 46.
  • Mice treated with ETD02282 (Group 3) did not show reduction of any genes compared to PBS treated mice (Group 2) except TIMP1.
  • pro-fibrotic genes COL1A1 and TIMP1 and pro-inflammatory genes TNF- ⁇ , IL1b, and CCL2 are shown in Table 47.
  • Mice treated with either siRNA did not show any significant change in expression of pro-inflammatory markers IL1b or CCL2 at 24 weeks.
  • Preventative siRNA-mediated knockdown of GPAM in a mouse model of non-alcoholic fatty liver disease [00396] The protective effects of siRNA-mediated knockdown of GPAM in the liver was investigated in a high fat, high fructose diet-based mouse model of non-alcoholic fatty liver disease (NAFLD).
  • NASH non-alcoholic fatty liver disease
  • Six- to eight-week-old C57BL/6NHsd female mice (Envigo) were given a Western high fat diet (Envigo TD.120330, 0.2% cholesterol, 45% fat by calories) and high fructose water (55% fructose, 45% glucose).
  • PBS phosphate buffered saline
  • Mice on the 8 week study received a total of 4 siRNA doses and mice on the 16 week study received a total of 16 siRNA doses.
  • the siRNA sequences that were used are shown in Table 48, where Nf (e.g.
  • Af, Cf, Gf, Tf, or Uf is a 2’ fluoro-modified nucleoside
  • n e.g. a, c, g, t, or u
  • s is a phosphorothioate linkage.
  • Table 48 The base sequences for each siRNA, with and without the 3’ UU extension, are shown in Table 49 Table 48.
  • Example siRNA BASE Sequences si ce N ET UU ET UU si ce N gs ET G ET G si ce N gs ET G ETD02284 13402 CAAUAGACGUUUCUUAUC 13723 GAUAAGAAACGUCUAUUG [00398] Body weights were recorded biweekly until the end of the study. Mice were fasted 4-6 hours for serum collection on Day 0 and then 1 day prior to each biweekly siRNA injection. Serum was sent for the following clinical chemistry assays performed at IDEXX Laboratories, Incorporated – ALP, ALT, BUN, cholesterol, glucose, total bilirubin, total protein, triglycerides, and beta-hydroxybutarate.
  • mice in Groups 1-6 were euthanized via cervical dislocation following isoflurane exposure with a final serum bleed at 8 weeks and all mice in Groups 7-12 were euthanized via cervical dislocation following isoflurane exposure with a final serum bleed at 16 weeks and the liver was separated into 4 sections.
  • One section was placed in RNAlater for qRT-PCR (Thermo Fisher #AM7020), one section was snap frozen in liquid nitrogen for liver triglyceride measurement, one section was placed into 10% formalin for fixation and paraffin embedding for H&E and Picrosirius Red staining, and one section was fixed with 10% formalin at 4C for 24 hours and then embedded in OCT for Oil Red O staining.
  • Total liver RNA was prepared by homogenizing the RNAlater liver tissue in homogenization buffer (Maxwell RSC simplyRNA Tissue Kit) using a Percellys 24 tissue homogenizer (Bertin Instruments) set at 5000 rpm for two 10 second cycles. Total RNA from the lysate was purified on a Maxwell RSC 48 platform (Promega Corporation) according to the manufacturer’s recommendations. Preparation of cDNA was performed using Quanta qScript cDNA SuperMix (VWR, Catalog# 95048-500) according to the manufacturer’s instructions.
  • liver GPAM mRNA were assessed by RT- qPCR in triplicate on a QuantStudioTM 6 Pro Real-Time PCR System using TaqMan assays for mouse GPAM (ThermoFisher, assay# Mm01261106_m1), mouse COL1A1 (ThermoFisher, assay # Mm00801666_g1), mouse TIMP1 (ThermoFisher, assay # Mm00801666_g1), mouse TGFB1 (ThermoFisher, assay # Mm01341361_m1), mouse Acta2/SMA (ThermoFisher, assay # Mm00725412_s1), mouse Tnf- ⁇ (ThermoFisher, assay # Mm00443258_m1), mouse Il1B (ThermoFisher, assay # Mm00434228_m1), mouse CCL2 (ThermoFisher, assay # Mm0000
  • the 8 -week GPAM mRNA levels are shown in Table . Data were normalized to the level in animals receiving PBS and on the Western Diet (Group 2). Mice on the Western Diet treated with ETD02282 (Groups 3 and 4) or ETD02284 (Groups 5 and 6) had reduced liver GPAM mRNA levels compared to mice on the Western Diet receiving PBS (Group 2). The mice on the Western Diet had elevated levels of GPAM compared to the mice on normal chow.
  • Mice on the Western Diet treated with ETD02282 (Groups 9 and 10) or ETD02284 (Groups 11 and 12) had reduced liver GPAM mRNA levels compared to mice on the Western Diet receiving PBS (Group 8).
  • the mice on the Western Diet had elevated levels of GPAM compared to the mice on normal chow.
  • Table 51 16-week GPAM mRNA Levels in Mice Treated with ETD02282 and ETD02284 Relative Liver GPAM Mean Relative Liver GPAM G 59 0.09 60 0.08 [00403]
  • the starting (Day 0) and ending (Day 56) body weights are shown in Table 52 for the 8-week study. There was reduced weight gain in the mice fed the Western Diet and treated with both doses of ETD02284 (Groups 5 and 6) compared to mice on the Western Diet and treated with PBS (Group 2) at 8 weeks.
  • the 8-week serum levels of ALT, cholesterol, and ketones are shown in Table 54.
  • mice treated with both doses of ETD02282 (Groups 3 and 4) and ETD02284 (Groups 5 and 6) were lower than mice treated with PBS (Group 2) on Western Diet.
  • the serum cholesterol levels at 8 weeks in mice treated with either ETD02282 (Groups 3 and 4) or ETD02284 (Groups 5 and 6) on Western Diet were not significantly decreased compared to mice treated with PBS (Group 2) on Western Diet.
  • mice at 8 weeks treated with both doses of ETD02284 (Groups 5 and 6) on Western Diet were higher than mice treated with PBS (Group 2) on Western Diet while the mice treated with both doses of ETD02282 (Groups 3 and 4) did not show an increase in ketones.
  • mice treated with either ETD02282 (Groups 9 and 10) or ETD02284 (Groups 11 and 12) on Western Diet increased compared to mice treated with PBS (Group 8) on Western Diet.
  • the (BHB) serum levels at 16 weeks were not increased with either siRNA treatment or dosage.
  • liver sections were formalin fixed and paraffin embedded then stained with hematoxylin and eosin (H&E).
  • H&E hematoxylin and eosin
  • the % steatotic area was calculated by measuring the white areas left behind by lipid droplets in the tissue divided by the total area of the tissue section. Five different liver serial sections were used per animal for quantification. The 16-week % steatotic area of the H&E stained formalin fixed liver sections is in Table 57.
  • Liver triglycerides were measured in a piece of flash frozen liver using a Triglyceride Colorimetric Kit (Elabscience #E-BC-K238) following the manufacturer’s instructions.
  • the levels of triglycerides in the flash frozen liver sections are shown in Table 59 with the data normalized to mice on normal diet treated with PBS (Group 7).
  • the level of triglycerides in the liver at 16 weeks were not significantly reduced in mice treated with either dose of ETD02282 (Groups 9 and 10) and 200 ug -230- Attorney Docket No.54462-742.601 ETD02284 (Group 11) compared to mice treated with PBS (Group 8) on Western Diet.
  • the samples from the 60 ug dose of ETD02284 (Group 12) could not be successfully processed for this experiment.
  • mice on Western Diet treated with 60 ug of ETD02282 had reduced expression of pro-fibrotic genes COL1A1 and TIMP1 and reduced expression of CCL2 compared to mice treated with PBS (Group 2).
  • mice on the Western Diet for 16 weeks and treated with 200 ug (Group 9) or 60 ug (Group 10) of -232- Attorney Docket No.54462-742.601 ETD02282 or 200 ug of ETD02284 (Group 11) did not show any significant reduction in expression of pro-fibrotic or pro-inflammatory genes compared to mice treated with PBS (Group 8).
  • the mice treated with 60 ug of ETD02284 (Group 12) showed significant reduction of all pro-fibrotic and pro- inflammatory genes measured at 16 weeks compared to mice on PBS (Group 8).
  • Nf e.g. Af, Cf, Gf, Tf, or Uf
  • n e.g.
  • a, c, g, t, or u is a 2’ O-methyl modified nucleoside, and “s” is a phosphorothioate linkage.
  • the base sequences for each siRNA, with and without the 3’ UU extension, are shown in Table 63.
  • RNAlaterTM Stabilization Solution (Thermo Fisher, Catalog# AM7020) in 20 seconds and stored for 24 hours at 4°C. The RNAlater was then removed and the liver tissue was stored in the freezer until they were shipped to Empirico. [00416] Total liver RNA was prepared by homogenizing the RNAlater liver tissue in homogenization buffer (Maxwell RSC simplyRNA Tissue Kit) using a Percellys 24 tissue homogenizer (Bertin Instruments) set at 5000 rpm for two 10 second cycles. Total RNA from the lysate was purified on a Maxwell RSC 48 platform (Promega Corporation) according to the manufacturer’s recommendations.
  • cDNA was prepared using Quanta qScript cDNA SuperMix (VWR, Catalog# 95048-500) according to the manufacturer’s instructions.
  • the levels of liver GPAM mRNA were assessed by RT- qPCR in triplicate on a QuantStudioTM 6 Pro Real-Time PCR System using TaqMan assays for monkey -234- Attorney Docket No.54462-742.601 GPAM (ThermoFisher, assay# Mf02878271_m1), and the monkey housekeeping gene GAPDH (ThermoFisher, assay# Mf04392546_g1) using PerfeCTa® qPCR FastMix®, Low ROXTM (VWR, Catalog# 101419-222).

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Abstract

Les troubles métaboliques tels que des troubles hépatiques et des troubles cardiovasculaires sont largement répandus, et peuvent toucher une grande diversité de gens. Des thérapeutiques améliorés sont requis pour traiter ces troubles. L'invention concerne des compositions comprenant un oligonucléotide qui cible le GPAM. L'oligonucléotide peut comprendre un petit ARN interférent (ARNsi) ou un oligonucléotide antisens (ASO). L'invention concerne également des méthodes de traitement d'états associés à des mutations du gène GPAM qui comprennent l'administration d'un oligonucléotide ciblant le GPAM chez un sujet. Certains exemples de maladies qui peuvent être traitées comprennent des maladies hépatiques ou des maladies cardiométaboliques.
EP23827986.3A 2022-06-21 2023-06-20 Traitement de maladies et de troubles liés au gpam Pending EP4544052A1 (fr)

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PCT/US2023/068741 WO2023250327A1 (fr) 2022-06-21 2023-06-20 Traitement de maladies et de troubles liés au gpam

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WO2025137167A2 (fr) * 2023-12-20 2025-06-26 Empirico Inc. Traitement de maladies et de troubles liés au gpam

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CA2685127C (fr) * 2007-04-23 2019-01-08 Alnylam Pharmaceuticals, Inc. Glycoconjugues d'agents d'interference arn
WO2021252649A2 (fr) * 2020-06-09 2021-12-16 Alnylam Pharmaceuticals, Inc. Compositions de petit arn interférent et procédés de silençage de l'expression de la gpam (glycérol-3-phosphate acyltransférase 1, mitochondriale)

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