WO2026006830A1 - Compositions et méthodes de traitement de maladies et de troubles caractérisés par une détérioration de la barrière hémato-encéphalique ou la génération d'espèces réactives de l'oxygène - Google Patents

Compositions et méthodes de traitement de maladies et de troubles caractérisés par une détérioration de la barrière hémato-encéphalique ou la génération d'espèces réactives de l'oxygène

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Publication number
WO2026006830A1
WO2026006830A1 PCT/US2025/035911 US2025035911W WO2026006830A1 WO 2026006830 A1 WO2026006830 A1 WO 2026006830A1 US 2025035911 W US2025035911 W US 2025035911W WO 2026006830 A1 WO2026006830 A1 WO 2026006830A1
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WO
WIPO (PCT)
Prior art keywords
pgdh
alkyl
subject
alkylene
brain
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Pending
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PCT/US2025/035911
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English (en)
Inventor
Sanford Markowitz
Andrew Pieper
Min-Kyoo SHIN
Yeojung Koh
Edwin VAZQUEZ-ROSA
Farrah GAO
Hongyun LI
Dawn Dawson
Ramachandra KATABATHULA
Salendra Singh
Bindu PAUL
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Case Western Reserve University
Johns Hopkins University
US Department of Veterans Affairs
University Hospitals Cleveland Medical Center
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Case Western Reserve University
Johns Hopkins University
US Department of Veterans Affairs
University Hospitals Cleveland Medical Center
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Application filed by Case Western Reserve University, Johns Hopkins University, US Department of Veterans Affairs, University Hospitals Cleveland Medical Center filed Critical Case Western Reserve University
Publication of WO2026006830A1 publication Critical patent/WO2026006830A1/fr
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Anticipated expiration legal-status Critical

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Classifications

    • 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
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/519—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim ortho- or peri-condensed with heterocyclic rings
    • 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
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/4353—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom ortho- or peri-condensed with heterocyclic ring systems
    • A61K31/4365—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom ortho- or peri-condensed with heterocyclic ring systems the heterocyclic ring system having sulfur as a ring hetero atom, e.g. ticlopidine

Definitions

  • a prominent and early feature of Alzheimer’s Disease (AD) is deterioration in blood-brain barrier (BBB) structure and function, which impairs neurovascular coupling and increases permeability of blood-borne substances and peripheral immune cells into the brain parenchyma, triggering perivascular inflammation.
  • BBB blood-brain barrier
  • BBB dysfunction occurs in the earliest pathological stages of AD and has been proposed as an early biomarker for the disease. Aging further contributes to BBB deterioration, especially in the hippocampus, a region essential for learning and memory that is particularly vulnerable in AD.
  • TBI traumatic brain injury
  • Common causes of TBI include falls, motor vehicle accidents, explosions, interpersonal violence, and sports injuries.
  • Embodiments described herein relate to compositions and methods of treating diseases or disorders characterized by structural deterioration of the blood-brain barrier and/or by increased generation of reactive oxygen species within the nervous system of a subject in need thereof.
  • 15-hydroxyprostaglandin dehydrogenase 15-hydroxyprostaglandin dehydrogenase
  • BBB blood brain barrier
  • 15-PGDH 15-hydroxyprostaglandin dehydrogenase
  • AD Alzheimer’s disease
  • TBI traumatic brain injury
  • aging 15-hydroxyprostaglandin dehydrogenase
  • Pathological increase in 15-PGDH correlates with pronounced oxidative stress, neuroinflammation, and neurodegeneration, alongside profound BBB structural degeneration characterized by astrocytic endfeet swelling and functional impairment.
  • Pharmacologic inhibition or genetic reduction of 15-PGDH in AD and TBI models strikingly mitigates cognitive decline, oxidative damage, suppresses neuroinflammation, and restores BBB integrity.
  • inhibiting 15-PGDH not only blocks damage to the brain by blocking microglial generation of reactive oxygen in 15-PGDH positive microglial cells and halts neurodegeneration but also preserves cognitive function at levels indistinguishable from healthy controls.
  • these neuroprotective effects in AD are achieved without affecting amyloid pathology, underscoring a non-canonical mechanism for treating AD.
  • our findings position 15-PGDH inhibition as a novel and broad-spectrum strategy to treat oxidative stress, neuroinflammation, and neurodegeneration, alongside profound BBB structural degeneration, associated with pathological increases in 15-PGDH.
  • a method of treating diseases or disorders characterized by structural deterioration of the blood-brain barrier in a subject in need thereof can include administering to the subject an amount of a 15-PGDH inhibitor and/or 15-PGDH substrate effective to prevent the onset of deterioration and/or the progression of further deterioration of the blood-brain barrier and/or prevent the neurodegeneration associated with blood-brain barrier deterioration and/or prevent the impaired neuronal function associated with blood- brain barrier deterioration and/or restore the integrity and/or structure and/or function of the blood-brain barrier.
  • a method of treating diseases or disorders characterized by increased generation of reactive oxygen species within the nervous system, and particularly, central nervous system, of a subject in need thereof can include administering to the subject an amount of a 15-PGDH inhibitor and/or 15-PGDH substrate effective to prevent and/or decrease reactive oxygen species generation and/or neurodegeneration and/or impaired neuronal function associated with reactive oxygen species.
  • the subject has memory loss and/or cognitive decline and the 15-PGDH inhibitor and/or 15-PGDH substrate is administered at an amount effective to improve memory and/or cognition.
  • the 15-PGDH inhibitor and/or 15-PGDH substrate is administered to the subject at an amount effective to block aberrantly increased 15-PGDH enzymatic activity and prevent perivascular inflammation.
  • the subject is asymptomatic for Alzheimer’s dementia, has preclinical Alzheimer’s disease, or prodromal Alzheimer’s disease.
  • the 15-PGDH inhibitor and/or 15-PGDH substrate is administered to the subject at an amount effective to decrease 4-hydroxy-2-nonenal (4-HNE) and/or 3-nitrotyrosine (3-NT) levels by at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30% at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75% or more.
  • the 15-PGDH inhibitor is a small molecule that binds to 15-PGDH protein.
  • the 15-PGDH inhibitor can inhibit enzymatic activity of recombinant 15-PGDH at an IC 50 of less than 1 ⁇ M, or preferably at an IC 50 of less than 250 nM, or more preferably at an IC 50 of less than 50 nM, or more preferably at an IC 50 of less than 10 nM, or more preferably at an IC 50 of less than 5 nM at a recombinant 15-PGDH concentration of about 5 nM to about 10 nM.
  • the 15-PGDH inhibitor has the following formula (V): (V) or a pharmaceutically acceptable salt, tautomer, or solvate thereof; wherein n is 0-2 X 6 is independently is N or CR c R 1 , R 6 , R 7 , and R c are the same or different each independently hydrogen or a substituted or unsubstituted group selected from C1-C24 alkyl, C2-C24 alkenyl, C2-C24 alkynyl, C 3 -C 20 aryl, heteroaryl, heterocycloalkenyl containing from 5-6 ring atoms, C 6 -C 24 alkaryl, C6-C24 aralkyl, halo, -Si(C1-C3 alkyl)3, hydroxyl, sulfhydryl, C1-C24 alkoxy, C2-C24 alkenyloxy, C 2 -C 24 alkynyloxy, C 5 -C 20
  • the 15-PGDH inhibitor is a molecule or generates a molecule that binds to 15-PGDH mRNA to promote degradation 15-PGDH mRNA and/or inhibit expression of 15-PGDH.
  • the molecule can include an RNA interference (RNAi) molecule, preferably at least one of a small interfering RNA (siRNA) or microRNA (miRNA).
  • RNAi RNA interference
  • the 15-PGDH inhibitor can include an agent or method of gene editing that inactivates or alters the HPGD gene that encodes 15-PGDH.
  • the 15-PGDH inhibitor can include at least one nuclease that can remove, cut, or alter a target nucleic acid sequence of the HPGD gene.
  • the 15-PGDH inhibitor is an agent that promotes the degradation of 15-PGDH protein.
  • the agent can include at least one of a small molecule-assisted shutoff (SMASh) system, a Shield-1-inducible degron, an auxin inducible degron, an IMid-inducible degron, a peptidic degron, a proteolysis targeting chimera, or an antibody for targeted 15-PGDH degradation.
  • the 15-PGDH substrate can include at least one of PGE2, PGF2 ⁇ , RvD1, LXA4, or 15-HETE.
  • the method can include measuring at least one biomarker of reactive oxygen species generation in the brain of the subject and comparing the at least one biomarker of reactive oxygen species generation to a control to determine the efficacy of the 15-PGDH inhibitor and/or PGDH substrate in treating neurodegeneration and/or impaired neuronal function in the subject.
  • the method can include measuring 4-hydroxy- 2-nonenal (4-HNE) and/or 3-nitrotyrosine (3-NT) levels of the subject following administration of the 15-PGDH inhibitor and/or 15-PGDH substrate and comparing the measured 4-hydroxy-2-nonenal (4-HNE) and/or 3-nitrotyrosine (3-NT) levels to a control, wherein a decrease in 4-hydroxy-2-nonenal (4-HNE) and/or 3-nitrotyrosine (3-NT) levels is indicative of efficacy of the 15-PGDH inhibitor and/or PGDH substrate in treating neurodegeneration and/or impaired neuronal function in the subject.
  • the control is the 4-hydroxy-2-nonenal (4-HNE) and/or 3-nitrotyrosine (3-NT) levels of the subject prior to administration of the 15-PGDH inhibitor and/or 15-PGDH substrate.
  • the method can include measuring blood-brain barrier (BBB) permeability of the subject following administration of the 15-PGDH inhibitor and/or 15-PGDH substrate and comparing the measured BBB permeability to a control, wherein a decrease in BBB permeability is indicative of efficacy of the 15-PGDH inhibitor and/or PGDH substrate in treating neurodegeneration and/or impaired neuronal function in the subject.
  • BBB blood-brain barrier
  • the control is the BBB permeability of the subject prior to administration of the 15-PGDH inhibitor and/or 15-PGDH substrate.
  • BRIEF DESCRIPTION OF THE DRAWINGS [0023] Figs.1(A-J) illustrate brain 15-PGDH increases in human and mouse AD, TBI, and aging, and is enriched in brain myeloid cells.
  • Figs.1(A-J) illustrate brain 15-PGDH increases in human and mouse AD, TBI, and aging, and is enriched in brain myeloid cells.
  • A qPCR data from human cortex reveals that 15-PGDH mRNA (HPGD expression) is increased in subjects with Alzheimer’s disease (AD), compared to control subjects without dementia. HPGD was normalized to the geometric mean of ACTB, POLR1B, and LDHA (* p ⁇ 0.05, unpaired t-test).
  • ERP015139 reveals that 15-PGDH mRNA (HPGD expression) is increased in subjects with chronic traumatic encephalopathy (CTE), compared to control subjects (* p ⁇ 0.05, unpaired t-test). Data are shown from both superior parietal cortex and posterior visual cortex.
  • E Mouse hippocampal 15-PGDH enzymatic activity is increased 3 weeks after TBI, relative to sham-injury (**** p ⁇ 0.0001, unpaired t-test).
  • FISH Fluorescent in situ hybridization
  • Hpgd expressing myeloid cells are composed of Mrc1 positive PVMs and Tmem119 positive microglia.
  • a heatmap of cortical myeloid cells depicting expression (log2(SCT)) of 15-PGDH mRNA (Hpgd), Mrc1 (perivascular macrophages), Lyve1 (perivascular macrophages), Aif1 (microglia and macrophages), and Tmem119 (microglia) is shown.
  • RNA FISH in mouse brain shows an increase in Hpgd-expressing microglia [double positive for 15-PGDH mRNA, Hpgd, (red) and Tmem119 mRNA (green)], in symptomatic 5xFAD mice (6 months old), compared to WT littermates, in cortex and hippocampus (** p ⁇ 0.01, unpaired t-test; 4-8 brain sections/mouse).
  • RNA FISH shows an increase in Hpgd-expressing PVMs [double positive for 15-PGDH mRNA, Hpgd (red) and Mrc1 mRNA (green)] in symptomatic 5xFAD mice (6 months old), compared to WT littermates, in cortex and hippocampus (* p ⁇ 0.05, ** p ⁇ 0.01, unpaired t-test; 2-7 brain sections/mouse). Expanded view of boxed region shown at lower right. In all graphs (A-J), datapoints indicate values of individual human subjects or individual mice. [0024] Figs.2(A-L) illustrate 15-PGDH inhibition protects 5xFAD mice from cognitive impairment and BBB deterioration without reducing amyloid pathology.
  • Tg-WT and Tg-5xFAD refer to 5xFAD genotypes, respectively designating absence or presence of the 5xFAD transgene, and Hpgd +/+ and Hpgd +/- denote Hpgd genotype.
  • Mice were injected with a bolus of BrdU (150 mg/kg) 1 month before sacrifice to label newborn hippocampal neurons. MWM testing of cognition was conducted one week before brain tissue was harvested for analysis at nine months of age.
  • Hpgd haploinsufficiency fully blocks the increase in 15-PGDH enzyme activity in symptomatic 5xFAD mice.
  • Brain 15-PGDH enzyme activity is highly elevated in Tg-5xFAD Hpgd +/+ mice compared to control Tg-WT Hpgd +/+ mice and is preserved at control levels in Tg-5xFAD Hpgd +/- mice, with no difference detected between Tg-WT Hpgd +/+ and Tg-5xFAD Hpgd +/- mice (** p ⁇ 0.01, one-way ANOVA and Tukey’s post hoc analysis).
  • Hpgd haploinsufficiency prevents cognitive impairment in 5xFAD mice in the MWM.
  • Tg-5xFAD Hpgd +/+ mice performed significantly worse than control Tg-WT Hpgd +/+ mice, as shown by significantly longer latency to first cross the platform area.
  • Tg-5xFAD Hpgd +/- mice were protected from memory deficits, as evidenced by latency to first cross the platform area in the memory test being significantly lower than Tg-5xFAD Hpgd +/+ mice and not significantly different from Tg-WT Hpgd +/+ mice (Interaction p ⁇ 0.05, * p ⁇ 0.05, **** p ⁇ 0.0001, two-way ANOVA and Tukey’s post hoc analysis).
  • TEM Transmission electron microscopy
  • (J) TEM shows structural damage of the BBB, as measured by the percentage of brain capillaries affected by astrocyte endfeet swelling in Tg-5xFAD Hpgd +/+ mice as compared to control Tg-WT Hpgd +/+ mice.
  • (L) IgG staining shows functional impairment and increased permeability of the BBB as evidenced by IgG infiltration into the brain parenchyma in Tg- 5xFAD Hpgd +/+ mice, compared to control Tg-WT Hpgd +/+ mice.
  • FIG.3(A-J) illustrate 15-PGDH inhibition protects 5xFAD mice from decreased survival of newborn hippocampal neurons, neuroinflammation, and oxidative stress.
  • FIG.3(A-J) illustrate 15-PGDH inhibition protects 5xFAD mice from decreased survival of newborn hippocampal neurons, neuroinflammation, and oxidative stress.
  • FIG.3(A-J) illustrate 15-PGDH inhibition protects 5xFAD mice from decreased survival of newborn hippocampal neurons, neuroinflammation, and oxidative stress.
  • FIG.3(A-J) illustrate 15-PGDH inhibition protects 5xFAD mice from decreased survival of newborn hippocampal neurons, neuroinflammation, and oxidative stress.
  • A 6- month-old symptomatic 5xFAD mice have impaired survival of BrdU-labeled newborn hippocampal neurons (black arrows), compared to WT littermates, as assessed 28 days post- BrdU injection.
  • Each graphed dot represents the average value of BrdU-labelled cells from an individual mouse as determined from 5-6 sections spaced at least 400 ⁇ m apart across the hippocampus.
  • Tg- 5xFAD Hpgd +/+ mice have impaired survival of BrdU-labeled newborn hippocampal neurons (black arrows), versus Tg-WT Hpgd +/+ littermates, as assessed 28 days post-BrdU injection.
  • (C) 5xFAD mice show increased hippocampal GFAP expression, which is significantly reduced by treatment with (+)-SW033291 (Interaction p ⁇ 0.001, ** p ⁇ 0.01, **** p ⁇ 0.0001, two-way ANOVA and Tukey’s post hoc analysis).
  • (D) Tg-5xFAD Hpgd +/+ mice show increased whole brain GFAP expression compared to Tg-WT Hpgd +/+ mice, which is significantly prevented in Tg-5xFAD Hpgd +/- mice (** p ⁇ 0.01, *** p ⁇ 0.001, one-way ANOVA and Tukey’s post hoc analysis).
  • FIGs.4(A-K) illustrate 15-PGDH inhibition protects mice from TBI-induced cognitive impairment, oxidative stress, neurodegeneration, and BBB damage.
  • FIG. 1 Schematic diagram of experimental procedure for evaluating the protective efficacy of genetic 15-PGDH inhibition in TBI.
  • Control 8-week-old Hpgd +/+ (WT) and Hpgd -/- (KO) mice were subjected to multimodal TBI. Brain tissue was harvested for analysis 3 weeks after TBI.
  • FIGs.5(A-E) illustrate Brain 15-PGDH is elevated in human and rodent Alzheimer’s disease.
  • A RNA sequencing data from the Allen brain atlas shows higher 15- PGDH mRNA (HPGD expression, 203913_s_at (ID_Ref)) in the hippocampus of human Alzheimer’s disease (AD) subjects, compared to no dementia matched control subjects (* p ⁇ 0.05, unpaired t-test).
  • B 15-PGDH whole brain enzymatic activity is increased in symptomatic TgF344-AD rats (6 months old), compared to WT littermates (* p ⁇ 0.05, unpaired t-test).
  • FIG. 6(A-G) illustrate Brain 15-PGDH expression is enriched in PVMs and microglia.
  • A UMAP depiction of single cell RNA sequencing data reanalyzed from Yao et al. reveals 15-PGDH mRNA (Hpgd expression) is most highly enriched in a brain PVM plus microglia cell cluster (bold arrow), along with lesser enrichment in a minority neuronal subpopulation (thin arrow). Increased Hpgd expression level is indicated by increased dot size and color intensity.
  • FIG. 7 The mean log2(SCT) expression level of Hpgd in each cluster of the hippocampus shows that the PVM/microglia population has the highest mean expression of Hpgd, followed by neurons in CA3 and IG-FC.
  • Figs.7(A-C) illustrate 15-PGDH expressing myeloid cells are proximal to blood vessels.
  • A Mouse brain 15-PGDH enzymatic activity in WT mice (6 months old) is enriched ⁇ 200-fold in CD11b (+) myeloid cells, compared to CD11b (-) non-myeloid cells (* p ⁇ 0.05, paired t-test). Data points indicate values of individual mice.
  • RNA FISH for Hpgd and Itgam combined with CD31 immunofluorescence (IF) in mouse brain shows 15- PGDH positive brain myeloid cells (Hpgd+ Itgam+ cells) close to blood vessel (CD31) in cortex and hippocampus of 6-month-old WT mouse. Histogram shows the number of Hpgd and Itgam double positive cells in each distance category from the nearest CD31 marked cell.
  • a total of 12 images per brain section (6 from the cortex, 6 from the hippocampus) were analyzed. Two brain sections from each mouse were used, with a total of 6 WT mice included in the study.
  • FIG.8(A-J) illustrate Pharmacokinetics of (+)-SW033291 in the plasma and the brain; 15-PGDH inhibition protects 5xFAD mice from cognitive impairment, with similar swim speed in Morris water maze (MWM) across all groups.
  • (+)-SW033291 fully prevents cognitive deficits in 5XFAD mice in the MWM, in which vehicle-treated 5xFAD mice performed significantly worse than vehicle-treated WT littermates, as evidenced by significantly fewer platform area crossings. This was preserved at normal WT littermate vehicle levels in 5xFAD mice treated with (+)-SW033291 (Treatment (vehicle vs. (+)- SW033291) p ⁇ 0.01, * p ⁇ 0.05, two-way ANOVA and Tukey’s post hoc analysis).
  • Tg- 5xFAD Hpgd +/+ mice performed significantly worse than control Tg-WT Hpgd +/+ mice, as evidenced by significantly fewer platform area crossings.
  • Hpgd haploinsufficient Tg-5xFAD Hpgd +/- mice were protected from memory deficit, as evidenced by a normal level of platform area crossings.
  • Tg-5xFAD Hpgd +/+ mice displayed learning deficit during the training phase, as evidenced by longer time to find the platform (escape latency) than the other groups on days 2 and 3 of training, which was significantly protected by Hpgd haploinsufficiency in Tg-5xFAD Hpgd +/+ mice.
  • Tg-5xFAD Hpgd +/- mice performed comparably to Tg-WT Hpgd +/+ and Tg-WT Hpgd +/- mice (Time x group Interaction p ⁇ 0.001, ** p ⁇ 0.01, **** p ⁇ 0.0001, repeated measures two-way ANOVA and Tukey’s post hoc analysis).
  • Tg-5xFAD mice showed significant difference in swim speed versus Tg-WT mice (Genotype (Tg-WT vs Tg- 5xFAD) p ⁇ 0.05, two-way ANOVA and Tukey’s post hoc analysis; no difference between other groups were observed, including no difference in swim speed as a function of haploinsuffiency of Hpgd).
  • Tg-5xFAD Hpgd +/+ and Tg-5xFAD Hpgd +/- mice weighed significantly less compared to Tg-WT Hpgd +/+ and Tg-WT Hpgd +/- mice (Genotype (Tg-WT vs Tg-5xFAD) p ⁇ 0.01, two-way ANOVA and Tukey’s post hoc analysis). Males are shown as filled circles or filled squares and females are shown as open circles or open squares, here and in all subsequent panels.
  • FIG. 10(A-E) illustrate pharmacologic or genetic 15-PGDH inhibition increases survival of newborn hippocampal neurons.
  • Hpgd -/- Genetic inhibition of 15-PGDH increases survival of BrdU-labeled newborn hippocampal neurons (black arrows) in male and female adult mice, assayed 15 days post-BrdU injection (Genotype p ⁇ 0.0001, **** p ⁇ 0.0001, two-way ANOVA and Tukey’s post hoc analysis). No difference was detected between males and females.
  • (+)-SW033291 inhibition of 15-PGDH increases survival of BrdU-labeled newborn hippocampal neurons (black arrows) in adult wild type (Hpgd +/+ ) mice.
  • Hpgd -/- mice show increased survival of BrdU-labeled new hippocampal neurons (versus control Hpgd +/+ ), but do not show any additional response to (+)-SW033291 treatment, confirming that (+)-SW033291 potentiates the net magnitude of hippocampal neurogenesis by an on-target effect of inhibiting 15-PGDH.
  • BrdU-labeled cells were quantified 28 days post-BrdU injection.
  • FIG.3C Western blot of GFAP protein, quantified in Fig.3C, with each lane on the gel representing an individual mouse.
  • a to B 5xFAD mice also show characteristic over-expression of APP and induction of IBA1, neither of which are prevented by treatment with (+)-SW033291 (Genotype p ⁇ 0.0001, ** p ⁇ 0.01, two-way ANOVA and Tukey’s post hoc analysis).
  • C Western blot of GFAP protein, quantified in Fig.3D, with each lane on the gel representing an individual mouse.
  • Tg-5xFAD Hpgd +/+ mice also show overexpression of APP and IBA1, as compared to control Tg-WT Hpgd +/+ mice, and neither of these are prevented in Tg-5xFAD Hpgd +/- mice. (* p ⁇ 0.05, one-way ANOVA and Tukey’s post hoc analysis). In all graphs (A-D), data points indicate values of individual mice.
  • Figs.12(A-C) illustrate A ⁇ oligomer-induced generation of reactive oxygen species (ROS) in BV2 microglia cells is suppressed by treatment with 15-PGDH substrates.
  • ROS reactive oxygen species
  • compositions are described as having, including, or comprising, specific components, it is contemplated that compositions also consist essentially of, or consist of, the recited components.
  • methods or processes are described as having, including, or comprising specific process steps, the processes also consist essentially of, or consist of, the recited processing steps.
  • order of steps or order for performing certain actions is immaterial so long as the compositions and methods described herein remains operable. Moreover, two or more steps or actions can be conducted simultaneously.
  • pharmaceutically acceptable means suitable for use in contact with the tissues of humans and animals without undue toxicity, irritation, allergic response, and the like, commensurate with a reasonable benefit/risk ratio, and effective for their intended use within the scope of sound medical judgment.
  • salts include those obtained by reacting the active compound functioning as a base, with an inorganic or organic acid to form a salt, for example, salts of hydrochloric acid, sulfuric acid, phosphoric acid, methanesulfonic acid, camphorsulfonic acid, oxalic acid, maleic acid, succinic acid, citric acid, formic acid, hydrobromic acid, benzoic acid, tartaric acid, fumaric acid, salicylic acid, mandelic acid, carbonic acid, etc.
  • acid addition salts may be prepared by reaction of the compounds with the appropriate inorganic or organic acid via any of a number of known methods.
  • salts also includes those obtained by reacting the active compound functioning as an acid, with an inorganic or organic base to form a salt, for example salts of ethylenediamine, N-methyl- glucamine, lysine, arginine, ornithine, choline, N,N'-dibenzylethylenediamine, chloroprocaine, diethanolamine, procaine, N-benzylphenethylamine, diethylamine, piperazine, tris-(hydroxymethyl)-aminomethane, tetramethylammonium hydroxide, triethylamine, dibenzylamine, ephenamine, dehydroabietylamine, N-ethylpiperidine, benzylamine, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, ethylamine, basic amino acids, and the like.
  • Non limiting examples of inorganic or metal salts include lithium, sodium, calcium, potassium, magnesium salts and the like.
  • the salts of the compounds described herein can exist in either hydrated or unhydrated (the anhydrous) form or as solvates with other solvent molecules.
  • Non-limiting examples of hydrates include monohydrates, dihydrates, etc.
  • Nonlimiting examples of solvates include ethanol solvates, acetone solvates, etc.
  • solvates means solvent addition forms that contain either stoichiometric or non-stoichiometric amounts of solvent. Some compounds have a tendency to trap a fixed molar ratio of solvent molecules in the crystalline solid state, thus forming a solvate.
  • the solvent is water the solvate formed is a hydrate
  • the solvate formed is an alcoholate. Hydrates are formed by the combination of one or more molecules of water with one of the substances in which the water retains its molecular state as H 2 O, such combination being able to form one or more hydrate.
  • the compounds and salts described herein can exist in several tautomeric forms, including the enol and imine form, and the keto and enamine form and geometric isomers and mixtures thereof. Tautomers exist as mixtures of a tautomeric set in solution. In solid form, usually one tautomer predominates. Even though one tautomer may be described, the present application includes all tautomers of the present compounds.
  • a tautomer is one of two or more structural isomers that exist in equilibrium and are readily converted from one isomeric form to another. This reaction results in the formal migration of a hydrogen atom accompanied by a switch of adjacent conjugated double bonds. In solutions where tautomerization is possible, a chemical equilibrium of the tautomers will be reached. The exact ratio of the tautomers depends on several factors, including temperature, solvent, and pH. The concept of tautomers that are interconvertable by tautomerizations is called tautomerism. [0047] Of the various types of tautomerism that are possible, two are commonly observed. In keto-enol tautomerism a simultaneous shift of electrons and a hydrogen atom occurs.
  • Tautomerizations can be catalyzed by: Base: 1. deprotonation; 2. formation of a delocalized anion (e.g., an enolate); 3. protonation at a different position of the anion; Acid: 1. protonation; 2. formation of a delocalized cation; 3. deprotonation at a different position adjacent to the cation.
  • Base 1. deprotonation; 2. formation of a delocalized anion (e.g., an enolate); 3. protonation at a different position of the anion
  • Acid 1. protonation; 2. formation of a delocalized cation; 3. deprotonation at a different position adjacent to the cation.
  • Amino refers to the -NH2 radical.
  • Cyano refers to the -CN radical.
  • Halo or halogen refers to bromo, chloro, fluoro or iodo radical.
  • An alkyl comprising up to 12 carbon atoms is a C1-C12 alkyl
  • an alkyl comprising up to 10 carbon atoms is a C 1 -C 10 alkyl
  • an alkyl comprising up to 6 carbon atoms is a C1-C6 alkyl
  • an alkyl comprising up to 5 carbon atoms is a C1-C5 alkyl.
  • a C1- C 5 alkyl includes C 5 alkyls, C 4 alkyls, C 3 alkyls, C 2 alkyls and C 1 alkyl (i.e., methyl).
  • a C 1 - C6 alkyl includes all moieties described above for C1-C5 alkyls but also includes C6 alkyls.
  • a C 1 -C 10 alkyl includes all moieties described above for C 1 -C 5 alkyls and C 1 -C 6 alkyls, but also includes C7, C8, C9 and C10 alkyls.
  • a C1-C12 alkyl includes all the foregoing moieties, but also includes C 11 and C 12 alkyls.
  • Non-limiting examples of C 1 -C 12 alkyl include methyl, ethyl, n-propyl, i-propyl, sec-propyl, n-butyl, i-butyl, sec-butyl, t-butyl, n-pentyl, t- amyl, n-hexyl, n-heptyl, n-octyl, n-nonyl, n-decyl, n-undecyl, and n-dodecyl.
  • an alkyl group can be optionally substituted.
  • Alkylene or “alkylene chain” refers to a fully saturated, straight or branched divalent hydrocarbon chain radical, and having from one to twelve carbon atoms.
  • C 1 -C 12 alkylene include methylene, ethylene, propylene, n-butylene, ethenylene, propenylene, n-butenylene, propynylene, n-butynylene, and the like.
  • the alkylene chain is attached to the rest of the molecule through a single bond and to the radical group through a single bond. The points of attachment of the alkylene chain to the rest of the molecule and to the radical group can be through one carbon or any two carbons within the chain.
  • alkylene chain can be optionally substituted.
  • alkenyl or “alkenyl group” refers to a straight or branched hydrocarbon chain radical having from two to twelve carbon atoms, and having one or more carbon-carbon double bonds. Each alkenyl group is attached to the rest of the molecule by a single bond. Alkenyl group comprising any number of carbon atoms from 2 to 12 are included.
  • An alkenyl group comprising up to 12 carbon atoms is a C 2 -C 12 alkenyl
  • an alkenyl comprising up to 10 carbon atoms is a C 2 -C 10 alkenyl
  • an alkenyl group comprising up to 6 carbon atoms is a C2-C6 alkenyl
  • an alkenyl comprising up to 5 carbon atoms is a C2-C5 alkenyl.
  • a C2- C 5 alkenyl includes C 5 alkenyls, C 4 alkenyls, C 3 alkenyls, and C 2 alkenyls.
  • a C 2 -C 6 alkenyl includes all moieties described above for C2-C5 alkenyls but also includes C6 alkenyls.
  • a C2- C 10 alkenyl includes all moieties described above for C 2 -C 5 alkenyls and C 2 -C 6 alkenyls, but also includes C7, C8, C9 and C10 alkenyls.
  • a C2-C12 alkenyl includes all the foregoing moieties, but also includes C 11 and C 12 alkenyls.
  • Non-limiting examples of C 2 -C 12 alkenyl include ethenyl (vinyl), 1-propenyl, 2-propenyl (allyl), iso-propenyl, 2-methyl-1- propenyl, 1-butenyl, 2-butenyl, 3-butenyl, 1-pentenyl, 2-pentenyl, 3-pentenyl, 4-pentenyl, 1- hexenyl, 2-hexenyl, 3-hexenyl, 4-hexenyl, 5-hexenyl, 1-heptenyl, 2-heptenyl, 3-heptenyl, 4- heptenyl, 5-heptenyl, 6-heptenyl, 1-octenyl, 2-octenyl, 3-octenyl, 4-octenyl, 5-octenyl, 6- octenyl, 7-octenyl, 1-nonenyl, 2-nonenyl, 3-nonenyl, 4-n
  • alkyl group can be optionally substituted.
  • alkenylene or “alkenylene chain” refers to a straight or branched divalent hydrocarbon chain radical, having from two to twelve carbon atoms, and having one or more carbon-carbon double bonds.
  • C 2 -C 12 alkenylene include ethene, propene, butene, and the like.
  • the alkenylene chain is attached to the rest of the molecule through a single bond and to the radical group through a single bond. The points of attachment of the alkenylene chain to the rest of the molecule and to the radical group can be through one carbon or any two carbons within the chain.
  • alkenylene chain can be optionally substituted.
  • alkynyl or “alkynyl group” refers to a straight or branched hydrocarbon chain radical having from two to twelve carbon atoms, and having one or more carbon-carbon triple bonds. Each alkynyl group is attached to the rest of the molecule by a single bond. Alkynyl group comprising any number of carbon atoms from 2 to 12 are included.
  • An alkynyl group comprising up to 12 carbon atoms is a C2-C12 alkynyl
  • an alkynyl comprising up to 10 carbon atoms is a C 2 -C 10 alkynyl
  • an alkynyl group comprising up to 6 carbon atoms is a C 2 -C 6 alkynyl
  • an alkynyl comprising up to 5 carbon atoms is a C 2 -C 5 alkynyl.
  • a C 2 -C 5 alkynyl includes C5 alkynyls, C4 alkynyls, C3 alkynyls, and C2 alkynyls.
  • a C2-C6 alkynyl includes all moieties described above for C 2 -C 5 alkynyls but also includes C 6 alkynyls.
  • a C 2 -C 10 alkynyl includes all moieties described above for C2-C5 alkynyls and C2-C6 alkynyls, but also includes C 7 , C 8 , C 9 and C 10 alkynyls.
  • a C 2 -C 12 alkynyl includes all the foregoing moieties, but also includes C11 and C12 alkynyls.
  • Non-limiting examples of C2-C12 alkenyl include ethynyl, propynyl, butynyl, pentynyl and the like.
  • alkynylene or “alkynylene chain” refers to a straight or branched divalent hydrocarbon chain radical, having from two to twelve carbon atoms, and having one or more carbon-carbon triple bonds.
  • C 2 -C 12 alkynylene include ethynylene, propargylene and the like.
  • the alkynylene chain is attached to the rest of the molecule through a single bond and to the radical group through a single bond. The points of attachment of the alkynylene chain to the rest of the molecule and to the radical group can be through one carbon or any two carbons within the chain.
  • alkynylene chain can be optionally substituted.
  • Alkoxy refers to a radical of the formula -OR a where R a is an alkyl, alkenyl or alknyl radical as defined above containing one to twelve carbon atoms. Unless stated otherwise specifically in the specification, an alkoxy group can be optionally substituted.
  • Alkylamino refers to a radical of the formula -NHR a or -NR a R a where each R a is, independently, an alkyl, alkenyl or alkynyl radical as defined above containing one to twelve carbon atoms.
  • alkylamino group can be optionally substituted.
  • R a is an alkyl, alkenyl or alkynyl radical as defined above.
  • a non-limiting example of an alkyl carbonyl is the methyl carbonyl (“acetal”) moiety.
  • Alkylcarbonyl groups can also be referred to as “C w -C z acyl” where w and z depicts the range of the number of carbon in Ra, as defined above.
  • C 1 -C 10 acyl refers to alkylcarbonyl group as defined above, where R a is C 1 -C 10 alkyl, C2-C10 alkenyl, or C2-C10 alkynyl radical as defined above. Unless stated otherwise specifically in the specification, an alkyl carbonyl group can be optionally substituted.
  • Aryl refers to a hydrocarbon ring system radical comprising hydrogen, 6 to 18 carbon atoms and at least one aromatic ring.
  • the aryl radical can be a monocyclic, bicyclic, tricyclic or tetracyclic ring system, which can include fused or bridged ring systems.
  • Aryl radicals include, but are not limited to, aryl radicals derived from phenyl (benzene), aceanthrylene, acenaphthylene, acephenanthrylene, anthracene, azulene, chrysene, fluoranthene, fluorene, as-indacene, s-indacene, indane, indene, naphthalene, phenalene, phenanthrene, pleiadene, pyrene, and triphenylene.
  • aryl is meant to include aryl radicals that are optionally substituted.
  • “Aralkyl” or “arylalkyl” refers to a radical of the formula -Rb-Rc where Rb is an alkylene group as defined above and R c is one or more aryl radicals as defined above.
  • Aralkyl radicals include, but are not limited to, benzyl, diphenylmethyl and the like. Unless stated otherwise specifically in the specification, an aralkyl group can be optionally substituted.
  • “Aralkenyl” or “arylalkenyl” refers to a radical of the formula -R b -R c where R b is an alkenylene group as defined above and Rc is one or more aryl radicals as defined above.
  • an aralkenyl group can be optionally substituted.
  • “Aralkynyl” or “arylalkynyl” refers to a radical of the formula -R b -R c where R b is an alkynylene group as defined above and R c is one or more aryl radicals as defined above. Unless stated otherwise specifically in the specification, an aralkynyl group can be optionally substituted.
  • Carbocyclyl “carbocyclic ring” or “carbocycle” refers to a ring structure, wherein the atoms which form the ring are each carbon.
  • Carbocyclic rings can comprise from 3 to 20 carbon atoms in the ring.
  • Carbocyclic rings include aryls and cycloalkyl. Cycloalkenyl and cycloalkynyl as defined herein. Unless stated otherwise specifically in the specification, a carbocyclyl group can be optionally substituted.
  • Cycloalkyl refers to a stable non-aromatic monocyclic or polycyclic fully saturated hydrocarbon radical consisting solely of carbon and hydrogen atoms, which can include fused, bridged, or spiral ring systems, having from three to twenty carbon atoms, preferably having from three to ten carbon atoms, and which is attached to the rest of the molecule by a single bond.
  • Monocyclic cycloalkyl radicals include, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl.
  • Polycyclic cycloalkyl radicals include, for example, adamantyl, norbornyl, decalinyl, 7,7-dimethyl-bicyclo[2.2.1]heptanyl, and the like. Unless otherwise stated specifically in the specification, a cycloalkyl group can be optionally substituted.
  • Cycloalkenyl refers to a stable non-aromatic monocyclic or polycyclic hydrocarbon radical consisting solely of carbon and hydrogen atoms, having one or more carbon-carbon double bonds, which can include fused, bridged, or spiral ring systems, having from three to twenty carbon atoms, preferably having from three to ten carbon atoms, and which is attached to the rest of the molecule by a single bond.
  • Monocyclic cycloalkenyl radicals include, for example, cyclopentenyl, cyclohexenyl, cycloheptenyl, cycloctenyl, and the like.
  • Polycyclic cycloalkenyl radicals include, for example, bicyclo[2.2.1]hept-2-enyl and the like. Unless otherwise stated specifically in the specification, a cycloalkenyl group can be optionally substituted.
  • Cycloalkynyl refers to a stable non-aromatic monocyclic or polycyclic hydrocarbon radical consisting solely of carbon and hydrogen atoms, having one or more carbon-carbon triple bonds, which can include fused, bridged, or spiral ring systems, having from three to twenty carbon atoms, preferably having from three to ten carbon atoms, and which is attached to the rest of the molecule by a single bond.
  • Monocyclic cycloalkynyl radicals include, for example, cycloheptynyl, cyclooctynyl, and the like. Unless otherwise stated specifically in the specification, a cycloalkynyl group can be optionally substituted.
  • “Cycloalkylalkyl” refers to a radical of the formula -R b -R d where R b is an alkylene, alkenylene, or alkynylene group as defined above and R d is a cycloalkyl, cycloalkenyl, cycloalkynyl radical as defined above. Unless stated otherwise specifically in the specification, a cycloalkylalkyl group can be optionally substituted.
  • Haloalkyl refers to an alkyl radical, as defined above, that is substituted by one or more halo radicals, as defined above, e.g., trifluoromethyl, difluoromethyl, trichloromethyl, 2,2,2-trifluoroethyl, 1,2-difluoroethyl, 3-bromo-2-fluoropropyl, 1,2-dibromoethyl, and the like. Unless stated otherwise specifically in the specification, a haloalkyl group can be optionally substituted.
  • Haloalkenyl refers to an alkenyl radical, as defined above, that is substituted by one or more halo radicals, as defined above, e.g., 1-fluoropropenyl, 1,1-difluorobutenyl, and the like. Unless stated otherwise specifically in the specification, a haloalkenyl group can be optionally substituted.
  • Haloalkynyl refers to an alkynyl radical, as defined above, that is substituted by one or more halo radicals, as defined above, e.g., 1-fluoropropynyl, 1-fluorobutynyl, and the like.
  • Heterocyclyl refers to a stable 3- to 20-membered non-aromatic, partially aromatic, or aromatic ring radical which consists of two to twelve carbon atoms and from one to six heteroatoms selected from the group consisting of nitrogen, oxygen and sulfur. Heterocyclycl or heterocyclic rings include heteroaryls as defined below.
  • the heterocyclyl radical can be a monocyclic, bicyclic, tricyclic or tetracyclic ring system, which can include fused, bridged, and spiral ring systems; and the nitrogen, carbon or sulfur atoms in the heterocyclyl radical can be optionally oxidized; the nitrogen atom can be optionally quaternized; and the heterocyclyl radical can be partially or fully saturated.
  • heterocyclyl radicals include, but are not limited to, aziridinyl, oextanyl, 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, thiazolidinyl, tetrahydrofuryl, trithianyl, tetrahydropyranyl, thiomorpholinyl, thia
  • Heterocyclylalkyl refers to a radical of the formula -R b -R e where R b is an alkylene group as defined above and R e is a heterocyclyl radical as defined above. Unless stated otherwise specifically in the specification, a heterocyclylalkyl group can be optionally substituted.
  • Heterocyclylalkenyl refers to a radical of the formula -R b -R e where R b is an alkenylene group as defined above and Re is a heterocyclyl radical as defined above. Unless stated otherwise specifically in the specification, a heterocyclylalkenyl group can be optionally substituted.
  • Heterocyclylalkynyl refers to a radical of the formula -R b -R e where R b is an alkynylene group as defined above and Re is a heterocyclyl radical as defined above. Unless stated otherwise specifically in the specification, a heterocyclylalkynyl group can be optionally substituted.
  • N-heterocyclyl refers to a heterocyclyl radical as defined above containing at least one nitrogen and where the point of attachment of the heterocyclyl radical to the rest of the molecule is through a nitrogen atom in the heterocyclyl radical. Unless stated otherwise specifically in the specification, a N-heterocyclyl group can be optionally substituted.
  • Heteroaryl refers to a 5- to 20-membered ring system radical one to thirteen carbon atoms and one to six heteroatoms selected from the group consisting of nitrogen, oxygen and sulfur, as the ring member.
  • the heteroaryl radical can be a monocyclic, bicyclic, tricyclic or tetracyclic ring system, which can include fused or bridged ring systems, wherein at least one ring containing a heteroatom ring member is aromatic.
  • the nitrogen, carbon or sulfur atoms in the heteroaryl radical can be optionally oxidized and the nitrogen atom can be optionally quaternized.
  • Examples include, but are not limited to, azepinyl, acridinyl, benzimidazolyl, benzothiazolyl, benzindolyl, benzodioxolyl, benzofuranyl, benzooxazolyl, benzothiazolyl, benzothiadiazolyl, benzo[b][1,4]dioxepinyl, 1,4-benzodioxanyl, benzonaphthofuranyl, benzoxazolyl, benzodioxolyl, benzodioxinyl, benzopyranyl, benzopyranonyl, benzofuranyl, benzofuranonyl, benzothienyl (benzothiophenyl), benzotriazolyl, benzo[4,6]imidazo[1,2-a]pyridinyl, carbazolyl, cinnolinyl, dibenzofuranyl, dibenzothiophenyl, furany
  • heteroaryl group can be optionally substituted.
  • N-heteroaryl refers to a heteroaryl radical as defined above containing at least one nitrogen and where the point of attachment of the heteroaryl radical to the rest of the molecule is through a nitrogen atom in the heteroaryl radical. Unless stated otherwise specifically in the specification, an N-heteroaryl group can be optionally substituted.
  • Heteroarylalkyl refers to a radical of the formula -R b -R f where R b is an alkylene chain as defined above and Rf is a heteroaryl radical as defined above.
  • heteroarylalkyl group can be optionally substituted.
  • “Heteroarylalkenyl” refers to a radical of the formula -R b -R f where R b is an alkenylene, chain as defined above and Rf is a heteroaryl radical as defined above. Unless stated otherwise specifically in the specification, a heteroarylalkenyl group can be optionally substituted.
  • “Heteroarylalkynyl” refers to a radical of the formula -R b -R f where R b is an alkynylene chain as defined above and Rf is a heteroaryl radical as defined above.
  • heteroarylalkynyl group can be optionally substituted.
  • “Thioalkyl” refers to a radical of the formula -SR a where R a is an alkyl, alkenyl, or alkynyl radical as defined above containing one to twelve carbon atoms. Unless stated otherwise specifically in the specification, a thioalkyl group can be optionally substituted.
  • substituted means any of the above groups (e.g., alkyl, alkylene, alkenyl, alkenylene, alkynyl, alkynylene, alkoxy, alkylamino, alkylcarbonyl, thioalkyl, aryl, aralkyl, carbocyclyl, cycloalkyl, cycloalkenyl, cycloalkynyl, cycloalkylalkyl, haloalkyl, heterocyclyl, N-heterocyclyl, heterocyclylalkyl, heteroaryl, N-heteroaryl, heteroarylalkyl, heteroarylalkenyl, heteroarylalkynyl, etc.) wherein at least one hydrogen atom is replaced by a bond to a non-hydrogen atoms such as, but not limited to: a halogen atom such as F, Cl, Br, and I; an oxygen atom in
  • “Substituted” also means any of the above groups in which one or more hydrogen atoms are replaced by a higher-order bond (e.g., a double- or triple-bond) to a heteroatom such as oxygen in oxo, carbonyl, carboxyl, and ester groups; and nitrogen in groups such as imines, oximes, hydrazones, and nitriles.
  • a higher-order bond e.g., a double- or triple-bond
  • nitrogen in groups such as imines, oximes, hydrazones, and nitriles.
  • Rg and R h are the same or different and independently hydrogen, alkyl, alkenyl, alkynyl, alkoxy, alkylamino, thioalkyl, aryl, aralkyl, cycloalkyl, cycloalkenyl, cycloalkynyl, cycloalkylalkyl, haloalkyl, haloalkenyl, haloalkynyl, heterocyclyl, N-heterocyclyl, heterocyclylalkyl, heteroaryl, N-heteroaryl and/or heteroarylalkyl.
  • “Substituted” further means any of the above groups in which one or more hydrogen atoms are replaced by a bond to an amino, cyano, hydroxyl, imino, nitro, oxo, thioxo, halo, alkyl, alkenyl, alkynyl, alkoxy, alkylamino, thioalkyl, aryl, aralkyl, cycloalkyl, cycloalkenyl, cycloalkynyl, cycloalkylalkyl, haloalkyl, haloalkenyl, haloalkynyl, heterocyclyl, N-heterocyclyl, heterocyclylalkyl, heteroaryl, N- heteroaryl and/or heteroarylalkyl group.
  • a point of attachment bond denotes a bond that is a point of attachment between two chemical entities, one of which is depicted as being attached to the point of attachment bond and the other of which is not depicted as being attached to the point of attachment bond.
  • “ ” indicates that the chemical entity “A” is bonded to another chemical entity via the point of attachment bond.
  • the specific point of attachment to the non-depicted chemical entity can be specified by inference.
  • parenteral administration and “administered parenterally” are art-recognized terms, and include modes of administration other than enteral and topical administration, such as injections, and include, without limitation, intravenous, intramuscular, intrapleural, intravascular, intrapericardial, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intra- articular, subcapsular, subarachnoid, intraspinal and intrastemal injection and infusion.
  • treating is art-recognized and includes inhibiting a disease, disorder or condition in a subject, e.g., impeding its progress; and relieving the disease, disorder or condition, e.g., causing regression of the disease, disorder and/or condition. Treating the disease or condition includes ameliorating at least one symptom of the particular disease or condition, even if the underlying pathophysiology is not affected.
  • preventing is art-recognized and includes stopping a disease, disorder or condition from occurring in a subject, which may be predisposed to the disease, disorder and/or condition but has not yet been diagnosed as having it.
  • Preventing a condition related to a disease includes stopping the condition from occurring after the disease has been diagnosed but before the condition has been diagnosed.
  • a "patient,” “subject,” or “host” to be treated by the subject method may mean either a human or non-human animal, such as a mammal, a fish, a bird, a reptile, or an amphibian.
  • the subject of the herein disclosed methods can be a human, non-human primate, horse, pig, rabbit, dog, sheep, goat, cow, cat, guinea pig or rodent.
  • the term does not denote a particular age or sex. Thus, adult and newborn subjects, as well as fetuses, whether male or female, are intended to be covered.
  • the subject is a mammal.
  • a patient refers to a subject afflicted with a disease or disorder.
  • the terms "prophylactic” or “therapeutic” treatment is art-recognized and includes administration to the host of one or more of the subject compositions. If it is administered prior to clinical manifestation of the unwanted condition (e.g., disease or other unwanted state of the host animal) then the treatment is prophylactic, i.e., it protects the host against developing the unwanted condition, whereas if it is administered after manifestation of the unwanted condition, the treatment is therapeutic (i.e., it is intended to diminish, ameliorate, or stabilize the existing unwanted condition or side effects thereof).
  • the unwanted condition e.g., disease or other unwanted state of the host animal
  • therapeutic agent include molecules and other agents that are biologically, physiologically, or pharmacologically active substances that act locally or systemically in a patient or subject to treat a disease or condition.
  • the terms include without limitation pharmaceutically acceptable salts thereof and prodrugs.
  • agents may be acidic, basic, or salts; they may be neutral molecules, polar molecules, or molecular complexes capable of hydrogen bonding; they may be prodrugs in the form of ethers, esters, amides and the like that are biologically activated when administered into a patient or subject.
  • terapéuticaally effective amount or “pharmaceutically effective amount” is an art-recognized term.
  • the term refers to an amount of a therapeutic agent that produces some desired effect at a reasonable benefit/risk ratio applicable to any medical treatment.
  • the term refers to that amount necessary or sufficient to eliminate, reduce or maintain a target of a particular therapeutic regimen.
  • the effective amount may vary depending on such factors as the disease or condition being treated, the particular targeted constructs being administered, the size of the subject or the severity of the disease or condition.
  • One of ordinary skill in the art may empirically determine the effective amount of a particular compound without necessitating undue experimentation.
  • a therapeutically effective amount of a therapeutic agent for in vivo use will likely depend on a number of factors, including: the rate of release of an agent from a polymer matrix, which will depend in part on the chemical and physical characteristics of the polymer; the identity of the agent; the mode and method of administration; and any other materials incorporated in the polymer matrix in addition to the agent.
  • ED50 is art-recognized. In certain embodiments, ED50 means the dose of a drug, which produces 50% of its maximum response or effect, or alternatively, the dose, which produces a pre-determined response in 50% of test subjects or preparations.
  • LD50 is art-recognized.
  • LD50 means the dose of a drug, which is lethal in 50% of test subjects.
  • therapeutic index is an art-recognized term, which refers to the therapeutic index of a drug, defined as LD50/ED50.
  • IC50 half maximal inhibitory concentration
  • IC50 half maximal inhibitory concentration
  • the phrase "optionally substituted” means that a non-hydrogen substituent may or may not be present on a given atom, and, thus, the description includes structures wherein a non-hydrogen substituent is present and structures wherein a non-hydrogen substituent is not present.
  • the term “or” as used herein should be understood to mean “and/or”, unless the context clearly indicates otherwise.
  • the term "about” or “approximately” refers to a quantity, level, value, number, frequency, percentage, dimension, size, amount, weight or length that varies by as much as 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2% or 1% to a reference quantity, level, value, number, frequency, percentage, dimension, size, amount, weight or length.
  • the term "about” or “approximately” refers a range of quantity, level, value, number, frequency, percentage, dimension, size, amount, weight or length ⁇ 15%, ⁇ 10%, ⁇ 9%, ⁇ 8%, ⁇ 7%, ⁇ 6%, ⁇ 5%, ⁇ 4%, ⁇ 3%, ⁇ 2%, or ⁇ 1% about a reference quantity, level, value, number, frequency, percentage, dimension, size, amount, weight or length. [0096] All percentages and ratios used herein, unless otherwise indicated, are by weight.
  • brain injury includes traumatic injuries and injuries as a result of disease, in particular neurodegenerative diseases and dementias.
  • brain injury includes, but is not limited to mild, moderate, or severe trauma to the brain such as that received in military conflict, sports injury, accidents and falls, and the like, and also includes but is not limited to injury to the brain as a result of any tauopathy or dementia.
  • the brain injury is accompanied by, associated with, or mediated by astrogliosis or astroglial activation.
  • Glasgow Coma Scale or “GCS” as used herein refers to a 15 point scale for estimating and categorizing the outcomes of brain injury on the basis of overall social capability or dependence on others.
  • the test measures the motor response, verbal response and eye opening response with these values: I. Motor Response (6--Obeys commands fully; 5--Localizes to noxious stimuli; 4--Withdraws from noxious stimuli; 3--Abnormal flexion, i.e., decorticate posturing; 2--Extensor response, i.e. decerebrate posturing; and 1--No response); II.
  • Verbal Response (5--Alert and Oriented; 4--Confused, yet coherent, speech; 3-- Inappropriate words and jumbled phrases consisting of words; 2--Incomprehensible sounds; and 1--No sounds); and III. Eye Opening (4--Spontaneous eye opening; 3--Eyes open to speech; 2--Eyes open to pain; and 1--No eye opening). The final score is determined by adding the values of I+II+III.
  • the final score can be categorized into four possible levels for survival, with a lower number indicating a more severe injury and a poorer prognosis: Mild (13-15); Moderate Disability (9-12) (Loss of consciousness greater than 30 minutes; Physical or cognitive impairments which may or may resolve: and Benefit from Rehabilitation); Severe Disability (3-8) (Coma: unconscious state. No meaningful response, no voluntary activities); and Vegetative State (Less Than 3) (Sleep wake cycles; Arousal, but no interaction with environment; No localized response to pain).
  • Moderate brain injury is defined as a brain injury resulting in a loss of consciousness from 20 minutes to 6 hours and a Glasgow Coma Scale of 9 to 12.
  • Severe brain injury is defined as a brain injury resulting in a loss of consciousness of greater than 6 hours and a Glasgow Coma Scale of 3 to 8.
  • "Glasgow Outcome Scale” as used herein refers to a global scale for functional outcome that rates patient status into one of five categories: Dead, Vegetative State, Severe Disability, Moderate Disability or Good Recovery.
  • Primary brain injuries occur during the initial insult and result from displacement of the physical structures of the brain. More specifically, a primary brain injury is the physical damage to parenchyma (tissue, vessels) that occurs during the traumatic event, resulting in shearing and compression of the surrounding brain tissue. Secondary brain injuries occur subsequent to the primary injury and may involve an array of cellular processes.
  • a secondary brain injury refers to the changes that evolve over a period of time (from hours to days) after the primary brain injury. It includes an entire cascade of cellular, chemical, tissue, or blood vessel changes in the brain that contribute to further destruction of brain tissue.
  • An injury to the head can be either closed or open (penetrating).
  • a closed head injury refers to a trauma to the scalp, skull or brain where there is no penetration of the skull by a striking object.
  • An open head injury refers a trauma to the scalp, skull or brain where there is penetration of the skull by a striking object.
  • An injury to the head may be caused by physical shaking of a person, by blunt impact by an external mechanical or other force that results in a closed or open head trauma (e.g., vehicle accident, such as with an automobile, plane, train, etc.; blow to the head, such as with a baseball bat, or from a firearm), a cerebral vascular accident (e.g., stroke), one or more falls (e.g., as in sports or other activities), explosions or blasts (collectively, "blast injuries”) and by other types of blunt force trauma.
  • a closed or open head trauma e.g., vehicle accident, such as with an automobile, plane, train, etc.; blow to the head, such as with a baseball bat, or from a firearm
  • a cerebral vascular accident e.g., stroke
  • one or more falls e.g., as in sports or other activities
  • explosions or blasts collectively, "blast injuries”
  • an injury to the head may be caused by the ingestion and/or exposure to a chemical, toxin or
  • Examples of such chemicals and/or toxins include fires, molds, asbestos, pesticides and insecticides, organic solvents, paints, glues, gases (such as carbon monoxide, hydrogen sulfide, and cyanide), organic metals (such as methyl mercury, tetraethyl lead and organic tin) and/or one or more drugs of abuse.
  • gases such as carbon monoxide, hydrogen sulfide, and cyanide
  • organic metals such as methyl mercury, tetraethyl lead and organic tin
  • an injury to the head may be caused as a result of a subject suffering from an autoimmune disease, a metabolic disorder, a brain tumor, one or more viruses, meningitis, hydrocephalus, hypoxia or any combinations thereof. In some cases, it is not possible to be certain whether any such event or injury has occurred or taken place.
  • TBI Traumatic Brain Injury
  • TBI can be classified as "mild,” “moderate,” or “severe.”
  • the causes of TBI are diverse and include, for example, physical shaking by a person, a car accident, injuries from firearms, cerebral vascular accidents (e.g., strokes), falls, explosions or blasts and other types of blunt force trauma.
  • Other causes of TBI include the ingestion and/or exposure to one or more chemicals or toxins (such as fires, molds, asbestos, pesticides and insecticides, organic solvents, paints, glues, gases (such as carbon monoxide, hydrogen sulfide, and cyanide), organic metals (such as methyl mercury, tetraethyl lead and organic tin), one or more drugs of abuse or combinations thereof).
  • TBI can occur in subjects suffering from an autoimmune disease, a metabolic disorder, a brain tumor, hypoxia, one or more viruses, meningitis, hydrocephalus or combinations thereof. Young adults and the elderly are the age groups at highest risk for TBI.
  • traumatic brain injury or TBI does not include and specifically excludes cerebral vascular accidents such as strokes.
  • Mild TBI refers to a brain injury where loss of consciousness is brief and usually a few seconds or minutes and/or confusion and disorientation is shorter than 1 hour. Mild TBI is also referred to as a concussion, minor head trauma, minor TBI, minor brain injury, and minor head injury.
  • Mild TBI is the most prevalent TBI and is often missed at time of initial injury. Typically, a subject has a Glasgow Coma scale number of between 13-15 (such as 13-15 or 14-15). Fifteen percent (15%) of people with mild TBI have symptoms that last 3 months or more. Mild TBI is defined as the result of the forceful motion of the head or impact causing a brief change in mental status (confusion, disorientation or loss of memory) or loss of consciousness for less than 30 minutes.
  • Modemize TBI refers to a brain injury where loss of consciousness and/or confusion and disorientation is between 1 and 24 hours and the subject has a Glasgow Coma scale number of between 9-12. The individual with moderate TBI have abnormal brain imaging results.
  • “Severe TBI” as used herein refers to a brain injury where loss of consciousness is more than 24 hours and memory loss after the injury or penetrating skull injury longer than 24 hours and the subject has a Glasgow Coma scale number between 3-8. The deficits range from impairment of higher level cognitive functions to comatose states. Survivors may have limited function of arms or legs, abnormal speech or language, loss of thinking ability or emotional problems. Individuals with severe injuries can be left in long-term unresponsive states. For many people with severe TBI, long-term rehabilitation is often necessary to maximize function and independence.
  • Common symptoms of moderate to severe TBI include cognitive deficits including difficulties with attention, concentration, distractibility, memory, speed of processing, confusion, perseveration, impulsiveness, language processing, and/or "executive functions", not understanding the spoken word (receptive aphasia), difficulty speaking and being understood (expressive aphasia), slurred speech, speaking very fast or very slow, problems reading, problems writing, difficulties with interpretation of touch, temperature, movement, limb position and fine, discrimination, the integration or patterning of sensory impressions into psychologically meaningful data, partial or total loss of vision, weakness of eye muscles and double vision (diplopia), blurred vision, problems judging distance, involuntary eye movements (nystagmus), intolerance of light (photophobia), hearing, such as decrease or loss of hearing, ringing in the ears (tinnitus), increased sensitivity to sounds, loss or diminished sense of smell (anosmia), loss or diminished sense of taste, the convulsions associated with epilepsy that can be several types and can
  • neurodegenerative disease is used interchangeably herein to refer to a varied assortment of central nervous system diseases, disorders, and conditions characterized by gradual and progressive loss of neural tissue and/or neural tissue function.
  • a neurodegenerative disease is a class of neurological disorder or disease, and where the neurological disease is characterized by a gradual and progressive loss of neural tissue, and/or altered neurological function, typically reduced neurological function as a result of a gradual and progressive loss of neural tissue.
  • vascular dementia is also referred to as "multi-infarct dementia” in the art refers to a group of syndromes caused by different mechanisms all resulting in vascular lesions in the brain.
  • vascular dementia The main subtypes of vascular dementia are, for example vascular mild cognitive impairment, multi-infarct dementia, vascular dementia due to a strategic single infarct (affecting the thalamus, the anterior cerebral artery, the parietal lobes or the cingulate gyrus), vascular dementia due to hemorrhagic lesions, small vessel disease (including, e.g., vascular dementia due to lacunar lesions and Binswanger disease), and mixed Alzheimer's Disease with vascular dementia.
  • vascular mild cognitive impairment vascular mild cognitive impairment
  • multi-infarct dementia vascular dementia due to a strategic single infarct (affecting the thalamus, the anterior cerebral artery, the parietal lobes or the cingulate gyrus)
  • vascular dementia due to hemorrhagic lesions
  • small vessel disease including, e.g., vascular dementia due to lacunar lesions and Binswanger disease
  • mixed Alzheimer's Disease with vascular dementia
  • disease refers to any alteration in state of the body or of some of the organs, interrupting or disturbing the performance of the functions and/or causing symptoms such as discomfort, dysfunction, distress, or even death to the person afflicted or those in contact with a person.
  • a disease or disorder can also relate to a distemper, ailing, ailment, malady, disorder, sickness, illness, complaint, or affectation.
  • blood-brain barrier or "BBB” are used interchangeably herein, and are used to refer to the permeability barrier that exists in blood vessels as they travel through the brain tissue that severely restricts and closely regulates what is exchanged between the blood and the brain tissue.
  • the blood brain barrier components include the endothelial cells that form the innermost lining of all blood vessels, the tight junctions between adjacent endothelial cells that are the structural correlate of the BBB, the basement membrane of endothelial cells and the expanded foot processes of nearby astrocytes which cover nearly all of the exposed outer surface of the blood vessel.
  • the BBB prevents most substances in the blood from entering brain tissue, including most large molecules such as Ig, antibodies, complement, albumin and drugs and small molecules.
  • abnormal BBB is used to refer to a dysfunctional BBB, for example, where the BBB does not allow transit of molecules that normally transit a functional BBB, for example nutrients and sugars such as glucose.
  • An abnormal BBB can also refer to when the BBB is permeable to molecules that a normally functioning BBB would typically exclude, which is typically referred to "BBB permeability" herein.
  • BBB permeability or “permeable BBB” are commonly referred to by persons in the art as “leaky BBB”. The terms are used interchangeably herein to refer to impaired BBB integrity and increased vascular permeability.
  • a permeable BBB allows transit of molecules through the BBB that an intact BBB would normally exclude from the brain tissue, for example, Ig molecules, complement proteins, serum albumin and numerous other proteins.
  • An assay to determine the presence of a permeable BBB can be, for example, to assess the presence of extravascular Ig in the brain tissue which is normally be restricted to the lumen of blood vessels when the BBB is functioning normally (i.e., when the BBB is not permeable).
  • 15-hydroxyprostaglandin dehydrogenase 15-hydroxyprostaglandin dehydrogenase
  • BBB blood brain barrier
  • 15-PGDH 15-hydroxyprostaglandin dehydrogenase
  • AD Alzheimer’s disease
  • TBI traumatic brain injury
  • aging 15-hydroxyprostaglandin dehydrogenase
  • Pathological increase in 15-PGDH correlates with pronounced oxidative stress, neuroinflammation, and neurodegeneration, alongside profound BBB structural degeneration characterized by astrocytic endfeet swelling and functional impairment.
  • Pharmacologic inhibition or genetic reduction of 15-PGDH in AD and TBI models strikingly mitigates cognitive decline, oxidative damage, suppresses neuroinflammation, and restores BBB integrity.
  • inhibiting 15-PGDH not only blocks damage to the brain by blocking microglial generation of reactive oxygen in 15-PGDH positive microglial cells and halts neurodegeneration but also preserves cognitive function at levels indistinguishable from healthy controls.
  • Generation of reactive oxygen species by microglial cells can also be blocked with multiple substrates that 15-PGDH enzymatically degrades and limits, including PGE2, PGF2a, 15-HETE, RvD1, and LXA4 (respectively prostaglandin E2, prostaglandin F2 alpha, 15-HETE, resolving D1, and lipoxin A4).
  • these neuroprotective effects in AD are achieved without affecting amyloid pathology, underscoring a non-canonical mechanism for treating AD.
  • the cognitive decline, neuroinflammation, neurodegeneration, and/or neurodegenerative and/or neuropsychiatric conditions, diseases and/or disorders characterized by structural deterioration of the blood-brain barrier and/or by increased generation of reactive oxygen species within the nervous system can also be associated with enhanced or aberrant 15-PGDH activity in a subject in need thereof.
  • a method of treating diseases or disorders characterized by structural deterioration of the blood-brain barrier can include administering to the subject an amount of a 15-PGDH inhibitor and/or 15-PGDH substrate effective to prevent the onset of deterioration and/or the progression of further deterioration of the blood-brain barrier and/or prevent the neurodegeneration associated with blood-brain barrier deterioration and/or prevent the impaired neuronal function associated with blood-brain barrier deterioration and/or restore the integrity and/or structure and/or function of the blood-brain barrier.
  • a method of treating diseases or disorders characterized by increased generation of reactive oxygen species within the nervous system or central nervous system of a subject in need thereof, including cognitive decline, neural injuries, neurodegeneration, and/or neurodegenerative and/or neuropsychiatric conditions can include administering to the subject an amount of a 15-PGDH inhibitor and/or 15-PGDH substrate effective to prevent and/or decrease reactive oxygen species generation and/or neurodegeneration and/or impaired neuronal function associated with reactive oxygen species.
  • the subject has memory loss and/or cognitive decline and the 15-PGDH inhibitor and/or 15-PGDH substrate is administered at amount effective to improve memory and/or cognition.
  • the 15-PGDH inhibitor and/or 15-PGDH substrate is administered to the subject at amount effective to block aberrantly increased 15-PGDH enzymatic activity and prevent perivascular inflammation.
  • Other embodiments described herein relate to a method of treating cognitive decline, neural injuries, neurodegeneration, and/or neurodegenerative and/or neuropsychiatric conditions, diseases, and/or disorders characterized by increased generation of reactive oxygen species within the nervous system in a subject in need thereof.
  • the 15-PGDH inhibitor and/or 15-PGDH substrate can be administered to the subject at an amount effective to decrease at least one biomarker of reactive oxygen species generation, such as 4-hydroxy-2-nonenal (4-HNE) and/or 3- nitrotyrosine (3-NT) levels, by at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30% at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75% or more.
  • at least one biomarker of reactive oxygen species generation such as 4-hydroxy-2-nonenal (4-HNE) and/or 3- nitrotyrosine (3-NT) levels
  • the method includes measuring at least one biomarker of reactive oxygen species generation in the brain of the subject and administering to the subject an amount of a 15-PGDH inhibitor and/or 15-PGDH substrate effective to decrease reactive oxygen species generation in the brain of the subject.
  • Methods of measuring reactive oxygen species generation in the brain can include, for example, measuring 4-hydroxy-2-nonenal (4- HNE) and/or 3-nitrotyrosine (3-NT) levels as described in Domenico et al., Free Radic. Biol. Med.111, 253–261 (2017); Shin et al., Biomedicines 8, 326 (2020); and Sultana, et al., Neurobiol. Dis.22, 76–87 (2006).
  • the method includes measuring at least one biomarker of reactive oxygen species generation, such as 4-hydroxy-2-nonenal (4-HNE) and/or 3- nitrotyrosine (3-NT) levels, in blood of the subject following administration of the 15-PGDH inhibitor and comparing the measured biomarker of reactive oxygen species generation to a control, wherein a decrease in 4-hydroxy-2-nonenal (4-HNE) and/or 3-nitrotyrosine (3-NT) levels is indicative of efficacy of the 15-PGDH inhibitor and/or PGDH substrate in treating neurodegeneration and/or impaired neuronal function in the subject.
  • a biomarker of reactive oxygen species generation such as 4-hydroxy-2-nonenal (4-HNE) and/or 3- nitrotyrosine (3-NT) levels
  • the control can be, for example, the 4-hydroxy-2-nonenal (4-HNE) and/or 3-nitrotyrosine (3-NT) levels of the subject prior to administration of the 15-PGDH inhibitor and/or 15-PGDH substrate.
  • the cognitive decline, neural injuries, neurodegeneration, and/or neurodegenerative and/or neuropsychiatric conditions, diseases and/or disorders are associated with an increase in reactive oxygen species in neurotissue, such as brain tissue, of at least about 5%, about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, about 90%, about 100%, about 110%, about 120%, about 130%, about 140%, about 150%, about 160%, about 170%, about 180%, about 190%, about 200%, about 250%, about 300%, about 350%, about 400%, about 450%, about 500%, about 600%, about 700%, about 800%, about 900%, or about 1000% relative to normal or healthy neurotissue.
  • administration of a 15-PGDH inhibitor and/or 15- PGDH substrate can be used to decrease reactive oxygen species in neurotissue of the subject from about 5% to about 200%, about 5% to about 180%, about 5% to about 160%, about 5% to about 140%, about 5% to about 120%, about 5% to about 100%, about 5% to about 80%, about 5% to about 60%, about 5% to about 40%, about 10% to about 200%, about 10% to about 180%, about 10% to about 160%, about 10% to about 140%, about 10% to about 120%, about 10% to about 100%, about 10% to about 80%, about 10% to about 60%, about 30% to about 200%, about 30% to about 180%, about 30% to about 160%, about 30% to about 140%, about 30% to about 120%, about 30% to about 100%, about 30% to about 80%, about 40% to about 200%, about 40% to about 180%, about 40% to about 160%, about 40% to about 140%, about 40% to about 120%, about 40% to about 100%, about 50% to about
  • Still other embodiments described herein relate to a method of treating cognitive decline, neural injuries, neurodegeneration, and/or neurodegenerative and/or neuropsychiatric conditions, diseases, and/or disorders characterized by structural deterioration of the blood- brain barrier in a subject in need thereof.
  • the 15-PGDH inhibitor and/or 15-PGDH substrate can be administered to the subject at an amount effective to decrease BBB permeability by at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30% at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75% or more.
  • the method includes measuring BBB permeability of the subject and administering to the subject an amount of a 15-PGDH inhibitor and/or 15-PGDH substrate effective to decrease the measured BBB permeability of the subject.
  • Methods of measuring BBB permeability can include, for example, tracer-based techniques using dyes, fluorescent tracers, or radiolabeled tracers, magnetic resonance imaging (MRI) techniques using dynamic contrast-enhanced MRI imaging and chelated metal contrast agents, PET/SPECT imaging using radiolabeled small molecules, and/or measurement of albumin CSF/serum ratio.
  • the method includes measuring BBB permeability of the subject following administration of the 15-PGDH inhibitor and/or 15-PGDH substrate and comparing the measured BBB permeability a control, wherein a decrease in BBB permeability is indicative of efficacy of the 15-PGDH inhibitor and/or PGDH substrate in treating neurodegeneration and/or impaired neuronal function in the subject.
  • the control can be, for example, the BBB permeability of the subject prior to administration of the 15-PGDH inhibitor and/or 15-PGDH substrate.
  • the cognitive decline, neural injuries, neurodegeneration, and/or neurodegenerative and/or neuropsychiatric conditions, diseases and/or disorders are associated with an increase in BBB permeability of at least about 5%, about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, about 90%, about 100%, about 110%, about 120%, about 130%, about 140%, about 150%, about 160%, about 170%, about 180%, about 190%, about 200%, about 250%, about 300%, about 350%, about 400%, about 450%, about 500%, about 600%, about 700%, about 800%, about 900%, or about 1000% relative to normal or healthy BBB.
  • administration of a 15-PGDH inhibitor and/or 15-PGDH substrate can be used to decrease BBB permeability of the subject from about 5% to about 200%, about 5% to about 180%, about 5% to about 160%, about 5% to about 140%, about 5% to about 120%, about 5% to about 100%, about 5% to about 80%, about 5% to about 60%, about 5% to about 40%, about 10% to about 200%, about 10% to about 180%, about 10% to about 160%, about 10% to about 140%, about 10% to about 120%, about 10% to about 100%, about 10% to about 80%, about 10% to about 60%, about 30% to about 200%, about 30% to about 180%, about 30% to about 160%, about 30% to about 140%, about 30% to about 120%, about 30% to about 100%, about 30% to about 80%, about 40% to about 200%, about 40% to about 180%, about 40% to about 160%, about 40% to about 140%, about 40% to about 120%, about 40% to about 100%, about 50% to about 200%,
  • the diseases or disorders characterized by structural deterioration of the blood-brain barrier and/or increased generation of reactive oxygen species include brain injuries.
  • Brain injuries that can be treated include any brain injury that is mediated by structural deterioration of the blood-brain barrier and/or increased generation of reactive oxygen species.
  • TBI traumatic brain injury
  • stroke ischemic and hemorrhagic
  • spinal cord injury SCI
  • brain hemorrhage for example intracerebral hemorrhage and subarachnoid hemorrhage
  • CTE chronic traumatic encephalopathy
  • AD Alzheimer’s disease
  • FTD frontotemporal dementia
  • PD Parkinson’s disease
  • MS multiple sclerosis
  • MS amyloid lateral sclerosis
  • ALS amyloid lateral sclerosis
  • CTE prodromal chronic traumatic encephalopathy
  • AD prodromal Alzheimer’s disease
  • TBI occurs due to a fall, vehicle collision, work injury, sports injury, violence, and the like. TBI can result in various physical, cognitive and behavioral symptoms, depending on the area of the brain affected and its severity, symptoms which may be permanent. It is a major cause of death and disability. Current treatment focusses on minimizing the damage caused, and prevention.
  • CTE also referred to as traumatic encephalopathy syndrome or dementia pugilisitca, is a neurodegenerative condition caused by repeated head injuries, and tends to get worse over time, resulting in dementia. The cause frequently is repeated injury in contact sports, the military, domestic violence, or repeated banging of the head. Firm diagnosis often is made only at autopsy, and no treatment is available and focusses on maintenance and support only.
  • Neurodegenerative diseases are those which involve progressive loss of neurons or their function, including death of neurons, and which result in a progressive loss of brain function.
  • Neurodegenerative diseases associated with a tauopathy include Alzheimer's disease, FTD, and the like.
  • CTE also sometimes also is classified as this type of neurodegenerative disease.
  • the cognitive decline, neural injuries, neurodegeneration, and/or neurodegenerative and/or neuropsychiatric conditions, diseases, and/or disorders characterized by structural deterioration of the blood-brain barrier and/or increased generation of reactive oxygen species can include at least one of subarachnoid hemorrhage, schizophrenia, depression, bipolar disorder, normal aging, epilepsy, traumatic brain injury and/or a visual symptom associated therewith, post-traumatic stress disorder, Parkinson’s disease, Parkinson Plus syndromes, Lewy Body Dementia, multiple system atrophy, corticobasal neurodegeneration, progressive supranuclear palsy, Alexander’s disease, Alzheimer’s disease, Alzheimer's disease related dementias, prodromal Alzheimer’s disease, Down syndrome, spinocerebellar ataxia, amyotrophic lateral sclerosis, Huntington’s disease, stroke, brain radiation therapy, chronic stress, abuse or cellular toxicity of a neuro-active drug, retinal degeneration, spinal cord injury, peripheral nerve
  • the cognitive decline, neural injuries, neurodegeneration, and/or neurodegenerative and/or neuropsychiatric conditions, diseases and/or disorders characterized by structural deterioration of the blood-brain barrier and/or increased generation of reactive oxygen species is also associated with an aberrant level (e.g., decrease or increase in the level) of at least one eicosanoid selected from PGE2, 15-keto-PGE2, PGF2 ⁇ , 6-keto-PGF1 ⁇ , PGD2, PGJ2, TN-E, TXB2, LTB4, 15-HETE, 12-HETE, 8-HETE, 5- HETE, 17-HDA, 12, 13-DiHOME, 9,10-DiHOME, 14,15-DHET, or 11,12-DHET in neurotissue of the subject.
  • an aberrant level e.g., decrease or increase in the level
  • the neurodegenerative condition, disease, or disorder can be characterized by structural deterioration of the blood-brain barrier and/or increased generation of reactive oxygen species and an increase or decrease in the level of at least one eicosanoid selected from PGE2, 15-keto-PGE2, PGF2 ⁇ , 6-keto-PGF1 ⁇ , PGD2, PGJ2, TN-E, TXB2, LTB4, 15-HETE, 12-HETE, 8-HETE, 5-HETE, 17-HDA, 12, 13-DiHOME, 9,10- DiHOME, 14,15-DHET, or 11,12-DHET of at least about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 100%, or more in neurotissue of the subject relative to a normal or healthy subject.
  • the 15-PGDH inhibitor and/or 15-PGDH substrate can be administered to the subject to decrease structural deterioration of the blood-brain barrier and/or generation of reactive oxygen species and modulate the level of the aberrant eicosanoid to a normal or healthy level in the neurotissue.
  • the neurodegeneration and/or neurodegenerative condition, disease, or disorder can be associated with structural deterioration of the blood-brain barrier and/or increased generation of reactive oxygen species and an increase in the level of at least one of 15-keto-PGE2, TN-E, TXB2, LTB4, 15-HETE, 12-HETE, 8-HETE, 5-HETE, 17-HDA, 12, 13-DiHOME, 9,10-DiHOME, or 14,15-DHET in neurotissue (e.g., brain tissue) of the subject, and the 15-PGDH inhibitor and/or 15-PGDH substrate can be administered to the subject at amount effective to decrease structural deterioration of the blood-brain barrier and/or generation of reactive oxygen species and the levels of the at least one of 15-keto-PGE2, TN-E, TXB2, LTB4, 15-HETE, 12-HETE, 8-HETE, 5-HETE, 17-HDA, 12, 13-DiHOME, 9,10-DiHOME, or 14,15-
  • the neurodegeneration and/or neurodegenerative condition, disease, or disorder can be associated with an increase in structural deterioration of the blood-brain barrier and/or increased generation of reactive oxygen species and a decrease in the levels of at least one of PGJ2, TNE, 15-HETE, or 9,10- DiHOME in neurotissue (e.g., brain tissue) of the subject, and the 15-PGDH inhibitor and/or 15-PGDH substrate can be administered to the subject at amount effective to decrease structural deterioration of the blood-brain barrier and/or generation of reactive oxygen species and increase the levels of the at least one of PGJ2, TNE, 15-HETE, or 9,10-DiHOME in neurotissue (e.g., brain tissue) of the subject.
  • neurotissue e.g., brain tissue
  • FIG. 1 Another embodiments described herein relate to a method of treating and/or inhibiting memory loss and/or cognitive decline associated with structural deterioration of the blood-brain barrier and/or increased generation of reactive oxygen species in a subject in need thereof by administering to the subject a therapeutically effective amount of a 15-PGDH inhibitor and/or 15-PGDH substrate.
  • the memory loss and/or cognitive decline associated with structural deterioration of the blood-brain barrier and/or increased generation of reactive oxygen species in a subject in need thereof can be caused by and/or associated with neural injuries, neurodegeneration, neurodegenerative and/or neuropsychiatric conditions, diseases, and/or disorders.
  • neurodegenerative condition, disease, or disorder can include at least one of Alzheimer's disease, prodromal Alzheimer’s disease, Lewy body dementia, prodromal Lewy body dementia, Vascular dementia, prodromal Vascular dementia, Age-related dementia, Frontotemporal dementia, prodromal Frontotemporal dementia, mixed dementia, prodromal mixed dementia, or traumatic brain injury.
  • the memory loss and/or cognitive decline associated with structural deterioration of the blood-brain barrier and/or increased generation of reactive oxygen species within the nervous system is also associated with an aberrant level (e.g., decrease or increase in the level) of at least one eicosanoid selected from PGE2, 15-keto-PGE2, PGF2 ⁇ , 6-keto-PGF1 ⁇ , PGD2, PGJ2, TN-E, TXB2, LTB4, 15-HETE, 12-HETE, 8-HETE, 5-HETE, 17-HDA, 12, 13-DiHOME, 9,10-DiHOME, 14,15-DHET, or 11,12-DHET in brain tissue of the subject.
  • an aberrant level e.g., decrease or increase in the level
  • the 15-PGDH inhibitor can be administered to the subject to lower GFAP levels and modulate the level of the aberrant eicosanoid to a normal or healthy level in the brain tissue.
  • the memory loss and/or cognitive decline can be associated with structural deterioration of the blood-brain barrier and/or increased generation of reactive oxygen species within the nervous system and an increase in the level of at least one of 15-keto-PGE2, TN-E, TXB2, LTB4, 15-HETE, 12-HETE, 8-HETE, 5-HETE, 17-HDA, 12, 13-DiHOME, 9,10-DiHOME, or 14,15-DHET in brain tissue of the subject, and the 15-PGDH inhibitor can be administered to the subject at amount effective to decrease the structural deterioration of the blood-brain barrier and/or generation of reactive oxygen species within the nervous system and the level of the at least one of 15-keto-PGE2, TN-E, TXB2, LTB4, 15-HETE, 12-HETE, 8-HETE,
  • the memory loss and/or cognitive decline can be associated with an increase structural deterioration of the blood-brain barrier and/or increased generation of reactive oxygen species within the nervous system and a decrease in the level of at least one of PGJ2, TNE, 15-HETE, or 9,10-DiHOME in brain tissue of the subject, and the 15-PGDH inhibitor can be administered to the subject at amount effective to decrease the structural deterioration of the blood-brain barrier and/or generation of reactive oxygen species within the nervous system and increase the levels of the at least one of PGJ2, TNE, 15-HETE, or 9,10-DiHOME in e.g., brain tissue of the subject.
  • small molecule 15-PGDH inhibitors used to decrease structural deterioration of the blood-brain barrier and/or generation of reactive oxygen species within the nervous system can be identified using assays in which putative inhibitor compounds are applied to cells expressing 15-PGDH and then the functional effects on 15- PGDH activity are determined. Samples or assays comprising 15-PGDH that are treated with a potential inhibitor are compared to control samples without the inhibitor to examine the extent of effect. Control samples (untreated with modulators) are assigned a relative 15- PGDH activity value of 100%.
  • 15-PGDH activity value relative to the control is about 80%, optionally 50% or 25%, 10%, 5% or 1%.
  • Small molecules tested as 15-PGDH inhibitors can be any small chemical molecule or compound.
  • the assays are designed to screen large chemical libraries by automating the assay steps and providing compounds from any convenient source to assays, which are typically run in parallel (e.g., in microtiter formats on microtiter plates in robotic assays).
  • the 15-PGDH inhibitor can include a compound having the following formula (I): pharmaceutically acceptable salt, tautomer, or solvate thereof; wherein n is 0-2; Y 1 , Y 2 , and R 1 are the same or different and are independently hydrogen or a substituted or unsubstituted group selected from C1-C24 alkyl, C2-C24 alkenyl, C2-C24 alkynyl, C 3 -C 20 aryl, heteroaryl, heterocycloalkenyl containing from 5-6 ring atoms, C 6 -C 24 alkaryl, C6-C24 aralkyl, halo, -Si(C1-C3 alkyl)3, hydroxyl, sulfhydryl, C1-C24 alkoxy, C 2 -C 24 alkenyloxy, C 2 -C 24 alkynyloxy, C 5 -C 20 aryloxy, acyl, acyloxy
  • the 15-PGDH inhibitor can include a compound having the following formula (II): pharmaceutically acceptable salt, tautomer, or solvate thereof; wherein n is 0-2 X 4 , X 5 , X 6 , and X 7 are independently N or CR c ; R 1 , R 6 , R 7 , and R c are the same or different and independently hydrogen or a substituted or unsubstituted group selected from C 1 -C 24 alkyl, C 2 -C 24 alkenyl, C 2 -C 24 alkynyl, C3-C20 aryl, heteroaryl, heterocycloalkenyl containing from 5-6 ring atoms, C6-C24 alkaryl, C 6 -C 24 aralkyl, halo, -Si(C 1 -C 3 alkyl) 3 , hydroxyl, sulfhydryl, C 1 -C 24 alkoxy, C2-C24 al
  • the 15-PGDH inhibitor can include a compound having the following formula (III) or (IV): (IV) or a pharmaceutically acceptable salt, tautomer, or solvate thereof; wherein n is 0-2 X 6 is independently N or CR c ; R 1 , R 6 , R 7 , and R c are the same or different and independently hydrogen or a substituted or unsubstituted group selected from C1-C24 alkyl, C2-C24 alkenyl, C2-C24 alkynyl, C3-C20 aryl, heteroaryl, heterocycloalkenyl containing from 5-6 ring atoms, C6-C24 alkaryl, C6-C24 aralkyl, halo, -Si(C1-C3 alkyl)3, hydroxyl, sulfhydryl, C1-C24 alkoxy, C2-C24 alkenyloxy, C2-C24 alkynyloxy,
  • R 6 and R 7 can each independently be one of the following:
  • R 73 , and R74 are the same or different and are independently selected from the group consisting of hydrogen, substituted or unsubstituted C1-C24 alkyl, C2-C24 alkenyl, C2-C24 alkynyl, C 3 -C 20 aryl, heterocycloalkenyl containing from 5-6 ring atoms, (wherein from 1-3 of the ring atoms is independently selected from N, NH, N(C1-C6 alkyl), NC(O)(C1-C6 alkyl), O, and S), heteroaryl or heterocyclyl containing from 5-14 ring atoms, (wherein from 1-6 of the ring atoms is independently selected from N, NH, N(C1-C3 alkyl), O, and S), C6-C24 alkaryl, C 6 -C 24 aralkyl, halo, silyl, hydroxyl, sulfhydryl, C 1 -C 24 alkoxy, C 2 -
  • R 6 and R 7 can independently be a group that improves aqueous solubility, for example, a phosphate ester (-OPO3H2), a phenyl ring linked to a phosphate ester (-OPO 3 H 2 ), a phenyl ring substituted with one or more methoxyethoxy groups, or a morpholine, or an aryl or heteroaryl ring substituted with such a group.
  • the 15-PGDH inhibitor can include a compound having the following formula (V): pharmaceutically acceptable salt, tautomer, or solvate thereof; wherein n is 0-2 X 6 is independently is N or CR c R 1 , R 6 , R 7 , and R c are the same or different each independently hydrogen or a substituted or unsubstituted group selected from C1-C24 alkyl, C2-C24 alkenyl, C2-C24 alkynyl, C 3 -C 20 aryl, heteroaryl, heterocycloalkenyl containing from 5-6 ring atoms, C 6 -C 24 alkaryl, C 6 -C 24 aralkyl, halo, -Si(C 1 -C 3 alkyl) 3 , hydroxyl, sulfhydryl, C 1 -C 24 alkoxy, C 2 -C 24 alkenyloxy, C2-C24 alkynyloxy, C5-
  • V pharmaceutically acceptable salt,
  • R 6 and R 7 can each independently be one of the following: each R 8 R9 R10 R11 R12 R13 R14 R15 16 17 18 19 20 21 22 23 24 25 26 27 28 , , , , , , , , R , R , R , R , R , R , R , R , R , R , R , R , R , R , R , R , R 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 , R , R , R , R , R , R , R , R , R , R , R , R , R , R , R , R , R , R , R , R , R , R
  • R 6 and R 7 can independently be a group that improves aqueous solubility, for example, a phosphate ester (-OPO3H2), a phenyl ring linked to a phosphate ester (-OPO 3 H 2 ), a phenyl ring substituted with one or more methoxyethoxy groups, or a morpholine, or an aryl or heteroaryl ring substituted with such a group.
  • the 15-PGDH inhibitor can include a compound having a structure of formula (IA): or a pharmaceutically acceptable salt, tautomer, or solvate thereof, wherein: R 1 is alkyl, haloalkyl, cycloalkyl, alkylene-cycloalkyl, alkylene-alkoxy, heterocyclyl, or alkylene-heterocyclyl; R 2 is –NH 2 , CN, or -NHC(O)alkyl; R 6 is heterocyclyl or heteroaryl, each of which is optionally substituted with one or more R 3 ; R 7 is alkyl, haloalkyl, cycloalkyl, aryl, heterocyclyl, heteroaryl, -C(O)-alkyl, -C(O)O-alkyl, or -C(O)NR 5 -alkyl, each of which is optionally substituted with one or more R 4 ; R 3 is ox
  • the 15-PGDH inhibitor can include a compound having a structure of formula (IIA): or a pharmaceutically acceptable salt, tautomer, or solvate thereof, wherein: R 1 is alkyl, haloalkyl, cycloalkyl, alkylene-cycloalkyl, alkylene-alkoxy, heterocyclyl, or alkylene-heterocyclyl; R 2 is –NH 2 , CN, or -NHC(O)alkyl; R 6 is heterocyclyl or heteroaryl, each of which is optionally substituted with one or more R 3 ; R 7 is alkyl, haloalkyl, cycloalkyl, aryl, heterocyclyl, heteroaryl, -C(O)-alkyl, -C(O)O-alkyl, or -C(O)NR 5 -alkyl, each of which is optionally substituted with one or more R 4 ; R 3 is ox
  • R 1 is C1-C6 alkyl, C1-C6 haloalkyl, 3- to 6-membered cycloalkyl, -(C 1 -C 6 alkylene)-(3- to 6-membered cycloalkyl), -(C 1 -C 6 alkylene)-(C 1 -C 6 alkoxy), 3- to 6-membered heterocyclyl, or -(C1-C6 alkylene)-(3- to 6-membered heterocyclyl).
  • R 1 is cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, -(CH 2 ) p -cyclopropyl, -(CH 2 ) p -cyclobutyl, -(CH 2 ) p -cyclopentyl, or -(CH 2 ) p -cyclohexyl; wherein p is 1, 2, or 3.
  • R 2 is –NH 2 .
  • R 6 is 5- to 6-membered heterocyclyl or 5- to 10- membered heteroaryl, each of which is optionally substituted with one or more R 3 .
  • R 6 is 5- to 6-membered heteroaryl optionally substituted with one or more R 3 .
  • R 6 is 8- to 10-membered bicyclic heteroaryl optionally substituted with one or more R 3 .
  • R 7 is C 1 -C 6 alkyl, C 1 -C 6 haloalkyl, 3- to 6-membered cycloalkyl, 6- to 10-membered aryl, 3- to 6-membered heterocyclyl, 5- to 10-membered heteroaryl, -C(O)(C 1 -C 6 alkyl), -C(O)O(C 1 -C 6 alkyl), or -C(O)NR 5 (C 1 -C 6 alkyl), each of which is optionally substituted with one or more R 4 .
  • R 7 is C 1 -C 6 alkyl, C 1 -C 6 haloalkyl, 3- to 6-membered cycloalkyl, phenyl, 3- to 6-membered heterocyclyl, or 5- to 10-membered heteroaryl, each of which is optionally substituted with one or more R 4 .
  • R 7 is C 1 -C 6 haloalkyl, 3- to 6-membered cycloalkyl, phenyl, 5- to 10-membered heteroaryl each of which is optionally substituted with one or more R 4 .
  • R 3 is -O-(C1-C6 alkylene)-N(R 5 )2, -N(R 5 )2, -N(R 5 )(C1-C6 alkylene-OH), -C(O)N(R 5 ) 2 , -C(O)N(R 5 )(C 1 -C 6 alkylene-OH), -C(O)(C 1 -C 6 alkyl), - C(O)O(C1-C6 alkyl), or -S(O)m(C1-C6 alkyl).
  • R 3 is -(C 1 -C 3 alkyl)OH, -NH 2 , -N(C 1 -C 3 alkyl) 2 , -NHCH2CH2OH, -N(C1-C3 alkyl)CH2CH2OH, N(CH2CH2OH)2, -NHCH2CH(CH2OH)2, -N(C 1 -C 3 alkyl)CH 2 CH(CH 2 OH) 2 , -NHCH 2 CH 2 OCH 2 CH 2 OH, -NHCH 2 CH 2 OCH 2 CH 2 NH 2 , -NHCH2CH2NH2, -N(C1-C3 alkyl)CH2CH2NH2, -NHCH2CH2NH(C1-C3 alkyl), -NHCH 2 CH 2 N(C 1 -C 3 alkyl) 2 , -N(C 1 -C 3 alkyl)CH 2 CH 2 NH(C 1 -C 3 alkyl)CH 2 CH 2 NH(
  • R 3 is -NH2, -N(C1-C3 alkyl)2, -NHCH2CH2OH, -N(C1-C3 alkyl)CH 2 CH 2 OH, N(CH 2 CH 2 OH) 2 , -NHCH 2 CH(CH 2 OH) 2 , -N(C 1 -C 3 alkyl)CH2CH(CH2OH)2, -NHCH2CH2OCH2CH2OH, -NHCH2CH2OCH2CH2NH2, -NHCH 2 CH 2 NH 2 , -N(C 1 -C 3 alkyl)CH 2 CH 2 NH 2 , -NHCH 2 CH 2 NH(C 1 -C 3 alkyl), -NHCH 2 CH 2 N(C 1 -C 3 alkyl) 2 , -N(C 1 -C 3 alkyl)CH 2 CH 2 NH(C 1 -C 3 alkyl), -N(C 1 -C 3 alkyl)CH 2 CH 2
  • R 3 is -NHCH 2 CH 2 OH or –N(CH 3 )CH 2 CH 2 OH.
  • R 4 is halogen, -CN, -N(R 5 ) 2 , -OH, -O-(C 1 -C 6 alkylene)- OH, -S(O) m (C 1 -C 6 alkyl), -C(O)(C 1 -C 6 alkyl), -C(O)-(3- to 6-membered cycloalkyl), C 1 -C 6 alkyl, C 1 -C 6 haloalkyl, 3- to 6-membered cycloalkyl, or 3- to 6-membered heterocyclyl.
  • n is 1.
  • the compound has the structure of formula (IIIA): or a pharmaceutically acceptable salt, tautomer, or solvate thereof, wherein: R 1 is 3- to 6-membered cycloalkyl, -(C 1 -C 6 alkylene)-(3- to 6-membered cycloalkyl), 3-to 6-membered heterocyclyl, or -(C1-C6 alkylene)-(3- to 6-membered heterocyclyl); R 2 is –NH2, CN, or -NHC(O)alkyl; R 6 is heterocyclyl or heteroaryl, each of which is optionally substituted with one or more R 3 ; R 7 is alkyl, haloalkyl, cycloalkyl, aryl, heterocyclyl, heteroaryl, -C(O)-alkyl, - C(O)O-alkyl, or -C(O)NR 5
  • R 1 is 3- to 5-membered cycloalkyl or -(C1-C6 alkylene)- (3- to 5-membered cycloalkyl).
  • R 1 is cyclobutyl.
  • R 1 is a bicyclic 4- to 6-membered cycloalkyl.
  • R 7 is C 1 -C 6 alkyl, C 1 -C 6 haloalkyl, 3- to 6-membered cycloalkyl, 6- to 10-membered aryl, 3- to 6-membered heterocyclyl, 5- to 10-membered heteroaryl, -C(O)(C 1 -C 6 alkyl), -C(O)O(C 1 -C 6 alkyl), or -C(O)NR 5 (C 1 -C 6 alkyl), each of which is optionally substituted with one or more R 4 .
  • R 7 is C 1 -C 4 alkyl, C 1 -C 6 haloalkyl, 3- to 6-membered cycloalkyl, phenyl, 3- to 6-membered heterocyclyl, or 5- to 10-membered heteroaryl, each of which is optionally substituted with one or more R 4 .
  • R 7 is C 1 -C 6 haloalkyl, 3- to 6-membered cycloalkyl, phenyl, 3- to 6-membered heterocyclyl, or 5- to 10- membered heteroaryl, each of which is optionally substituted with one or more R 4 .
  • R 7 is C1-C3 alkyl, C1-C3 haloalkyl, 3-membered cycloalkyl, phenyl, 4-membered heterocyclyl, or 5- to 6-membered heteroaryl, each of which is optionally substituted with one or more R 4 .
  • R 7 is C1-C3 haloalkyl, 3-membered cycloalkyl, phenyl, 4-membered heterocyclyl, or 5- to 6-membered heteroaryl, each of which is optionally substituted with one or more R 4 .
  • R 7 is -CF 3 , isopropyl, cyclopropyl, phenyl, pyridyl, pyrazole, or triazole, each of which is optionally substituted with one or more R 4 .
  • R 7 is -CF 3 , cyclopropyl, phenyl, pyridyl, pyrazole, or triazole, each of which is optionally substituted with one or more R 4 .
  • R 3 is -O-(C 1 -C 6 alkylene)-N(R 5 ) 2 , -N(R 5 ) 2 , -N(R 5 )(C 1 -C 6 alkylene-OH), -C(O)N(R 5 ) 2 , -C(O)N(R 5 )(C 1 -C 6 alkylene-OH), -C(O)(C 1 -C 6 alkyl), -C(O)O(C 1 -C 6 alkyl), or -S(O) m (C 1 -C 6 alkyl).
  • R 3 is -NH 2 , -N(C 1 -C 3 alkyl) 2 , -NHCH 2 CH 2 OH, -N(C 1 -C 3 alkyl)CH 2 CH 2 OH, N(CH 2 CH 2 OH) 2 , -NHCH 2 CH(CH 2 OH) 2 , -N(C 1 -C 3 alkyl)CH 2 CH(CH 2 OH) 2 , -NHCH 2 CH 2 OCH 2 CH 2 OH, -NHCH 2 CH 2 OCH 2 CH 2 NH 2 , -NHCH 2 CH 2 NH 2 , -NHCH 2 CH 2 NH 2 , -N(C 1 -C 3 alkyl)CH 2 CH 2 NH 2 , -NHCH 2 CH 2 NH(C 1 -C 3 alkyl), -NHCH 2 CH 2 N(C 1 -C 3 alkyl) 2 , -N(C 1 -C 3 alkyl)CH 2 CH 2 NH(C 1
  • R 3 is -NHCH2CH2OH or –N(CH3)CH2CH2OH.
  • substituents at the R 7 position could be modified to improve hERG activity, including hERG inhibition (IC 50 ), blockade, and efflux ratio.
  • IC 50 hERG inhibition
  • certain 6- to 10-membered aryls e.g., optionally substituted phenyl
  • 5-to 10-membered heteroaryls e.g., optionally substituted pyridyls, pyrazoles, and triazoles
  • the compound has the structure of formula (IVA): or a pharmaceutically acceptable salt, tautomer, or solvate thereof, wherein: R 1 is cycloalkyl, alkylene-cycloalkyl, alkylene-alkoxy, heterocyclyl, or alkylene-heterocyclyl; R 2 is –NH 2 , CN, or -NHC(O)alkyl; R 6 is heterocyclyl or heteroaryl, each of which is optionally substituted with one or more R 3 ; R 7 is C1-C6 haloalkyl, aryl or heteroaryl, each of which is optionally substituted with one or more R 4 ; R 3 is oxo, –OH, -O-alkylene-N(R 5 ).
  • R 7 is –CF 3 , pyridyl, pyrazole, phenyl, or triazole, each of which is optionally substituted with R 4 .
  • R 7 is –CF 3 , pyridyl, fluorophenyl, or a triazole optionally substituted with halogen or methyl.
  • R 7 is –CF 3. 7
  • R is .
  • R7 is [00190]
  • R 6 is 8- to 10-membered bicyclic heteroaryl optionally substituted with one or more R 3 .
  • the compound has the structure of formula (VA): (VA) or a pharmaceutically acceptable salt, tautomer, or solvate thereof, wherein: R 1 is cycloalkyl, -alkylene-cycloalkyl, -alkylene-alkoxy, heterocyclyl, or -alkylene-heterocyclyl; R 2 is –NH 2 , CN, or -NHC(O)alkyl; R 6 is heterocyclyl or heteroaryl, each of which is optionally substituted with one or more R 3 ; R 7 is 3- to 6-membered cycloalkyl, optionally substituted with one or more R 4 ; R 3 is oxo, –OH, -O-alkylene-N(R 5 ).
  • R 7 is cyclopropyl.
  • R 1 is 3- to 6-membered cycloalkyl, -(C1-C6 alkylene)-(3- to 6-membered cycloalkyl), -(C1-C6 alkylene)-(C1-C6 alkoxy), 3- to 6-membered heterocyclyl, or -(C1-C6 alkylene)-(3- to 6-membered heterocyclyl).
  • R 1 is cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, -(CH2)p-cyclopropyl, -(CH2)p-cyclobutyl, -(CH2)p-cyclopentyl, or -(CH2)p-cyclohexyl; wherein p is 1, 2, or 3.
  • R 3 is -NH2, -N(C1-C3 alkyl)2, -NHCH2CH2OH, -N(C1-C3 alkyl)CH 2 CH 2 OH, N(CH 2 CH 2 OH) 2 , -NHCH 2 CH(CH 2 OH) 2 , -N(C 1 -C 3 alkyl)CH2CH(CH2OH)2, -NHCH2CH2OCH2CH2OH, -NHCH2CH2OCH2CH2NH2, -NHCH 2 CH 2 NH 2 , -N(C 1 -C 3 alkyl)CH 2 CH 2 NH 2 , -NHCH 2 CH 2 NH(C 1 -C 3 alkyl), -NHCH2CH2N(C1-C3 alkyl)2, -N(C1-C3 alkyl)CH2CH2NH(C1-C3 alkyl), -N(C1-C3 alkyl)CH 2 CH2NH(C1-C3
  • R 3 is -NHCH 2 CH 2 OH or –N(CH 3 )CH 2 CH 2 OH.
  • R 3 is -O-(C1-C6 alkylene)-N(R 5 )2, -N(R 5 )2 or -N(R 5 )(C1- C 6 alkylene-OH).
  • R 5 is H, C1-C6 alkyl, -(C1-C6 alkylene)-OH, or -S(O)2(C1- C 3 alkyl).
  • R 3 is -NH 2 , -N(C 1 -C 3 alkyl) 2 , -NHCH 2 CH 2 OH, -N(C 1 -C 3 alkyl)CH 2 CH 2 OH, N(CH 2 CH 2 OH) 2 , -NHCH 2 CH(CH 2 OH) 2 , -N(C 1 -C 3 alkyl)CH 2 CH(CH 2 OH) 2 , -NHCH 2 CH 2 OCH 2 CH 2 OH, -NHCH 2 CH 2 OCH 2 CH 2 NH 2 , -NHCH 2 CH 2 NH 2 , -NHCH 2 CH 2 NH 2 , -N(C 1 -C 3 alkyl)CH 2 CH 2 NH 2 , -NHCH 2 CH 2 NH(C 1 -C 3 alkyl), -NHCH 2 CH 2 N(C 1 -C 3 alkyl) 2 , -N(C 1 -C 3 alkyl)CH 2 CH 2 NH(C 1
  • R 3 is -NHCH 2 CH 2 OH or –N(CH 3 )CH 2 CH 2 OH. [00205] In still other embodiments, R 3 is –NHCH 2 CH 2 OH. [00206] In some embodiments, R 6 is 5- to 6-membered heterocyclyl or 5- to 10- membered heteroaryl, each of which is optionally substituted with one or more R 3 . [00207] In other embodiments, R 6 is 5- to 6-membered heteroaryl optionally substituted with one or more R 3 .
  • R 6 is furan, thiophene, pyrrole, thiazole, isothiazole, oxazole, isooxazole, pyrazole, imidazole, triazole, pyridine, pyrimidine, pyridazine, or pyrazine, each optionally substituted with one or more R 3 .
  • R 6 is thiazole, imidazole, oxazole, pyridine, or pyrimidine.
  • R 6 is 8- to 10-membered bicyclic heteroaryl optionally substituted with one or more R 3 .
  • R 6 is 5- to 6-membered heterocyclyl, optionally substituted with one or more R 3 , selected from morpholine, pyridine-one, or piperidine.
  • R 7 is C1-C3 haloalkyl, 3-membered cycloalkyl, phenyl, 4-membered heterocyclyl, or 5- to 6-membered heteroaryl, each of which is optionally substituted with one or more R 4 .
  • R 7 is -CF 3 , cyclopropyl, phenyl, pyrazole, pyridyl, or triazole, each of which is optionally substituted with one or more R 4 .
  • the compound has the structure of formula (VIIA): or a pharmaceutically acceptable salt, tautomer, or solvate thereof, wherein: R 1 is cycloalkyl, alkylene-cycloalkyl, alkylene-alkoxy, heterocyclyl, or alkylene-heterocyclyl; R 2 is –NH2, CN, or -NHC(O)alkyl; R 6 is fused bicyclic heterocyclyl or fused bicyclic heteroaryl, each of which is optionally substituted with one or more R 3 ; R 7 is alkyl, haloalkyl, cycloalkyl, aryl, heterocyclyl, heteroaryl, -C(O)-alkyl, -C(O)O-alkyl, or -C(O)NR 5 -alkyl, each of which is optionally substituted with one or more R 4 ; R 3 is oxo, -OH, -O-alkyl, or a pharmaceutically acceptable
  • R 6 is 8- to 10-membered fused bicyclic heteroaryl, each of which is optionally substituted with one or more R 3 .
  • R 7 is C 1 -C 6 alkyl, C 1 -C 6 haloalkyl, 3- to 6-membered cycloalkyl, 6- to 10-membered aryl, 3- to 6-membered heterocyclyl, 5- to 10-membered heteroaryl, -C(O)(C 1 -C 6 alkyl), -C(O)O(C 1 -C 6 alkyl), or -C(O)NR 5 (C 1 -C 6 alkyl), each of which is optionally substituted with one or more R 4 .
  • R 7 is C 1 -C 4 alkyl, C 1 -C 6 haloalkyl, 3- to 6-membered cycloalkyl, phenyl, 3- to 6-membered heterocyclyl, or 5- to 10-membered heteroaryl, each of which is optionally substituted with one or more R 4 .
  • R 7 is C 1 -C 6 haloalkyl, 3- to 6-membered cycloalkyl, phenyl, 3- to 6-membered heterocyclyl, or 5- to 10- membered heteroaryl, each of which is optionally substituted with one or more R 4 .
  • R 7 is C 1 -C 3 alkyl, C 1 -C 6 haloalkyl, 3- to 6-membered cycloalkyl, phenyl, 3- to 6-membered heterocyclyl, or 5- to 6-membered heteroaryl, each of which is optionally substituted with one or more R 4 .
  • R 7 is C1-C3 alkyl, C1-C6 haloalkyl, 3- to 6-membered cycloalkyl, phenyl, pyrazole, pyridyl, or triazole, each of which is optionally substituted with one or more R 4 .
  • R 7 is C1-C6 haloalkyl, 3- to 6-membered cycloalkyl, phenyl, pyrazole, pyridyl, or triazole, each of which is optionally substituted with one or more R 4 .
  • R 7 is -CF 3 , isopropyl, cyclopropyl, phenyl, pyridyl, or triazole, each of which is optionally substituted with one or more R 4 .
  • R 7 is -CF 3 , cyclopropyl, phenyl, pyridyl, or triazole, each of which is optionally substituted with one or more R 4 .
  • R 1 is C 1 -C 6 alkyl, C 1 -C 6 haloalkyl, 3- to 6-membered cycloalkyl, -(C1-C6 alkylene)-(3- to 6-membered cycloalkyl), -(C1-C6 alkylene)-(C1-C6 alkoxy), 3- to 6-membered heterocyclyl, or -(C 1 -C 6 alkylene)-(3- to 6-membered heterocyclyl).
  • R 1 is cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, -(CH2)p-cyclopropyl, -(CH2)p-cyclobutyl, -(CH2)p-cyclopentyl, or -(CH2)p-cyclohexyl; wherein p is 1, 2, or 3.
  • R 3 is -O-(C1-C6 alkylene)-N(R 5 )2, -N(R 5 )2, -N(R 5 )(C1-C6 alkylene-OH), -C(O)N(R 5 ) 2 , -C(O)N(R 5 )(C 1 -C 6 alkylene-OH), -C(O)(C 1 -C 6 alkyl), -C(O)O(C1-C6 alkyl), or -S(O)m(C1-C6 alkyl).
  • R 3 is -NH 2 , -N(C 1 -C 3 alkyl) 2 , -NHCH 2 CH 2 OH, -N(C 1 -C 3 alkyl)CH2CH2OH, N(CH2CH2OH)2, -NHCH2CH(CH2OH)2, -N(C1-C3 alkyl)CH 2 CH(CH 2 OH) 2 , -NHCH 2 CH 2 OCH 2 CH 2 OH, -NHCH 2 CH 2 OCH 2 CH 2 NH 2 , -NHCH 2 CH 2 NH 2 , -N(C 1 -C 3 alkyl)CH 2 CH 2 NH 2 , -NHCH 2 CH 2 NH(C 1 -C 3 alkyl), -NHCH 2 CH 2 N(C 1 -C 3 alkyl) 2 , -N(C 1 -C 3 alkyl)CH 2 CH 2 NH(C 1 -C 3 alkyl), -N(C 1 -C 3 alky
  • R 3 is -NHCH 2 CH 2 OH or –N(CH 3 )CH 2 CH 2 OH.
  • R 4 is halogen, -CN, -N(R 5 ) 2 , -OH, -O-(C 1 -C 6 alkylene)- OH, -S(O) m (C 1 -C 6 alkyl), -C(O)(C 1 -C 6 alkyl), -C(O)-(3- to 6-membered cycloalkyl), C 1 -C 6 alkyl, C 1 -C 6 haloalkyl, 3- to 6-membered cycloalkyl, or 3- to 6-membered heterocyclyl.
  • the compound has the structure of formula (VIIIA): or a pharmaceutically acceptable salt, tautomer, or solvate thereof, wherein: R 1 is cyclobutyl or -(C1-C4 alkylene)-(C1-C3 alkoxy); R 2 is –NH 2 , CN, or -NHC(O)alkyl; R 6 is heterocyclyl or heteroaryl, each of which is optionally substituted with one or more R 3 ; R 3 is oxo, -OH, -O-alkylene-OH, -O-alkylene-N(R 5 )2, -N(R 5 )2, - N(R 5 )(alkylene-OH), -N(R 5 )(alkylene-O-alkyl), alkyl, -alkylene-OH, haloalkyl, cycloalkyl, heterocyclyl, -C(O)N(R 5 )2,
  • R 2 is –NH2.
  • R 6 is 5- to 6-membered heterocyclyl or 5- to 10- membered heteroaryl, each of which is optionally substituted with one or more R 3 .
  • R 6 is 5- to 6-membered heteroaryl optionally substituted with one or more R 3 .
  • R 6 is 8- to 10-membered bicyclic heteroaryl optionally substituted with one or more R 3 .
  • R 3 is -O-(C 1 -C 6 alkylene)-N(R 5 ) 2 , -N(R 5 ) 2 , -N(R 5 )(C 1 -C 6 alkylene-OH), -C(O)N(R 5 )2, -C(O)N(R 5 )(C1-C6 alkylene-OH), -C(O)(C1-C6 alkyl), -C(O)O(C 1 -C 6 alkyl), or -S(O) m (C 1 -C 6 alkyl).
  • R 3 is -(C1-C3 alkyl)OH, -NH2, -N(C1-C3 alkyl)2, -NHCH 2 CH 2 OH, -N(C 1 -C 3 alkyl)CH 2 CH 2 OH, N(CH 2 CH 2 OH) 2 , -NHCH 2 CH(CH 2 OH) 2 , -N(C1-C3 alkyl)CH2CH(CH2OH)2, -NHCH2CH2OCH2CH2OH, -NHCH2CH2OCH2CH2NH2, -NHCH 2 CH 2 NH 2 , -N(C 1 -C 3 alkyl)CH 2 CH 2 NH 2 , -NHCH 2 CH 2 NH(C 1 -C 3 alkyl), -NHCH2CH2N(C1-C3 alkyl)2, -N(C1-C3 alkyl)CH2CH2NH(C1-C3 alkyl),
  • R 3 is -NHCH 2 CH 2 OH or –N(CH 3 )CH 2 CH 2 OH.
  • R 4 is halogen, -CN, -N(R 5 ) 2 , -OH, -O-(C 1 -C 6 alkylene)- OH, -S(O) m (C 1 -C 6 alkyl), -C(O)(C 1 -C 6 alkyl), -C(O)-(3- to 6-membered cycloalkyl), C 1 -C 6 alkyl, C 1 -C 6 haloalkyl, 3- to 6-membered cycloalkyl, or 3- to 6-membered heterocyclyl.
  • n is 1.
  • the compound has the structure of formula (IXA): or a pharmaceutically acceptable salt, tautomer, or solvate thereof, wherein: R 1 is cyclobutyl or -(C1-C4 alkylene)-(C1-C3 alkoxy); R 2 is –NH2, CN, or -NHC(O)alkyl;
  • R 1 is cyclobutyl.
  • R 3 is - NH(C 1 -C 4 alkylene)-OH (e.g., -NH(C 2 -C 4 alkylene)-OH). In some embodiments of Formula (IXA), R 3 is C1-C3 alkyl (e.g., methyl or ethyl). [00242] In some embodiments, of Formula (IXA), R 7 is –CF 3 , isopropyl, cyclopropyl, or cyclobutyl. In some embodiments, of Formula (IXA), R 7 is isopropyl. In some embodiments of Formula (VII), R7 is , , each of which is optionally substituted with one or more R 4 .
  • each R 4 is independently selected from methyl or ethyl.
  • X is –CH.
  • Examples of compounds having formulas (I), (II), (III), (IV), (V), (IA), (IIA), (IIIA), (IVA), (VA), (VIA), (VIIA), (VIIIA), and (IXA) are described in U.S. Patent Application Publication Nos.2015/0072998, 2017/0165241, 2017/0173028, 2018/0118756, WO2018/218251, and WO2020/106998, all of which are incorporated by reference in their entirety.
  • the 15-PGDH inhibitor can include a compound selected from the group consisting of: , , , , , ,
  • R 7 and R 8 are same or different and are each independently selected from the group consisting of H, a substituted or unsubstituted aryl, a substituted or unsubstituted heteroaryl, a substituted or unsubstituted cycloalkyl, and a substituted or unsubstituted heterocyclyl, and at least one of R 7 or R 8 is not H;
  • R 9 , R 10 , R 11 , R 12 , R 13 , and R 15 are the same or different and are independently selected from the group consisting of hydrogen, substituted or unsubstituted C1-C24 alkyl, C 2 -C 24 alkenyl, C 2 -C 24 alkynyl, C 3 -C 20 aryl, heterocycloalkenyl containing from 5-7 ring atoms, (wherein from 1-3 of the ring atoms is independently selected
  • Examples of 15-PGDH inhibitors having formulas (IB), (IBa), (IBb), (IBc), (IBd), (IBe), (IIB), (IIBa), (IIBb), (IIBc), (IIBd), (IBe), or (IIBf) can include the following compounds:
  • 15-PGDH inhibitors include compounds described in WO2022/032230A1, WO2021/236779A1, WO2020/106998A1, WO2018/218251A1, WO2018/145080A1, WO2016/168472A1, WO2015/065716A1, WO2013/158649A1, WO2024/233550A1, U.S. Patent Application Publication No.2024/0294519A1, U.S. Patent Application Publication No.2024/0374579A1, and U.S. Patent Application Publication No. 2024/0409557 all of which are herein incorporated by reference in its entirety.
  • the 15-PGDH inhibitor can be a 5,6-fused and 6,6-fused bicyclic alcohol or ether described in U.S. Patent Application Publication No.2024/0409557.
  • a 5,6- fused and 6,6-fused bicyclic alcohol or ether can be a compound of Formula (A): or pharmaceutically acceptable salts thereof; m is 0 or 1; X is N or CH; Y is N or CR 3 ; Z is N or CR 5 ; b is N or CR 4 ; R 1a is H, deuterium, C 1-6 alkyl, or C 1-6 haloalkyl; wherein, when R 1a is C1-6 alkyl or C1-6 haloalkyl, then R 1a may be unsubstituted or substituted with one or more substituents, each of which independently is halogen, -OH, -N(R b ) 2 , C 1-3 alkyl, or C1-3 alkoxy; R 1b is
  • the small molecule 15-PGDH inhibitor can be selected that can ia) at 2.5 ⁇ M concentration, stimulate a Vaco503 reporter cell line expressing a 15- PGDH luciferase fusion construct to a luciferase output level of greater than 70 (using a scale on which a value of 100 indicates a doubling of reporter output over baseline); iia) at 2.5 ⁇ M concentration stimulate a V9m reporter cell line expressing a 15-PGDH luciferase fusion construct to a luciferase output level of greater than 75; iiia) at 7.5 ⁇ M concentration stimulate a LS174T reporter cell line expressing a 15-PGDH luciferase fusion construct to a luciferase output level of greater than 70; and iva) at 7.5 ⁇ M concentration, does not activate a negative control V9m cell line expressing TK-renilla luciferase reporter to a level greater than
  • the small molecule 15-PGDH inhibitor can ib) at 2.5 ⁇ M concentration, stimulate a Vaco503 reporter cell line expressing a 15-PGDH luciferase fusion construct to increase luciferase output; iib) at 2.5 ⁇ M concentration stimulate a V9m reporter cell line expressing a 15-PGDH luciferase fusion construct to increase luciferase output; iiib) at 7.5 ⁇ M concentration stimulate a LS174T reporter cell line expressing a 15-PGDH luciferase fusion construct to increase luciferase output; ivb) at 7.5 ⁇ M concentration, does not activate a negative control V9m cell line expressing TK-renilla luciferase reporter to a luciferase level greater than 20% above background; and vb) inhibits the enzymatic activity of recombinant 15-PGDH protein at an IC 50 of less than 1
  • the small molecule 15-PGDH inhibitor can inhibit the enzymatic activity of recombinant 15-PGDH at an IC 50 of less than 1 ⁇ M, or preferably at an IC50 of less than 250 nM, or more preferably at an IC50 of less than 50 nM, or more preferably at an IC 50 of less than 10 nM, or more preferably at an IC 50 of less than 5 nM at a recombinant 15-PGDH concentration of about 5 nM to about 10 nM.
  • 15-PGDH inhibitors can be used in the methods described herein.
  • 15-PGDH inhibitors can include known 15-PGDH inhibitors including, for example, tetrazole compounds of formulas (I) and (II), 2-alkylideneaminooxyacetamide compounds of formula (I), heterocyclic compounds of formulas (VI) and (VII), and pyrazole compounds of formula (III) described in U.S. Patent Application Publication No.2006/0034786 and U.S. Patent No.7,705,041; benzylidene-1,3- thiazolidine compounds of formula (I) described in U.S.
  • Patent Application Publication No.2007/0071699 phenylfurylmethylthiazolidine-2,4-dione and phenylthienylmethylthiazolidine-2,4-dione compounds described in U.S. Patent Application Publication No.2007/0078175; thiazolidenedione derivatives described in U.S. Patent Application Publication No.2011/0269954; phenylfuran, phenylthiophene, or phenylpyrrazole compounds described in U.S. Patent No.7,294,641, 5-(3,5-disubstituted phenylazo)-2-hydroxybenzene-acetic acids and salts and lactones described in U.S.
  • PubMed PMID 21650226; Duveau DY et al. Structure-activity relationship studies and biological characterization of human NAD(+)- dependent 15-hydroxyprostaglandin dehydrogenase inhibitors. Bioorg Med Chem Lett. 2014;24(2):630-5. doi: 10.1016/j.bmcl.2013.11.081. PubMed PMID: 24360556; PMCID: PMC3970110; Duveau DY et al. Discovery of two small molecule inhibitors, ML387 and ML388, of human NAD+-dependent 15-hydroxyprostaglandin dehydrogenase. Probe Reports from the NIH Molecular Libraries Program.
  • the 15-PGDH inhibitor can include an inhibitory nucleic acid, e.g., antisense DNA or RNA, morpholino oligomer, small interfering RNA (siRNA), microRNA (miRNA), or short hairpin RNA (shRNA).
  • an inhibitory nucleic acid e.g., antisense DNA or RNA, morpholino oligomer, small interfering RNA (siRNA), microRNA (miRNA), or short hairpin RNA (shRNA).
  • the inhibitory RNA targets a sequence that is identical or substantially identical (e.g., at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 91%, at least about 92%, at least about 93%, at least about 94%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identical) to a target sequence in a 15-PGDH polynucleotide (e.g., a portion comprising at least 20, at least 30, at least 40, at least 50, at least 60, at least 70, at least 80, at least 90, or at least 100 contiguous nucleotides, e.g., from 20-500, 20-250, 20-100, 50-500, or 50-250 contiguous nucleotides of a 15-PGDH-encoding polynucleotide sequence (e.g., the human HPGD gene, Gene ID: 3248, including
  • the methods described herein comprise treating a subject with diseases or disorders characterized by structural deterioration of the blood-brain barrier and/or by increased generation of reactive oxygen species within the nervous system using an shRNA or siRNA.
  • a shRNA is an artificial RNA molecule with a hairpin turn that can be used to silence target gene expression via the siRNA it produces in cells. See, e.g., Fire et.
  • a method of treating a subject comprises administering to the subject a therapeutically effective amount of a modified RNA or a vector comprising a polynucleotide that encodes an shRNA or siRNA capable of hybridizing to a portion of a 15-PGDH mRNA (e.g., a portion of the human 15-PGDH-encoding polynucleotide sequence set forth in any of GenBank Accession Nos. NM_000860.6.
  • the vector further comprises appropriate expression control elements known in the art, including, e.g., promoters (e.g., inducible promoters or tissue specific promoters), enhancers, and transcription terminators.
  • promoters e.g., inducible promoters or tissue specific promoters
  • enhancers e.g., promoters, enhancers, and transcription terminators.
  • the 15-PGDH inhibitor is a 15-PGDH-specific microRNA (miRNA or miR).
  • miRNA is a small non-coding RNA molecule that functions in RNA silencing and post-transcriptional regulation of gene expression. miRNAs base pair with complementary sequences within the mRNA transcript.
  • the mRNA transcript may be silenced by one or more of the mechanisms, such as cleavage of the mRNA strand, destabilization of the mRNA through shortening of its poly(A) tail, and decrease in the translation efficiency of the mRNA transcript into proteins by ribosomes.
  • the 15-PGDH inhibitor may be a morpholino oligomer, an antisense oligonucleotide, e.g., an RNase H-dependent antisense oligonucleotide (ASO), or combinations thereof.
  • ASOs are single-stranded, chemically modified oligonucleotides that bind to complementary sequences in target mRNAs and reduce gene expression both by RNase H-mediated cleavage of the target RNA and by inhibition of translation by steric blockade of ribosomes.
  • the oligonucleotide is capable of hybridizing to a portion of a 15-PGDH mRNA (e.g., a portion of a human 15-PGDH-encoding polynucleotide sequence as set forth in any of GenBank Accession Nos.
  • the oligonucleotide has a length of about 10-30 nucleotides (e.g., 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, or 30 nucleotides). In some embodiments, the oligonucleotide has 100% complementarity to the portion of the mRNA transcript it binds.
  • the DNA oligonucleotide has less than 100% complementarity (e.g., about 95%, about 90%, about 85%, about 80%, about 75%, or about 70% complementarity) to the portion of the mRNA transcript it binds, but can still form a stable RNA:DNA duplex for the RNase H to cleave the mRNA transcript.
  • Suitable antisense molecules, siRNA, miRNA, and shRNA can be produced by standard methods of oligonucleotide synthesis or by ordering such molecules from a contract research organization or supplier by providing the polynucleotide sequence being targeted.
  • Inhibitory nucleic acids can also include morpholino oligomers (DNA bases attached to a backbone of methylenemorpholine rings linked through phosphordiamidate groups), which block access to short (approximately 25 bases) specific sequences of the base- pairing surfaces of ribonucleic acid and are used to knock down gene function. Also included are RNA aptamers, which are short, synthetic oligonucleotide sequences that bind to proteins (see, e.g., Li et al., Nuc. Acids Res. (2006), 34:6416-24). Both are notable for both high affinity and specificity for the targeted molecule, and have the additional advantage of being smaller than antibodies (usually less than 6 kD).
  • RNA aptamers with a desired specificity are generally selected from a combinatorial library, and can be modified to reduce vulnerability to ribonucleases, using methods known in the art.
  • the 15-PGDH inhibitor is an anti-15-PGDH antibody or an antigen-binding fragment thereof.
  • the antibody is a blocking antibody (e.g., an antibody that binds to a target and directly interferes with the target's function, e.g., 15-PGDH enzyme activity).
  • the antibody is a neutralizing antibody (e.g., an antibody that binds to a target and negates the downstream cellular effects of the target).
  • the antibody binds to human 15-PGDH.
  • the antibody is a monoclonal antibody. In some embodiments, the antibody is a polyclonal antibody. In some embodiments, the antibody is a chimeric antibody. In some embodiments, the antibody is a humanized antibody. In some embodiments, the antibody is a human antibody. In some embodiments, the antibody is an antigen-binding fragment, such as a F(ab′)2, Fab′, Fab, scFv, and the like. The term “antibody or antigen-binding fragment” can also encompass multi-specific and hybrid antibodies, with dual or multiple antigen or epitope specificities.
  • an anti-15-PGDH antibody comprises a heavy chain sequence or a portion thereof, and/or a light chain sequence or a portion thereof, of an antibody sequence disclosed herein.
  • an anti-15-PGDH antibody comprises one or more complementarity determining regions (CDRs) of an anti-15-PGDH antibody as disclosed herein.
  • an anti-15-PGDH antibody is a nanobody, or single-domain antibody (sdAb), comprising a single monomeric variable antibody domain, e.g., a single VHH domain.
  • sdAb single-domain antibody
  • antibodies are prepared by immunizing an animal or animals (such as mice, rabbits, or rats) with an antigen for the induction of an antibody response.
  • the antigen is administered in conjugation with an adjuvant (e.g., Freund's adjuvant).
  • an adjuvant e.g., Freund's adjuvant
  • one or more subsequent booster injections of the antigen can be administered to improve antibody production.
  • antigen-specific B cells are harvested, e.g., from the spleen and/or lymphoid tissue. For generating monoclonal antibodies, the B cells are fused with myeloma cells, which are subsequently screened for antigen specificity.
  • genes encoding the heavy and light chains of an antibody of interest can be cloned from a cell, e.g., the genes encoding a monoclonal antibody can be cloned from a hybridoma and used to produce a recombinant monoclonal antibody.
  • Gene libraries encoding heavy and light chains of monoclonal antibodies can also be made from hybridoma or plasma cells.
  • phage or yeast display technology can be used to identify antibodies and heteromeric Fab fragments that specifically bind to selected antigens (see, e.g., McCafferty et al., Nature 348:552-554 (1990); Marks et al., Biotechnology 10:779-783 (1992); Lou et al. m PEDS 23:311 (2010); and Chao et al., Nature Protocols, 1:755-768 (2006)).
  • antibodies and antibody sequences may be isolated and/or identified using a yeast-based antibody presentation system, such as that disclosed in, e.g., Xu et al., Protein Eng Des Sel, 2013, 26:663-670; WO 2009/036379; WO 2010/105256; and WO 2012/009568. Random combinations of the heavy and light chain gene products generate a large pool of antibodies with different antigenic specificity (see, e.g., Kuby, Immunology (3rd ed.1997)). Techniques for the production of single chain antibodies or recombinant antibodies (U.S. Pat. Nos.4,946,778, 4,816,567) can also be adapted to produce antibodies.
  • Antibodies can be produced using any number of expression systems, including prokaryotic and eukaryotic expression systems.
  • the expression system is a mammalian cell, such as a hybridoma, or a CHO cell. Many such systems are widely available from commercial suppliers.
  • the VH and VL regions may be expressed using a single vector, e.g., in a di-cistronic expression unit, or be under the control of different promoters. In other embodiments, the VH and VL region may be expressed using separate vectors.
  • an anti-15-PGDH antibody comprises one or more CDR, heavy chain, and/or light chain sequences that are affinity matured.
  • chimeric antibodies methods of making chimeric antibodies are known in the art.
  • chimeric antibodies can be made in which the antigen binding region (heavy chain variable region and light chain variable region) from one species, such as a mouse, is fused to the effector region (constant domain) of another species, such as a human.
  • “class switched” chimeric antibodies can be made in which the effector region of an antibody is substituted with an effector region of a different immunoglobulin class or subclass.
  • an anti-15-PGDH antibody comprises one or more CDR, heavy chain, and/or light chain sequences that are humanized.
  • humanized antibodies methods of making humanized antibodies are known in the art. See, e.g., U.S. Pat. No. 8,095,890.
  • a humanized antibody has one or more amino acid residues introduced into it from a source which is non-human.
  • human antibodies can be generated.
  • transgenic animals e.g., mice
  • mice can be produced that are capable, upon immunization, of producing a full repertoire of human antibodies in the absence of endogenous immunoglobulin production.
  • JH antibody heavy-chain joining region
  • antibody fragments such as a Fab, a Fab′, a F(ab′)2, a scFv, nanobody, or a diabody
  • Various techniques have been developed for the production of antibody fragments, such as proteolytic digestion of intact antibodies (see, e.g., Morimoto et al., J. Biochem. Biophys. Meth., 24:107-117 (1992); and Brennan et al., Science, 229:81 (1985)) and the use of recombinant host cells to produce the fragments.
  • antibody fragments can be isolated from antibody phage libraries.
  • Fab′-SH fragments can be directly recovered from E. coli cells and chemically coupled to form F(ab′)2 fragments (see, e.g., Carter et al., BioTechnology, 10:163-167 (1992)).
  • F(ab′)2 fragments can be isolated directly from recombinant host cell culture.
  • Other techniques for the production of antibody fragments will be apparent to those skilled in the art. [00273] Methods for measuring binding affinity and binding kinetics are known in the art.
  • the nucleic acid molecules may be used in a TALEN (transcription activator-like effector nuclease technology), zinc-finger nuclease or CRISPR- Cas system to target a nucleic acid sequence for genetic screening, targeted transcriptional regulation, targeted knock-in, and targeted genome editing, including base editing, epigenetic editing, and introducing double strand breaks (DSBs) for homologous recombination- mediated insertion of a nucleotide sequence.
  • TALEN transcription activator-like effector nuclease technology
  • zinc-finger nuclease or CRISPR- Cas system to target a nucleic acid sequence for genetic screening, targeted transcriptional regulation, targeted knock-in, and targeted genome editing, including base editing, epigenetic editing, and introducing double strand breaks (DSBs) for homologous recombination- mediated insertion of a nucleotide sequence.
  • Genome editing can refer to the targeted modification of a DNA sequence, including but not limited to, adding, removing, replacing, or modifying existing DNA sequences, and inducing chromosomal rearrangements or modifying transcription regulation elements (e.g., methylation/demethylation of a promoter sequence of a gene) to alter gene expression.
  • inhibiting 15-PGDH expression comprises inserting a silencer sequence near a polynucleotide sequence encoding 15-PGDH.
  • upregulating PGE2 synthesis comprises inserting an enhancer sequence near a polynucleotide sequence encoding PGE2.
  • CRISPR-Cas system requires a guide system that can locate Cas protein to the target DNA site in the genome.
  • the guide system comprises a crispr RNA (crRNA) with a 17-20 nucleotide sequence that is complementary to a target DNA site and a trans-activating crRNA (tracrRNA) scaffold recognized by the Cas protein (e.g., Cas9).
  • the 17-20 nucleotide sequence complementary to a target DNA site is referred to as a spacer while the 17-20 nucleotide target DNA sequence is referred to a protospacer.
  • crRNAs and tracrRNAs exist as two separate RNA molecules in nature, single guide RNA (sgRNA or gRNA) can be engineered to combine and fuse crRNA and tracrRNA elements into one single RNA molecule.
  • the CRISPR guide system comprises two or more RNAs, e.g., crRNA and tracrRNA.
  • the CRISPR guide system comprises a sgRNA comprising a spacer sequence for genomic targeting and a scaffold sequence for Cas protein binding.
  • the guide system naturally comprises a sgRNA.
  • Cas12a/Cpf1 utilizes a guide system lacking tracrRNA and comprising only a crRNA containing a spacer sequence and a scaffold for Cas12a/Cpf1 binding. While the spacer sequence can be varied depending on a target site in the genome, the scaffold sequence for Cas protein binding can be identical for all gRNAs.
  • CRISPR-Cas systems described herein can comprise different CRISPR enzymes.
  • the CRISPR-Cas system can comprise Cas9, Cas12a/Cpf1, Cas12b/C2c1, Cas12c/C2c3, Cas12d/CasY, Cas12e/CasX, Cas12g, Cas12h, or Cas12i.
  • Non- limiting examples of Cas enzymes include, but are not limited to, Cas1, CasiB, Cas2, Cas3, Cas4, Cas5, Cas5d, Cas5t, Cas5h, Cas5a, Cas6, Cas7, Cas8, Cas8a, Cas8b, Cas8c, Cas9 (also known as Csn1 or Csx12), Cas10, Cas10d, Cas12a/Cpf1, Cas12b/C2c1, Cas12c/C2c3, Cas12d/CasY, Cas12e/CasX, Cas12f/Cas14/C2c10, Cas12g, Cas12h, Cas12i, Cas12k/C2c5, Cas13a/C2c2, Cas13b, Cas13c, Cas13d, C2c4, C2c8, C2c9, Csy1, Csy2, Csy3, Csy4, Cse1, Cs
  • Type II Cas effector proteins Type V Cas effector proteins
  • Type VI Cas effector proteins CARF, DinG, homologues thereof, or modified or engineered versions thereof such as dCas9 (endonuclease-dead Cas9) and nCas9 (Cas9 nickase that has inactive DNA cleavage domain).
  • compositions, methods, devices, and systems, described herein may use the Cas9 nuclease from Streptococcus pyogenes, of which amino acid sequences and structures are well known to those skilled in the art.
  • Binding of a Cas protein to its target DNA sequence i.e. target dsDNA
  • PAM protospacer adjacent motif
  • Cas proteins from different bacterial species recognize different PAM sequences and make cuts 3-4 nucleotides upstream (e.g., Cas9) or 18-23 nucleotides downstream (e.g., Cas12a/Cpf1) of the PAM sequence.
  • Cas9 from Streptococcus pyogene recognizes the PAM sequence 5′-NGG-3′ (wherein “N” can be any nucleotide) and cleaves 3 nucleotide upstream of the PAM sequence.
  • Cas9 from Staphylococcus aureus recognizes the PAM sequence 5′-NNGRRN-3′ (wherein “N” can be any nucleotide) and cleaves 4 nucleotide upstream of the PAM sequence.
  • N can be any nucleotide
  • the PAM sequence is essential for Cas-mediated cleavage, in some instances, the gRNA sequence does not comprise a PAM sequence.
  • nucleic acid molecules comprising a target sequence comprising a spacer that is complementary to a sequence at a target site in the genome.
  • a spacer, as described herein can comprise about 10 to about 25 nucleotides in length.
  • a spacer sequence that is complementary to a target site sequence in the genome can be about 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 22, 23, 24, 25, 26, 27, 27, 28, 30, or more nucleotides in length.
  • a target site, as described herein can comprise a sequence of about 20 nucleotides immediately upstream or 5′ of the first nucleotide of the PAM.
  • guide system nucleic acid molecules wherein at least 1, at least 2, or at least 3 consecutive nucleotides at the 5′ terminus of the guide system nucleic acid molecules are 100% identical to the 3′ terminus of the target sequence in a genome.
  • guide system nucleic acid molecules exhibiting enhanced editing efficiency of a target sequence when guide system nucleic acid molecules are contacted with the target sequence in a complex with a CRISPR-Cas system (e.g., CRISPR- Cas9, CRISPR-Cas12a/Cpf1, etc.).
  • gRNA guide RNA
  • gRNA can exhibit enhanced editing efficiency of a target sequence when in complex with a CRISPR- Cas system compared to a gRNA comprising additional one or more 5′ terminal G nucleotides, wherein the additional one or more 5′ terminal G nucleotides are not present in the protospacer sequence or not complementary to one or more nucleotides at the 3′ terminus of the target sequence in a genome.
  • the term “codon” generally refers to three consecutive nucleotides, which may or may not encode an amino acid.
  • the efficiency of editing can refer to the ability of a guide system nucleic acid molecule directed effector protein (e.g., CRISPR-Cas protein) to modify a target DNA sequence.
  • modification of a target sequence can include introducing a double stranded break, modifying a nucleobase, inducing chromosomal rearrangements, and modifying methylation/demethylation of a promoter sequence of a gene.
  • a target sequence may be located in a gene or in a promoter region in a genome.
  • An effector protein may be a gRNA directed nuclease, e.g., Cas protein such as Cas9 or any other Cas protein described herein.
  • the editing efficiency can be measured by using any methods well known to one skilled in the art.
  • the efficiency of genome editing or editing efficiency can be measured by using tracking of indels by decomposition (TIDE) analysis, surveyor nuclease assay, junction PCR, droplet digital PCR (ddPCR), denaturing high- performance liquid chromatography (DHPLC), PCR single-stranded conformational polymorphism (SSCP), high-resolution melting (HRM), restriction enzyme digestion- suppressed PCR (RE-PCR), engineered nuclease-induced translocations (ENIT), restriction enzyme digestion, Sanger DNA sequencing, deep sequencing such as next generation sequencing (NGS), or any combination thereof.
  • TIDE indels by decomposition
  • ddPCR droplet digital PCR
  • DPLC denaturing high- performance liquid chromatography
  • SSCP PCR single-stranded conformational polymorphism
  • HRM high-resolution melting
  • RE-PCR restriction enzyme digestion- suppressed PCR
  • ENIT engine
  • the efficiency of genome editing e.g., generating a double-strand break
  • TIDE analysis a three-step method whereby the region targeted by the nuclease (e.g., Cas9) is PCR-amplified from DNA isolated from cells transfected with CRISPR-Cas system and gRNAs. Amplicons of 500-1500 bp generated around the target site are subject to conventional Sanger DNA sequencing followed by analysis using the web-based TIDE software.
  • the editing efficiency e.g., the efficiency of generating a DNA break in the intended target site, is at least 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 99%.
  • guide system nucleic acid molecules that exhibit reduced off-target editing (e.g., editing of a non-target sequence) when guide system nucleic acid molecules are contacted with the target sequence in a complex with a CRISPR-Cas system (e.g., CRISPR-Cas9, CRISPR-Cas12a/Cpf1, etc.).
  • off-target editing e.g., editing of a non-target sequence
  • CRISPR-Cas system e.g., CRISPR-Cas9, CRISPR-Cas12a/Cpf1, etc.
  • gRNA provided herein can exhibit reduced off-target editing when in complex with a CRISPR-Cas system compared to a gRNA comprising additional one or more 5′ terminal G nucleotides, wherein the additional one or more 5′ terminal G nucleotides are not present in the protospacer sequence or not complementary to one or more nucleotides at the 3′ terminus of the target sequence in a genome.
  • the off-target editing can be reduced by at least 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 99%.
  • the scaffold region of the guide system nucleic acid molecule recognized by the Cas protein may form a secondary structure such as a stem, a hairpin, and/or a loop.
  • Stems or hairpins, described herein can be about 3-10 nucleotides in length.
  • Loops can be about 6-20 nucleotides in length.
  • Stems may comprise one or more bulges of 1-10 nucleotides in length.
  • guide system nucleic acid molecules comprising a target sequence comprising one or more mismatched nucleotide
  • the spacer sequence may comprise one or more nucleotides that are not complementary to the target site sequence in the genome.
  • Spacers described herein may harbor various number of mismatches, for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 mismatches.
  • a spacer comprises at most 1, 2, 3, 4, or 5 mismatches.
  • a spacer does not comprise any mismatch as compared to a protospacer sequence at the target site, i.e., the spacer hybridizes with the target sequence at 100%.
  • Spacers described herein may comprise at least 1 to at least 5 mismatched nucleotides.
  • the spacer may comprise at least 1, at least 2, at least 3, at least 4, or at least 5 mismatched nucleotides.
  • the spacer may comprise at most 3, at most 4, at most 5, at most 6, or at most 7 mismatched nucleotides.
  • the one or more mismatched nucleotides may be located at the 5′ terminus of the spacer sequence.
  • the one or more mismatched nucleotides may be located at the 3′ terminus of the spacer sequence.
  • the one or more mismatched nucleotides may be internally located in the spacer sequence.
  • a sequence extension can be on the 5′ or 3′ terminus or can be added internally.
  • the 5′ terminus of the gRNA Cas12a/Cpf1 e.g., crRNA
  • the 5′ terminus of the gRNA Cas12a/Cpf1 can be extended by 2- 59 nucleotides.
  • Extending the 5′ terminus of the Cas12a/Cpf1 gRNA, which comprises a scaffold sequence for Cas12a/Cpf1 binding on the 5′ terminus and a target sequence on the 3′ terminus can increase the editing efficiency and delivery of Cas12a/Cpf1 in vitro and in vivo.
  • a gRNA of Cas9 comprising an internal extension of 2-10 nucleotides to extend the stem region of the stem loop structure can increases gene knockout efficiency in CRISPR-Cas9-mediated genome editing.
  • a gRNA may comprise two or more of crRNA sequences and tracrRNA sequence and bind two or more Cas proteins and a target DNA sites at two or more distinct regions in the genome.
  • the gRNAs described herein may comprise a 5′ sequence extension.
  • the gRNAs described herein may comprise a 3′ sequence extension.
  • the gRNAs described herein may comprise an internal sequence extension.
  • the sequence extension can comprise about at least 1 to at least 70 nucleotides.
  • the sequence extension can comprise at least 1, at least 5, at least 10, at least 20, at least 30, at least 40, at least 50, at least 60, or at least 70 nucleotides.
  • G guanine
  • a nucleotide analog comprises an alteration in a phosphate backbone, a sugar, and/or nucleobases.
  • Non-limiting examples of nucleotide analog include 2-aminoadenosine, 2-thiothymidine, inosine, pyrrolo-pyrimidine, 3-methyl adenosine, 5-methylcytidine, 2-aminoadenosine, C5-bromouridine, C5-fluorouridine, C5- iodouridine, C5-propynyl-uridine, C5-propynyl-cytidine, C5-methylcytidine, 2- aminoadenosine, 7-deazaadenosine, 7-deazaguanosine, 8-oxoadenosine, 8-oxoguanosine, 0(6)-methylguanine, 2-thiocytidine, 2′-fluororibose, ribose, 2′-deoxyribose, arabinose, hexose, phosphorothioate linkages, 5′-N-phosphoramidite link
  • gRNAs comprising a 5′ terminal G analog that can exhibit enhanced editing efficiency of a target sequence when in complex with a CRISPR-Cas system compared to a gRNA lacking 5′ terminal G analog.
  • nucleic acid molecules comprising 10 to 100 nucleotides in length.
  • the synthesized nucleic acid molecules comprises 10, 20, 30, 40, 50, 60, 70, 80, 90, or 100 nucleotides in length.
  • the synthesized nucleic acid molecules comprises 10 to 20 nucleotides, 10 to 30 nucleotides, 10 to 40 nucleotides, 10 to 50 nucleotides, 10 to 60 nucleotides, 10 to 70 nucleotides, 10 to 80 nucleotides, 10 to 90 nucleotides, 10 to 100 nucleotides, 20 to 30 nucleotides, 20 to 40 nucleotides, 20 to 50 nucleotides, 20 to 60 nucleotides, 20 to 70 nucleotides, 20 to 80 nucleotides, 20 to 90 nucleotides, 20 to 100 nucleotides, 30 to 40 nucleotides, 30 to 50 nucleotides, 30 to 60 nucleotides, 30 to 70 nucleotides, 30 to 80 nucleotides, 30 to 90 nucleotides, 30 to 100 nucleotides, 40 to 50 nucleotides, 40 to 60 nucleotides, 30 to 70 nucleotides, 30
  • the synthesized nucleic acid molecules comprises at least 10, at least 20, at least 30, at least 40, at least 50, at least 60, at least 70, at least 80, or at least 90 nucleotides in length. In some embodiments, the synthesized nucleic acid molecules comprises at most 20, at most 30, at most 40, at most 50, at most 60, at most 70, at most 80, at most 90, or at most 100 nucleotides in length. In some embodiments, the synthesized nucleic acid molecules comprises at least 20 nucleotides in length. In some embodiments, the synthesized nucleic acid molecules comprises at least 80 nucleotides in length. In some embodiments, the synthesized nucleic acid molecules comprises at most 30 nucleotides in length.
  • the synthesized nucleic acid molecules can comprise one or more modifications.
  • the synthesized nucleic acid molecules can comprise synthetic nucleotide, synthetic nucleotide analog, nucleotide derivatives, and/or modified nucleotides.
  • the one or more modification can increase stability of the synthesized nucleic acid molecules.
  • the one or more modification can enhance biological activity of the synthesized nucleic acid molecules.
  • a modification of internucleotide linkage using phosphorothioate (PS) bond substitutes can be introduced to inhibit exonuclease-mediated degradation of nucleic acid molecules.
  • PS phosphorothioate
  • the one or more modifications can be made at any location of the synthesized nucleic acid molecule.
  • the synthesized nucleic acid molecule can comprise natural nucleosides (e.g., adenosine, guanosine, cytidine, and uridine), nucleoside analogs (e.g., 2-aminoadenosine, 2-thiothymidine, inosine, pyrrolo-pyrimidine, 3-methyl adenosine, 5-methylcytidine, 2-aminoadenosine, C5-bromouridine, C5-fluorouridine, C5- iodouridine, C5-propynyl-uridine, C5-propynyl-cytidine, C5-methylcytidine, 2- aminoadenosine, 7-deazaadenosine, 7-deazaguanosine, 8-oxoadenosine, 8-ox
  • nucleic acid molecule comprising a chemical modification.
  • Chemical modifications as described herein may comprise one or more 5′ modifications selected from the group consisting of 5′ triphosphate, 5′ diphosphate, 5′ monophosphate, and 5′ hydroxyl.
  • the chemical modification comprises one or more ribose modifications selected from the group consisting of 2′-O-methylation (2′OMe), 2′-O- methoxy-ethyl (2′-MOE), 2′-fluoro (2′F), 2-deoxy-2′-thio, and 2′-azido.
  • the chemical modification comprises one or more internucleotide linkage modifications selected from the group consisting of phosphorothioate, methylphosphonate, phosphonocarboxylate phosphonothiocarboxylate, boranophosphonate, alkylphosphonate, and alkylphosphonate.
  • Chemical modifications can further comprise modified nucleotides comprising one or more heterocyclic modifications selected from the group consisting of 2,6- Diaminopurine, 2-Aminopurine, inosine, 2-aminoadenosine, N6-methyladenosine, N6,2′-O- dimethyladenosine, N1-methyladenosine, 2-amino-6-chloropurineriboside, 5-methylcytidine, 5-hydroxymethylcytidine, 8-oxo-7,8-dihydroguanosine, pseudouridine, N4-acetylcytidine, 5- bromo-uridine, 5-methyluridine, and 5-nitroindole.
  • heterocyclic modifications selected from the group consisting of 2,6- Diaminopurine, 2-Aminopurine, inosine, 2-aminoadenosine, N6-methyladenosine, N6,2′-O- dimethyladenosine, N1-methyladeno
  • the chemical modification comprises, but is not limited to, modified nucleotides comprising one or more 5′ cap modifications selected from the group consisting of GpppG, 7-methylguanylate (m7GpppG), m2,2,7GpppG, and m7-3′-OGpppG (ARCA).
  • the chemical modification comprises modified nucleotides comprising one or more 5′ cap modifications selected from the group consisting of an attachment chemistry (e.g., biotin), a dye, a cell targeting moiety, an active chemistry, and an amino modifier.
  • the attachment chemistry can comprise biotin.
  • the dye comprises fluorescein.
  • the cell targeting moiety comprises digoxigenin.
  • the active chemistry comprises azides, acrydite, thiols, or alkynes.
  • the amino modifier comprises aminoallyl.
  • the 15-PGDH inhibitor can provide regulated degradation of 15-PGDH by incorporating a degron into the amino acid sequence of 15- PGDH that allows recruitment to the endogenous protein turnover machinery. Mechanisms for targeted protein degradation include, but are not limited to, recruitment to an E3 ligase for ubiquitination and subsequent proteasomal degradation, direct recruitment to the proteasome, and recruitment to the lysosome.
  • methods for inducible protein degradation by a degron includes, but is not limited to, ligand induced degradation (LID) using a SMASH tag, ligand induced degradation using Shield-1, ligand induced degradation using auxin, ligand induced degradation using rapamycin, peptidic degrons (e.g., IKZF3 based degrons), and proteolysis- targeting chimeras (PROTACs).
  • LID ligand induced degradation
  • Shield-1 ligand induced degradation using auxin
  • ligand induced degradation using rapamycin peptidic degrons (e.g., IKZF3 based degrons)
  • PROTACs proteolysis- targeting chimeras
  • a degron tag that is held in an inactive conformation but is induced to adopt a conformation capable of recognition by the proteasome upon binding of a specific molecule, such as but not limited to, a Shield-1 molecule.
  • a specific molecule such as but not limited to, a Shield-1 molecule.
  • the inducible degron element is selected from the group consisting of a ligand inducible degron element such as a small molecule-assisted shutoff (SMASH) degron element, Shield-1 responsive degron element, auxin responsive degron element, and rapamycin responsive degron element; a peptidic degron element; and a peptidic proteolysis targeting chimera (PROTAC) element.
  • SMASH small molecule-assisted shutoff
  • PROTAC peptidic proteolysis targeting chimera
  • the ligand inducible degron element is a small molecule-assisted shutoff (SMASH) degron element and the exogenous factor for controlling immunogenicity is asunaprevir.
  • SMASH small molecule-assisted shutoff
  • a peptide tag is used that confers small molecule-mediated recruitment to an E3 ligase.
  • the peptide tag comprises the lymphoid-restricted transcription factor IKZF3 that is recruited to the E3 ligase receptor (CRBN) in an immunomodulatory drug (IMiD) dependent manner, as described in Koduri et al., Proc Natl Acad Sci, 2019, 116(7), 2539-2544, which is herein incorporated by reference in its entirety.
  • the degron is capable of targeting immunosuppressive factors for degradation (e.g., through a ubiquitination pathway), inducing protein degradation, or degrading proteins.
  • the 15-PGDH inhibitors and/or 15-PGDH substrates described herein can be used to treat, prevent, or reduce the symptoms or severity of any neural injury, neurodegeneration, neurodegenerative and/or neuropsychiatric conditions, diseases, and/or disorders characterized by structural deterioration of the blood-brain barrier and/or by increased generation of reactive oxygen species within the nervous system of a subject in need thereof.
  • a subject having the neural injury, neurodegeneration, neurodegenerative and/or neuropsychiatric conditions, diseases, and/or disorders characterized by structural deterioration of the blood-brain barrier and/or by increased generation of reactive oxygen species within the nervous system of a subject in need thereof can have or be at risk of memory loss, cognitive decline, axonal degeneration, neuronal cell death, glia cell damage, and/or blood brain barrier permeability, and the 15-PGDH inhibitor can be administered to the subject at an amount effective to decrease structural deterioration of the blood-brain barrier and/or generation of reactive oxygen species within the nervous system of a subject in need thereof.
  • Subjects amenable to treatment by 15-PGDH inhibitors and/or 15-PGDH substrates as disclosed herein include subjects having structural deterioration of the blood- brain barrier and/or increased generation of reactive oxygen species within the nervous system and at risk of a neurodegenerative condition, disease, or disorder but not showing symptoms (for example asymptomatic subjects), as well as subjects presently showing symptoms.
  • the present methods can be administered prophylactically to the general population characterized by structural deterioration of the blood-brain barrier and/or by increased generation of reactive oxygen species within the nervous system without any assessment of the risk of the subject patient.
  • Subjects can be further screened for their likelihood of having or developing a neurodegenerative condition, disease, or disorder based on a number of biochemical and genetic markers in addition to structural deterioration of the blood-brain barrier and/or increased generation of reactive oxygen species within the nervous system. For example, one can also diagnose a subject with increased risk of developing Alzheimer's Disease using genetic markers for Alzheimer's Disease. Genetic abnormality in a few families has been traced to chromosome 21 (St.
  • One genetic marker is, for example mutations in the APP gene, particularly mutations at position 717 and positions 670 and 671 referred to as the Hardy and Swedish mutations respectively (see Hardy, TINS, supra).
  • Other markers of risk are mutations in the presenilin genes, PS1 and PS2, and ApoE4, family history of Alzheimer's Disease, hypercholesterolemia or atherosclerosis. Subjects with APP, PS1 or PS2 mutations are highly likely to develop Alzheimer's disease. ApoE is a susceptibility gene, and subjects with the e4 isoform of ApoE (ApoE4 isoform) have an increased risk of developing Alzheimer's disease.
  • Alzheimer disease susceptibility genes include, for example ACE, CHRNB2, CST3, ESR1, GAPDH, IDE, MTHFR, NCSTN, PRNP, PSEN1, TF, TFAM and TNF and be used to identify subjects with increased risk of developing Alzheimer's disease (Bertram et al, Nat. Genet.2007 January; 39(1):17-23), as well as variances in the alpha-T catenin (VR22) gene (Bertram et al, J Med. Genet.2007 January; 44(1):e63) and Insulin-degrading enzyme (IDE) and Kim et al, J Biol. Chem.2007; 282:7825-32).
  • Neurodegenerative conditions, disease, or disorders associated with cognitive decline or memory loss can be diagnosed using standard practice and the progression can be monitored over an extended period of time.
  • One such method includes at least one of the following; (i) a memory assessment, (ii) an extensive neuropsychological exam, (iii) an examination by a geriatric neurologist and (iv) MRI imaging of the brain.
  • Disease progression can be documented by changes in these parameters over time.
  • changes in the parameters of at least one of these assessments can be used to assess the efficacy of the 15-PGDH inhibitors and/or 15-PGDH substrates in the subject over time.
  • Other methods to diagnose a patient at risk of or having a neurodegenerative condition, disease or disorder includes measurement of blood-brain barrier permeability and/or generation of reactive oxygen species within the nervous system and 15-PGDH activity and/or expression in the neurotissue, such as brain tissue, wherein increased blood-brain barrier permeability and/or increased generation of reactive oxygen species within the nervous system of the subject and 15-PGDH activity and/or expression compared to a control (e.g., normal or healthy neurotissue) is indicative of the subject having or at increased risk of the neurodegenerative condition, disease, or disorder.
  • a control e.g., normal or healthy neurotissue
  • direct detection of BBB breakdown can be assessed using MRI and injection of contrasting agent. Improvements in the resolution of MRIs and in the use of special contrasting agents can be used to detect BBB permeability.
  • subjects are administered a contrasting agent immediately prior to brain imaging, such as MRI imaging.
  • MRI imaging In cases of intact BBB, the contrasting agents are confined to brain blood vessels whereas, in subjects with a disrupted BBB, the contrasting agent is "sprayed out" into the brain tissue, which can be visualized.
  • the brain locations, the size of BBB breakdown and extent of BBB compromise, such as extent of vascular leak in subjects can be directly and quantitatively assessed as measurable parameters of BBB permeability.
  • such methods can be used to assess any improvements in these parameters resulting from treatment of the subject with an agent that inhibits 15-PGDH activity.
  • Direct visualization of BBB breakdown and its associated vascular leak into the brain is useful in the methods as disclosed herein for monitoring the beneficial effects of treating a subject with BBB permeability with a 15-PGDH inhibitor and/or 15-PGDH substrate.
  • An improvement in at least one measurable parameter of BBB permeability, such as location, size of BBB breakdown and extent of vascular leak in subjects administered a 15-PGDH inhibitor and/or 15-PGDH substrate indicates a positive outcome from administration of an inhibitor of 15- PGDH.
  • Parameters of BBB permeability can be monitored by direct visualization and quantified by MRI-associated image analysis and are useful in the methods as disclosed herein.
  • the 15-PGDH inhibitors and/or 15-PGDH substrates described herein are useful in preventing and treating neurodegenerative conditions, diseases and disorders, such as those characterized by structural deterioration of the blood-brain barrier and/or increased generation of reactive oxygen species within the nervous system, aberrant 15-PGDH activity, and risk of memory loss, cognitive decline, axonal degeneration, neuronal cell death, or glia cell damage.
  • neurodegenerative conditions, diseases, or disorders can include subarachnoid hemorrhage, schizophrenia, depression, bipolar disorder, normal aging, epilepsy, traumatic brain injury and/or a visual symptom associated therewith, post-traumatic stress disorder, Parkinson’s disease, Parkinson Plus syndromes, Lewy Body Dementia, multiple system atrophy, corticobasal neurodegeneration, progressive supranuclear palsy, Alexander’s disease, Alzheimer’s disease, Alzheimer's disease related dementias, Down syndrome, spinocerebellar ataxia, amyotrophic lateral sclerosis, Huntington’s disease, stroke, brain radiation therapy, chronic stress, abuse or cellular toxicity of a neuro-active drug, retinal degeneration, spinal cord injury, peripheral nerve injury, idiopathic peripheral neuropathy, cognitive decline and/or general frailty associated with normal aging and/or chemotherapy, chemotherapy induced neuropathy, concussive injury, peripheral nerve crush injury, peripheral neuropathy, diabetic neuropathy, post-traumatic headache, multiple
  • Additional examples of neurodegenerative diseases or disorders associated with structural deterioration of the blood-brain barrier and/or increased generation of reactive oxygen species within the nervous system include, for example, polyglutamine repeat disorders such as Spinocerebellar ataxias (e.g., types 1, 2, 3, 6, 7 and 17), Machado-Joseph disease, Spinal and Bulbar muscular atrophy (SBMA or Kennedy's disease), Dentatorubral Pallidoluysian Atrophy (DRPLA) and other neurological conditions arising from polyglutamine expansions, or disease arising from non-coding DNA repeat expansions such as Fragile X syndrome, Fragile XE mental retardation, Friedreich ataxia, myotonic dystrophy, Spinocerebellar ataxias (types 8, 10 and 12) or other neurodegenerative diseases such as spinal muscular atrophy (Werdnig-Hoffman disease, Kugelberg-Welander disease), and spongiform encephalopathies.
  • polyglutamine repeat disorders such
  • Additional neurodegenerative diseases associated with structural deterioration of the blood-brain barrier and/or increased generation of reactive oxygen species within the nervous system for which agents inhibiting 15-PGDH can be useful include, for example, age-related memory impairment, agyrophilic grain dementia, Parkinsonism-dementia complex of Guam, auto-immune conditions (eg Guillain-Barre syndrome, Lupus), Biswanger's disease, brain and spinal tumors (including neurofibromatosis), cerebral amyloid angiopathies (Journal of Alzheimer's Disease vol 3, 65-73 (2001)), cerebral palsy, chronic fatigue syndrome, corticobasal degeneration, conditions due to developmental dysfunction of the CNS parenchyma, conditions due to developmental dysfunction of the cerebrovasculature, dementia--multi infarct, dementia--subcortical, dementia with Lewy bodies, dementia of human immunodeficiency virus (HIV), dementia lacking distinct histology, Dementia Pugilistica, diffies neurodegenerative diseases
  • Additional neurodegenerative diseases associated with structural deterioration of the blood-brain barrier and/or increased generation of reactive oxygen species within the nervous system for which 15-PGDH inhibitors and/or 15-PGDH substrates are useful include other dementias not listed above, such as but without limitation, other mixed dementia, frontotemporal dementia, progressive supranuclear palsy (PSP), Parkinson's Disease with associated dementia, corticobasal degeneration, multiple system atrophy, HIV-induced dementia, white matter disease-associated dementias, mild cognitive impairment (MCI).
  • the 15-PGDH inhibitors and/or 15-PGDH substrates described herein are useful in preventing and treating blood brain barrier (BBB) permeability in a subject.
  • BBB blood brain barrier
  • diseases and disorders where BBB permeability occurs include, for example, multiple sclerosis, cerebral amyloid angiopathy, diabetic retinopathy, prion disorders, amyotrophic lateral sclerosis (ALS), Stiff-person Syndrome, Spinocerebellar Ataxias, Friedreich Ataxia, Ataxia Telangiectasia, Bulbospinal Atrophy (Kennedy Syndrome), Spinal Muscular Atrophy, Neuronal storage diseases (lipofuscinoses), Mitochondrial encephalomyopathies, Leukodystrophies, Neural sequelae of spinal shock/blunt trauma, Hypertensive Cerebrovascular disease, such as Lacunar Infarcts, Slit hemorrhages, Hypertensive encephalopathy.
  • ALS amyotrophic lateral sclerosis
  • Stiff-person Syndrome Spinocerebellar Ataxias
  • Friedreich Ataxia Friedreich Ataxia
  • Ataxia Telangiectasia Bulbospinal At
  • the 15-PGDH inhibitor and/or 15-PGDH substrate can be administered to a subject with structural deterioration of the blood-brain barrier and/or increased generation of reactive oxygen species within the nervous system associated with or caused by administration of a drug or therapeutic agent to the subject.
  • the drug or therapeutic agent can be one that interacts with brain-located cellular receptors (membranous and cytosolic) or one that interacts with physiological pathways in the brain (action on a transporter), or a drug targeted to an organ other than the brain that interacts with brain- located cellular receptors or impacts/interferes physiological pathways of the brain.
  • the drug is for use in treating a brain-related pathology; the brain-related pathology may include depression, anxiety, stress, panic disorder, pain, epilepsy, dementia, suicidal ideation, Alzheimer's disease, Parkinson's disease.
  • the drug may include a selective- serotonin reuptake inhibitor (SSRI), a selective norepinephrine reuptake inhibitor (SNRI), a selective dopamine reuptake inhibitor (SDRI), serotonin receptor agonist/antagonists (interacting with the ‘5-HT’ receptor family), dopamine receptor agonist/antagonists (interacting with the ‘DT’ receptor family), adrenergic receptor agonist/antagonists (interacting with the ‘alpha’ and ‘beta’ receptor families, including beta-2-adrenergic receptor agonists), GABA receptor agonist/antagonists (interacting with the ‘GABAA’ and ‘GABAB’ receptor families), acetylcholine receptor agonist/antagonists (interacting with the ‘muscarinic’ and ‘nicotinic’ receptor families), glutamate receptor agonists/antagonists (e.g., NMDA receptor agonist/antagonists), opioid receptor agonist/antagonists (interacting with the ‘delta’, ‘kappa’
  • the drug can result in a change in homeostasis to neurotransmitter-related physiological pathways.
  • the drug can be intended for use in or is used in treating a neuropsychiatric condition.
  • the condition can be, for example, depression, anxiety, panic disorder, suicidal ideation or stress.
  • the drug can be any drug tested for use in or used to treat a neuropsychiatric condition but is preferably a neurotransmitter reuptake inhibitor such as an SSRI, SNRI, or SDRI or a neurotransmitter receptor agonist or antagonist (includes partial agonists/antagonists), especially a serotonin or dopamine receptor.
  • the 15-PGDH inhibitors and/or 15-PGDH substrates described herein can be provided in a pharmaceutical composition.
  • a pharmaceutical composition containing the 15-PGDH inhibitors and/or 15-PGDH substrates described herein as an active ingredient may be manufactured by mixing the derivative with a pharmaceutically acceptable carrier(s) or an excipient(s) or diluting the 15-PGDH inhibitors and/or 15-PGDH substrates with a diluent in accordance with conventional methods.
  • the pharmaceutical composition may further contain fillers, anti-cohesives, lubricants, wetting agents, flavoring agents, emulsifying agents, preservatives and the like.
  • the pharmaceutical composition may be formulated into a suitable formulation in accordance with the methods known to those skilled in the art so that it can provide an immediate, controlled or sustained release of the 15-PGDH inhibitors and/or 15-PGDH substrates after being administered into a mammal.
  • the pharmaceutical composition may be formulated into a parenteral or oral dosage form.
  • the solid dosage form for oral administration may be manufactured by adding excipient, if necessary, together with binder, disintegrants, lubricants, coloring agents, and/or flavoring agents, to the 15-PGDH inhibitors and shaping the resulting mixture into the form of tablets, sugar-coated pills, granules, powder or capsules.
  • the additives that can be added in the composition may be ordinary ones in the art.
  • examples of the excipient include lactose, sucrose, sodium chloride, glucose, starch, calcium carbonate, kaolin, microcrystalline cellulose, silicate and the like.
  • exemplary binders include water, ethanol, propanol, sweet syrup, sucrose solution, starch solution, gelatin solution, carboxymethylcellulose, hydroxypropyl cellulose, hydroxypropyl starch, methylcellulose, ethylcellulose, shellac, calcium phosphonate and polypyrrolidone.
  • examples of the disintegrant include dry starch, sodium arginate, agar powder, sodium bicarbonate, calcium carbonate, sodium lauryl sulfate, stearic monoglyceride and lactose.
  • purified talc, stearates, sodium borate, and polyethylene glycol may be used as a lubricant; and sucrose, bitter orange peel, citric acid, tartaric acid, may be used as a flavoring agent.
  • the pharmaceutical composition can be made into aerosol formulations (e.g., they can be nebulized) to be administered via inhalation.
  • the 15-PGDH inhibitors and/or 15-PGDH substrates described herein may be combined with flavoring agents, buffers, stabilizing agents, and the like and incorporated into oral liquid dosage forms such as solutions, syrups or elixirs in accordance with conventional methods.
  • One example of the buffers may be sodium citrate.
  • the stabilizing agents include tragacanth, acacia and gelatin.
  • the 15-PGDH inhibitors and/or 15-PGDH substrates described herein may be incorporated into an injection dosage form, for example, for a subcutaneous, intramuscular or intravenous route by adding thereto pH adjusters, buffers, stabilizing agents, relaxants, topical anesthetics.
  • the pH adjusters and the buffers include sodium citrate, sodium acetate and sodium phosphate.
  • the stabilizing agents include sodium pyrosulfite, EDTA, thioglycolic acid and thiolactic acid.
  • the topical anesthetics may be procaine HCl, lidocaine HCl and the like.
  • the relaxants may be sodium chloride, glucose and the like.
  • the 15-PGDH inhibitors and/or 15-PGDH substrates described herein may be incorporated into suppositories in accordance with conventional methods by adding thereto pharmaceutically acceptable carriers that are known in the art, for example, polyethylene glycol, lanolin, cacao butter or fatty acid triglycerides, if necessary, together with surfactants such as Tween.
  • the pharmaceutical composition may be formulated into various dosage forms as discussed above and then administered through various routes including an oral, inhalational, transdermal, subcutaneous, intravenous or intramuscular route.
  • the dosage can be a pharmaceutically or therapeutically effective amount.
  • Therapeutically effective dosages of 15-PDGH inhibitor is one that reduces activity and/or expression of 15-PGDH and structural deterioration of the blood-brain barrier and/or generation of reactive oxygen species within the nervous system and generates the maximum protective effect in preventing a neurodegenerative disease or disorder, or reduces a symptom of a neurodegenerative disease or disorder.
  • an optimum dosage of the 15-PGDH inhibitor and/or 15- PGDH substrate is one generating the maximum beneficial effect on damaged tissue.
  • An effective dosage causes at least a statistically or clinically significant attenuation of at least one marker, symptom, or histological evidence characteristic of neurodegenerative condition, disease, or disorder.
  • Markers, symptoms and histological evidence characteristic of neurodegenerative condition, disease, or disorder include memory loss, confusion, disturbances in axonal transport, demyelination, induction of metalloproteinases (MMPs), activation of glial cells, infiltration of lymphocytes, edema and immunological reactions that lead to tissue damage and further vascular injury. Stabilization of symptoms or diminution of tissue damage, under conditions wherein control patients or animals experience a worsening of symptoms or tissue damage, is one indicator of efficacy of a suppressive treatment. [00315] Therapeutically effective dosage amounts of the 15-PGDH inhibitor and/or 15- PGDH substrate may be present in varying amounts in various embodiments.
  • a therapeutically effective amount of the 15-PGDH inhibitor may be an amount ranging from about 10-1000 mg (e.g., about 20 mg-1,000 mg, 30 mg-1,000 mg, 40 mg-1,000 mg, 50 mg-1,000 mg, 60 mg-1,000 mg, 70 mg-1,000 mg, 80 mg-1,000 mg, 90 mg- 1,000 mg, about 10-900 mg, 10-800 mg, 10-700 mg, 10-600 mg, 10-500 mg, 100-1000 mg, 100-900 mg, 100-800 mg, 100-700 mg, 100-600 mg, 100-500 mg, 100-400 mg, 100-300 mg, 200-1000 mg, 200-900 mg, 200-800 mg, 200-700 mg, 200-600 mg, 200-500 mg, 200-400 mg, 300-1000 mg, 300-900 mg, 300-800 mg, 300-700 mg, 300-600 mg, 300-500 mg, 400 mg-1,000 mg, 500 mg-1,000 mg, 100 mg-900 mg, 200 mg-800 mg, 300 mg-700 mg, 300-600 mg, 300-500 mg, 400 mg-1,000 mg, 500 mg-
  • the 15-PGDH inhibitor is present in an amount of or greater than about 10 mg, 50 mg, 100 mg, 150 mg, 200 mg, 250 mg, 300 mg, 350 mg, 400 mg, 450 mg, 500 mg, 550 mg, 600 mg, 650 mg, 700 mg, 750 mg, 800 mg. In some embodiments, the 15-PGDH inhibitor is present in an amount of or less than about 1000 mg, 950 mg, 900 mg, 850 mg, 800 mg, 750 mg, 700 mg, 650 mg, 600 mg, 550 mg, 500 mg, 450 mg, 400 mg, 350 mg, 300 mg, 250 mg, 200 mg, 150 mg, or 100 mg.
  • a therapeutically effective dosage amount may be, for example, about 0.001 mg/kg weight to 500 mg/kg weight, e.g., from about 0.001 mg/kg weight to 400 mg/kg weight, from about 0.001 mg/kg weight to 300 mg/kg weight, from about 0.001 mg/kg weight to 200 mg/kg weight, from about 0.001 mg/kg weight to 100 mg/kg weight, from about 0.001 mg/kg weight to 90 mg/kg weight, from about 0.001 mg/kg weight to 80 mg/kg weight, from about 0.001 mg/kg weight to 70 mg/kg weight, from about 0.001 mg/kg weight to 60 mg/kg weight, from about 0.001 mg/kg weight to 50 mg/kg weight, from about 0.001 mg/kg weight to 40 mg/kg weight, from about 0.001 mg/kg weight to 30 mg/kg weight, from about 0.001 mg/kg weight to 25 mg/kg weight, from about 0.001 mg/kg weight to 20 mg/kg weight, from about 0.00 0.001 mg/kg weight to
  • a therapeutically effective dosage amount may be, for example, about 0.0001 mg/kg weight to 0.1 mg/kg weight, e.g. from about 0.0001 mg/kg weight to 0.09 mg/kg weight, from about 0.0001 mg/kg weight to 0.08 mg/kg weight, from about 0.0001 mg/kg weight to 0.07 mg/kg weight, from about 0.0001 mg/kg weight to 0.06 mg/kg weight, from about 0.0001 mg/kg weight to 0.05 mg/kg weight, from about 0.0001 mg/kg weight to about 0.04 mg/kg weight, from about 0.0001 mg/kg weight to 0.03 mg/kg weight, from about 0.0001 mg/kg weight to 0.02 mg/kg weight, from about 0.0001 mg/kg weight to 0.019 mg/kg weight, from about 0.0001 mg/kg weight to 0.018 mg/kg weight, from about 0.0001 mg/kg weight to 0.017 mg/kg weight, from about 0.0001 mg/kg weight to 0.016 mg/kg weight, from
  • the therapeutically effective dose may be 0.0001 mg/kg weight, 0.0002 mg/kg weight, 0.0003 mg/kg weight, 0.0004 mg/kg weight, 0.0005 mg/kg weight, 0.0006 mg/kg weight, 0.0007 mg/kg weight, 0.0008 mg/kg weight, 0.0009 mg/kg weight, 0.001 mg/kg weight, 0.002 mg/kg weight, 0.003 mg/kg weight, 0.004 mg/kg weight, 0.005 mg/kg weight, 0.006 mg/kg weight, 0.007 mg/kg weight, 0.008 mg/kg weight, 0.009 mg/kg weight, 0.01 mg/kg weight, 0.02 mg/kg weight, 0.03 mg/kg weight, 0.04 mg/kg weight, 0.05 mg/kg weight, 0.06 mg/kg weight, 0.07 mg/kg weight, 0.08 mg/kg weight, 0.09 mg/kg weight, or 0.1 mg/kg weight.
  • a therapeutically effective dosage may be a dosage of 10 ⁇ g/kg/day, 50 ⁇ g/kg/day, 100 ⁇ g/kg/day, 250 ⁇ g/kg/day, 500 ⁇ g/kg/day, 1000 ⁇ g/kg/day or more.
  • the amount of the 15-PGDH inhibitor or pharmaceutical salt thereof is sufficient to provide a dosage to a patient of between 0.01 ⁇ g/kg and 10 ⁇ g/kg; 0.1 ⁇ g/kg and 5 ⁇ g/kg; 0.1 ⁇ g/kg and 1000 ⁇ g/kg; 0.1 ⁇ g/kg and 900 ⁇ g/kg; 0.1 ⁇ g/kg and 900 ⁇ g/kg; 0.1 ⁇ g/kg and 800 ⁇ g/kg; 0.1 ⁇ g/kg and 700 ⁇ g/kg; 0.1 ⁇ g/kg and 600 ⁇ g/kg; 0.1 ⁇ g/kg and 500 ⁇ g/kg; or 0.1 ⁇ g/kg and 400 ⁇ g/kg.
  • Particular doses or amounts to be administered in accordance with the present invention may vary, for example, depending on the nature and/or extent of the desired outcome, on particulars of route and/or timing of administration, and/or on one or more characteristics (e.g., weight, age, personal history, genetic characteristic, lifestyle parameter, severity of cardiac defect and/or level of risk of cardiac defect, etc., or combinations thereof). Such doses or amounts can be determined by those of ordinary skill. In some embodiments, an appropriate dose or amount is determined in accordance with standard clinical techniques.
  • an appropriate dose or amount is a dose or amount sufficient to reduce a disease severity index score by 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100% or more.
  • an appropriate dose or amount is a dose or amount sufficient to reduce a disease severity index score by 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100%.
  • an appropriate dose or amount is determined through use of one or more in vitro or in vivo assays to help identify desirable or optimal dosage ranges or amounts to be administered.
  • Various embodiments may include differing dosing regimen.
  • the 15-PGDH inhibitor and/or 15-PGDH substrate can be administered via continuous infusion.
  • the continuous infusion is intravenous.
  • the continuous infusion is subcutaneous.
  • the 15-PGDH inhibitor can be administered bimonthly, monthly, twice monthly, triweekly, biweekly, weekly, twice weekly, thrice weekly, daily, twice daily, or on another clinically desirable dosing schedule.
  • the composition can be administered in the form of aqueous, alcoholic, aqueous-alcoholic or oily solutions or suspensions, or of a dispersion of the lotion or serum type, of emulsions that have a liquid or semi-liquid consistency or are pasty, obtained by dispersion of a fatty phase in an aqueous phase (O/W) or vice versa (W/O) or multiple emulsions, of a free or compacted powder to be used as it is or to be incorporated into a physiologically acceptable medium, or else of microcapsules or microparticles, or of vesicular dispersions of ionic and/or nonionic type.
  • aqueous, alcoholic, aqueous-alcoholic or oily solutions or suspensions or of a dispersion of the lotion or serum type, of emulsions that have a liquid or semi-liquid consistency or are pasty, obtained by dispersion of a fatty phase in an aqueous phase (O/W) or vice versa
  • It may thus be in the form of a salve, a tincture, milks, a cream, an ointment, a powder, a patch, an impregnated pad, a solution, an emulsion or a vesicular dispersion, a lotion, aqueous or anhydrous gels, a spray, a suspension, a shampoo, an aerosol or a foam. It may be anhydrous or aqueous. It may also comprise solid preparations constituting soaps or cleansing cakes.
  • compositions including the 15-PGDH inhibitor described herein can additionally contain, for example, at least one 15-PGHD substrate chosen from prostaglandins, in particular prostaglandin PGE 1 , PGE 2 , their salts, their esters, their analogues and their derivatives, in particular those described in WO 98/33497, WO 95/11003, JP 97-100091, JP 96-134242, in particular agonists of the prostaglandin receptors.
  • at least one 15-PGHD substrate chosen from prostaglandins, in particular prostaglandin PGE 1 , PGE 2 , their salts, their esters, their analogues and their derivatives, in particular those described in WO 98/33497, WO 95/11003, JP 97-100091, JP 96-134242, in particular agonists of the prostaglandin receptors.
  • the composition can include at least one 15-PGDH inhibitor as defined above and at least one prostaglandin or one prostaglandin derivative such as for example the prostaglandins of series 2 including in particular PGF 2 ⁇ and PGE 2 in saline form or in the form of precursors, in particular of the esters (example isopropyl esters), their derivatives such as 16,16-dimethyl PGE 2 , 17-phenyl PGE 2 and 16,16-dimethyl PGF 2 ⁇ 17- phenyl PGF2 ⁇ , prostaglandins of series 1 such as 11-deoxyprostaglandin E1, 1- deoxyprostaglandin E1 in saline or ester form, is their analogues, in particular latanoprost, travoprost, fluprostenol, unoprostone, bimatoprost, cloprostenol, viprostol, butaprost, misoprostol, their salts or their esters, in particular of the est
  • TBI Alzheimer’s Disease
  • BBB traumatic brain injury
  • 15-PGDH is a short chain dehydrogenase/reductase that catalyzes nicotinamide adenine dinucleotide (NAD + )-dependent oxidative degradation of various substrates throughout the body, including prostaglandin E2 (PGE2), PGF2 ⁇ , resolvin D1 (RvD1), lipoxin A4 (LXA4), and 15-hydroxyeicosatetranenoic acid (15-HETE).
  • PGE2 prostaglandin E2
  • PGF2 ⁇ PGF2 ⁇
  • RvD1 resolvin D1
  • LXA4 lipoxin A4
  • 15-HETE 15-hydroxyeicosatetranenoic acid
  • mice overexpress the K670N/M671L (Swedish), I716V (Florida), and V717I (London) mutations in human amyloid precursor protein (APP) isoform 695, as well as M146L and L286V mutations in human presenilin 1 (PS1) and were generated as described previously.
  • APP amyloid precursor protein
  • PS1 human presenilin 1
  • mice Male and female 5xFAD (C57BL/6J background, Stock No: 034848, Jackson Laboratory) were bred with Hpgd -/- mice (C57BL/6J background) to generate Tg-WT Hpgd +/+ , Tg-WT Hpgd +/- , Tg-5xFAD Hpgd +/+ , and Tg-5xFAD Hpgd +/- mice. Genotypes of the animals were blinded until they were euthanized. For TBI genetic studies, Hpgd -/- mice were also backcrossed for 14 generations onto an FVB background for generation of complete knockout animals for studies with TBI.
  • mice were randomly assigned to the treatment groups until we have similar number in each treatment groups.
  • mice had unrestricted access to food and water, and they were randomly allocated to various experimental groups. Hpgd -/- mice, the kind gift of Dr. Beverly Koller in whose laboratory the mice were derived, were backcrossed for 12 generations onto a C57BL/6J background for studies involving crosses to 5xFAD mice. In the AD rat study, six-month-old male TgF344-AD rats and WT littermates were used.
  • Human subjects [00328] Experiments with human brain samples were approved by Institutional Review Board (IRB): Northwestern ADRC IRB Number STU00023196 and Case Western Reserve University IRB Number 03-00-26. Human subjects or their representative provided written informed consent. Sex and age information of samples related to Fig.1A.
  • (+)-SW033291 compound was custom synthesized by WuXi AppTec. The compound dissolved in two parts pre-warmed ethanol at 10 mg/ml concentration by incubation at 37°C for at least 15 minutes with vortexing every 5 minutes. One part of Kolliphor EL was added, followed by further dilution with 17 parts of 5% D-Glucose water. The final working concentration of (+)-SW033291 compound was 1 mg/ml in 10% ethanol, 5.0% Kolliphor EL, and 4.25% d-glucose water. Behavioral analysis [00330] The Morris water maze (MWM) test was conducted in a 128-cm diameter tank filled with 22-23°C water.
  • the Congo Red kit (Sigma- Aldrich, HT60) was used according to the manufacturer’s instruction. Briefly, sections were mounted on slides and dried overnight at room temperature. Tissues were quickly rinsed with tap water 3 times and then incubated with hematoxylin solution for 10 mins.
  • sections were incubated in 50% formamide and 2x SSC at 65°C for 2 hours to denature DNA, followed by 5 minute washing with 2xSSC.
  • sections were incubated in 2M HCl at 37°C for 30 minutes for additional denaturation.
  • sections were rinsed in 0.1M borate buffer followed by PBS washing.
  • For blocking sections were incubated in 5% BSA and 5% normal horse serum in 0.1% T-PBS for 1 hour at room temperature.
  • Anti-BrdU antibody was used at 1:1,000 in 5% BSA and 5% normal horse serum in T-PBS and sections were incubated at 4°C overnight.
  • sections of 40 ⁇ m thickness were prepared, rinsed in PBS containing 0.3% Triton X-100, and blocked with 5% normal goat serum. They were then rinsed with PBS and left to incubate overnight at 4°C with rabbit anti-4-HNE (1:500, HNE11S, Alpha diagnostic International) or rabbit anti-3-NT (1:500, AB5411, Millipore Sigma). The following day, sections were washed with PBS and then incubated for two hours at room temperature with either goat anti-rabbit Alexa 594 or 488 (Invitrogen). After another rinse in PBS, sections were mounted using an antifade aqueous media (Vectashield® Plus with DAPI, Vector Laboratories).
  • Protocol for multiplex fluorescent detection (ACD, #323110) of probe for mouse 15-PGDH (Hpgd, #456681), mouse CD206 (Mrc1, #437511), mouse Tmem119 (Tmem119, #472901) was performed according to manufacturer specifications. Fluorescent reagents were employed as per protocol from Akoya BiosciencesTM for the Fluorescein green (#FP1487991KT) and Texas red (#FP1495001KT) to detect Tmem119, CD206, and 15-PGDH, respectively. All sections were counterstained with Akoya Spectral DAPI mounting medium. Slides were stored at 4°C in the dark until high magnification perivascular imaging could be performed.
  • mice were anesthetized and euthanized by transcardial perfusion with a fixative solution (1/4 strength Karnovsky’s fixative) at a flow rate of 10 ml/min for 10 min.
  • the brains were carefully removed and then sliced into thin slices (4 mm x 4 mm x 2 mm). Tissues were fixed with 2.5% glutaraldehyde, 2% paraformaldehyde in 0.1 M HEPES buffer, pH 7.4 for 2h at room temperature.
  • specimens were thoroughly rinsed in 0.1 M phosphate buffer, pH7.4, and then postfixed for 2 h in an unbuffered 1:1 mixture of 2% osmium tetroxide and 3% potassium ferricyanide. After rinsing with distilled water, specimens were soaked overnight in an acidified solution of 0.25% uranyl acetate. After another rinse in distilled water, specimens were dehydrated in ascending concentrations of ethanol, passed through propylene oxide, and embedded in EMbed 812 resin mixture (Electron Microscopy Sciences, PA). For genetic study in AD and TBI studies, free-floating sections were used.
  • Thin sections (70 nm) were cut on an RMC MT6000-XL ultramicrotome and mounted on Gilder square 300 mesh nickel grids (Electron Microscopy Sciences, PA). Thin sections were sequentially stained with acidified uranyl acetate followed by a modification of Sato’s triple lead stain and examined in a FEI Tecnai Spirit (T12) with a Gatan US40004k x 4k CCD.
  • Microglia and non-microglia isolation for western blot Each mouse was perfused with DPBS for 5 min at 15.5rpm and whole brain tissue was collected in DPBS and processed using the MACS Adult Brain dissociation kit (Miltenyi Biotec, 130-107-677) to generate a single cell suspension. Each sample was digested using a combination of enzymatic and mechanical dissociation after being finely sliced with a scalpel. For enzymatic dissociation, enzyme mixes 1 and 2 (provided in the kit) were used for mechanical dissociation.
  • CD11b positive cells labeled by microbead-CD11b antibodies
  • the CD11b positive cell population was collected by removing the column from the magnetic stand and placing the plunger into the column to flush out the cells. After centrifugation, cells were lysed in in RIPA buffer (Sigma-Aldrich, R0278) containing protease and phosphatase inhibitor cocktail (Thermo Scientific, #1861284) for subsequent western blot analysis.
  • mice were euthanized 30 minutes after the last dose of vehicle or (+)-SW033291 injection and brain tissues were excised. Then, tissues were further rinsed in ice-cold PBS only, and snap-frozen in liquid nitrogen. Frozen samples were pulverized over liquid nitrogen.
  • the powder was transferred to an Eppendorf tube with 500 ⁇ L of cold lysis buffer (50 mM Tris-HCl, pH 7.5, 0.1 mM DTT, 0.1 mM EDTA) and then homogenized using Kimble Kontes Pellet Pestle Cordless Motor (Fisher, K749540-0000) with Kimble Kontes blue pellet pestles (Fisher, K749521-1590) for ⁇ 30s on ice.
  • the suspension was centrifuged for 10 min at 12,000rpm.
  • the enzyme activity was measured by transfer of tritium from a tritiated PGE2 substrate to glutamate by coupling 15-PGDH to glutamate dehydrogenase in a 1 hour reaction with 1mM NAD+, 5 mM NH4Cl, 1 mM ⁇ - keto-glutarate (Sigma#75890), and 16 U of bovine liver glutamic dehydrogenase (Sigma, cat. #G-2501), as described previously.
  • the activity was normalized to protein concentration measured by BCA assay (Thermo Scientific, cat. #23225) and expressed as CPM/Hour/mg protein.
  • the head is secured in a cushioned holder without any restraints, while the body is entirely protected by a metal tube.
  • Microglia and non-microglia isolation for 15-PGDH activity assays [00337] Whole brain tissue was collected in DPBS with calcium, magnesium, glucose, and pyruvate (Thermo Fisher Scientific, 14287080) and processed using the MACS Adult Brain dissociation kit (Miltenyi Biotec, 130-107-677) to generate a single cell suspension. Each sample was digested using a combination of enzymatic and mechanical dissociation after being finely sliced with a scalpel. For enzymatic dissociation, enzyme mixes 1 and 2 (provided in the kit) were used for mechanical dissociation.
  • Protocol for multiplex fluorescent detection (ACD, #323110) of probe for mouse 15-PGDH (Hpgd, #456681), mouse CD11b (Itgam, #311491) was performed according to manufacturer specifications. Fluorescent reagents of Opal 570 (Akoya Biosciences, #FP1488001KT) and Opal 650 (Akoya Biosciences, #FP1495001KT) were used to detect 15-PGDH and CD11b mRNA, respectively. Next, brain sections were blocked in 5% normal donkey serum (NDS) and 5% bovine serum albumin (BSA) in PBS with 0.25% Triton-X (PBS-T) for 1 hour.
  • NDS normal donkey serum
  • BSA bovine serum albumin
  • tissues were incubated with anti-CD31 antibody (BD #550274) in 1:50 dilution in 1% NHS, 1% BSA, in PBS with 0.25% Triton-X for 1hr. After 3 times of PBS wash, tissues were incubated with Donkey anti-rat 488 antibody (1:500, #A21208, Invitrogen). Then, tissues were washed with PBS three times and stained with DAPI (Thermo Scientific, D3571).0.1% Sudan Black B (Sigma-Aldrich, #199694) in 70% ethanol was applied to the sections to reduce autofluorescence signals. After rising with PBS, slides were cover slipped with Vectashield antifade mounting medium (Vector Laboratories, H-1000-10).
  • Fluorescence images were acquired using a z-stack approach with a 0.5 ⁇ m interval, spanning a total depth of 2 ⁇ m, across five optical sections. Orthogonal projection was performed for visualization and analysis. Cell detection and quantification was performed using QuPath. Nuclei were identified based on DAPI staining, and the proximity of each detected cell to the nearest CD31+ endothelial cell to determine the distance to the closest blood vessel. Quantification of immunohistochemistry [00339] Images were acquired with Axiolmager.M2 with monochromatic digital camera (Zeiss Ax- ioCam MRm Rev.3) and Zeiss Axio Scan.Z1.
  • ImageJ version 1.42 software National Institutes of Health, Bethesda, MD was used to analyze the integrated density of silver staining with the plugin of the color deconvolution method, as previously described. BrdU-labelled cells were counted manually using AxioImager.M2 and normalized by the total area of hippocampus. The percentage of Congo Red-positive area was analyzed by measuring total percent area in ImageJ. 4-HNE and 3-NT staining images were captured on LSM 880, and the mean fluorescence intensities and number of colocalized cells were quantified using Fiji Image J.
  • mice were carefully removed from the skull to include the entire brain, cerebellum, and the proximal brainstem, and placed in fresh 10% neutral buffered formalin (NBF) and stored at 4°C for 24 hours.
  • NBF neutral buffered formalin
  • the fixed brains were then rinsed in PBS and placed in an adult mouse Zivic instrument device (Pittsburgh, PA) (coronal 1 mm divisions) (BSMAA001-1) and sectioned to provide seven to eight 2 mm thick sections ranging from immediately posterior to the olfactory lobe to the proximal brain stem.
  • the 2 mm sections were then placed serially on three glass slides to preserve left-right orientation and anterior to posterior order and coated with a thin layer of 2% Sea Plaque Agarose (Lonza, #50104) to maintain optimal flat sections.
  • the composite slabs of brain sections were wrapped in Histowrap®, placed in routine depth processing cassettes, returned to fresh cold 10% NBF, and stored for up to 2 days at 4°C.
  • the fixed tissue slabs were processed for paraffin embedding (FFPE) using an 8-hour standard schedule at the shared Tissue Resource Core within the Case Comprehensive Cancer Center. Paraffin blocks were sectioned at 5 ⁇ m for initial hematoxylin and eosin staining.
  • the paraffin tissue block containing optimal hippocampus morphology was used for further sectioning at 5 ⁇ m for mRNA in situ hybridization.
  • Quantification of in situ hybridization [00341] Whole section scans were acquired with Keyence BZ-X810, which is an inverted fluorescence phase contrast microscope with 2/3 inch, 2.83-million-pixel monochrome CCD (colorized with LC filter). Each field of view was stitched together with BZ-H4XD/Advanced Observation Module. QuPath version 0.3.2 software with the built in Cell detection method was used to identify DAPI positive nuclei, and the QuPath built in Subcellular detection method was used to quantify 15-PGDH, CD206, and TMEM119 positive cells.
  • Proteins were heated at 95°C in either 2x or 4x Laemmli Sample Buffer (Bio-Rad Laboratories, Inc., #1610737 or #1610747) with beta-mercaptoethanol (Bio-Rad Laboratories, Inc., #1610710) for 5 min, and then resolved in 4%–20% Criterion TGX Stain- Free gels (Bio-Rad Laboratories, Inc., #5678095).
  • Proteins were transferred onto 0.2 mm polyvinylidene fluoride (Bio-Rad Laboratories, Inc., #1704157) or nitrocellulose membranes (Bio-Rad Laboratories, Inc., #1704271) with the Trans-Blot Turbo system (Bio-Rad Laboratories, Inc.). Membranes were blocked with 5% nonfat dry milk (Research Products International, #M17200) in tris-buffered saline-tween 20 (TBS-T) for 1 hour at room temperature, and then incubated with primary antibodies at 4°C overnight.
  • TBS-T tris-buffered saline-tween 20
  • mice anti-15-PGDH Markowitz laboratory MM1, 1:1,000
  • mouse anti-GFAP Thermo Fisher, MA5-12023, 1:1,000
  • rabbit anti-IBA1 Wako, #019-19741, 1:1,000
  • rabbit anti-APP Sigma Aldrich, #A8717, 1:5,000
  • mouse anti-GAPDH EMD Millipore Corp, MAB 374, 1:5,000.
  • RNA concentrations, 260/230 ratio, and 260/280 ratio were determined by UV visible absorption spectra, using Nanodrop 2000 (Thermo Scientific, USA).
  • First-strand cDNA was synthesized from total RNA (500ng) using iScript cDNA Synthesis Kit (Bio-Rad Laboratories Inc., #1708891), according to the manufacturer’s instruction.
  • Quantitative PCR was performed in triplicate using Fast Taqman Master Mix (Thermo Fisher Scientific, #444964) on the QuantStudio 6 Pro real-time PCR systems (Applied Biosystems).
  • Taqman probes were used to examine the relative expression of human HPGD gene (Thermo Fisher Scientific, #Hs00168359_m1) and mouse Hpgd (Thermo Fisher Scientific, #Mm00515121_m1) gene. Fold change of gene expression was calculated by the comparative CT quantification method. Mouse gene expression was normalized to expression of Gapdh (Thermo Fisher Scientific, #Mm99999915_g1) and human gene expression was normalized to the geometric mean of three housekeeping genes, ACTB (Thermo Fisher Scientific, #Hs01060665_g1), LDHA (Thermo Fisher Scientific, #Hs01378790_g1), and POLR1B (Thermo Fisher Scientific, #Hs00219263_m1).
  • ACTB Thermo Fisher Scientific, #Hs01060665_g1
  • LDHA Thermo Fisher Scientific, #Hs01378790_g1
  • POLR1B Thermo Fisher Scientific, #Hs002192
  • the single cell transcriptomic profiles of the HPF region are divided into two main parts: (1) the hippocampal region, including fields CA1, CA2, CA3, and dentate gyrus; and (2) the retrohippocampal region, including lateral entorhinal area, medial entorhinal area, parasubiculum, postsubiculum, presubiculum, subiculum, prosubiculum, and the immune/vascular region that includes endothelial cells, smooth muscle cells and pericytes, vascular/leptomeningeal cells, and microglia/perivascular macrophages (Micro-PVM).
  • the hippocampal region including fields CA1, CA2, CA3, and dentate gyrus
  • retrohippocampal region including lateral entorhinal area, medial entorhinal area, parasubiculum, postsubiculum, presubiculum, subiculum, prosubiculum, and the immune/vascular region that includes endothelial cells, smooth muscle cells and pericytes, vascular/leptomen
  • CTX In the cortex (CTX), the following areas were profiled for single cell transcriptomics: frontal pole, primary motor, secondary motor, primary somatosensory (SSp), and supplemental somatosensory.
  • SSp primary somatosensory
  • SSp single cell transcriptomic profiles with 23 different cell subpopulations that mainly included astrocytes, glutamatergic neurons (L2/3 IT, L4/5 IT, Car3, and L6b), vascular/leptomeningeal cells (VLMC), and microglia/perivascular macrophages (Micro-PVM).
  • the Seurat pipeline v4 was employed to study the single cell transcriptomic profile. The preprocessing was performed by removing low-quality cells or empty droplets and dying cells.
  • Expression profiles were normalized using sctransform (SCT) and subsequently scaled to the expression of each gene, so that the mean expression across cells was 0 and variance across cells was 1.
  • SCT sctransform
  • the SCT computes Pearson residuals from regularized negative binomial regression, where sequencing depth is utilized as a covariate in a generalized linear model to remove the influence of technical characteristics while preserving biological heterogeneity.
  • principal component analysis was performed on the top 2000 highly variable genes and the top 10 principal components were inputted to visualization methodologies, including uniform manifold approximation and projection (UMAP) and t-distributed stochastic neighbor embedding (TSNE). Differentially expressed genes have been computed over log2SCT normalized profiles.
  • UMAP uniform manifold approximation and projection
  • TSNE stochastic neighbor embedding
  • a ⁇ peptide preparation [00345] The one milligram of A ⁇ 1-42 peptide (Anygen, Cat. No. AGP-8338) was dissolved in 1 mL of 1,1,1,3,3,3,-hexafluoro-2-propanol (HFIP; Sigma, Cat. No.105228) and incubated at room temperature in the dark for three days with daily vertexing. The solution was then aliquoted into 20 tubes (50 ⁇ g per tube), dried using a SpeedVac, sealed, and stored at ⁇ 80°C until further use.
  • HFIP 1,1,1,3,3,3,3,-hexafluoro-2-propanol
  • a ⁇ oligomers were prepared by dissolving the A ⁇ peptide stock in 11.07 ⁇ L of anhydrous dimethyl sulfoxide and subsequently diluting it with ice-cold 1 ⁇ phosphate-buffered saline to a final concentration of 0.1 mM. The solution was then incubated at 4°C in the dark for 24 hours.
  • ROS measurement [00346] BV2 cells (1.5 ⁇ 104 cells/well) were seeded in a 24-well plate using DMEM supplemented with 10% FBS and 1% Antibiotics/antimycotic.
  • 15-PGDH substrates PGE 2 , PGF 2 ⁇ , RvD1, RvD2, LXA 4 , 15-HETE
  • the culture medium was then replaced with 500 ⁇ L of phenol red-free medium containing 5.7 ⁇ M CM-H2DCFDA, followed by treatment with A ⁇ oligomers (5 ⁇ M).
  • ROS levels were immediately measured using the Incucyte live-cell imaging system (IncuCyte S3, Sartorius, Germany).
  • (+)-SW033291 pharmacokinetics [00347] Studies to evaluate (+)-SW033291 pharmacokinetics were approved and conducted under the oversight of the UT Soiled IACUC. Twenty-one female CD-1 mice were dosed IP with 10 mg/kg (+)-SW033291. For plasma measurements, whole blood was collected, and plasma was processed from whole blood by centrifugation of the ACD anticoagulant-treated blood for 10 minutes at 9600 x g in a microcentrifuge. For measurement of brain levels, brain tissues were collected, rinsed to remove surface adhering blood, weighed, and snap frozen in liquid nitrogen.
  • brain tissue was homogenized in a 3-fold volume of PBS so that the total volume of homogenate in mL was 4 times the weight of tissue in g.
  • Samples were stored at -80°C until analysis.
  • plasma or brain homogenate was diluted with 2 parts methanol containing 0.15% formic acid, 3mM ammonium acetate (final conc.2mM), and 37.5ng/ml internal standard (IS, final conc. 25ng/ml, n-benzylbenzamide, Sigma Aldrich, 02914LH).
  • (+)-SW033291 was detected with the mass spectrometer in multiple reaction monitoring (MRM) mode with compound transition 413.1 to 339.2 and n-benzylbenzamide internal standard transition 212.1 to 91.1.
  • concentration of (+)-SW033291 in each time-point sample was quantified using Sciex Analyst software.
  • a value of 3-fold above the signal obtained from blank plasma or tissue homogenate was designated as the limit of detection (LOD).
  • LOD limit of quantitation
  • LOQ limit of quantitation
  • Brain concentrations were adjusted to remove the amount of compound present in murine brain vasculature (30 ⁇ l/g brain tissue) according to literature values and measured plasma concentrations.
  • TBI is a major risk factor for AD, and these conditions also display various common pathological features.
  • RNA-fluorescence in situ hybridization RNA-FISH
  • Fig.1F RNA-fluorescence in situ hybridization
  • RNA-sequencing sc- RNAseq
  • sc- RNAseq public single-cell RNA-sequencing
  • the next highest Hpgd-expressing population comprised a rare subset of Layer L2/3 glutamatergic cortical neurons, with only 9% of these neurons showing expression (average expression level of 0.1 log2(SCT) (Fig.6B-E).
  • 15-PGDH enrichment was similarly observed in the hippocampal myeloid cell (micro-PVM) cluster (Fig.6F and G).
  • Bead fractionation of murine cortical cells into CD11b + and CD11b- populations confirmed an approximately 200-fold enrichment of 15-PGDH enzyme activity in the CD11b + myeloid cell population (Fig.7A).
  • Dual RNA-FISH confirmed 15-PGDH colocalization with both microglia (Hpgd + red and Tmem119 + green, double positive) and PVMs (Hpgd + red and Mrc1 + green, double positive) in both 5xFAD and WT mice (Fig.1I and J).
  • 5xFAD mice exhibited a two-fold increase in both 15-PGDH-expressing microglia and PVMs within both the cortical and hippocampal regions, as compared to WT littermates (Fig.1I and J).
  • our findings demonstrate that 15-PGDH is predominantly enriched in brain myeloid cells tightly associated with the BBB, with 15-PGDH level and cellular prevalence markedly elevated in AD mouse brain.
  • 15-PGDH inhibition prevents cognitive impairment in AD without reducing amyloid pathology
  • both pharmacologic and genetic inhibition of 15-PGDH robustly prevented BBB structural degradation in 5xFAD mice (Fig.2I and J).
  • the glial limitans barrier formed through astrocytic endfeet interactions with endothelial cells, normally prevents infiltration of blood-derived components into brain tissue and represents a region of functional compromise associated with astrocytic endfeet swelling.
  • IgG immunoglobulin
  • pharmacologic 15- PGDH inhibition fully rescued this neurodegeneration and attenuated axonal degeneration in the dorsal fornix, a major hippocampal output tract (Fig.3A and Fig.10E).
  • Genetic 15- PGDH haploinsufficiency similarly restored survival of adult-born neurons in 5xFAD mice to WT levels (Fig.3B), showing that even partial 15-PGDH suppression suffices for neuroprotection.
  • 15-PGDH inhibition attenuates astrocyte reactivity and microglial oxidative stress in AD
  • 15-PGDH inhibition attenuates astrocyte reactivity and microglial oxidative stress in AD
  • GFAP a marker of astrocyte reactivity
  • IBA1 a marker of microglia activation
  • lipid peroxidation is known to rapidly accumulate within hours of TBI, thereby driving the acute phase of pathology, and also to persist chronically and increase the risk of developing other forms of aging related neurodegeneration, such as AD. Consistent with our previous reports in this model, we detected no changes in IBA1 or GFAP expression after TBI (Fig.13E-G). [00369] To validate target specificity, we further performed TBI studies in Hpgd +/+ and Hpgd -/- mice (Fig.4H). Genetic ablation of 15-PGDH mirrored pharmacologic inhibition, conferring complete protection against axonal neurodegeneration (Fig.4I), BBB damage (Fig.4J), and oxidative stress (Fig.4K and Fig.13H).
  • This Example identifies 15-PGDH as a novel therapeutic target for AD and TBI, demonstrating that its inhibition preserves BBB integrity and cognitive function.
  • 15-PGDH was observed elevated 15-PGDH in aging, TBI, and AD in both human and mouse brains, with prominent localization to brain myeloid cells, including microglia and PVMs at the BBB.
  • Genetic or pharmacologic inhibition of 15-PGDH in mouse models of AD and TBI mitigated perivascular inflammation, protected BBB integrity, and prevented downstream neuroinflammation, oxidative stress, and neurodegeneration.
  • 15-PGDH inhibition is critically positioned to therapeutically interrupt the self-amplifying pathogenic cycle created as ROS generation and BBB degradation reciprocally drive and reinforce one another.
  • 15-PGDH acts to catalyze the degradation of a diverse array of bioactive molecules with context-dependent pro- or anti-inflammatory effects.
  • Our attempts to define global eicosanoid profile changes after 15-PGDH inhibition were confounded by the rapid induction and lability of these signaling molecules.
  • 15-PGDH inhibition represents a paradigm-shifting therapeutic strategy for AD, TBI, and potentially other neurodegenerative conditions marked by BBB dysfunction.

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Abstract

L'invention concerne une méthode de traitement de maladies ou de troubles caractérisés par une détérioration structurale de la barrière hémato-encéphalique ou une génération accrue d'espèces réactives de l'oxygène dans le système nerveux d'un sujet en ayant besoin, comprenant l'administration au sujet d'une quantité d'un inhibiteur de la 15-PGDH et/ou d'un substrat pour la 15-PGDH.
PCT/US2025/035911 2024-06-28 2025-06-30 Compositions et méthodes de traitement de maladies et de troubles caractérisés par une détérioration de la barrière hémato-encéphalique ou la génération d'espèces réactives de l'oxygène Pending WO2026006830A1 (fr)

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Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20230082516A1 (en) * 2019-01-08 2023-03-16 Kyorin Pharmaceutical Co., Ltd. 15-pgdh inhibitor
US20230210829A1 (en) * 2020-06-11 2023-07-06 The Board Of Trustees Of The Leland Stanford Junior University Rejuvenation of aged tissues by inhibition of the pge2 degrading enzyme, 15-pgdh
US20230310390A1 (en) * 2020-08-07 2023-10-05 Case Western Reserve University Inhibitors of short-chain dehydrogenase activity for treating neurodegeneration
US20240043443A1 (en) * 2012-04-16 2024-02-08 Case Western Reserve University Compositions and methods of modulating 15-pgdh activity
US20250034134A1 (en) * 2021-11-18 2025-01-30 Wuhan Humanwell Innovative Drug Research and Development Center Limited Company 15-pgdh inhibitor and use thereof
US20250154162A1 (en) * 2022-01-28 2025-05-15 Scinnohub Pharmaceutical Co., Ltd. Compound for regulating and controlling 15-pgdh activity and preparation method therefor

Patent Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20240043443A1 (en) * 2012-04-16 2024-02-08 Case Western Reserve University Compositions and methods of modulating 15-pgdh activity
US20230082516A1 (en) * 2019-01-08 2023-03-16 Kyorin Pharmaceutical Co., Ltd. 15-pgdh inhibitor
US20230210829A1 (en) * 2020-06-11 2023-07-06 The Board Of Trustees Of The Leland Stanford Junior University Rejuvenation of aged tissues by inhibition of the pge2 degrading enzyme, 15-pgdh
US20230310390A1 (en) * 2020-08-07 2023-10-05 Case Western Reserve University Inhibitors of short-chain dehydrogenase activity for treating neurodegeneration
US20250034134A1 (en) * 2021-11-18 2025-01-30 Wuhan Humanwell Innovative Drug Research and Development Center Limited Company 15-pgdh inhibitor and use thereof
US20250154162A1 (en) * 2022-01-28 2025-05-15 Scinnohub Pharmaceutical Co., Ltd. Compound for regulating and controlling 15-pgdh activity and preparation method therefor

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