WO2025212851A2 - Compositions d'arnm et leurs utilisations dans des vaccins contre le virus varicelle-zona - Google Patents

Compositions d'arnm et leurs utilisations dans des vaccins contre le virus varicelle-zona

Info

Publication number
WO2025212851A2
WO2025212851A2 PCT/US2025/022899 US2025022899W WO2025212851A2 WO 2025212851 A2 WO2025212851 A2 WO 2025212851A2 US 2025022899 W US2025022899 W US 2025022899W WO 2025212851 A2 WO2025212851 A2 WO 2025212851A2
Authority
WO
WIPO (PCT)
Prior art keywords
mrna
sequence
lipid
seq
vzv
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Pending
Application number
PCT/US2025/022899
Other languages
English (en)
Other versions
WO2025212851A3 (fr
Inventor
Gilles BESIN
Kapil BAHL
Luis Alberto BARRERA
Jason GEHRKE
Benjamin Mahler GEILICH
Nahal HABIBI
Kristen OTT
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Orbital Therapeutics Inc
Original Assignee
Orbital Therapeutics Inc
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Orbital Therapeutics Inc filed Critical Orbital Therapeutics Inc
Publication of WO2025212851A2 publication Critical patent/WO2025212851A2/fr
Publication of WO2025212851A3 publication Critical patent/WO2025212851A3/fr
Pending legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/12—Viral antigens
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/12—Antivirals
    • A61P31/20—Antivirals for DNA viruses
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/51—Medicinal preparations containing antigens or antibodies comprising whole cells, viruses or DNA/RNA
    • A61K2039/53—DNA (RNA) vaccination
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/54—Medicinal preparations containing antigens or antibodies characterised by the route of administration
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/545—Medicinal preparations containing antigens or antibodies characterised by the dose, timing or administration schedule
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/555—Medicinal preparations containing antigens or antibodies characterised by a specific combination antigen/adjuvant
    • A61K2039/55511—Organic adjuvants
    • A61K2039/55555—Liposomes; Vesicles, e.g. nanoparticles; Spheres, e.g. nanospheres; Polymers
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/57—Medicinal preparations containing antigens or antibodies characterised by the type of response, e.g. Th1, Th2
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/57—Medicinal preparations containing antigens or antibodies characterised by the type of response, e.g. Th1, Th2
    • A61K2039/572—Medicinal preparations containing antigens or antibodies characterised by the type of response, e.g. Th1, Th2 cytotoxic response
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/57—Medicinal preparations containing antigens or antibodies characterised by the type of response, e.g. Th1, Th2
    • A61K2039/575—Medicinal preparations containing antigens or antibodies characterised by the type of response, e.g. Th1, Th2 humoral response
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2710/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA dsDNA viruses
    • C12N2710/00011—Details
    • C12N2710/16011—Herpesviridae
    • C12N2710/16711—Varicellovirus, e.g. human herpesvirus 3, Varicella Zoster, pseudorabies
    • C12N2710/16734—Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein

Definitions

  • Shingles also known as herpes zoster (HZ), zoster, or zona, is a viral disease characterized by painful skin lesions (typically, a blistering rash). Shingles is caused by the varicella zoster virus (VZV).
  • HZ herpes zoster
  • VZV varicella zoster virus
  • VZV initial infection with VZV causes chickenpox, which generally occurs in children. Once an episode of chickenpox has resolved, the VZV is not eliminated from the body. VZV becomes latent in the nerve cell bodies and, less frequently, in non-neuronal satellite cells of dorsal root, cranial nerve, or autonomic ganglion, without causing any symptoms. Years or decades after a chickenpox infection, the virus may reactivate as shingles. The virus breaks out of nerve cell bodies and travels down nerve axons to cause viral infection of the skin in the region of the nerve (i.e., the dermatome), causing painful skin lesions.
  • the dermatome the region of the nerve
  • Shingles can cause pain, itching, or tingling of the skin followed by a painful rash of blister- like sores, fever, headache, chills, and upset stomach. This reactivation of the VZV causes more than five million cases of shingles annually. According to the Centers for Disease Control and Prevention, an estimated one million people get shingles each year in the United States, and about one out of every three people in the United States will develop shingles in their lifetime. [05] Shingles vaccination is the only way to protect against shingles and postherpetic neuralgia, the most common complication of shingles.
  • Shingrix® is an FDA-approved vaccine indicated for prevention of herpes zoster (HZ)(shingles). Shingrix® is not indicated for prevention of primary varicella infection (chickenpox). Shingrix® is an adjuvanted recombinant VZV that unfortunately causes unwanted side effects, such as pain and swelling at injection sites, muscle pain, headache, fever and shivering. According to the Centers for Disease Control and Prevention, it is estimated that about one out of six people receiving the Shingrix® vaccine had symptoms severe enough to prevent them from performing regular activities.
  • the codon-optimized ORF sequence comprises a sequence at least 97% identical to any one of the sequences of SEQ ID NO: 1-95. In some embodiments, the codon-optimized ORF sequence comprises a sequence at least 98% identical to any one of the sequences of SEQ ID NO: 1-95. In some embodiments, the codon-optimized ORF sequence comprises a sequence at least 99% identical to any one of the sequences of SEQ ID NO: 1-95. In some embodiments, the codon-optimized ORF sequence comprises any one of the sequences of SEQ ID NO: 1-95.
  • the mRNA further comprises a 5’ untranslated region (5’ UTR), a 3’ untranslated region (3’ UTR), and/or a poly(adenine) (poly(A)) sequence.
  • the mRNA further comprises a 5’ UTR, a 3’ UTR, or a poly(A) sequence.
  • the mRNA further comprises a 5’ UTR and a 3’ UTR.
  • the mRNA further comprises a 5’ UTR and a poly(A) sequence.
  • the mRNA further comprises a 3’ UTR and a poly(A) sequence.
  • the mRNA further comprises a 5’ UTR, a 3’ UTR, and a poly(A) sequence.
  • the mRNA comprises, from the 5’ end to the 3’ end, (a) 5’ UTR,(b) the ORF sequence, (c) the 3’ UTR, and (d) the poly(A) sequence.
  • the mRNA further comprises a 5’ cap.
  • the mRNA comprises, from the 5’ end to the 3’ end, (a) 5’ cap, (b) 5’ UTR, (c) the ORF sequence, (d) 3’ UTR and (e) poly(A) sequence.
  • the 5’ cap is CAPO, CAP1 or CAP2. In some embodiments, the 5’ cap is CAPO. In some embodiments, the 5’ cap is CAP1 or a modified CAP1. In some embodiments, the 5’ cap is CAP2 or a modified CAP2. In some embodiments, the 5’ cap is a modified CAP1 comprising (7’ methyl)Guanosine-ppp- (2’-O-methyl)Adenosine-Guanosine (m7G-ppp-AmG).
  • the 5’ cap is a modified CAP1 comprising (7’ methyl-3’-O-methyl)Guanosine- ppp-(N-6 methyl- 2’-O-methyl) Adenosine-Guanosine ((m7-3OMe)G-ppp-(m6-2OMe)AG).
  • the 5’ cap is a modified CAP1 comprising (7’ methyl-3’-O- methyl)Guanosine-ppp- (2’-O-methyl) Adenosine-Guanosine ((m7-3OMe)G-ppp-AmG) .
  • the mRNA comprises a sequence at least 90% identical to any one of SEQ ID NO: 96-190.
  • the mRNA comprises a sequence at least 90% identical to SEQ ID NO: 116. In some embodiments, the mRNA comprises a sequence at least 95% identical to SEQ ID NO: 116. In some embodiments, the mRNA comprises a sequence at least 96% identical to SEQ ID NO: 116. In some embodiments, the mRNA comprises a sequence at least 97% identical to SEQ ID NO: 116. In some embodiments, the mRNA comprises a sequence at least 98% identical to SEQ ID NO: 116. In some embodiments, the mRNA comprises a sequence at least 99% identical to SEQ ID NO: 116. In some embodiments, the mRNA comprises the sequence of SEQ ID NO: 116.
  • the mRNA comprises a sequence at least 90% identical to SEQ ID NO: 119. In some embodiments, the mRNA comprises a sequence at least 95% identical to SEQ ID NO: 119. In some embodiments, the mRNA comprises a sequence at least 96% identical to SEQ ID NO: 119. In some embodiments, the mRNA comprises a sequence at least 97% identical to SEQ ID NO: 119. In some embodiments, the mRNA comprises a sequence at least 98% identical to SEQ ID NO: 119. In some embodiments, the mRNA comprises a sequence at least 99% identical to SEQ ID NO: 119. In some embodiments, the mRNA comprises the sequence of SEQ ID NO: 119.
  • the mRNA is unmodified. In some embodiments, the mRNA is chemically unmodified.
  • At least 98% of the uracil nucleosides are chemically modified. In some embodiments, at least 99% of the uracil nucleosides are chemically modified. In some embodiments, at least 100% of the uracil nucleosides are chemically modified. In some embodiments, the chemical modification is in the sugar subunit of the nucleoside.
  • the present invention provides mRNA formulated in a delivery vehicle.
  • the mRNA is formulated in lipid nanoparticles (LNPs).
  • the mRNA is formulated without a delivery or carrier vehicle.
  • the LNP formulation is stored in a storage buffer.
  • the storage buffer comprises saline. In some embodiments, the storage buffer does not comprise saline.
  • the LNP comprises an ionizable lipid, a helper lipid, cholesterol, and a polyethylene glycol (PEG)-modified lipid.
  • PEG polyethylene glycol
  • the ionizable lipid comprises a compound selected from Table Cl, C2, or C3.
  • the ionizable lipid comprises 3-((((l- ethylpiperidin-3-yl)methoxy)carbonyl)oxy)-2-(((4-(((Z)-oct-5-en-l-yl)oxy)-4-(((Z)-oct-5-en- l-yl)oxy)butanoyl)oxy)methyl)propyl (9Z,12Z)-octadeca-9,12-dienoate (Compound 1).
  • the ionizable lipid comprises ((3-hydroxypropyl)azanediyl)bis(heptane- 7,1-diyl) bis(4,4-bis(((E)-oct-5-en-l-yl)oxy)butanoate) (Compound 37). In some embodiments, the ionizable lipid comprises ((2-hydroxyethyl)azanediyl)bis(hexane-6,l-diyl) bis(6,6-bis(hexyloxy)hexanoate) (Compound 49).
  • the ionizable lipid comprises ((2-hydroxyethyl)azanediyl)bis(heptane-7, 1-diyl) bis(4,4-bis(((Z)-oct-5-en- 1- yl)oxy)butanoate) (Compound 36). In some embodiments, the ionizable lipid comprises nonyl 8-((6-((4,4-bis(octyloxy)butanoyl)oxy)hexyl)(2-hydroxyethyl)amino)octanoate (Compound 82).
  • the ionizable lipid comprises nonyl 8-((2- hydroxyethyl)(6-((4-(((Z)-oct-5-en-l-yl)oxy)-4-(((Z)-oct-5-en-l- yl)oxy)butanoyl)oxy)hexyl)amino)octanoate (Compound 81).
  • the ionizable lipid comprises Compound 1:
  • the ionizable lipid comprises Compound 37: (Compound 37).
  • the ionizable lipid comprises Compound 49:
  • the ionizable lipid comprises Compound 82:
  • the ionizable lipid comprises Compound 81:
  • the LNP comprises a molar ratio of 47.5 % ionizable lipid, 10 % helper lipid, 40-41% cholesterol, and 1.5-2.5 % PEG-modified lipid.
  • the LNP comprises a molar ratio of 47.5 % ionizable lipid, 10 % helper lipid, 40.75% cholesterol, and 1.75 % PEG-modified lipid. In some embodiments, the LNP comprises a molar ratio of 47.5 % ionizable lipid, 10 % helper lipid, 41% cholesterol, and 1.5% PEG-modified lipid.
  • the LNP comprises a molar ratio of 47.5 % Compound 1, 10 % DSPC, 40.75% cholesterol, and 1.75 % DMG-PEG2K. In some embodiments, the LNP comprises a molar ratio of 47.5 % Compound 49, 10 % DSPC, 40.75% cholesterol, and 1.75 % DMG-PEG2K. In some embodiments, the LNP comprises a molar ratio of 47.5 % Compound 1, 10 % DOPE, 40.75% cholesterol, and 1.75 % DMG-PEG2K. In some embodiments, the LNP comprises a molar ratio of 47.5 % Compound 49, 10 % DOPE, 40.75% cholesterol, and 1.75 % DMG-PEG2K.
  • the present invention comprises a vector for making the mRNA described herein.
  • the vector is a non-viral DNA vector.
  • the vector is a viral vector.
  • the vector further comprises a promoter sequence.
  • the present invention provides a composition
  • a composition comprising a messenger ribonucleic acid (mRNA) comprising an open reading frame (ORF) sequence encoding a varicella-zoster virus (VZV) glycoprotein E (gE) polypeptide, wherein the mRNA is formulated in a lipid nanoparticle comprising an ionizable lipid comprising Compound 49.
  • the LNP further comprises a helper lipid, cholesterol, and a PEG-modified lipid.
  • the present invention provides a kit comprising the mRNA described herein. In some embodiments, the present invention provides a kit comprising the compositions described herein.
  • the present invention provides a vaccine comprising an mRNA described herein. In some embodiments, the present invention provides a vaccine comprising the compositions described herein. In some embodiments, the vaccine further comprises an adjuvant. In some embodiments, the vaccine is a prophylactic vaccine.
  • the mRNA comprises a sequence at least 95% identical to any one of SEQ ID NO: 96-190. In some embodiments, the mRNA comprises a sequence at least 96% identical to any one of SEQ ID NO: 96-190. In some embodiments, the mRNA comprises a sequence at least 97% identical to any one of SEQ ID NO: 96-190. In some embodiments, the mRNA comprises a sequence at least 98% identical to any one of SEQ ID NO: 96-190. In some embodiments, the mRNA comprises a sequence at least 99% identical to any one of SEQ ID NO: 96-190. In some embodiments, the mRNA comprises a sequence 100% identical to any one of SEQ ID NO: 96-190.
  • the codon-optimized ORF sequence comprises a sequence at least 90% identical to SEQ ID NO: 24. In some embodiments, the codon-optimized ORF sequence comprises a sequence at least 95% identical to SEQ ID NO: 24. In some embodiments, the codon-optimized ORF sequence comprises a sequence at least 96% identical to SEQ ID NO: 24. In some embodiments, the codon-optimized ORF sequence comprises a sequence at least 97% identical to SEQ ID NO: 24. In some embodiments, the codon-optimized ORF sequence comprises a sequence at least 98% identical to SEQ ID NO: 24. In some embodiments, the codon-optimized ORF sequence comprises a sequence at least 99% identical to SEQ ID NO: 24. In some embodiments, the codon-optimized ORF sequence comprises SEQ ID NO: 24.
  • the administration is for prophylactic treatment of a disease caused by VZV, such as shingles.
  • the immune response is a neutralizing antibody response to the glycoprotein E (gE) polypeptide encoded by the mRNA and a cellular immune response to the gE polypeptide encoded by the mRNA.
  • the immune response is a neutralizing antibody response to the gE polypeptide encoded by the mRNA or a cellular immune response to the gE polypeptide encoded by the mRNA.
  • the immune response is a neutralizing antibody response to the gE polypeptide encoded by the mRNA.
  • the immune response is a cellular immune response to the gE polypeptide encoded by the mRNA.
  • the present invention provides a method for treating or preventing a disease caused by varicella-zoster virus (VZV) comprising administering to a human subject in need thereof the mRNA or the composition described herein.
  • VZV varicella-zoster virus
  • the method is for treating a disease caused by VZV.
  • the method is for preventing a disease caused by VZV.
  • the administration is to prevent primary VZV infection in the human subject. In some embodiments, the administration is to prevent herpes zoster (shingles) in an adult human subject.
  • the mRNA is administered in a single dose. In some embodiments, the mRNA is administered in multiple doses. In some embodiments, the mRNA is administered in two doses. In some embodiments, the mRNA is administered in three doses.
  • the mRNA is administered to the human subject via intramuscular administration. In some embodiments, the mRNA is administered to the human subject via intradermal administration. In some embodiments, the mRNA is administered to the human subject via subcutaneous administration.
  • FIG. 1 is a diagram of the gE antigen polypeptide, and shows the extracellular domain, transmembrane domain (TM), and intracellular domain.
  • exemplary mRNA VZV vaccines encode a truncated gE antigen polypeptide.
  • the gE antigen polypeptide is truncated at residue 562, 568, or 574 as marked in FIG. 1.
  • exemplary gE antigen polypeptides comprise an alanine (A) substituted in place of tyrosine (Y) at residue position 569 (Y569A mutation).
  • FIG. 2 shows the in vitro expression of exemplary mRNA VZV gE constructs.
  • the “Fold-over-Control” (FOC) values were calculated by dividing the geometric mean value of the VZV gE fluorescence intensity of each sample by the geometric mean value of negative control samples.
  • FIG. 4A - FIG 4D show the IgG titers from enzyme-linked immunosorbent assays (ELISAs) performed on serum samples collected after administration of exemplary mRNA VZV vaccines in mice.
  • the exemplary mRNA VZV vaccines were administered intramuscularly on Day 0 and Day 21, and serum samples were collected on Day 21 and Day 42.
  • the IgG titers include total IgG (FIG. 4A) alongside IgGl (FIG. 4B), IgG2b (FIG. 4C), and IgG2c (FIG. 4D) specific titers.
  • FIG. 5 A - FIG. 5B show the ratio of IgG2c titers to IgGl titers in serum samples collected after administration of exemplary mRNA VZV vaccines in mice.
  • the exemplary mRNA VZV vaccines were administered intramuscularly on Day 0 and Day 21, and serum samples were collected on Day 21 (FIG. 5 A) and Day 42 (FIG. 5B).
  • Higher IgG2c/IgGl ratios are indicative of T helper cell 1 (TH1) response in mice.
  • FIG. 6 shows the level of VZV gE-specific neutralizing antibodies titers (PRNT50 titer) in serum samples collected after administration of exemplary mRNA VZV vaccines in mice.
  • FIG. 7A - FIG. 7E summarize the VZV gE-specific CD4 T cell responses following administration of exemplary mRNA VZV gE vaccines in mice (Day 42). Quantification of CD4 T cell responses include interferon-gamma (IFNy)- secreting CD4 T cells (FIG. 7A), Tumor Necrosis Factor-alpha (TNFa)-secreting CD4 T cells (FIG. 7B), interleukin-2 (IL-2) secreting CD4 T cells (FIG. 7C), interleukin-4 (IL-4) secreting CD4 T cells (FIG. 7D), and interleukin-5 (IL-5) secreting CD4 T cells (FIG. 7E).
  • IFNy interferon-gamma
  • TNFa Tumor Necrosis Factor-alpha
  • IL-2 interleukin-2
  • IL-4 interleukin-4
  • IL-5 interleukin-5 secreting CD4 T cells
  • FIG. 11 A - FIG. 1 IB show the total IgG titers as measured by ELISA assays in serum samples collected from mice administered a high (10 pg) dose of exemplary mRNA VZV vaccines formulated in LNP at Day 21 (FIG. 11 A) and Day 35 (FIG. 1 IB).
  • FIG. 12A - FIG. 12B show the total IgG titers as measured by ELISA assays in serum samples collected from mice administered a low (1 pg) dose of exemplary mRNA VZV vaccines formulated in LNP at Day 21 (FIG. 12A) and Day 35 (FIG. 12B).
  • FIG. 13A - FIG. 13C summarize the CD4 T cell response to administration of a high dose of exemplary mRNA VZV vaccines formulated in LNP in mice. Quantification of CD4 T cell responses include IFNy-secreting CD4 T cells (FIG. 13A), TNF a- secreting CD4 T cells (FIG. 13B), and IL-2 secreting CD4 T cells (FIG. 13C).
  • FIG. 15A - FIG. 15B show the body weight in mice administered a high (FIG. 15A) or low (FIG. 15B) dose of exemplary mRNA VZV vaccines formulated in LNP.
  • FIG. 16A - FIG. 16B summarize injection site reactions to administration of a high dose of exemplary mRNA VZV vaccines formulated in LNP in mice at Day 1 (FIG. 16 A) and Day 22 (FIG. 16B). Increased scores represent more severe injection site reactions, while a score of 0 indicated no observed reaction.
  • FIG. 18A - FIG. 18B show the total IgG titers in response to one or two doses of exemplary mRNA VZV vaccines in mice in Experimental Groups as described in Table 4.
  • FIG. 19A - FIG. 19B show the total IgG titers as measured by ELISA assays in serum samples collected from mice administered a high (10 pg) dose of exemplary mRNA VZV vaccines with different 5’ caps in Experimental Groups as described in Table 4.
  • FIG. 20A - FIG. 20B show the total IgG titers as measured by ELISA assays in serum samples collected from mice administered a low (1 pg) dose of exemplary mRNA VZV vaccines with different 5’ caps in Experimental Groups as described in Table 4.
  • FIG. 21A - FIG. 21B show the ratio of IgG2c titers to IgGl titers in serum samples collected after administration of a high (10 pg) dose of exemplary mRNA VZV vaccines with different 5’ caps in mice in Experimental Groups as described in Table 4. Higher IgG2c/IgGl ratios are indicative of T helper cell 1 (TH1) response.
  • TH1 T helper cell 1
  • FIG. 22A - FIG. 22B show the ratio of IgG2c titers to IgGl titers in serum samples collected after administration of a low (1 pg) dose of exemplary mRNA VZV vaccines with different 5’ caps in mice in Experimental Groups as described in Table 4 (FIG. 22B). Higher IgG2c/IgGl ratios are indicative of T helper cell 1 (TH1) response.
  • TH1 T helper cell 1
  • FIG. 23A - FIG. 23B show the total IgG titers as measured by ELISA assays in serum samples collected from mice administered a high (10 pg) dose of exemplary mRNA VZV vaccines prepared with different buffer concentrations in Experimental Groups as described in Table 4.
  • FIG. 24A - FIG. 24B show the total IgG titers as measured by ELISA assays in serum samples collected from mice administered a low (1 pg) dose of exemplary mRNA VZV vaccines prepared with different buffer concentrations in Experimental Groups as described in Table 4.
  • FIG. 25 summarizes the size, polydispersity index (PDI), and encapsulation efficiency (EE%) of LNPs in different buffers after 0, 1, 3, and 5 freeze-thaw (FT) cycles.
  • FIG. 26 shows the percent change in the size, PDI, and mRNA encapsulation efficiency (EE%) of LNPs in various buffers after 1 FT cycle.
  • FIG. 27 shows the size, PDI, and EE% of LNPs with different buffer compositions after storage at 4°C for 0 days, 7 days, or 30 days.
  • FIG. 28 shows the size, PDI, and EE% of LNPs formulated with Compound 49 after a FT cycle.
  • FIG. 29 shows the effect of acidification buffer and nitrogemphosphate (N/P) ratio on the size, PDI, and EE% of LNPs formulated with Compound 1.
  • FIG. 30 shows the effect of acidification buffer and nitrogemphosphate (N/P) ratio on the size, PDI, and EE% of LNPs formulated with Compound 49.
  • FIG. 31A - FIG. 3 IB summarize the in vitro expression of VZV gE, as shown by the percent of VZV gE positive cells, after delivering exemplary mRNA VZV vaccines formulated in LNP formulations to K562 cells (FIG. 31 A), and the area under the curve (AUC) of VZV gE positive cells over time (FIG. 3 IB).
  • FIG. 32A - FIG. 32B summarize the in vitro expression of VZV gE, as shown by the geometric mean fluorescence intensity (gMFI), after delivering exemplary mRNA VZV vaccines formulated in LNP formulations to K562 cells (FIG. 32A), and the area under the curve (AUC) of gMFI over time (FIG. 3 IB).
  • gMFI geometric mean fluorescence intensity
  • FIG. 34A - FIG. 34B summarize injection site reactions to administration of a high (10 pg) dose of exemplary mRNA VZV vaccines formulated in LNP formulations in mice at Day 1 (FIG. 34A) and Day 22 (FIG. 34B). Increased scores represent more severe injection site reactions, with a score of 0 indicating no observed reaction.
  • FIG. 35A - FIG. 35B show the total IgG titers as measured by ELISA assays in serum samples collected from mice administered a high (10 pg) dose of exemplary mRNA VZV vaccines formulated in LNP formulations at Day 21 (FIG. 35A) and Day 35 (FIG. 35B).
  • FIG. 36A - FIG. 36C show the titers of IgGl (FIG. 36A), IgG2b (FIG. 36B), and IgG2c (FIG. 36C) as measured by ELISA assays in serum samples collected from mice administered a high (10 pg) dose of exemplary mRNA VZV vaccines formulated in LNP formulations.
  • FIG. 37 shows the ratio of IgG2c titers to IgGl titers in serum samples collected after administration of a high (10 pg) dose of exemplary mRNA VZV vaccines formulated in LNP formulations in mice. Higher IgG2c/IgGl ratios are indicative of T helper cell 1 (TH1)- response.
  • TH1 T helper cell 1
  • FIG. 39A - FIG. 39C show the percent of IFNy-secreting (FIG. 39A), IL2- secreting (FIG. 39B), and TNFa- secreting CD8 T cells (FIG. 39C) following administration of exemplary mRNA VZV vaccines formulated in LNP formulations in mice (Day 35).
  • FIG. 40A - FIG. 40C show the pharmacokinetic profiles after exemplary mRNA VZV vaccines formulated in LNP formulations comprising Compound 1 and Compound 49 were administered to mice.
  • FIG. 40A shows the pharmacokinetic profile after administration in serum.
  • FIG. 40B shows the pharmacokinetic profile after administration in liver
  • FIG. 40C shows the pharmacokinetic profile after administration in injection site muscle.
  • the pharmacokinetic profiles demonstrated that the biodegradation profiles of the mRNA VZV vaccines formulated in both LNP formulations containing Compound land Compound 49 ionizable lipids were appropriate for safe and efficacious administration.
  • FIG. 41A - FIG. 41D show the anti-gE VZV total IgG in serum as measured by ELISA following administration of exemplary mRNA VZV vaccines formulated in LNP formulations in mice.
  • the anti-gE VZV total IgG in serum was quantified at Day 28 and Day 49 (FIG. 41 A), Day 53 and Day 67 (FIG. 41B), Day 81 and Day 95 (FIG. 41C), and Day 124 (FIG. 41D).
  • Adjuvant refers to agents which confer immunity by themselves.
  • An adjuvant assists the immune system un- specifically to enhance the antigen- specific immune response by e.g. promoting presentation of an antigen to the immune system or induction of an unspecific innate immune response.
  • an adjuvant may preferably e.g. modulate the antigen-specific immune response by e.g. shifting the dominating Th2-based antigen specific response to a more Th 1 -based antigen specific response or vice versa. Accordingly, an adjuvant may favorably modulate cytokine expression/secretion, antigen presentation, type of immune response etc.
  • An adjuvant or an adjuvant component in the broadest sense is typically a (e.g. pharmacological or immunological) agent or composition that may modify, e.g. enhance, the efficacy of other agents, such as a drug or vaccine.
  • a (e.g. pharmacological or immunological) agent or composition that may modify, e.g. enhance, the efficacy of other agents, such as a drug or vaccine.
  • the term refers in the context of the invention to a compound or composition that serves as a carrier or auxiliary substance for immunogens and/or other pharmaceutically active compounds. It is to be interpreted in a broad sense and refers to a broad spectrum of substances that are able to increase the immunogenicity of antigens incorporated into or co-administered with an adjuvant in question.
  • an adjuvant will preferably enhance the specific immunogenic effect of the active agents of the present invention.
  • Adaptive immune response As used herein, the “adaptive immune response” is typically understood to be antigen- specific. Antigen specificity allows for the generation of responses that are tailored to specific antigens, pathogens or pathogen-infected cells. The ability to mount these tailored responses is maintained in the body by “memory cells”. Should a pathogen infect the body more than once, these specific memory cells are used to quickly eliminate it. In this context, the first step of an adaptive immune response is the activation of naive antigen- specific T cells or different immune cells able to induce an antigen- specific immune response by antigen-presenting cells.
  • Dendritic cells take up antigens by phagocytosis and macropinocytosis and are stimulated by contact with e.g. a foreign antigen to migrate to the local lymphoid tissue, where they differentiate into mature dendritic cells.
  • Macrophages ingest particulate antigens such as bacteria and are induced by infectious agents or other appropriate stimuli to express major histocompatibility complex (MHC) molecules.
  • MHC major histocompatibility complex
  • T cells presenting the antigen on MHC molecules leads to activation of T cells which induces their proliferation and differentiation into armed effector T cells.
  • effector T cells The most important function of effector T cells is the killing of infected cells by CD8+ cytotoxic T cells and the activation of macrophages by Thl cells which together make up cell-mediated immunity, and the activation of B cells by both Th2 and Thl cells to produce different classes of antibody, thus driving the humoral immune response.
  • T cells recognize an antigen by their T cell receptors which do not recognize and bind antigen directly, but instead recognize short peptide fragments e.g., of pathogen-derived protein antigens, which are bound to MHC molecules on the surfaces of other cells.
  • Cellular immunity /cellular immune response As used herein, the phrase “cellular immunity” relates typically to the activation of macrophages, natural killer cells (NK), antigen- specific cytotoxic T-lymphocytes, and the release of various cytokines in response to an antigen. In a more general way, cellular immunity is not related to antibodies but to the activation of cells of the immune system. A cellular immune response is characterized e.g.
  • cytotoxic T-lymphocytes that are able to induce apoptosis in body cells displaying epitopes of an antigen on their surface, such as virus-infected cells, cells with intracellular bacteria, and cancer cells displaying tumor antigens; activating macrophages and natural killer cells, enabling them to destroy pathogens; and stimulating cells to secrete a variety of cytokines that influence the function of other cells involved in adaptive immune responses and innate immune responses.
  • Humoral immunity /humoral immune response refers typically to antibody production and the accessory processes that may accompany it.
  • a humoral immune response may be typically characterized, e.g., by Th2 activation and cytokine production, germinal center formation and isotype switching, affinity maturation and memory cell generation.
  • Humoral immunity also typically may refer to the effector functions of antibodies, which include pathogen and toxin neutralization, classical complement activation, and opsonin promotion of phagocytosis and pathogen elimination.
  • Innate immune response As used herein, the term “innate immune response”, also known as non-specific immune response, comprises the cells and mechanisms that defend the host from infection by other organisms in a non-specific manner. The cells of the innate system recognize and respond to pathogens in a generic way, but unlike the adaptive immune system, it does not confer long-lasting or protective immunity to the host.
  • the innate immune system may be e.g. activated by ligands of pathogen-associated molecular patterns (PAMP) receptors, e.g.
  • PAMP pathogen-associated molecular patterns
  • TLRs Toll-like receptors
  • auxiliary substances such as lipopolysaccharides, TNF-alpha, CD40 ligand, or cytokines, monokines, lymphokines, interleukins or chemokines, IL-1, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL- 10, IL- 12, IL-13, IL-14, IL-15, IL-16, IL-17, IL-18, IL-19, IL-20, IL-21, IL-22, IL-23, IL-24, IL-25, IL- 26, IL-27, IL-28, IL-29, IL-30, IL-31, IL-32, IL-33, IFN-alpha, IFN-beta, IFN-gamma, GM- CSF, G-CSF, M-CSF, LT-beta, TNF-alpha, growth factors, and hGH,
  • Subject As used herein, the terms “subject,” “individual,” and “patient,” are used interchangeably herein and refer to any mammalian subject for whom diagnosis, prognosis, treatment, or therapy is desired, particularly humans.
  • Treat or Treatment refers to obtaining a desired pharmacologic and/or physiologic effect.
  • the effect may be prophylactic in terms of completely or partially preventing a disease or symptom thereof and/or may be therapeutic in terms of a partial or complete cure for a disease and/or adverse effect attributable to the disease.
  • Treatment covers any treatment of a disease in a mammal, particularly in a human, and includes: (a) increasing survival time: (b) decreasing the risk of death due to the disease; (c) preventing the disease from occurring in a subject which may be predisposed to the disease but has not yet been diagnosed as having it; (d) inhibiting the disease, i.e., arresting its development (e.g., reducing the rate of disease progression); and (e) relieving the disease, e.g., causing regression of the disease.
  • Vaccine refers to a prophylactic or therapeutic material providing at least one antigen or antigenic function.
  • the antigen or antigenic function may stimulate the body's adaptive immune system to provide an adaptive immune response.
  • Vehicle refers to an agent, e.g., a carrier, that may typically be used within a pharmaceutical composition or vaccine for facilitating administering of the components of the pharmaceutical composition or vaccine to an individual.
  • the present invention provides mRNA vaccines for treating herpes zoster.
  • the present invention relates to mRNA vaccines comprising at least one mRNA molecule encoding at least one VZV antigen for use in the treatment and/or prevention of herpes zoster in an elderly patient.
  • the elderly patient preferably exhibits an age of at least 50 years. Accordingly, vaccination of the patient with mRNA vaccines described herein elicits an immune response in the patient to treat herpes zoster.
  • VZV Varicella zoster virus
  • Chickenpox is associated with the primary infection by VZV. After chickenpox, VZV can permanently rest in the body’s nerves until reactivation leads to shingles. It is estimated that over 99% of Americans bom before 1980 had chickenpox and have the potential to develop shingles (also known as herpes zoster).
  • a weakened immune system is one of the most common “triggers” for shingles and can be caused by, for example and without limitation, increased age, immune suppression, and/or stress. Around one in three people in the United States will likely develop shingles in their lifetime.
  • a vaccine when injected, for example, intramuscularly, is taken up by dendritic cells and trafficked to the draining lymph node.
  • MHC molecules on the dendritic cells present protein antigens from the vaccine and activate T cells.
  • T cells then drive B cell development in the lymph node, resulting in the maturation of the antibody response. Maturation of the antibody response is associated with increases in antibody affinity and induction of different antibody isotypes.
  • the present invention provides, among other things, mRNA molecules comprising coding sequences for mRNA VZV vaccine constructs.
  • the coding sequence also known as open reading frame (ORF), is defined herein as the region beginning with a start codon and ending in an in-frame stop codon.
  • the mRNA molecule includes a region to stop translation.
  • This region may include any translation termination sequence or signal including a stop codon.
  • the region includes a stop codon.
  • the stop codon may be “TGA,” “TAA,” “TGA,” “TAG,” “UGA,” “UAA,” “UGA” or “UAG.”
  • the regions to initiate or terminate translation independently range from 3 to 40 nucleotides, e.g., at least 4, 5-30, 10-20, 10-15, or 4-30 nucleotides in length. Additionally, in some embodiments, these regions comprise, in addition to a start and/or stop codon, one or more signal and/or restriction sequences.
  • a masking agent is used to mask a first start codon or alternative start codon to increase the chance that translation will initiate on a start codon or alternative start codon downstream to the masked start codon or alternative start codon.
  • the mRNA molecule of the present invention comprises a coding sequence.
  • the mRNA molecule of the present invention comprises an open reading frame (ORF).
  • the antigen polypeptide comprises an amino acid sequence selected from Table A, or variants thereof.
  • Table A Exemplary sequences of mRNA VZV vaccines.
  • the mRNA molecule encodes a wild-type VZV gE antigen comprising an amino acid sequence of SEQ ID NO: 199, including the underlined signal peptide.
  • the mRNA molecule encodes a mutated VZV gE antigen.
  • the mRNA molecule encodes a VZV gE polypeptide variant.
  • the VZV gE polypeptide variant is truncated.
  • the truncated VZV gE polypeptide variant lacks the carboxy terminal domain.
  • the truncated VZV gE antigen comprises residues 1-561 of the wild-type VZV antigen.
  • the truncated VZV gE antigen comprises residues 1-567 of the wild-type VZV antigen.
  • the truncated VZV GE antigen comprises residues 1-573 residues of the wild-type VZV antigen. In some embodiments, the mutated VZV antigen comprises a Y569A mutation relative to the wildtype VZV antigen. In some embodiments, the truncated VZV antigen further comprises a
  • the mRNA molecule encodes, in part, a signal peptide. In some embodiments, the mRNA molecule encodes a signal peptide derived from VZV gE. In some embodiments, the mRNA molecule encodes a signal peptide derived from IgGK. In some embodiments, the signal peptide comprises an amino acid sequence of SEQ ID NO: 196:
  • the mRNA molecules of the present invention may be codon-optimized.
  • the coding nucleic acid sequences are codon-optimized.
  • the coding sequences of the mRNA molecules are codon-optimized.
  • the ORF sequences of the mRNA molecules are codon-optimized. Codon optimization methods are known in the art and may be useful in efforts to achieve one or more of several goals.
  • These goals include, but are not limited to, match codon frequencies in target and host organisms to ensure proper folding, alter GC content to increase mRNA stability or reduce secondary structures, minimize tandem repeat codons or base runs that may impair gene construction or expression, customize transcriptional and translational control regions, insert or remove protein trafficking sequences, remove/add post translation modification sites in encoded protein (e.g. glycosylation sites), add, remove or shuffle protein domains, insert or delete restriction sites, modify ribosome binding sites and mRNA degradation sites, to adjust translational rates to allow the various domains of the protein to fold properly, or to reduce or eliminate problem secondary structures within the polynucleotide.
  • match codon frequencies in target and host organisms to ensure proper folding, alter GC content to increase mRNA stability or reduce secondary structures, minimize tandem repeat codons or base runs that may impair gene construction or expression, customize transcriptional and translational control regions, insert or remove protein trafficking sequences, remove/add post translation modification sites in encoded protein (e.g. glycosylation sites
  • a codon-optimized sequence may be one in which codons in a polynucleotide encoding a polypeptide have been substituted to increase the expression, stability and/or activity of the polypeptide.
  • Factors that influence codon optimization include, but are not limited to one or more of the following: (i) variation of codon biases between two or more organisms or genes or synthetically constructed bias tables, (ii) variation in the degree of codon bias within an organism, gene, or set of genes, (iii) systematic variation of codons including context, (iv) variation of codons according to their decoding tRNAs, (v) variation of codons according to GC %, either overall or in one position of the triplet, (vi) variation in degree of similarity to a reference sequence for example a naturally occurring sequence, (vii) variation in the codon frequency cutoff, (viii) structural properties of mRNAs transcribed from the DNA sequence, (ix) prior knowledge about the function of the DNA sequences upon which design of the codon substitution set is to be based, and/or (x) systematic variation of codon sets for each amino acid.
  • Codon optimization tools, algorithms and services are known in the art; nonlimiting examples include, but are not limited to, software from GeneArt (Life Technologies), DNA2.0 (Menlo Park Calif.), and/or proprietary methods.
  • the coding sequence is optimized using optimization algorithms. Codon options for each amino acid are given in Table B.
  • the coding sequences were codon-optimized to increase expression of VZV gE. In some embodiments, the coding sequences were codon-optimized to increase immune response to VZV gE. In some embodiments, the coding sequences were codon-optimized to increase mRNA stability. In some embodiments, the coding sequences were codon-optimized to increase mRNA half-life.
  • the ORF comprises a sequence at least 97% identical to any one of SEQ ID NO: 1-95. In some embodiments, the ORF comprises a sequence at least 98% identical to any one of SEQ ID NO: 1-95. In some embodiments, the ORF comprises a sequence at least 99% identical to any one of SEQ ID NO: 1-95. In some embodiments, the ORF comprises the sequence of any one of SEQ ID NO: 1-95.
  • the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 55. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 56. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 57.
  • the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 61. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 62. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 63.
  • the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 64. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 65. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 66.
  • the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 67. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 68. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 69.
  • the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 70. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 71. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 72.
  • the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 73. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 74. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 75.
  • the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 76. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% , or 100% identical to SEQ ID NO: 77. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% , or 100% identical to SEQ ID NO: 78.
  • the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% , or 100% identical to SEQ ID NO: 79. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% , or 100% identical to SEQ ID NO: 80. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% , or 100% identical to SEQ ID NO: 81.
  • the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% , or 100% identical to SEQ ID NO: 82. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% , or 100% identical to SEQ ID NO: 83. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 84.
  • the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% , or 100% identical to SEQ ID NO: 85. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% , or 100% identical to SEQ ID NO: 86. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% , or 100% identical to SEQ ID NO: 87.
  • the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% , or 100% identical to SEQ ID NO: 88. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% , or 100% identical to SEQ ID NO: 89. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% , or 100% identical to SEQ ID NO: 90.
  • the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% , or 100% identical to SEQ ID NO: 91. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% , or 100% identical to SEQ ID NO: 92. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% , or 100% identical to SEQ ID NO: 93.
  • the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% , or 100% identical to SEQ ID NO: 94. In some embodiments, the ORF comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 95.
  • the ORF comprises a sequence at least 90% identical to SEQ ID NO: 24. In some embodiments, the ORF comprises a sequence at least 95% identical to SEQ ID NO: 24. In some embodiments, the ORF comprises a sequence at least 96% identical to SEQ ID NO: 24. In some embodiments, the ORF comprises a sequence at least 97% identical to SEQ ID NO: 24. In some embodiments, the ORF comprises a sequence at least 98% identical to SEQ ID NO: 24. In some embodiments, the ORF comprises a sequence at least 99% identical to SEQ ID NO: 24. In some embodiments, the ORF comprises a sequence identical to SEQ ID NO: 24.
  • the mRNA molecules of the present disclosure comprise regions that are partially or substantially not translatable, e.g., having a noncoding region. Such noncoding regions are different from the non-coding functional sequences and are located in any region of the mRNA molecule.
  • the non-coding regions include but are not limited to the linker, the spacer and/or the flanking regions. In some embodiments, the noncoding regions are located in more than one region of the mRNA molecule.
  • UTRs Untranslated regions
  • the mRNA molecule comprises at least one untranslated region (UTR), such as a 5’ UTR and/or 3’ UTR.
  • UTRs untranslated regions
  • the 5' UTR starts at the transcription start site and continues to the start codon but does not include the start codon; whereas the 3 'UTR starts immediately following the stop codon and continues until the transcription termination signal.
  • the regulatory features of a UTR can be incorporated into the polynucleotides, primary constructs and/or mRNA molecules of the present invention to enhance the stability of the molecule.
  • Kozak sequences have the consensus GCCRCCAUG, where R is a purine (adenosine or guanosine) three bases upstream of the start codon (AUG).
  • R is a purine (adenosine or guanosine) three bases upstream of the start codon (AUG).
  • the 5’UTR is an engineered 5’UTR.
  • RBPs RNA Binding Proteins
  • 5' UTR of a gene encompasses the DNA sequence and the RNA sequence of the 5' UTR.
  • Exemplary 5’ UTR sequences include SEQ ID NO: 191 and SEQ ID NO: 193.
  • the mRNA molecule comprises a 5’ UTR sequence comprising GGGAAAUAAGAGAGAAAAGAAGAGUAAGAAGAAAUAUAAGAGCCACC (SEQ ID NO: 191).
  • the mRNA molecule comprises a 5’ UTR sequence comprising AGGAAAUAAGAGAGAAAAGAAGAGUAAGAAGAAAUAUAAGAGCCACC (SEQ ID NO: 193).
  • the mRNA molecule provided herein comprises 3' UTR.
  • a 3' UTR is typically the part of an mRNA which is located between the protein coding region (i.e., the open reading frame) and the poly(A) sequence of the mRNA.
  • a 3' UTR of the mRNA is not translated into an amino acid sequence.
  • the 3' UTR sequence is generally encoded by the gene which is transcribed into the respective mRNA during the gene expression process.
  • a 3' UTR corresponds to the sequence of a mature mRNA which is located 3' to the stop codon of the protein coding region, preferably immediately 3' to the stop codon of the protein coding region, and which extends to the 5'- side of the poly (A) sequence, preferably to the nucleotide immediately 5' to the poly (A) sequence.
  • the term “corresponds to” means that the 3' UTR sequence may be an RNA sequence, such as in the mRNA sequence used for defining the 3' UTR sequence, or a DNA sequence which corresponds to such RNA sequence.
  • the 3' UTR may be derived from human beta-globin, human alpha-globin, xenopus beta-globin, xenopus alpha-globin.
  • the 3’ UTR is an engineered UTR sequence.
  • Exemplary 3’ UTR sequences include SEQ ID NO: 192 and SEQ ID NO: 194.
  • the mRNA molecule comprises a 3’ UTR sequence comprising GCUGGAGCCUCGGUGGCCAUGCUUCUUGCCCCUUGGGCCUCCCCCCAGCCCCU CCUCCCCUUCCUGCACCCGUACCCCCGUGGUCUUUGAAUAAAGUCUGAGUGGG CGGC (SEQ ID NO: 192).
  • the mRNA molecule comprises a 3’ UTR sequence comprising GCUGGAGCCUCGGUGGCCAUGCUUCUUGCCCCUUGGGCCUCCCCCCAGCCCCU CCUCCCCUUCCUGCACCCGUACCCCCGUGGUCUUUGAAUAAAGUCUGAGUGGG CGGC (SEQ ID NO: 194).
  • the mRNA molecule comprises a poly A sequence (also called as poly (A) tail).
  • a poly(A) sequence is typically a long sequence of adenosine nucleotides of up to about 500 adenosine nucleotides, e.g., from about 25 to about 400, preferably from about 50 to about 400, more preferably from about 50 to about 300, even more preferably from about 50 to about 250, most preferably from about 60 to about 250 adenosine nucleotides, added to the 3'-end of an RNA.
  • poly(A) sequences, or poly(A) tails may be generated in vitro by enzymatic polyadenylation of the RNA, e.g., using Poly(A)polymerases derived from E. coli or yeast.
  • the length of a poly(A) tail is greater than 30 nucleotides in length. In some embodiments, the poly A tail is greater than 25 nucleotides in length (e.g., at least or greater than about 25, 40, 45, 50, 55, 60, 70, 80, 90, 100, 120, 140, 160, 180, 200, 250, 300, 350, 400, 450, 500, 600, 700, 800, 900, 1000, 1100, 1200, 1300, 1400, 1500, 1600, 1700, 1800, 1900, 2000, 2500, and 3000 nucleotides).
  • the poly A tail is greater than 25 nucleotides in length (e.g., at least or greater than about 25, 40, 45, 50, 55, 60, 70, 80, 90, 100, 120, 140, 160, 180, 200, 250, 300, 350, 400, 450, 500, 600, 700, 800, 900, 1000, 1100, 1200, 1300, 1400, 1500, 1600, 1700, 1800, 1900, 2000, 2500,
  • the poly A tail includes from about 30 to about 3,000 nucleotides (e.g., from 30 to 50, from 30 to 100, from 30 to 250, from 30 to 500, from 30 to 750, from 30 to 1,000, from 30 to 1,500, from 30 to 2,000, from 30 to 2,500, from 50 to 100, from 50 to 250, from 50 to 500, from 50 to 750, from 50 to 1,000, from 50 to 1,500, from 50 to 2,000, from 50 to 2,500, from 50 to 3,000, from 100 to 500, from 100 to 750, from 100 to 1,000, from 100 to 1,500, from 100 to 2,000, from 100 to 2,500, from 100 to 3,000, from 500 to 750, from 500 to 1,000, from 500 to
  • the poly(A) tail is further modified with 1, 2, 3, 4, or 5 guanosines. In some embodiments, the poly(A) tail is further modified with 1 guanosine. In some embodiments, the poly(A) tail is further modified with 2 guanosines. In some embodiments, the poly(A) tail is further modified with 3 guanosines. In some embodiments, the poly(A) tail is further modified with 4 guanosines. In some embodiments, the poly(A) tail is further modified with 5 guanosines.
  • the poly(A) tail is further modified with 1, 2, 3, 4, or 5 cytosines. In some embodiments, the poly(A) tail is further modified with 1 cytosine. In some embodiments, the poly(A) tail is further modified with 2 cytosines. In some embodiments, the poly(A) tail is further modified with 3 cytosines. In some embodiments, the poly(A) tail is further modified with 4 cytosines. In some embodiments, the poly(A) tail is further modified with 5 cytosines.
  • the poly(A) tail is further modified with 1, 2, 3, 4, or 5 uracils. In some embodiments, the poly(A) tail is further modified with 1 uracil. In some embodiments, the poly(A) tail is further modified with 2 uracils. In some embodiments, the poly(A) tail is further modified with 3 uracils. In some embodiments, the poly(A) tail is further modified with 4 uracils. In some embodiments, the poly(A) tail is further modified with 5 uracils. vi. 5’ Capping
  • a 5' cap is typically a modified nucleotide, particularly a guanosine nucleotide containing a methyl group at the 7 position, added to the 5 '-end of an mRNA molecule through a 5 '-5 '-triphosphate linkage, denoted as m7GpppN, where N is the first transcribed base of the mRNA).
  • N is alanine.
  • N is guanosine.
  • N is uracil.
  • N is cytosine.
  • N is an unmodified nucleotide.
  • N is a modified nucleotide.
  • Synthetic mRNA can be capped enzymatically (e.g., with vaccinia capping enzyme) or through co-transcription with a di- or tri-nucleotide cap analog (e.g., m7GpppApG).
  • a di- or tri-nucleotide cap analog e.g., m7GpppApG.
  • RNA 7(10): 1486-495 Further cap analogues have been described previously (U.S. Pat. No. 7,074,596, W02008/016473, WO2008/157688, WO2009/149253, WO2011/015347, and WO2013/059475).
  • the 5’ cap comprises m7GpppAG, m7GpppGG, 3’0Me,m7Gppp-m6AG, or 3’0Me,m7Gppp-AG. In some embodiments, the 5’ cap comprises m7GpppAG. In some embodiments, the 5’ cap comprises m7GpppGG. In some embodiments, the 5’ cap comprises 3’0Me,m7Gppp-m6AG. In some embodiments, the 5’ cap comprises 3’0Me,m7Gppp-AG. vii. Modified Nucleotides
  • the mRNA molecule comprises one or more modified nucleotides. In some embodiments, the mRNA molecule is unmodified.
  • the mRNA molecules of the present invention may include one, two, three, or more modifications.
  • the modified nucleotides are located in coding region(s). In some embodiments, the modified nucleotides are in the untranslated region(s).
  • the modifications stabilize the mRNA molecule and enhance resistance to degradation as compared to unmodified nucleotides.
  • modified nucleotides enhance biological functions of nucleic acid molecules, for example, increase binding to a RNA binding protein or increasing translation.
  • the modified nucleotide is one or more of Nl- methylpseudouridine, 5-methoxyuridine, N6-methyladenosine, pseudouridine or 5- methylcytosine.
  • the modified nucleotide is Nl-methylpseudouridine. In some embodiments, the modified nucleotide is 5-methoxyuridine. In some embodiments, the modified nucleotide is N6-methyladenosine. In some embodiments, the modified nucleotide is pseudouridine. In some embodiments, the modified nucleotide is 5-methylcytosine.
  • the mRNA molecules of the present disclosure may contain from about 0% to about 100% modified nucleotides (either in relation to overall nucleotide content, or in relation to one or more types of nucleotide, i.e. any one or more of A, G, T/U or C) or any intervening percentage (e.g., from 1% to 20%, from 1% to 25%, from 1% to 50%, from 1% to 60%, from 1% to 70%, from 1% to 80%, from 1% to 90%, from 1% to 95%, from 10% to 20%, from 10% to 25%, from 10% to 50%, from 10% to 60%, from 10% to 70%, from 10% to 80%, from 10% to 90%, from 10% to 95%, from 10% to 100%, from 20% to 25%, from 20% to 50%, from 20% to 60%, from 20% to 70%, from 20% to 80%, from 20% to 90%, from 20% to 95%, from 20% to 100%, from 50% to 60%, from 50% to 70%, from 50% to 80%, from 50% to 90%, from 20% to 95%, from 20% to 100%
  • nucleotide sugar, base and phosphate moiety, e.g., linkage
  • any modification to any portion of a nucleotide, or nucleoside will constitute a modification.
  • the mRNA molecules are designed with a patterned array of sugar, nucleobase or linkage modifications.
  • the mRNA molecules comprise modifications to maximize stability.
  • the mRNA molecules comprise modifications to decrease stability.
  • nucleobases and nucleosides having a modified uracil include pseudouridine (y), pyridin-4- one ribonucleoside, 5-aza-uridine, 6-aza-uridine, 2-thio-5-aza-uridine, 2-thio-uridine (s 2 U), 4- thio-uridine (s 4 U), 4-thio-pseudouridine, 2-thio-pseudouridine, 5-hydroxy-uridine (ho 5 U), 5- aminoallyl-uridine, 5-halo-uridine (e.g., 5-iodo-uridine (I 5 U) or 5-bromo-uridine (br 5 U)), 3- methyl-uridine (m 3 U), 5-methoxy-uridine (mo 5 U), uridine 5-oxyacetic acid (cmo 5 U), uridine 5-oxyacetic acid methyl ester (mcmo 5 U), 5-carboxymethyl-uridine (cm 5 U), 1-carboxymethyl- pseudouridine, 5-
  • 2-thio- 1 -methyl-pseudouridine 1 -methyl- 1 -deaza-pseudouridine, 2-thio- 1 -methyl- 1 -deaza- pseudouridine, dihydrouridine (D), dihydropseudouridine, 5,6-dihydrouridine, 5-methyl- dihydrouridine (m 5 D), 2-thio-dihydrouridine, 2-thio-dihydropseudouridine, 2-methoxy- uridine, 2-methoxy-4-thio-uridine, 4-methoxy-pseudouridine, 4-methoxy-2-thio- pseudouridine, N1 -methyl-pseudouridine (also known as 1 -methylpseudouridine (m , 3- (3-amino-3-carboxypropyl)uridine (acp 3 U), l-methyl-3-(3-amino-3- carboxypropyl)pseudouridine (acp 3 y), 5-
  • the modified nucleobase is a modified adenosine(A).
  • exemplary nucleobases and nucleosides having a modified adenosine include 2-amino- purine, 2, 6-diaminopurine, 2-amino-6-halo-purine (e.g., 2-amino-6-chloro-purine), 6-halo- purine (e.g., 6-chloro-purine), 2-amino-6-methyl-purine, 8-azido-adenosine, 7-deaza- adenosine, 7-deaza-8-aza- adenosine, 7-deaza-2-amino-purine, 7-deaza-8-aza-2-amino-purine,
  • the modified nucleobase is a modified guanosine(G).
  • exemplary nucleobases and nucleosides having a modified guanosine include inosine (I), 1- methyl-inosine (m 1 !), wyosine (imG), methylwyosine (mimG), 4-demethyl-wyosine (imG- 14), isowyosine (imG2), wybutosine (yW), peroxywybutosine (02yW), hydroxywybutosine (OHyW), undermodified hydroxywybutosine (OHyW*), 7-deaza-guanosine, queuosine (Q), epoxy queuo sine (oQ), galactosyl-queuosine (galQ), mannosyl-queuosine (manQ), 7-cyano-7- deaza-guanosine (preQo), 7-aminomethyl-7-
  • the nucleobase of the nucleotide is independently selected from a purine, a pyrimidine, a purine or pyrimidine analog.
  • the nucleobase and/or analog is each independently selected from adenosine, cytosine, guanosine, uracil, naturally-occurring and synthetic derivatives of a base, including but not limited to pyrazolo[3,4-d]pyrimidines, 5-methylcytosine (5-me-C), 5 -hydroxymethyl cytosine, xanthine, hypoxanthine, 2-aminoadenosine, 6-methyl and other alkyl derivatives of adenosine and guanosine, 2-propyl and other alkyl derivatives of adenosine and guanosine, 2- thiouracil, 2-thiothymine and 2-thiocytosine, 5-propynyl uracil and cytosine, 6-azo
  • the mRNA molecule comprises a nucleoside modification.
  • one or more atoms of a pyrimidine nucleobase is replaced or substituted, for example, with optionally substituted amino, optionally substituted thiol, optionally substituted alkyl (e.g., methyl or ethyl), optionally substituted or halo (e.g., chloro or fluoro) atoms or groups.
  • uracil nucleosides of the mRNA molecule of the present disclosure are all modified. In some embodiments, at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, or 100% of the uracil nucleosides of the mRNA molecule are modified. In some embodiments, the uracil nucleosides of the mRNA molecule comprise modifications in the sugar subunit.
  • modifications of the modified nucleosides and nucleotides are present in the sugar subunit.
  • the mRNA molecule described herein comprise at least one sugar modification.
  • mRNA includes the sugar subunit: ribose, which is a 5-membered ring having an oxygen.
  • the 2' hydroxyl group (OH) can be modified or replaced with a number of different substituents.
  • exemplary sugar modifications include replacement of the oxygen(O) in ribose (e.g., with S, Se, or alkylene, such as methylene or ethylene); addition of a double bond (e.g., to replace ribose with cyclopentenyl or cyclohexenyl); ring contraction of ribose (e.g., to form a 4-membered ring of cyclobutane or oxetane); ring expansion of ribose (e.g., to form a 6- or 7-membered ring having an additional carbon or heteroatom, such as for anhydrohexitol, altritol, mannitol, cyclohexanyl, cyclohexenyl, and morpholino that also has a phosphoramidate backbone); multicyclic forms (e.g., tricyclo; and “unlocked” forms, such as glycol nucleic acid (GNA) (e.
  • GAA
  • the sugar subunit contains one or more carbons that possess the opposite stereochemical configuration than that of the corresponding carbon in ribose.
  • mRNA molecules as described herein include nucleotides containing, e.g., arabinose, as the sugar.
  • one or more modifications are present in the intemucleoside linkage (the linking phosphate or the phosphodiester linkage or the phosphodiester backbone).
  • the phrases “phosphate” and “phosphodiester” are used interchangeably.
  • backbone phosphate groups are modified by replacing one or more of the oxygen atoms with a different substituent.
  • modified nucleosides and nucleotides include replacement of an unmodified phosphate moiety with another intemucleoside linkage as described herein.
  • modified phosphate groups include, but are not limited to, phosphorothioate, methylphosphonates phosphoroselenates, boranophosphates, boranophosphate esters, hydrogen phosphonates, phosphoramidates, phosphorodiamidates, alkyl or aryl phosphonates, and phosphotriesters.
  • Phosphorodithioates have both non-linking oxygens replaced by sulfur.
  • the phosphate linker is also modified by the replacement of a linking oxygen with nitrogen (bridged phosphoramidates), sulfur (bridged phosphorothioates), and carbon (bridged methylene-phosphonates).
  • the a-thio substituted phosphate moiety is provided to confer stability to RNA and DNA polynucleotides through unnatural phosphorothioate backbone linkages.
  • Phosphorothioate DNA and RNA have increased nuclease resistance and subsequently a longer half-life in a cellular environment.
  • Phosphorothioate linked polynucleotide molecules are expected to also reduce the innate immune response through weaker binding/activation of cellular innate immune molecules.
  • modified nucleotides incorporated in the mRNA molecules include, for example, 2’-O-Methyl-modified or 2’-O-Methoxyethyl-modified nucleotides (2’- OMe and 2’-M0E modifications, respectively), an alpha-thio-nucleoside (e.g., 5'-O-(l- thiophosphate)-adenosine, 5'-O-(l-thiophosphate)-cytidine (a-thio-cytidine), 5'-O-(l- thiophosphate)-guanosine, 5'-O-(l-thiophosphate)-uridine, or 5'-O-(l-thiophosphate)- pseudouridine.
  • an alpha-thio-nucleoside e.g., 5'-O-(l- thiophosphate)-adenosine, 5'-O-(l-thiophosphate)-cytidine (a
  • Additional modifications to mRNA molecules of the present disclosure include, for example, modification or deletion of nucleotides (or codons) encoding one or more N-linked glycosylation site in a translated polypeptide.
  • nucleobase modifications e.g., nucleobase modifications, and/or intemucleoside linkages (e.g., backbone structures) are introduced at various positions in a polynucleotide described herein.
  • nucleotide analogs or other modification(s) may be located at any position(s) of a polynucleotide such that the function of the polynucleotide is not substantially decreased.
  • the one or more modified nucleotides is a 2' O-methyl or a phosphorothioate modified nucleotide. Accordingly, in some embodiments, the one or more modified nucleotides comprises a 2' O-methyl modification. In some embodiments, the one or more modified nucleotides comprises a phosphorothioate modification.
  • RNA bases include for example, 2'-O- methoxy-ethyl bases (2'-MOE) such as 2-MethoxyEthoxy A, 2-MethoxyEthoxy MeC, 2- MethoxyEthoxy G, 2-MethoxyEthoxy T.
  • Other modified bases include for example, 2'-O- Methyl RNA bases, and fluoro bases.
  • fluoro bases are known, and include for example, Fluoro C, Fluoro U, Fluoro A, Fluoro G bases.
  • 2'OMethyl modifications can also be used with the methods described herein.
  • the mRNA molecule comprises one or more of the following modifications: phosphorothioates, 2'0-methyls, 2' fluoro (2'F), DNA.
  • the mRNA molecule comprises 2'OMe modifications at the 3' and 5 '-ends.
  • the mRNA molecule comprises one or more of the following modifications: 2' -O-2-Methoxyethyl (MOE), locked nucleic acids, bridged nucleic acids, unlocked nucleic acids, peptide nucleic acids, morpholino nucleic acids.
  • MOE 2' -O-2-Methoxyethyl
  • the mRNA molecule comprises one or more of the following base modifications: 2,6-diaminopurine, 2-aminopurine, pseudouracil, Nl-methyl-psuedouracil, 5' methyl cytosine, N6-methyladenosine, 2'pyrimidinone (zebularine), thymine.
  • the mRNA molecule can comprise a modified base such as, for example, 5', Int, 3' Azide (NHS Ester); 5' Hexynyl; 5', Int, 3' 5-Octadiynyl dU; 5', Int Biotin (Azide); 5', Int 6- FAM (Azide); and 5', Int 5-TAMRA (Azide).
  • modified base such as, for example, 5', Int, 3' Azide (NHS Ester); 5' Hexynyl; 5', Int, 3' 5-Octadiynyl dU; 5', Int Biotin (Azide); 5', Int 6- FAM (Azide); and 5', Int 5-TAMRA (Azide).
  • Other examples of RNA nucleotide modifications that can be used with the methods described herein include for example phosphorylation modifications, such as 5 '-phosphorylation and 3 '-phosphoryl
  • the mRNA molecule can also have one or more of the following modifications: an amino modification, biotinylation, thiol modification, alkyne modifier, adenylation, Azide (NHS Ester), Cholesterol-TEG, and Digoxigenin (NHS Ester).
  • the mRNA molecule is synthetic RNA.
  • the mRNA molecule is chemically synthesized.
  • RNA synthesis is carried out in synthesizer machines using nucleotide triphosphate derivatives known as phosphoramidites, which are building blocks of linear oligonucleotides. Nucleoside phosphoramidites use inert substituents to protect reactive moieties such as hydroxyl and amino groups from undesirable reactions and promote phosphodiester bond formation leading to greater homogenous yields. Once synthesis is complete, these groups are removed to generate RNA oligonucleotides of high purity.
  • the mRNA described herein can be purified by methods commonly known in the art.
  • the mRNA molecule is in vitro transcribed RNA.
  • An in vitro transcription (IVT) reaction typically comprises a double-stranded DNA (dsDNA) template, ribonucleotide triphosphates, and a DNA-dependent RNA polymerase.
  • dsDNA double-stranded DNA
  • ribonucleotide triphosphates ribonucleotide triphosphates
  • DNA-dependent RNA polymerase is derived from bacteriophage.
  • the DNA-dependent RNA polymerase is a T7 RNA polymerase, SP6 RNA polymerase, or T3 RNA polymerase.
  • the present invention provides DNA templates for the RNA molecules described herein.
  • the DNA template contains a promoter sequence to which the polymerase binds and catalyzes downstream transcription.
  • the promoter is about 20-40 nucleotides (nt) long.
  • the DNA template is a double-stranded PCR product.
  • the DNA template is a linearized plasmid containing a promoter upstream of the DNA sequence to be transcribed.
  • viral vectors are used to package the constructs for producing the mRNA molecules described herein.
  • AAV vectors are used to construct the mRNA molecules.
  • non- viral vectors such as plasmids, cosmids and artificial chromosomes are used to construct the mRNA molecules.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NO: 96-190 (Table 9).
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 96.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 97.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 101. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 102. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 103.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 104. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 105. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 106.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 107. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 108. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 109.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 113. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 114. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 115.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 116. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 117. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 118.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 122. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 123. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 124.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 125. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 126. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 127.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 128. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 129. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 130.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 131. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 132. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 133.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 134. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 135. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 136.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 137. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 138. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 139.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 140. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 141. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 142.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 143. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 144. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 145.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 152. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 153. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 154.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 155. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 156. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 157.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 161. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 162. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 163.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 167. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 168. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 169.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 170. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 171. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 172.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 173. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 174. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 175.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 176. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 177. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 178.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 179. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 180. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 181.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 182. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 183. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 184.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 185. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 186. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 187.
  • an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 188. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 189. In some embodiments, an mRNA molecule for VZV vaccine comprises a sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 190.
  • the mRNA molecule comprises a sequence at least 90% identical to any one of SEQ ID NO: 96-190. In some embodiments, the mRNA molecule comprises a sequence at least 95% identical to any one of SEQ ID NO: 96-190. In some embodiments, the mRNA molecule comprises a sequence at least 96% identical to any one of SEQ ID NO: 96-190. In some embodiments, the mRNA molecule comprises a sequence at least 97% identical to any one of SEQ ID NO: 96-190. In some embodiments, the mRNA molecule comprises a sequence at least 98% identical to any one of SEQ ID NO: 96-190. In some embodiments, the mRNA molecule comprises a sequence at least 99% identical to any one of SEQ ID NO: 96-190.
  • the mRNA molecule comprises a sequence at least 90% identical to SEQ ID NO: 116. In some embodiments, the mRNA molecule comprises a sequence at least 95% identical to SEQ ID NO: 116. In some embodiments, the mRNA molecule comprises a sequence at least 96% identical to SEQ ID NO: 116. In some embodiments, the mRNA molecule comprises a sequence at least 97% identical to SEQ ID NO: 116. In some embodiments, the mRNA molecule comprises a sequence at least 98% identical to SEQ ID NO: 116. In some embodiments, the mRNA molecule comprises a sequence at least 99% identical to SEQ ID NO: 116. In some embodiments, the mRNA molecule comprises a sequence identical to SEQ ID NO: 116.
  • the mRNA molecule comprises a sequence at least 90% identical to SEQ ID NO: 119. In some embodiments, the mRNA molecule comprises a sequence at least 95% identical to SEQ ID NO: 119. In some embodiments, the mRNA molecule comprises a sequence at least 96% identical to SEQ ID NO: 119. In some embodiments, the mRNA molecule comprises a sequence at least 97% identical to SEQ ID NO: 119. In some embodiments, the mRNA molecule comprises a sequence at least 98% identical to SEQ ID NO: 119. In some embodiments, the mRNA molecule comprises a sequence at least 99% identical to SEQ ID NO: 119. In some embodiments, the mRNA molecule comprises a sequence identical to SEQ ID NO: 119.
  • the mRNA of the present invention is formulated to improve delivery.
  • the mRNA can be formulated in lipid nanoparticles (LNPs), polymer-based nanoparticles, lipid-polymer based nanoparticles virus like particles (VLPs), and engineered exosomes.
  • LNPs lipid nanoparticles
  • VLPs lipid-polymer based nanoparticles virus like particles
  • the mRNA is delivered without any carrier molecules, or naked.
  • the mRNA is delivered in complex with cationic peptides or polymers.
  • an mRNA molecule of the present invention is formulated in lipid nanoparticles (LNPs).
  • LNP components are selected based on the desired target, cargo (e.g., mRNA molecules), size, and/or other desired feature.
  • LNP components include, for example, ionizable lipids, helper lipids, sterols, and/or PEG-lipids. The relative amounts, or molar ratios, of ionizable lipid, helper lipid, cholesterol, and PEG-lipid are optimized for a given target or administration route.
  • the LNPs do not contain a targeting ligand.
  • the LNPs contain a targeting ligand.
  • LNPs are small solid or semi-solid particles possessing an exterior lipid layer with a hydrophilic exterior surface that is exposed to the non-LNP environment, an interior space which may aqueous (vesicle like) or non-aqueous (micelle like), and at least one hydrophobic inter-membrane space.
  • LNP membranes may be lamellar or non-lamellar and may be comprised of 1, 2, 3, 4, 5 or more layers.
  • the LNPs of the present invention can be prepared with any method commonly known in the art.
  • the LNPs comprise a storage buffer.
  • the storage buffer comprises saline.
  • the storage buffer does not comprise saline.
  • LNP sizes vary.
  • the LNPs for formulating the mRNA molecules of the present disclosure have an average hydrodynamic diameter of 10-1000 nm (i.e., 10-100 nm, 10-150 nm, 10-200 nm, 50-100 nm, 50-120 nm, 50-150 nm, 60-90 nm, 60- 120 nm, 80-100 nm, 80-120 nm, 100-200 nm, 100-500 nm, 200-800 nm, 100-1000 nm, or 500-1000 nm).
  • 10-1000 nm i.e., 10-100 nm, 10-150 nm, 10-200 nm, 50-100 nm, 50-120 nm, 50-150 nm, 60-90 nm, 60- 120 nm, 80-100 nm, 80-120 nm, 100-200 nm, 100-500 nm, 200-800 nm, 100-1000 nm, or 500
  • the average hydrodynamic diameter is at least 10 nm, at least 20 nm, at least 50 nm, at least 60 nm, at least 70 nm, at least 80 nm, at least 90 nm, at least 100 nm, at least 110 nm, at least 120 nm, or at least 150 nm. In some embodiments, the average hydrodynamic diameter is less than 20 nm, less than 50 nm, less than 60 nm, less than 70 nm, less than 80 nm, less than 90 nm, less than 100 nm, less than 110 nm, less than 120 nm, or less than 150 nm.
  • the average hydrodynamic diameter is about 10 nm, about 20 nm, about 50 nm, about 60 nm, about 70 nm, about 80 nm, about 90 nm, about 100 nm, about 110 nm, about 120 nm, or about 150 nm.
  • the LNP comprises at least one ionizable lipid.
  • Ionizable lipids generally contain an amine-containing group on the head group.
  • the ionizable lipid comprises an ionizable cationic lipid.
  • the ionizable lipid comprises an ionizable anionic lipid.
  • the ionizable lipid comprises a compound disclosed in WO 2021/141969 Al (Hamilton et al.), the entirety of which is incorporated by reference herein. In some embodiments, the ionizable lipid comprises a compound of Formula (I) of WO 2021/141969 Al (Hamilton et al.).
  • R 1 in Formula (I) comprises C9-C20 alkyl or C9-C20 alkenyl with 1-3 units of unsaturation.
  • R 1 comprises a C9-C20 alkenyl with 2 units of unsaturation, such as, without limitation, a C17 alkenyl with 2 units of unsaturation.
  • X 3 , X 5 , and X 6 in Formula (I) are independently absent.
  • X 1 is -O-. In some embodiments, X 1 is absent.
  • X 2 is X 7 .
  • X 2 is -(CH2) a - or - CH(OH)-.
  • a is an integer between 0 and 6.
  • a is 0, 1, 2, 3, 4, 5, or 6.
  • a is 0 and X 2 is absent.
  • a is 1.
  • X 7 is independently hydrogen or hydroxyl. In some embodiments, X 7 is hydroxyl. In some embodiments, X 7 is hydrogen.
  • X 4 is a 6-membered heterocyclyl optionally substituted with 1 or 2 Ci-Ce alkyl groups.
  • the heterocyclyl comprises at least one nitrogen.
  • X 4 is piperidinyl.
  • X 4 is ethylpiperidinyl.
  • nl is an integer between 1 and 6. In some embodiments, nl is 1, 2, 3, 4, 5, or 6. In some embodiments, nl is 2.
  • the ionizable lipid is a compound selected from Table Cl. Table Cl. Exemplary ionizable lipids of Formula (I).
  • L 1 is C2-C6 heteroalkylenyl comprising at least 1 heteroatom.
  • the heteroatom is oxygen.
  • L 1 is a C4 heteroalkylenyl comprising 1 oxygen atom, such as, for example and without limitation, - OCH2CH2CH2-.
  • L 1 is a C3 heteroalkylenyl comprising 1 oxygen atom, such as, for example and without limitation, - OCH2CH2-.
  • each L is independently C1-C5 alkylenyl.
  • each L 2 is independently C4-C8 alkylenyl.
  • the ionizable lipid is a compound selected from Table C2. Table C2. Exemplary ionizable lipids of Formula (III-a-i).
  • the ionizable lipid is a compound of Formula (III-a-i) comprising ((3-hydroxypropyl)azanediyl)bis(heptane-7, 1-diyl) bis(4,4-bis(((E)-oct-5-en- 1- yl)oxy)butanoate) (Compound 37).
  • the ionizable lipid is a compound of Formula (III-a-i) comprising ((2-hydroxyethyl)azanediyl)bis(hexane-6, 1-diyl) bis(6,6-bis(hexyloxy)hexanoate)
  • the ionizable lipid is a compound of Formula (III-a-i) comprising ((2-hydroxyethyl)azanediyl)bis(heptane-7, 1-diyl) bis(4,4-bis(((Z)-oct-5-en-l- yl)oxy)butanoate) (Compound 36).
  • the ionizable lipid comprises a compound of Formula (I’ ’-a iii) of WO 2022/140252 Al (Patwardhan et al.).
  • R 1 is hydrogen
  • E 1 is C2-C6 heteroalkylenyl comprising at least 1 heteroatom.
  • the heteroatom is oxygen.
  • E 1 is a C3 heteroalkylenyl comprising 1 oxygen atom, such as, for example and without limitation, - OCH2CH2-.
  • each R is independently C6-C12 alkyl or C6-C12 alkenyl with 1-3 units of unsaturation.
  • each L is independently C1-C5 alkylenyl.
  • each L 2 is independently C4-C8 alkylenyl.
  • the ionizable lipid is a compound of Formula (I”-a-iii) comprising nonyl 8-((2-hydroxyethyl)(6-((4-(((Z)-oct-5-en-l-yl)oxy)-4-(((Z)-oct-5-en-l- yl)oxy)butanoyl)oxy)hexyl)amino)octanoate (Compound 81).
  • the ionizable lipid may be selected from any lipid known in the art, such as, but not limited to, DLin-MC3-DMA, DLin-DMA, Cl 2-200 and DLin-KC2-DMA.
  • the ionizable lipid constitutes 40 mol% to 50 mol% of the total moles of components in the LNP. In some embodiments, the ionizable lipid constitutes 45 mol% to 50 mol% of the total moles of components in the LNP. In some embodiments, the ionizable lipid constitutes 47 mol% to 48 mol% of the total moles of components in the LNP. ii. Sterols
  • Sterols can aid in stability and promote membrane fusion of the LNPs.
  • the LNP comprises at least one sterol.
  • the sterol comprises cholesterol.
  • the sterol is unmodified cholesterol.
  • the sterol comprises a variant of cholesterol.
  • the LNP comprises a derivative of cholesterol.
  • Cholesterol variants may be side-chain or ring oxidized from enzymes acting on unmodified cholesterol.
  • the cholesterol is oxidized on the beta-ring structure or on the hydrocarbon tail structure.
  • Cholesterols include, for example and without limitation, 25 -hydroxy cholesterol (25-OH), 20a-hydroxycholesterol (20a-OH), 27 -hydroxy cholesterol, 6-keto-5a-hydroxycholesterol, 7-ketocholesterol, 7 - hydroxycholesterol, 7a-hydroxycholesterol, 7 P-25 -dihydroxy cholesterol, beta-sitosterol, stigmasterol, brassicasterol, campesterol, or combinations thereof.
  • the sterol constitutes 40.25% of the total moles of components in the LNP. In some embodiments, the sterol constitutes 40.5% of the total moles of components in the LNP. In some embodiments, the sterol constitutes 40.75% of the total moles of components in the LNP. iii. PEG-Lipids
  • PEG-lipids can reduce LNP aggregation, shield the LNPs from non-specific endocytosis, and reduce opsonization by serum proteins and reticuloendothelial clearance.
  • the LNP comprises at least one PEG or PEG-modified lipid.
  • the LNP comprises at least one PEG-modified lipids.
  • the LNP comprises PEG.
  • the PEG-lipids may be referred to as PEGylated lipids or PEG-lipids.
  • the PEGylation is reversible with the PEG moiety gradually releasing into blood circulation.
  • PEG-lipids include, for example and without limitation, PEG conjugated to saturated or unsaturated alkyl chains having a length of C6-C20, PEG-modified phosphatidylethanolamines, PEG-modified phosphatidic acids, PEG- modified ceramides (PEG-CER), PEG-modified dialkylamines, PEG-modified diacylglycerols (PEG-DAG), PEG-modified dialkylglycerols, and mixtures thereof.
  • Additional examples of PEG-lipids include, without limitation, PEG-c-DOMG, PEG-DMG, PEG-DLPE, PEG-DMPE, PEG-DPPE, PEG-DSG, or PEG-DSPE lipids.
  • the PEGylation comprises PEGlk, PEG2k, PEG5k, or PEGlOk.
  • the LNP comprises PEG-DMG.
  • the PEGylation comprises PEG2k.
  • the LNP comprises DMG-PEG2k.
  • the PEG-lipid constitutes 0 mol% to 5 mol% of the total moles of components in the LNP. In some embodiments, the PEG-lipid constitutes 1 mol% to 5 mol% of the total moles of components in the LNP. In some embodiments, the PEG-lipid constitutes 1 mol% to 3 mol% of the total moles of components in the LNP. In some embodiments, the PEG-lipid constitutes 1 mol% to 2 mol% of the total moles of components in the LNP. In some embodiments, the PEG-lipid constitutes 2 mol% to 3 mol% of the total moles of components in the LNP.
  • the PEG-lipid constitutes 1 mol% of the total moles of components in the LNP. In some embodiments, the PEG-lipid constitutes 1.25 mol% of the total moles of components in the LNP. In some embodiments, the PEG-lipid constitutes 1.5 mol% of the total moles of components in the LNP. In some embodiments, the PEG-lipid constitutes 1.75 mol% of the total moles of components in the LNP. In some embodiments, the PEG-lipid constitutes 2 mol% of the total moles of components in the LNP. In some embodiments, the PEG-lipid constitutes 2.25 mol% of the total moles of components in the LNP.
  • the PEG-lipid constitutes 2.5 mol% of the total moles of components in the LNP. In some embodiments, the PEG-lipid constitutes 2.75 mol% of the total moles of components in the LNP. In some embodiments, the PEG-lipid constitutes 3 mol% of the total moles of components in the LNP.
  • the PEG is replaced by a different polymeric compound such as, but not limited to, polyethenes, poly(l-lysine)(PLL), PEG grafted to PLL, cationic lipopolymer, biodegradable cationic lipopolymer, polyethyleneimine (PEI), cross-linked branched poly (alkylene imines), a poly amine derivative, a modified poloxamer, a biodegradable polymer, elastic biodegradable polymer, biodegradable block copolymer, biodegradable random copolymer, biodegradable polyester copolymer, biodegradable polyester block copolymer, biodegradable polyester block random copolymer, multiblock copolymers, linear biodegradable copolymer, poly[a-(4-aminobutyl)-L-glycolic acid) (PAGA), biodegradable cross-linked cationic multi-block copolymers, polycarbonates, poly anhydrides,
  • PAGA biodegradable
  • Helper lipids in LNPs may contribute to stability and delivery efficiency and /or mitigate any toxicity from the ionizable lipids.
  • the helper lipid is a phospholipid (also known as neutral phospholipid).
  • the phospholipid comprises a phospholipid moiety and at least one fatty acid moiety.
  • the phospholipid comprises one or more (poly)unsaturated lipids. Phospholipids may assemble into one or more lipid bilayers.
  • Exemplary phospholipid moieties include, but are not limited to, phosphatidyl choline, phosphatidyl ethanolamine, phosphatidyl glycerol, phosphatidyl serine, phosphatidic acid, 2-lysophosphatidyl choline, and sphingomyelin.
  • the fatty acid moiety includes, for example and without limitation, lauric acid, myristic acid, myristoleic acid, palmitic acid, palmitoleic acid, stearic acid, oleic acid, linoleic acid, a-linoleic acid, erucic acid, phytanic acid, arachidic acid, arachidonic acid, eicosapentaenoic acid, behenic acid, docosapentaenoic acid, and docasohexaenoic acid.
  • Non-natural species or natural species with modification and substitutions, including branching, oxidation, cyclization, and alkynes, are included.
  • a phospholipid may be modified with an alkyne, which could allow for copper-catalyzed cycloaddition, or click-chemistry, with an azide to functionalize the lipid bilayer.
  • the lipid bilayer is functionalized.
  • the lipid bilayer is not functionalized.
  • the helper lipid is a lipid having cone-shape geometry, e.g., dioleoylphosphatidylethanolamine (DOPE).
  • DOPE dioleoylphosphatidylethanolamine
  • the helper lipid is a cylindrical- shaped lipid such as phosphatidylcholine.
  • Helper lipids include, for example and without limitation, distearoylphosphatidylcholine (DSPC), dioleoylphosphatidylethanolamine (DOPE), 1,2- dilauroyl-sn-glycero-3-phosphocholine (DLPC), l,2-dimyristoyl-sn-glycero-3- phosphocholine (DMPC), l,2-Dioleoyl-sn-glycero-3-phosphocholine (DOPC), dipalmitoylphosphatidylcholine (DPPC), l,2-(cis-cis-9,12-octadecadienoyl)-sn-glycero-3- phosphatidylcholine (DUPC), l-palmitoyl-2-oleoyl-glycero-3-phosphocholine (POPC), 1,2- di-O-octadecyl-sn-glycero-3-phosphocholine (18:0
  • the helper lipid is non-cationic lipid.
  • the helper lipid constitutes 5 mol% to 15 mol% of the total moles of components in the LNP. In some embodiments, the helper lipid constitutes 5 mol% to 10 mol% of the total moles of components in the LNP. In some embodiments, the helper lipid constitutes 10 mol% to 15 mol% of the total moles of components in the LNP. In some embodiments, the helper lipid constitutes 10 mol% of the total moles of components in the LNP. v. Exemplary LNP Formulations
  • the LNP formulation comprises about 40 mol% to about 50 mol% ionizable lipid, about 5 mol% to about 15 mol% helper lipid, about 35 mol% to about 45 mol% sterol, and about 0 mol% to about 5 mol% PEG-lipid. In some embodiments, the LNP formulation comprises about 45 mol% to about 50 mol% ionizable lipid, about 5 mol% to about 15 mol% helper lipid, about 40 mol% to about 45 mol% sterol, and about 1 mol% to about 3 mol% PEG-lipid.
  • the LNP comprises about 47.5 mol% ionizable lipid, about 10 mol% helper lipid, about 40 mol% sterol, and about 2.5 mol% PEG-lipid. In some embodiments, the LNP comprises about 47.5 mol% ionizable lipid, about 10 mol% helper lipid, about 40.75 mol% sterol, and about 1.75 mol% PEG-lipid. In some embodiments, the LNP comprises about 47.5 mol% ionizable lipid, about 10 mol% helper lipid, about 41 mol% sterol, and about 1.5 mol% PEG-lipid. In some embodiments, the LNP comprises about 47.5 mol% ionizable lipid, about 10 mol% helper lipid, about 40.5 mol% sterol, and about 2 mol% PEG-lipid.
  • the LNP comprises Compound 1 as the ionizable lipid, DSPC as the helper lipid, cholesterol as the sterol lipid, and DMG-PEG2k as the PEG-lipid.
  • the LNP comprises Compound 37 as the ionizable lipid, DSPC as the helper lipid, cholesterol as the sterol lipid, and DMG-PEG2k as the PEG-lipid.
  • the LNP comprises Compound 49 as the ionizable lipid, DSPC as the helper lipid, cholesterol as the sterol lipid, and DMG-PEG2k as the PEG-lipid.
  • the LNP comprises Compound 36 as the ionizable lipid, DSPC as the helper lipid, cholesterol as the sterol lipid, and DMG-PEG2k as the PEG-lipid.
  • the LNP comprises Compound 82 as the ionizable lipid, DSPC as the helper lipid, cholesterol as the sterol lipid, and DMG-PEG2k as the PEG-lipid.
  • the LNP comprises Compound 81 as the ionizable lipid, DSPC as the helper lipid, cholesterol as the sterol lipid, and DMG-PEG2k as the PEG-lipid.
  • Exosomes are tiny vesicles smaller than 50 nm secreted by mature reticulocytes, which are associated with transferrin receptors and function in antigen presentation during the regulation of immune cells.
  • engineered exosomes act as cargo carriers and deliver small hydrophilic or lipophilic molecules, including some therapeutic drugs to cells, participating in the regulation of many major diseases. Exosomes can improve bioavailability of some drugs when taken orally, reducing the total dose required for administration, and minimizing side effects.
  • the mRNA molecules discussed herein are delivered using engineered exosomes. ii. Viral Like Particles (VLPs)
  • mRNA molecules discussed herein are delivered using viral delivery particles.
  • Viral particles include recombinant viruses and viral like particles (VLPs).
  • VLPs viral like particles
  • Virus-like particles are molecules that closely resemble viruses, but are non-infectious because they contain no viral genetic material. They can be naturally occurring or synthesized through the individual expression of viral structural proteins, which can then self-assemble into the virus-like structure. Combinations of structural capsid proteins from different viruses can be used to create recombinant VLPs.
  • VLPs can be produced from different viruses, such as adeno-associated viruses, retroviruses, lentiviruses and vesiculoviruses.
  • the VLP is derived from a Vesiculovirus.
  • the VLP is derived from VSV (Indiana vesiculovirus, formerly Vesicular stomatitis Indiana virus (VSIV or VSV).
  • the virus like particle comprises a mutated VSV-G protein.
  • VSV-G protein is a single transmembrane glycoprotein (G) which plays a critical role during the initial steps of virus infection, it is responsible for virus attachment to specific receptor, LDL-R. In the cell, G protein triggers the fusion between the viral and endosomal membranes, which releases the viral genome in the cytosol for the subsequent steps of infection.
  • VSV-G protein is mutated to abolish its binding to LDL-R receptor.
  • a VSV-G envelope protein may be a mutated at one or more of any one of H8, K47, Y209, and/or R354.
  • a VLP may comprise a mutated VSV-G protein described in the PCT patent application Publication No. WO2019057974; the contents of which are incorporated herein by reference in their entireties.
  • the VLP for delivery of mRNA molecules is a viral particle disclosed in the PCT Publication NOs. WO2020236263 and WO2023107886; the contents of each of which are incorporated herein by reference in their entireties.
  • the virus like particle is pseudotyped.
  • the virus like particle is VSV-G-pseudotyped lentiviruses (VSV-G-LVs).
  • the viral particle for delivering mRNA molecules is a retrovirus, a recombinant AAV, or an adenovirus.
  • compositions comprising one or more mRNA molecules described herein.
  • Pharmaceutical compositions may include mRNA molecules as described herein in combination with one or more pharmaceutically or physiologically acceptable carrier, diluents, or excipients.
  • Such compositions may include buffers such as neutral buffered saline or phosphate buffered saline; carbohydrates such as glucose, mannose, sucrose or dextrans, mannitol; proteins; polypeptides or amino acids such as glycine; antioxidants; chelating agents such as EDTA or glutathione; adjuvants (e.g., aluminum hydroxide); and preservatives.
  • Cryopreservation solutions which may be used in the pharmaceutical compositions of the disclosure include, for example, DMSO.
  • Compositions can be formulated for any suitable administration, e.g., for intravenous administration.
  • compositions described herein may be prepared by any method known or hereafter developed in the art of pharmacology.
  • preparatory methods include the step of bringing the active ingredient (e.g., mRNA molecules) into association with an excipient and/or one or more other accessory ingredients, and then, if necessary and/or desirable, shaping and/or packaging the product into a desired single- or multi-dose unit.
  • a pharmaceutical composition in accordance with the present disclosure may be prepared, packaged, and/or sold in bulk, as a single unit dose, and/or as a plurality of single unit doses.
  • a “unit dose” is discrete amount of the pharmaceutical composition comprising a predetermined amount of the active ingredient.
  • the amount of the active ingredient is generally equal to the dosage of the active ingredient which would be administered to a subject and/or a convenient fraction of such a dosage such as, for example, one-half or one-third of such a dosage.
  • compositions in accordance with the present disclosure will vary, depending upon the identity, size, and/or condition of the subject treated and further depending upon the route by which the composition is to be administered.
  • the composition may comprise between 0.1% and 100% (w/w) mRNA molecules described herein.
  • a pharmaceutical composition of the present disclosure may be prepared, packaged, and/or sold in a formulation suitable for administration by one or more of a variety of routes, including oral, intravenous, intramuscular, intra-arterial, intramedullary, intrathecal, subcutaneous, intraventricular, transdermal, interdermal, rectal, intravaginal, intraperitoneal, topical (e.g., by powders, ointments, creams, gels, lotions, and/or drops), mucosal, nasal, buccal, enteral, vitreal, intratumoral, sublingual; by intratracheal instillation, bronchial instillation, and/or inhalation; as an oral spray, nasal spray, and/or aerosol, and/or through a portal vein catheter.
  • routes including oral, intravenous, intramuscular, intra-arterial, intramedullary, intrathecal, subcutaneous, intraventricular, transdermal, interdermal, rectal, intravaginal, intraperitoneal, topical (e
  • RNAs, and/or pharmaceutical compositions thereof are administered by systemic intravenous injection.
  • mRNA molecules and/or pharmaceutical compositions thereof may be administered intravenously and/or orally.
  • mRNA molecules and/or pharmaceutical compositions thereof may be administered intramuscularly.
  • compositions may optionally comprise one or more additional therapeutically active substances.
  • pharmaceutical compositions of the present invention may optionally comprise one or more additional prophylactic compounds.
  • the therapeutically active substance is an adjuvant.
  • the prophylactic compound is an adjuvant.
  • the pharmaceutical composition does not include an adjuvant.
  • the adjuvant may be used to enhance antibody response and can comprise any acceptable immuno stimulatory compound.
  • adjuvants include cytokines, toxins, or synthetic compositions.
  • Adjuvants may be formulated as oil-in-water emulsions, water-in- oil emulsions, mineral salts, polynucleotides, or natural substances.
  • the adjuvant may be encoded by a second mRNA molecule.
  • the pharmaceutical composition comprises an mRNA molecule of the present invention encoding the VZV gE antigen and a second mRNA molecule encoding an adjuvant.
  • compositions are administered to humans.
  • pharmaceutical compositions are principally directed to pharmaceutical compositions which are suitable for administration to humans, it will be understood by the skilled artisan that such compositions are generally suitable for administration to animals of all sorts.
  • the mRNAs encoding one or more VZV antigen polypeptides, compositions and vaccines comprising mRNA encoding one or more VZV antigen polypeptides can be used to induce an VZV-specific immune response, including any humoral and/or cellular immune response.
  • the patients are adults aged 18 years and older who are or will be at increased risk of HZ due to immunodeficiency or immunosuppression caused by known disease or therapy.
  • Immunocompromised patients may include patients having but not limited to Autologous Hematopoeitic Stem Cell Transplant (auHSCT) recipients, hematologic malignancies, renal transplant, solid malignant tumors, and HIV.
  • auHSCT Autologous Hematopoeitic Stem Cell Transplant
  • the mRNAs encoding one or more VZV antigen polypeptides, compositions and vaccines comprising mRNA encoding one or more VZV antigen polypeptides are for prevention of chickenpox.
  • the vaccine may be administered to the patient by any routes of administration.
  • the vaccine may be administered to the patient by intramuscular, intradermal, or subcutaneous administration.
  • the vaccine is administered by intradermal administration.
  • the vaccine is administered by intramuscular administration.
  • the vaccine is administered by subcutaneous administration. [0302] In some embodiments, the vaccine is administered intramuscularly.
  • the vaccine is administered in a single dose.
  • the vaccine is administered in multiple doses. In some embodiments, the vaccine is administered in two separate doses. As non-limiting examples, the vaccine is administered according to the following schedules: a first dose at Month 0 followed by a second dose administered 2 to 6 months later. In some embodiments, for patients who are or will be immunodeficient or immunosuppressed and who would benefit from a shorter vaccination schedule: A first dose at Month 0 followed by a second dose administered 1 to 2 months later. [0306] Accordingly, the compositions and vaccines significantly reduce the risk of developing HZ (shingles) by at least 50% in subjects aged 50 years and older. In some embodiments, the compositions and vaccines significantly reduce the risk of developing HZ (shingles) by at least 60% in subjects aged 50 years and older.
  • the compositions and vaccines significantly reduce the risk of developing HZ (shingles) by at least 70% in subjects aged 50 years and older. In some embodiments, the compositions and vaccines significantly reduce the risk of developing HZ (shingles) by at least 80% in subjects aged 50 years and older. In some embodiments, the compositions and vaccines significantly reduce the risk of developing HZ (shingles) by at least 90% in subjects aged 50 years and older. In some embodiments, the compositions and vaccines significantly reduce the risk of developing HZ (shingles) by at least 95% in subjects aged 50 years and older.
  • compositions and vaccines significantly reduce the risk of developing HZ in patients aged 18 years and older who are or will be immunodeficient or immunosuppressed.
  • the vaccine efficacy against HZ maintains the same the first year, the second year, the third year, the fourth year and later after vaccination.
  • compositions and vaccines may be administered concomitantly with another vaccine.
  • mRNA VZV vaccine constructs and “mRNA VZV gE constructs” are used interchangeably throughout the application and represent the mRNA described herein that encode a VZV gE polypeptide.
  • mRNA VZV vaccine refers to a composition comprising an mRNA VZV vaccine construct.
  • Glycoprotein E is one of the viral binding proteins for varicella zoster virus (VZV), making gE a strong target for the neutralizing antibodies induced by vaccines.
  • the gE protein comprises a signal domain, an extracellular domain, a transmembrane domain, and an intracellular domain.
  • the intracellular domain is responsible for receptor trafficking, including targeting to the trans-Golgi network, endocytosis, and phosphorylation while the extracellular domain is the domain responsible for viral attachment. Therefore, when optimizing the design of the antigen, the gE protein was truncated and/or mutated at various points to retain the extracellular domain and eliminate the regulatory motifs from the intracellular domain. The truncation points are visualized in FIG. 1.
  • a mutation (Y569A) was included. Although not bound by any theory, the Y569A mutation was designed to disrupt trans-Golgi network targeting.
  • Expression of exemplary mRNA VZV gE constructs was assessed in K562 lymphoblast cells electroporated with the respective mRNA constructs encoding VZV gE. Expression was determined by measuring VZV fluorescence intensity by flow cytometry. Fold-over-control (FOC) values were calculated for each construct by dividing the geometric mean of the sample by the geometric mean of the negative controls. As shown in FIG. 2, expression of exemplary constructs (gE A562, gE A562 IgGK, gE A568, gE A568 IgGK, and gE A574 Y569A described in Table A) outperformed a reference construct across the tested time points.
  • VZV gE percent of cells expressing VZV gE was higher for the exemplary constructs than the reference construct.
  • mRNA VZV gE constructs with high expression were selected for further testing in a second trial with similar methodology. Results from the second trial can be found in FIG. 3.
  • exemplary mRNA VZV gE constructs, including codon-optimized constructs, of the present invention displayed higher gE expression than a reference construct.
  • Blood samples were collected on Day 0, Day 28, Day 49, Day 53, Day 67, Day 81, Day 95, and Day 123/124 timepoints and analyzed for binding and neutralizing antibodies by ELISA and PRNT. Spleen samples were collected at the end of the study. At the terminal timepoint (Day 123/124), cytokine production following ex vivo stimulation of splenocytes with VZV gE peptide were detected by FACS analysis. Spleen samples were assessed by T- cell flow panels for VZV- specific T cell responses.
  • FIG 41A-FIG. 41D shows the total IgG anti- VZV gE in serum from tested timepoints. From the Day 67 timepoint until the end of the study, higher responses were observed after a second dose administration in Groups 2, 6, 8, and 10 compared to their respective single dose counterparts in Groups 1, 5, 7, and 9.
  • the treated samples were diluted in dT buffer and run by Oligo-dT Liquid Chromatography (LC) (Oligo(dT) affinity chromatography).
  • LC Oligo-dT Liquid Chromatography
  • the oligo(dT) specifically hybridizes to the polyA tails of mRNA.
  • the contaminants were removed during the washing steps.
  • the immobilized mRNAs were released from the column using an elution buffer.
  • the collected eluates were prepared again using RP buffer and run by RP- HPLC-FT (reversed-phase high performance liquid chromatography) for further purification.
  • the collected samples were diluted in PBS. Tangential flow filtration (TFF) was applied to further purify the mRNA samples.
  • IVT samples after DNase treatment were directly purified by Oligo-dT Liquid Chromatography (LC) (Oligo(dT) affinity chromatography), followed by molecular weight-cutoff spin filtration.
  • Exemplary mRNA VZV constructs were prepared with different 5’ caps and analyzed for the effect of 5’ capping on immunogenicity.
  • the caps were added using CleanCap® methodology, a one-pot co-transcriptional capping method. Any method or reagent known in the art may be equivalently used for generating the capped mRNA.
  • the preparation process was further optimized by reducing the concentration of CleanCap® reagent without significantly impairing capping efficiency.
  • the study design, with different exemplary 5’ caps and buffer concentrations are shown in Table 5. The study was then repeated with an mRNA dose of 1 pg.
  • This Example describes the optimization of exemplary LNP formulations for mRNA VZV vaccines. Additional LNP formulations can be found in WO 2021/141969 Al (Hamilton et al.) and WO 2022140252 Al (Patwardhan et al.), the disclosure of each of which is hereby incorporated by reference in its entirety.
  • the stability of the LNPs in each buffer was assessed by characterizing LNP size, polydispersity index (PDI), and encapsulation efficiency (EE%) after freeze thaw (FT) cycles and long-term storage.
  • stability was compared after 0, 1, 3, or 5 FT cycles.
  • increasing buffer concentration to 100 mM generally resulted in an increase in ENP size after FT cycles.
  • the 20 mM HEPES, MOPS, and TES buffers maintained the EE% better than their 100 mM counterparts.
  • the percent change in LNP size, PDI, and EE% was calculated for the exemplary buffer compositions, comparing the stability at a constant 4°C to that after one FT cycle at -80°C.
  • the 100 mM HEPES Saline buffer had the highest increase in size after the FT cycle. Sucrose concentrations did not appear to affect stability.
  • exemplary LNP formulations in various buffers were tested in vivo in C57BI/6J mice. Acidification buffers were modified and screened to optimize LNP characteristics and stability while ensuring the immune response was not impacted. Saline was also included in some exemplary buffers to improve potency for local administration.
  • ELISA assays were performed on the samples from Day 21 and Day 35. The resulting total IgG titers are displayed in FIG. 35A-FIG. 35B. The IgGl, IgG2b, and IgG2c titers from Day 35 are shown in FIG. 36A-FIG. 36B.
  • Exemplary LNP formulation LNP029, with DOPE as the helper lipid resulted in high IgG titers on Day 35.
  • exemplary LNP formulations with saline in the storage buffer surprisingly showed increased IgG titers on Day 35. Increasing the pH of the acidification buffer caused the LNP particle size to decrease, however, the smaller particles still maintained potency.
  • the IgG2c/IgGl ratios for each group are shown in FIG. 37.
  • the IgG2c/IgGl ratio is representative of the type of immune response induced by the exemplary mRNA VZV vaccine. A higher ratio indicates a Thl type cellular immune response, whereas a lower ratio is indicative of a Th2 type humoral immune response. The balance of the Thl and Th2 immune responses may improve the protective effect of the vaccine.
  • Exemplary mRNA VZV vaccine formulations with LNP007 generally have a higher Thl type cellular immune response on Day 35.

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Virology (AREA)
  • Medicinal Chemistry (AREA)
  • Veterinary Medicine (AREA)
  • Public Health (AREA)
  • General Health & Medical Sciences (AREA)
  • Animal Behavior & Ethology (AREA)
  • Chemical & Material Sciences (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Communicable Diseases (AREA)
  • Biotechnology (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Organic Chemistry (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Oncology (AREA)
  • Molecular Biology (AREA)
  • General Chemical & Material Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Immunology (AREA)
  • Microbiology (AREA)
  • Mycology (AREA)
  • Epidemiology (AREA)
  • Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)

Abstract

La présente divulgation concerne des séquences d'ARNm codant pour un antigène du virus varicelle-zona. Les séquences d'ARNm décrites ici sont utilisées en tant que vaccins et compositions pour prévenir ou traiter des maladies résultant d'un virus varicelle-zona, y compris le zona. L'invention concerne également des procédés de formulation et de préparation des vaccins.
PCT/US2025/022899 2024-04-03 2025-04-03 Compositions d'arnm et leurs utilisations dans des vaccins contre le virus varicelle-zona Pending WO2025212851A2 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US202463574160P 2024-04-03 2024-04-03
US63/574,160 2024-04-03

Publications (2)

Publication Number Publication Date
WO2025212851A2 true WO2025212851A2 (fr) 2025-10-09
WO2025212851A3 WO2025212851A3 (fr) 2025-11-13

Family

ID=95519096

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US2025/022899 Pending WO2025212851A2 (fr) 2024-04-03 2025-04-03 Compositions d'arnm et leurs utilisations dans des vaccins contre le virus varicelle-zona

Country Status (1)

Country Link
WO (1) WO2025212851A2 (fr)

Citations (11)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US7074596B2 (en) 2002-03-25 2006-07-11 Board Of Supervisors Of Louisiana State University And Agricultural And Mechanical College Synthesis and use of anti-reverse mRNA cap analogues
WO2008016473A2 (fr) 2006-07-28 2008-02-07 Applera Corporation ANALOGUES DE COIFFES DE NUCLÉOTIDE D'ARNm
WO2008157688A2 (fr) 2007-06-19 2008-12-24 Board Of Supervisors Of Louisiana State University And Agricultural And Mechanical College Synthèse et utilisation d'analogues de phosphorothioate anti-inverse de la coiffe d'arn messager
WO2009149253A2 (fr) 2008-06-06 2009-12-10 Uniwersytet Warszawski Analogues d'arnm cap
WO2011015347A1 (fr) 2009-08-05 2011-02-10 Biontech Ag Composition vaccinale contenant de l'arn dont la coiffe en 5' est modifiée
WO2013059475A1 (fr) 2011-10-18 2013-04-25 Life Technologies Corporation Analogues de coiffes à dérivation alcynyle, préparation et utilisations associées
WO2019057974A1 (fr) 2017-09-22 2019-03-28 Centre National De La Recherche Scientifique (Cnrs) Glycoprotéine mutée du virus de la stomatite vésiculaire
WO2020236263A1 (fr) 2019-05-23 2020-11-26 Massachusetts Institute Of Technology Découverte de ligands et apport de gènes par l'intermédiaire d'une présentation à la surface de rétrovirus
WO2021141969A1 (fr) 2020-01-09 2021-07-15 Guide Therapeutics, Inc. Nanomatériaux
WO2022140252A1 (fr) 2020-12-21 2022-06-30 Beam Therapeutics Inc. Nanomatériaux comprenant des acétals à liaison ester
WO2023107886A1 (fr) 2021-12-06 2023-06-15 The Board Of Trustees Of The Leland Stanford Junior University Méthodes et compositions pour la découverte d'une spécificité de ligand de récepteur par une entrée de cellule modifiée

Family Cites Families (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
TW202327646A (zh) * 2021-10-15 2023-07-16 美商輝瑞大藥廠 Rna分子
CN118497228A (zh) * 2023-02-15 2024-08-16 上海臻上医药科技有限公司 一种水痘-带状疱疹病毒mRNA疫苗及其应用

Patent Citations (11)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US7074596B2 (en) 2002-03-25 2006-07-11 Board Of Supervisors Of Louisiana State University And Agricultural And Mechanical College Synthesis and use of anti-reverse mRNA cap analogues
WO2008016473A2 (fr) 2006-07-28 2008-02-07 Applera Corporation ANALOGUES DE COIFFES DE NUCLÉOTIDE D'ARNm
WO2008157688A2 (fr) 2007-06-19 2008-12-24 Board Of Supervisors Of Louisiana State University And Agricultural And Mechanical College Synthèse et utilisation d'analogues de phosphorothioate anti-inverse de la coiffe d'arn messager
WO2009149253A2 (fr) 2008-06-06 2009-12-10 Uniwersytet Warszawski Analogues d'arnm cap
WO2011015347A1 (fr) 2009-08-05 2011-02-10 Biontech Ag Composition vaccinale contenant de l'arn dont la coiffe en 5' est modifiée
WO2013059475A1 (fr) 2011-10-18 2013-04-25 Life Technologies Corporation Analogues de coiffes à dérivation alcynyle, préparation et utilisations associées
WO2019057974A1 (fr) 2017-09-22 2019-03-28 Centre National De La Recherche Scientifique (Cnrs) Glycoprotéine mutée du virus de la stomatite vésiculaire
WO2020236263A1 (fr) 2019-05-23 2020-11-26 Massachusetts Institute Of Technology Découverte de ligands et apport de gènes par l'intermédiaire d'une présentation à la surface de rétrovirus
WO2021141969A1 (fr) 2020-01-09 2021-07-15 Guide Therapeutics, Inc. Nanomatériaux
WO2022140252A1 (fr) 2020-12-21 2022-06-30 Beam Therapeutics Inc. Nanomatériaux comprenant des acétals à liaison ester
WO2023107886A1 (fr) 2021-12-06 2023-06-15 The Board Of Trustees Of The Leland Stanford Junior University Méthodes et compositions pour la découverte d'une spécificité de ligand de récepteur par une entrée de cellule modifiée

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
STEPINSKI ET AL., RNA, vol. 7, no. 10, 2001, pages 1486 - 495

Also Published As

Publication number Publication date
WO2025212851A3 (fr) 2025-11-13

Similar Documents

Publication Publication Date Title
EP3917503B1 (fr) Procédés de préparation de nanoparticules lipidiques
US11510977B2 (en) Nucleic acid vaccines for coronavirus
EP3852728B1 (fr) Préparation de nanoparticules lipidiques et leurs méthodes d'administration
US20230053437A1 (en) Lipid compounds and lipid nanoparticle compositions
US20240218399A1 (en) Lipid compounds and lipid nanoparticle compositions
AU2021301922A1 (en) Lipid compounds and lipid nanoparticle compositions
CN101827613A (zh) 在包括连续疏水相的载体中的脂质体在体内输送多核苷酸中的应用
WO2022152141A2 (fr) Composés lipidiques conjugués polymères et compositions de nanoparticules lipidiques
WO2024037578A1 (fr) Composition de nanoparticules lipidiques
KR102948789B1 (ko) 항원 제시 세포를 표적으로 하는 지질 조성물 및 이의 용도
DE112024001143T5 (de) Neue lipid-nanopartikel-formeln für die abgabe von nukleinsäuren
WO2023151570A1 (fr) Vaccins à base d'acides nucléiques contre la rage
WO2023056912A1 (fr) Vaccins à base d'acide nucléique pour vzv
WO2024240125A1 (fr) Composés lipidiques et compositions de nanoparticules lipidiques
WO2025212851A2 (fr) Compositions d'arnm et leurs utilisations dans des vaccins contre le virus varicelle-zona
US20250235403A1 (en) Compositions and methods of the delivery of active agents including nucleic acids
CN120158462A (zh) 编码水痘-带状疱疹病毒糖蛋白E的mRNA疫苗
WO2026006687A1 (fr) Compositions d'arn circulaire et leurs utilisations pour des maladies provoquées par le virus varicelle-zona
WO2025153053A1 (fr) Composition et procédés pour induire une immunité spécifique à un antigène contre des cellules mutantes kras
WO2024037577A1 (fr) Composition de nanoparticules lipidiques
WO2024140624A1 (fr) Composés lipidiques et compositions de nanoparticules lipidiques
HK40041286A (en) Lipid nanoparticles
HK1233171A1 (en) Use of liposomes in a carrier comprising a continuous hydrophobic phase for delivery of polynucleotides in vivo