Immunogenicity of CPG-adjuvanted varicella zoster vaccine

Abstract

The present disclosure relates to a method for increasing cell-mediated immunity against Varicella-Zoster Virus (VZV) in a human subject in need thereof by administration of an immunogenic composition comprising an effective amount of a VZV glycoprotein E antigen and an oligonucleotide comprising an unmethylated cytidine-phospho-guanosine (CpG) motif. The immunogenic composition is suitable for the prevention of varicella zoster and / or post-herpetic neuralgia.

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims priority to and the benefit of U.S. Provisional Application No. 63 / 293,510, filed December 23, 2021, the disclosure of which is incorporated herein by reference in its entirety.

[0002] Reference to Electronic Sequence Listing The contents of the electronic sequence listing (377882008440SEQLIST.xml; size: 10,729 bytes; and creation date: December 20, 2022) are incorporated herein by reference in their entirety.

[0003] The present disclosure relates to a method for increasing cell-mediated immunity against Varicella-Zoster Virus (VZV) in a human subject in need thereof by administration of an immunogenic composition comprising an effective amount of a VZV glycoprotein E antigen and an oligonucleotide comprising an unmethylated cytidine-phospho-guanosine (CpG) motif. The immunogenic composition is suitable for the prevention of varicella zoster and / or post-herpetic neuralgia. [Background technology]

[0004] Herpes zoster (HZ), also known as shingles, is caused by reactivation of latent varicella zoster virus (VZV) and typically manifests as a localized dermatomal rash (Eshleman et al., Future Virology, 6(3):341-355, 2011; and Cohen, N Engl J Med, 369(3):255-263, 2013). Immunosenescence (age-dependent decrease in immunological responsiveness) and immunodeficiency (caused by disease or medication) are the most important risk factors for the development of HZ. A person's risk for shingles, and complications such as postherpetic neuralgia and hospitalization, increases sharply after age 50, closely linked to an age-related decline in VZV-specific T cell-mediated immunity (CMI), which may be important in preventing reactivation of latent VZV and preventing the proliferation of reactivated virus (Levin et al., J Infect Dis, 197(6):825-835, 2008; Weinberg et al., J Infect Dis, 201(7):1024-1030, 2010; and Weinberg and Levin, Curr Top Microbiol Immunol, 342:341-357, 2010). HZ vaccines are thought to boost VZV-specific memory T cells, preventing their decline below a currently unknown threshold required for protection against HZ (Oxman et al., Vaccine 29(20):3625-3627, 2011).

[0005] Prior to the availability of the HZ vaccine, nearly 1 million people in the United States suffered from HZ annually. In 2020, the incidence of HZ in unvaccinated adults was 9.92 per 1000 people annually (95% CI, 9.82-10.01). This generally increased with age, from 7.20 / 1000 in the 50-54 age group to 13.99 / 1000 in the ≥80 age group. Half of all HZ cases occur in people aged 60 years or older, and it is estimated that individuals living to age 85 years have a 50% chance of having HZ during their lifetime (Dooling et al., MMWR, 67:103-108, 2018).

[0006] In the United States, two vaccines have been available for the prevention of shingles in individuals aged 50 years or older. ZOSTAVAX® is a live attenuated virus vaccine marketed by Merck & Co., Inc. (Whitehouse Station, NJ). SHINGRIX® is a recombinant AS01B-adjuvanted subunit vaccine marketed by GlaxoSmithKline (Research Triangle Park, NC). Although no head-to-head clinical trials have been conducted, the Advisory Committee on Immunization Practices in 2017 reluctantly recommended preferential use of SHINGRIX® over ZOSTAVAX® based on evaluation of improved efficacy against shingles and postherpetic neuralgia (Dooling et al., MMWR, 67:103-108, 2018). The narrow vote in favor of SHINGRIX® (8-7) was related to concerns about reactogenicity. Phase III studies of SHINGRIX® reported that 16.5% of vaccine recipients experienced grade 3 adverse events compared to 3.1% of placebo recipients, and 10.8% of vaccine recipients experienced grade 3 systemic events (myalgia, fatigue, headache, shivering, fever, and gastrointestinal symptoms) compared to 2.4% of placebo recipients (Lal et al., N Eng J Med 372:2087-2096, 2015; and Cunningham et al., N Eng J Med 375:1019-1032, 2016). It was recognized that the relatively high level of serious adverse reactions following SHINGRIX® vaccination could negatively impact patient compliance levels for the second dose (Bharucha et al., Human Vaccines & Immunotherapeutics, 13:1789-1797, 2017), a persistent concern given the need for patients to receive both doses to be protected against HZ. ZOSTAVAX® was withdrawn from the U.S. market in 2020 as a result of lawsuits alleging that it caused shingles and severe shingles-related illness in some patients. Summary of the Invention [Problem to be solved by the invention]

[0007] Therefore, there is an unmet medical need for an HZ vaccine with a good safety profile. [Means for solving the problem]

[0008] The present disclosure relates to a method for increasing cell-mediated immunity against Varicella-Zoster Virus (VZV) in a human subject in need thereof by administration of an immunogenic composition comprising an effective amount of a VZV glycoprotein E antigen and an oligonucleotide comprising an unmethylated cytidine-phospho-guanosine (CpG) motif. The immunogenic composition is suitable for the prevention of varicella zoster and / or post-herpetic neuralgia. [Brief description of the drawings]

[0009] [Figure 1] Figure 1 shows the frequency of VZV gE-reactive CD4+ T cells expressing two, three and four markers of activation following incubation of PBMCs in the presence of a series of overlapping peptides spanning VZV gE with unstimulated (background) responses subtracted. PBMCs were isolated from blood samples obtained from study subjects at week 0 (baseline). [Diagram 2] Figure 2 shows the frequency of VZV gE-reactive CD4+ T cells expressing two, three and four markers of activation following incubation of PBMCs in the presence of a series of overlapping peptides spanning VZV gE, with unstimulated (background) responses subtracted. PBMCs were isolated from blood samples obtained from study subjects at week 8 (approximately 4 weeks after the first injection). [Diagram 3]Figure 3 shows the frequency of VZV gE-reactive CD4+ T cells expressing two, three and four markers of activation following incubation of PBMCs in the presence of a series of overlapping peptides spanning VZV gE with unstimulated (background) responses subtracted. PBMCs were isolated from blood samples obtained from study subjects at week 12 (approximately 4 weeks after the second injection). DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0010] The present disclosure relates to a method for increasing cell-mediated immunity against Varicella Zoster Virus (VZV) in a human subject in need thereof by administration of an immunogenic composition comprising an effective amount of a VZV glycoprotein E antigen and an oligonucleotide comprising an unmethylated cytidine-phospho-guanosine (CpG) motif. The immunogenic composition is suitable for the prevention of Herpes Zoster (HZ) and / or Postherpetic Neuralgia (PHN).

[0011] The reactogenicity of protein subunit vaccines can be influenced by a variety of factors, the most important of which is the nature of the adjuvant included to improve immunogenicity. Different adjuvant systems (AS) adjuvants combined with a model protein antigen were compared by immunization of healthy human subjects. The AS01B adjuvant was found to induce higher levels of local and systemic reactogenicity than the control adjuvants (AS01E, AS03A, AS04 and alum) (Leroux-Roels et al., Clin Immunol, 169:16-27, 2016). This suggests that AS01B is a major contributor to the reactogenicity profile of SHINGRIX® (a recombinant AS01B-adjuvanted subunit zoster vaccine marketed by GlaxoSmithKline, Research Triangle Park, NC).

[0012] Z-1018 is an exemplary non-infectious vaccine that has been developed for active booster immunization for the prevention of Herpes Zoster (HZ) (shingles) as two intramuscular (IM) doses administered two months apart in individuals aged 50-69 years as described in Example 1. Z-1018 is composed of glycoprotein E (gE) antigen and is adjuvanted by mixing with CpG 1018® adjuvant, with and without aluminum hydroxide (alum). CpG 1018® adjuvant (Dynavax Technologies Corporation) is a synthetic 22-base phosphorothioate oligodeoxynucleotide (PS ODN) containing cytidine-phospho-guanosine (CpG) immunostimulatory sequences that are agonists for Toll-like receptor 9 (TLR9).

[0013] General Techniques and Definitions The practice of the present disclosure will employ, unless otherwise indicated, conventional techniques of molecular biology (including recombinant techniques), microbiology, cell biology, biochemistry and immunology, which are within the skill of the art.

[0014] As used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless otherwise indicated. For example, "an" excipient includes one or more excipients.

[0015] As used herein, the phrase "comprising" is open ended and indicates that such embodiments may include additional elements. In contrast, the phrase "consisting of" is closed and indicates that such embodiments do not include additional elements (except for trace impurities). The phrase "consisting essentially of" is partially closed and indicates that such embodiments may include further elements that do not significantly alter the basic characteristics of such embodiments.

[0016] The term "about" as used herein in reference to a value includes 90% to 110% of that value (e.g., about 3000 μg of CpG 1018® Adjuvant refers to 2700 μg to 3300 μg of CpG 1018® Adjuvant).

[0017] As used interchangeably herein, the terms "polynucleotide" and "oligonucleotide" include single-stranded DNA (ssDNA), double-stranded DNA (dsDNA), single-stranded RNA (ssRNA) and double-stranded RNA (dsRNA), modified oligonucleotides and oligonucleosides, or combinations thereof. Oligonucleotides can be linear or cyclic in configuration, or they can contain both linear and cyclic segments. Oligonucleotides are generally polymers of nucleosides connected through phosphodiester linkages, although alternative linkages, such as phosphorothioate esters, may also be used in oligonucleotides. Nucleosides consist of a purine (adenine (A) or guanine (G), or derivatives thereof) or pyrimidine (thymine (T), cytosine (C), or uracil (U), or derivatives thereof) base attached to a sugar. The four nucleoside units (or bases) in DNA are called deoxyadenosine, deoxyguanosine, thymidine, and deoxycytidine. Nucleotides are phosphate esters of nucleosides.

[0018] The terms "CpG", "CpG motif" and "cytosine-phosphate-guanosine" as used herein refer to unmethylated cytidine-phospho-guanosine dinucleotides, which, when present in an oligonucleotide, contribute to a measurable immune response in vitro, in vivo and / or ex vivo. Examples of measurable immune responses include, but are not limited to, antigen-specific antibody production, secretion of cytokines, activation or expansion of lymphocyte populations, such as NK cells, CD4+ T lymphocytes, CD8+ T lymphocytes, B lymphocytes, and the like. Preferably, CpG oligonucleotides preferentially activate Th1-type responses.

[0019] An "effective amount" or "sufficient amount" of a substance is an amount sufficient to bring about a beneficial or desired result, including a clinical result, and, for example, an "effective amount" will depend on the context in which it is applied. In the context of administering an immunogenic composition, an effective amount will contain sufficient antigen and TLR9 agonist to stimulate an immune response (preferably serum protective levels of antibody to the antigen).

[0020] In this disclosure, the terms "individual" and "subject" refer to humans.

[0021] The term "dose" as used herein in reference to an immunogenic composition refers to the measured portion of an immunogenic composition that is taken by a subject (administered to a subject or received by a subject) at any one time.

[0022] As used herein, the terms "isolated" and "purified" refer to material that is removed from at least one component with which it is naturally associated (e.g., removed from its original environment). The term "isolated," when used in reference to a recombinant protein, refers to a protein that has been removed from the culture medium of a host cell that produced the protein.

[0023] "Stimulation" of a response or parameter includes eliciting and / or enhancing the response or parameter when compared to otherwise the same conditions except for the parameter of interest, or alternatively compared to another condition (e.g., increased TLR signaling in the presence of a TLR agonist compared to the absence of a TLR agonist). For example, "stimulation" of an immune response means an increase in the response. Depending on the parameter being measured, the increase can be 5-500-fold or more, or from 5-, 10-, 50-, or 100-fold to 500-, 1,000-, 5,000-, or 10,000-fold.

[0024] As used herein, the term "immunization" refers to the process of increasing a mammalian subject's response to an antigen, thus improving its ability to resist or overcome infection and / or resist disease.

[0025] As used herein, the term "vaccination" refers to the introduction of a vaccine into the body of a mammalian subject.

[0026] "Adjuvant" refers to a substance that, when added to a composition containing an antigen, enhances or potentiates the immune response to the antigen in a mammalian recipient upon exposure.

[0027] I. Immunogenic Compositions and Kits The present disclosure relates to an immunogenic composition for stimulating an immune response to Varicella-Zoster Virus (VZV), comprising a VZV glycoprotein E (gE) antigen and a Toll-like receptor 9 (TLR9) agonist, wherein the TLR9 agonist is an oligonucleotide of 8-35 nucleotides in length that contains an unmethylated cytidine-phospho-guanosine (also referred to as CpG or cytosine-phosphate-guanosine) motif, and wherein the gE antigen and the oligonucleotide are present in the immunogenic composition in an amount effective to stimulate an immune response to the gE antigen in a human subject. In some embodiments, the immunogenic composition further comprises an aluminum salt adjuvant to which the VZV gE antigen is adsorbed. In some preferred embodiments, the present disclosure relates to an immunogenic composition for increasing cell-mediated immunity to VZV.

[0028] In particular, the present disclosure relates to an immunogenic composition comprising: i) an unmethylated cytidine-phospho-guanosine (CpG)-containing oligonucleotide comprising the sequence 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO:1); ii) a truncated VZV glycoprotein E (gE) antigen; and iii) at least one excipient, wherein the gE antigen is a recombinant protein produced in a mammalian cell and is present in the immunogenic composition in an amount of about 25 μg to about 150 μg, the oligonucleotide is present in the immunogenic composition in an amount of about 750 μg to about 6000 μg, and the at least one excipient comprises a pharma- ceutically acceptable buffer.

[0029] In an exemplary embodiment, the immunogenic composition comprises i) an unmethylated cytidine-phospho-guanosine (CpG)-containing oligonucleotide comprising the sequence 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO:1); ii) a truncated VZV glycoprotein E (gE) antigen; and iii) at least one excipient, wherein the gE antigen is a recombinant protein produced in a mammalian cell and is present in the immunogenic composition in an amount of about 100 μg, the oligonucleotide is present in the immunogenic composition in an amount of about 3000 μg or about 6000 μg, and the at least one excipient comprises a pharma- ceutically acceptable buffer.

[0030] A. Oligonucleotide Toll-like Receptor 9 (TLR9) Agonists Toll-like receptors (TLRs) are expressed in and on dendritic cells and other innate immune cells and are one of the most important receptors for stimulating responses to the presence of invading pathogens. Humans have multiple types of TLRs that are similar in structure but recognize different parts of viruses or bacteria. By activating specific TLRs, it is possible to stimulate and control specific types of innate immune responses that can be utilized to enhance adaptive responses.

[0031] TLR9 (CD289) recognizes unmethylated cytidine-phospho-guanosine (CpG) motifs found in microbial DNA, which can be mimicked using synthetic CpG-containing oligodeoxynucleotides (CpG-ODNs). CpG-ODNs are known to enhance antibody production and stimulate T helper 1 (Th1) cell responses (Coffman et al., Immunity, 33:492-503, 2010). Based on structure and biological function, CpG-ODNs have been divided into three general classes: CpG-A, CpG-B, and CpG-C (Campbell, Methods Mol Biol, 1494:15-27, 2017). The degree of B cell activation varies between the classes of weak CpG-A ODNs, good CpG-C ODNs, and strong CpG-B ODNs. Oligonucleotide TLR9 agonists of the present disclosure are preferably good B cell activators (CpG-C ODN), or more preferably strong B cell activators (CpG-B ODN).

[0032] Optimal oligonucleotide TLR9 agonists often contain palindromic sequences following the general formula: 5'-purine-purine-CG-pyrimidine-pyrimidine-3', or 5'-purine-purine-CG-pyrimidine-pyrimidine-CG-3' (U.S. Patent No. 6,589,940). TLR9 agonism is also observed with certain non-palindromic CpG-rich phosphorothioate oligonucleotides, but may be affected by changes in the nucleotide sequence. In addition, TLR9 agonism is abolished by methylation of cytosines within CpG dinucleotides. Thus, in some embodiments, the TLR9 agonist is an oligonucleotide of 8-35 nucleotides in length that includes the sequence 5'-AACGTTCG-3'. In some embodiments, the oligonucleotide is greater than 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 nucleotides in length and the oligonucleotide is less than 35, 34, 33, 32, 31, 30, 29, 28, 27, 26, 25, or 24 nucleotides in length. In some embodiments, the TLR9 agonist is an oligonucleotide 10-35 nucleotides in length comprising the sequence 5'-AACGTTCGAG-3' (SEQ ID NO:3). In some embodiments, the oligonucleotide is greater than 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 nucleotides in length and the oligonucleotide is less than 35, 34, 33, 32, 31, 30, 29, 28, 27, 26, 25, or 24 nucleotides in length.

[0033] Researchers at Dynavax Technologies Corporation (Emeryville, CA) have identified the CpG 1018® adjuvant, a 22-base long phosphorothioate-linked oligodeoxynucleotide that contains a specific sequence that can substantially enhance immune responses to co-administered antigens across species (Campbell, Methods Mol Biol, 1494:15-27, 2017). The CpG 1018® adjuvant (5'-TGACTGTGAA CGTTCGAGAT GA-3', set forth as SEQ ID NO:1) was selected after screening an extensive panel of oligonucleotides for in vitro and in vivo immune stimulatory activity. The CpG 1018® adjuvant is a CpG-B ODN that is active in mice, rabbits, dogs, baboons, cynomolgus monkeys, and humans. Thus, in some preferred embodiments, the TLR9 agonist is an oligonucleotide of 22-35 nucleotides in length that contains the sequence of SEQ ID NO:1.

[0034] An exemplary oligonucleotide TLR9 agonist, CpG 1018® adjuvant, is a CpG-ODN, but the disclosure is not limited to a complete DNA molecule. That is, in some embodiments, the TLR9 agonist is a DNA / RNA chimeric molecule in which the CpG and palindromic sequences are deoxyribonucleic acids, and one or more nucleic acids outside these regions are ribonucleic acids. In some embodiments, the CpG oligonucleotide is linear. In other embodiments, the CpG oligonucleotide is circular or contains a hairpin loop. The CpG oligonucleotide can be single-stranded or double-stranded.

[0035] In some embodiments, the CpG oligonucleotide may contain modifications. Modifications include, but are not limited to, modifications of the 3'OH or 5'OH groups, modifications of the nucleotide base, modifications of the sugar moiety, and modifications of the phosphate group. Modified bases may be included in the palindromic sequence of the CpG oligonucleotide, so long as the modified base maintains the same specificity to its natural complement by Watson-Crick base pairing (e.g., the palindromic portion is still self-complementary). In some embodiments, the CpG oligonucleotide includes a non-canonical base. In some embodiments, the CpG oligonucleotide includes a modified nucleoside. In some embodiments, the modified nucleoside is selected from the group consisting of 2'-deoxy-7-deazaguanosine, 2'-deoxy-6-thioguanosine, arabinoguanosine, 2'-deoxy-2'-substituted-arabinoguanosine, and 2'-O-substituted-arabinoguanosine. In some embodiments, the TLR9 agonist is a TLR9 agonist having the sequence 5'-TCG 1 AACG 1 TTCG 1 -3' (SEQ ID NO:2), where G 1 is 2'-deoxy-7-deazaguanosine. In some embodiments, the oligonucleotide has the sequence 5'-TCG 1 AACG 1 TTCG 1 -XG 1 CTTG 1 CAAG 1 CT-5', where G 1 is 2'-deoxy-7-deazaguanosine and X is glycerol (5'-SEQ ID NO:2-3'-X-3'-SEQ ID NO:2-5').

[0036] CpG oligonucleotides may contain modifications of the phosphate group. For example, in addition to phosphodiester linkages, phosphate modifications include, but are not limited to, methylphosphonates, phosphorothioates, phosphoramidates (bridged or unbridged), phosphotriesters, and phosphorodithioates, and may be used in any combination. Other non-phosphate linkages may also be used. In some embodiments, the oligonucleotides contain only phosphorothioate backbones. In some embodiments, the oligonucleotides contain only phosphodiester backbones. In some embodiments, the oligonucleotides contain a combination of phosphate linkages in the phosphate backbone, such as a combination of phosphodiester and phosphorothioate linkages. Oligonucleotides with phosphorothioate backbones may be more immunogenic than those with phosphodiester backbones and appear to be more resistant to degradation after injection into a host (Braun et al., J Immunol, 141:2084-2089, 1988; and Latimer et al., Mol Immunol, 32:1057-1064, 1995). The CpG oligonucleotide of the present disclosure comprises at least one, two or three internucleotide phosphorothioate ester linkages.In some embodiments, when a plurality of CpG oligonucleotide molecules are present in a pharmaceutical composition that comprises at least one excipient, both stereoisomers of phosphorothioate ester linkages are present in a plurality of CpG oligonucleotide molecules.In some embodiments, all of the internucleotide linkages of CpG oligonucleotide are phosphorothioate linkages, or in the above-mentioned alternative, CpG oligonucleotide has phosphorothioate backbone.

[0037] A unit dose of the immunogenic composition, which is a 1.0 ml dose in an exemplary embodiment, may contain about 750 μg to about 6000 μg of CpG oligonucleotide, preferably about 3000 μg or about 6000 μg of CpG oligonucleotide. In some embodiments, a 1.0 ml dose of the immunogenic composition contains more than about 750, 1000, 1250 or 1500 μg of CpG oligonucleotide, and less than about 6000, 5000, 4000 or 3000 μg of CpG oligonucleotide. In some embodiments, a 1.0 ml dose of the immunogenic composition contains about 750 μg, 1500 μg, 3000 μg or 6000 μg of CpG oligonucleotide. In some embodiments, a 1.0 ml dose of the immunogenic composition contains about 3000 μg of CpG oligonucleotide. In some embodiments, a 1.0 ml dose of the immunogenic composition contains about 6000 μg of CpG oligonucleotide.

[0038] The CpG oligonucleotides described herein are in the form of their pharma-ceutically acceptable salts, unless otherwise indicated.Exemplary basic salts include ammonium salts, alkali metal salts such as sodium, lithium and potassium salts, alkaline earth metal salts such as calcium and magnesium salts, zinc salts, salts with organic bases (e.g., organic amines) such as N-Me-D-glucamine, N-[1-(2,3-dioleoyloxy)propyl]-N,N,N-trimethylammonium chloride, choline, tromethamine, dicyclohexylamine, t-butylamine, and salts with amino acids such as arginine, lysine, and the like.In some embodiments, the CpG oligonucleotides are in the form of ammonium, sodium, lithium or potassium salts.In one preferred embodiment, the CpG oligonucleotides are in the form of sodium salts.

[0039] B. Varicella-zoster virus (VZV) glycoprotein E (gE) antigen The VZV gE antigen of the immunogenic composition of the present disclosure comprises gE or a fragment thereof. In preferred embodiments, the gE antigen is recognized by VZV-reactive antibodies and / or a peptide fragment of gE is recognized by VZV-reactive T cells. In some embodiments, the gE antigen is a recombinant protein, while in other embodiments, the gE antigen is purified from VZV virions. In some preferred embodiments, the gE antigen is an isolated antigen. In some embodiments, the gE antigen is not a fusion protein. In some preferred embodiments, the gE antigen of the immunogenic composition of the present disclosure is not part of live attenuated VZV or whole inactivated VZV.

[0040] In some embodiments, the VZV gE antigen is a truncated recombinant protein that lacks the transmembrane and intravirion domains of the full-length VZV gE antigen. In some embodiments, the recombinant protein is produced in mammalian cells, such as Chinese Hamster Ovary (CHO) cells. A representative gE amino acid sequence is set forth as GenBank No. AQT34120.1. In some embodiments, the gE antigen comprises an amino acid sequence from residues 39-585 of GenBank No. AQT34120.1, or an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical thereto.

[0041] In some embodiments, the amino acid sequence of gE is set forth as SEQ ID NO:4. MGTVNKPVVG VLMGFGIITG TLRITNPVRA SVLRYDDFHI DEDKLDTNSV YEPYYHSDHA ESSWVNRGES SRKAYDHNSP YIWPRNDYDG FLENAHEHHG VYNQGRGIDS GERLMQPTQM SAQEDLGDDT GIHVIPTLNG DDRHKIVNVD QRQYGDVFKG DLNPKPQGQR LIEVSVEENH PFTLRAPIQR IYGVRYTETW SFLPSLTCTG DAAPAIQHIC LKHTTCFQDV VVDVDCAENT KEDQLAEISY RFQGKKEADQ PWIVVNTSTL FDELELDPPE IEPGVLKVLR TEKQYLGVYI WNMRGGSDGTS TYATFLVTWK GDEKTRNPTP AVTPQPRGAE FHMWNYHSHV FSVGDTFSLA MHLQYKIHEA PFDLLLEWLY VPIDPTCQPM RLYSTCLYHP NAPQCLSHMN SGCTFTSPHL AQRVASTVYQ NCEHADNYTA YCLGISHMEP SFGLILHDGG TTLKFVDTPE SLSGLYVFVV YFNGHVEAVA YTVVSTVDHF VNAIEERGFP PTAGQPPATT KPKEITPVNP GTSPLIRYAA WTGGLA In some embodiments, the gE antigen comprises the amino acid sequence of SEQ ID NO:4, or an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:4.

[0042] In some embodiments, the amino acid sequence of gE is set forth as SEQ ID NO:5. MGTVNKPVVG VLMGFGIITG TLRITNPVRA SVLRYDDFHT DEDKLDTNSV YEPYYHSDHA ESSWVNRGES SRKAYDHNSP YIWPRNDYDG FLENAHEHHG VYNQGRGIDS GERLMQPTQM SAQEDLGDDT GIHVIPTLNG DDRHKIVNVD QRQYGDVFKG DLNPKPQGQR LIEVSVEENH PFTLRAPIQR IYGVRYTETW SFLPSLTCTG DAAPAIQHIC LKHTTCFQDV VVDVDCAENT KEDQLAEISY RFQGKKEADQ PWIVVNTSTL FDELELDPPE IEPGVLKVLR TEKQYLGVYI WNMRGGSDGTS TYATFLVTWK GDEKTRNPTP AVTPQPRGAE FHMWNYHSHV FSVGDTFSLA MHLQYKIHEA PFDLLLEWLY VPIDPTCQPM RLYSTCLYHP NAPQCLSHMN SGCTFTSPHL AQRVASTVYQ NCEHADNYTA YCLGISHMEP SFGLILHDGG TTLKFVDTPE SLSGLYVFVV YFNGHVEAVA YTVVSTVDHF VNAIEERGFP PTAGQPPATT KPKEITPVNP GTSPLLR In some embodiments, the gE antigen comprises the amino acid sequence of SEQ ID NO:5, or an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:5.

[0043] In some embodiments, the amino acid sequence of gE corresponds to a truncated gE (see U.S. Patent Application Publication No. 2021 / 0187099). In some embodiments, the amino acid sequence of gE is SEQ ID NO:6: MGTVNKPVVG VLMGFGIITG TLRITNPVRA SVLRYDDFHX DEDKLDTNSV YEPYYHSDHA ESSWVNRGES SRKAYDHNSP YIWPRNDYDG FLENAHEHHG VYNQGRGIDS GERLMQPTQM SAQEDLGDDT GIHVIPTLNG DDRHKIVNVD QRQYGDVFKG DLNPKPQGQR LIEVSVEENH PFTLRAPIQR IYGVRYTETW SFLPSLTCTG DAAPAIQHIC LKHTTCFQDV VVDVDCAENT KEDQLAEISY RFQGKKEADQ PWIVVNTSTL FDELELDPPE IEPGVLKVLR TEKQYLGVYI WNMRGGSDGTS TYATFLVTWK GDEKTRNPTP AVTPQPRGAE FHMWNYHSHV FSVGDTFSLA MHLQYKIHEA PFDLLLEWLY VPIDPTCQPM RLYSTCLYHP NAPQCLSHMN SGCTFTSPHL AQRVASTVYQ NCEHADNYTA YCLGISHMEP SFGLILHDGG TTLKFVDTPE SLSGLYVFVV YFNGHVEAVA YTVVSTVDHF VNAIEERGFP PTAGQPPATT KPKEIXXXXX XXXXXXXXXX XXXXXX where: X at position 40 is I or T; Any one of X's at positions 526 to 546 is present or absent; X at position 526, if present, is T; X at position 527, if present, is P; X at position 528, if present, is V; X at position 529, if present, is N; X at position 530, if present, is P; X at position 531, if present, is G; X at position 532, if present, is T; X at position 533, if present, is S; X at position 534, if present, is P; X at position 535, if present, is L; X at position 536, if present, is I or L; X at position 537, if present, is R; X at position 538, if present, is Y; X at position 539, if present, is A; X at position 540, if present, is A; X at position 541, if present, is W; X at position 542, if present, is T; X at position 543, if present, is G; X at position 544, if present, is G; X at position 545, if present, is L; X at position 546, if present, is A; As a result, the amino acid sequence of SEQ ID NO:6 is 525, 526, 527, 528, 529, 530, 531, 532, 533, 534, 535, 536, 537, 538, 539, 540, 541, 542, 543, 544, 545 or 546 amino acids in length. In some embodiments, the gE antigen comprises the amino acid sequence of SEQ ID NO:6, or an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:6.

[0044] A unit dose of the immunogenic composition, which in an exemplary embodiment is a 1.0 ml dose, may contain about 15 μg to about 150 μg of gE antigen, preferably about 25 μg to about 125 μg of gE antigen, preferably about 80 to about 120 μg of gE antigen, or about 100 μg of gE antigen. In some embodiments, a 1.0 ml unit dose of the immunogenic composition may contain about 25 μg, about 50 μg, about 75 μg, about 100 μg, about 125 μg, or about 150 μg of gE antigen.

[0045] C. Additional Components The immunogenic compositions of the disclosure may include one or more additional components, such as one or more excipients, another adjuvant, and / or additional antigens.

[0046] 1. Excipients Pharmaceutically acceptable excipients of the present disclosure include, for example, solvents, bulking agents, buffering agents, osmolality modifiers, and preservatives (Pramanick et al., Pharma Times, vol. 45:65-77, 2013). In some embodiments, the immunogenic composition may include an excipient that functions as one or more of a solvent, bulking agent, buffering agent, and osmolality modifier (e.g., sodium chloride in saline can function as both an aqueous medium and an osmolality modifier).

[0047] In some embodiments, the immunogenic composition comprises an aqueous medium as a solvent. Suitable vehicles include, for example, sterile water, saline solution, phosphate-buffered saline, and Ringer's solution. In some embodiments, the composition is isotonic.

[0048] The immunogenic composition may include a buffering agent. The buffering agent controls the pH and inhibits degradation of the active agent during processing, storage, and optionally reconstitution. Suitable buffering agents include salts, including, for example, acetate, citrate, phosphate, sulfate, or Tris. In preferred embodiments, the buffering agent is not a phosphate-containing buffer. In some preferred embodiments, the buffering agent is a Tris buffer. Other suitable buffering agents include amino acids, such as, for example, arginine, glycine, histidine, and lysine. The buffering agent may further include hydrochloric acid or sodium hydroxide. In some embodiments, the buffering agent maintains the pH of the composition within the range of 6 to 9. In some embodiments, the pH is greater than 6, 7, or 8 (lower limit). In some embodiments, the pH is less than 9, 8, or 7 (upper limit). That is, the pH is in the range of about 6 to 9, with the lower limit being less than the upper limit. In some embodiments, the pH is about 6.5, about 7.0, or about 7.5.

[0049] The immunogenic composition may also include a osmolality modifier. Suitable osmolality modifiers include, for example, dextrose, glycerol, sodium chloride, glycerin, and mannitol.

[0050] The immunogenic composition may contain a bulking agent. Bulking agents are particularly useful when the pharmaceutical composition is freeze-dried before administration. In some embodiments, bulking agents are protective agents that help stabilize and prevent the degradation of active agents during freeze-drying or spray-drying and / or during storage. Suitable bulking agents are sugars (monosaccharides, disaccharides and polysaccharides), such as sucrose, lactose, trehalose, mannitol, sorbitol, glucose and raffinose.

[0051] The immunogenic composition may contain a preservative. Suitable preservatives include, for example, antioxidants and antimicrobial agents. However, in a preferred embodiment, the immunogenic composition is prepared under sterile conditions and is in a single-use container, so the inclusion of a preservative is not required.

[0052] 2. Additional adjuvants Adjuvants are known in the art and include, but are not limited to, alum (aluminum salts), oil-in-water emulsions, water-in-oil emulsions, liposomes, and microparticles, such as poly(lactide-co-glycoside) microparticles (Shah et al., Methods Mol Biol, 1494:1-14, 2017). In some embodiments, the immunogenic composition further comprises an aluminum salt adjuvant to which the gE antigen is adsorbed. In some embodiments, the aluminum salt adjuvant comprises one or more of the group consisting of amorphous aluminum hydroxyphosphate sulfate, aluminum hydroxide, aluminum phosphate, and aluminum potassium sulfate. In some embodiments, the aluminum salt adjuvant comprises one or both of aluminum hydroxide and aluminum phosphate. In some embodiments, the aluminum salt adjuvant consists of aluminum hydroxide.

[0053] In some embodiments, a unit dose, which in an exemplary embodiment is a 1.0 ml dose of the immunogenic composition, contains about 0.25 to about 1.25 mg of Al. 3+, preferably about 0.50 to about 1.00 mg of Al 3+ In some embodiments, the immunogenic composition comprises about 0.50 mg, about 0.75 mg, or about 1.00 mg of Al. 3+ , preferably about 0.75 mg of Al 3+ Includes.

[0054] In other embodiments, the immunogenic composition further comprises an additional adjuvant. Additional suitable adjuvants include, but are not limited to, squalene-in-water emulsions (e.g., MF59 or AS03), TLR3 agonists (e.g., poly-IC or poly-ICLC), TLR5 agonists (bacterial flagellin), and TLR7 and / or TLR8 agonists (imidazoquinoline derivatives, e.g., imiquimod and resiquimod) (Coffman et al., Immunity, 33:492-503, 2010). For veterinary use and for the production of antibodies in non-human animals, the mitogenic components of Freund's adjuvant (both complete and incomplete) can be used. However, in a preferred embodiment, the immunogenic composition does not include one or more of saponin, TLR4 agonist, and liposomes. In some preferred embodiments, the immunogenic compositions do not include one or more of QS21, 3-O-deacylated monophosphoryl lipid A (3D-MPL1), dioleoylphosphatidylcholine, and / or cholesterol.

[0055] D.Kit The present disclosure also provides a kit comprising: i) an immunogenic composition comprising a VZV gE antigen and a CpG oligonucleotide; and ii) a set of instructions for administering the immunogenic composition to stimulate an immune response against the gE antigen in a human subject in need thereof. In addition, the present disclosure provides a kit comprising: i) a first composition comprising a VZV gE antigen; (ii) a second composition comprising a CpG oligonucleotide; iii) instructions for mixing the first composition with the second composition to prepare an immunogenic composition; and optionally, iv) a further set of instructions for administering the immunogenic composition to stimulate an immune response against the gE antigen in a human subject in need thereof. In some preferred embodiments, the CpG oligonucleotide comprises the sequence 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO: 1). In some preferred embodiments, stimulating an immune response against the gE antigen comprises increasing cell-mediated immunity against VZV in the human subject.

[0056] In yet further preferred embodiments, the present disclosure provides a kit comprising: i) a first composition comprising a VZV gE antigen; ii) a second composition comprising a CpG oligonucleotide; iii) a third composition comprising an aluminum salt adjuvant; iv) instructions for preparing an immunogenic composition from the first, second and third compositions; and optionally v) a further set of instructions for administering the immunogenic composition to stimulate an immune response against the gE antigen in a human subject in need thereof. In some preferred embodiments, the CpG oligonucleotide comprises the sequence 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO: 1). In some preferred embodiments, stimulating an immune response against the gE antigen comprises increasing cell-mediated immunity against VZV in the human subject.

[0057] The kit may include the immunogenic composition in a suitable package. For example, when the immunogenic composition is a lyophilized powder, a vial with a flexible stopper is usually used so that the powder can be easily resuspended by injection liquid (e.g., sterile water, saline, etc.) through the flexible stopper. In some embodiments, the kit includes a device for administration (e.g., a syringe and a needle). The instructions for use of the immunogenic composition generally include information on the dosage, schedule and route of administration for the intended method of use. In some embodiments, the immunogenic composition is for stimulating an immune response against VZV, for example, to increase cell-mediated immunity against VZV in a human subject.

[0058] II. Method of Use The present disclosure relates to a method for stimulating an immune response against VZV, comprising administering an immunogenic composition comprising a VZV glycoprotein E (gE) antigen and a CpG oligonucleotide to a human subject to stimulate an immune response against the gE antigen in the human subject. In a preferred embodiment, the immunogenic composition should be administered by intramuscular injection, optionally in a volume (e.g., unit dose) of about 1.0 mL. In some embodiments, the intramuscular injection is into the deltoid muscle of the upper arm of the human subject in need thereof.

[0059] Stimulating an immune response means increasing an immune response, which may result from inducing a de novo immune response (e.g., as a result of a primary vaccination regimen) or enhancing an existing immune response (e.g., as a result of a booster vaccination regimen). In some embodiments, stimulating an immune response includes, but is not limited to, one or more of the group consisting of stimulating cytokine production, stimulating B lymphocyte proliferation, stimulating antibody production, stimulating interferon pathway-related gene expression, stimulating chemoattractant-related gene expression, and stimulating maturation of plasmacytoid dendritic cells.

[0060] In some preferred embodiments, the method of the present disclosure is suitable for increasing cell-mediated immunity to VZV in a subject, compared to the cell-mediated immunity to VZV in the subject before administration of the immunogenic composition. In some embodiments, the increase in cell-mediated immunity to VZV comprises an increase in the frequency of gE antigen-reactive activated CD4+ T cells, and the activated CD4+ T cells express two or more activation markers selected from interleukin-2, interferon-gamma, tumor necrosis factor-alpha and CD40L (CD154). In some embodiments, the cell-mediated immunity is measured by flow cytometry. In some embodiments, the gE antigen-reactive activated CD4+ T cells are present in peripheral blood mononuclear cells of the subject.

[0061] In some preferred embodiments, the method of the present disclosure is suitable for increasing both cell-mediated immunity to VZV in a subject and humoral immunity to VZV in a subject. In some embodiments, administration of the immunogenic composition results in an increase in humoral immunity to VZV in a subject, compared to the humoral immunity to VZV in the subject before administration of the immunogenic composition. In some embodiments, the increase in humoral immunity to VZV comprises an increase in the concentration of gE antigen-reactive IgG. In some embodiments, gE antigen-reactive IgG is measured by ELISA. In some embodiments, the increase in humoral immunity to VZV comprises an increase in the concentration of VZV-reactive antibodies. In some embodiments, the increase in the concentration of VZV-reactive antibodies comprises an increase in the concentration of gE and / or VZV neutralizing antibodies.

[0062] The human subject is typically 18 years of age or older. In some embodiments, the human subject is a healthy male or female adult. In some embodiments, the human subject is 50 years of age or older. In some embodiments, the human subject is 50-69 years of age. In some embodiments, the human subject is suspected of having a latent VZV infection. In some embodiments, the human subject suffered from chickenpox as a child (12 years of age or younger), adolescent (13-17 years of age) or young adult (18-25 years of age) prior to administration of the immunogenic composition. In other embodiments, the human subject is not infected with VZV or is not known to have been infected with VZV. In some embodiments, the human subject has not been previously vaccinated against chickenpox or shingles. In some embodiments, the human subject is 18 years of age or older and suffers from immunodeficiency or immunosuppression caused by a known disease or therapy.

[0063] In some methods of the disclosure, a first and a second dose of the immunogenic composition are administered to a human subject, with the second dose being administered one month to one year after the first dose. In some embodiments, the second dose is administered two months to six months after the first dose. In some embodiments, the second dose is administered about two months after the first dose. In an exemplary embodiment, each dose is administered by intramuscular injection, a 1 mL dose.

[0064] Enumeration of embodiments 1. A method for increasing cell-mediated immunity to Varicella Zoster Virus (VZV) in a human subject in need thereof, comprising: A method comprising administering to a human subject an immunogenic composition comprising: i) an unmethylated cytidine-phospho-guanosine (CpG)-containing oligonucleotide comprising the sequence 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO: 1); and ii) a truncated VZV glycoprotein E (gE) antigen, present in the immunogenic composition in an amount effective to increase cell-mediated immunity to VZV compared to cell-mediated immunity to VZV in the subject prior to administration of the immunogenic composition. 2. The method of embodiment 1, wherein the immunogenic composition comprises about 25 μg to about 150 μg of gE antigen and about 750 μg to about 6000 μg of oligonucleotide, optionally, the immunogenic composition comprises about 50 μg to about 100 μg of gE antigen and about 1500 μg to about 6000 μg of oligonucleotide. 3. The method of embodiment 1, wherein the immunogenic composition comprises about 25 μg, about 50 μg, about 75 μg, about 100 μg, about 125 μg or about 150 μg of gE antigen and about 750 μg, about 1500 μg, about 3000 μg or about 6000 μg of oligonucleotide. 4. The method of embodiment 3, wherein the immunogenic composition comprises about 100 μg of gE antigen and about 3000 μg or 6000 μg of oligonucleotide. 5. A method for increasing an immune response to Varicella Zoster Virus (VZV) in a human subject in need thereof, comprising: 1. A method comprising administering to a human subject an immunogenic composition comprising: i) an unmethylated cytidine-phospho-guanosine (CpG)-containing oligonucleotide comprising the sequence 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO:1); and ii) a truncated VZV glycoprotein E (gE) antigen, wherein the immunogenic composition comprises about 100 mcg of gE antigen and about 3000 mcg or about 6000 mcg of the oligonucleotide. 6. The method of any one of embodiments 1 to 5, wherein the oligonucleotide comprises at least one phosphorothioate linkage or all nucleotide linkages are phosphorothioate linkages. 7. The method of any one of embodiments 1 to 6, wherein the oligonucleotide is a single-stranded oligodeoxynucleotide. 8. The method according to any one of the preceding embodiments, wherein the gE antigen is a recombinant protein lacking the transmembrane and intravirion domains of the full-length VZV gE antigen. 9. The method of embodiment 8, wherein the recombinant protein is produced in mammalian cells. 10. The method of any one of embodiments 1-9, wherein the immunogenic composition further comprises an aluminum salt adjuvant. 11. The method of embodiment 10, wherein the aluminum salt adjuvant comprises one or more of the group consisting of amorphous aluminum hydroxyphosphate sulfate, aluminum hydroxide, aluminum phosphate and potassium aluminum sulfate. 12. The method of embodiment 10, wherein the aluminum salt adjuvant comprises aluminum hydroxide. 13. The immunogenic composition comprises about 0.25 mg to about 1.25 mg of Al. 3+ 13. The method according to any one of embodiments 10 to 12, comprising: 14. The immunogenic composition comprises about 0.50 mg, about 0.75 mg, or about 1.00 mg of Al. 3+ 13. The method according to any one of embodiments 10 to 12, comprising: 15. The immunogenic composition comprises about 0.75 mg of Al 3+ 15. The method of embodiment 14, comprising: 16. The method of any one of the preceding embodiments, wherein the immunogenic composition does not comprise a saponin, a TLR4 agonist and / or a liposome. 17. The method of any one of embodiments 1-15, wherein the immunogenic composition does not comprise QS21, 3-O-deacylated monophosphoryl lipid A (3D-MPL1), dioleoylphosphatidylcholine and / or cholesterol. 18. The method according to any one of the preceding embodiments, wherein the gE antigen and the oligonucleotide are present in the immunogenic composition in a ratio of 1:15 to 1:120 (wt / wt), and optionally, the gE antigen and the oligonucleotide are present in the immunogenic composition in a ratio of 1:15 to 1:60 (wt / wt). 19. The method according to any one of the preceding embodiments, wherein the gE antigen and the oligonucleotide are present in the immunogenic composition in a ratio of 1:15, 1:30, 1:60, 1:20, 1:40, 1:80, 1:30, 1:60 or 1:120 (wt / wt), optionally wherein the gE antigen and the oligonucleotide are present in the immunogenic composition in a ratio of 1:30 (wt / wt), and optionally wherein the gE antigen and the oligonucleotide are present in the immunogenic composition in a ratio of 1:60 (wt / wt). 20. Oligonucleotides and Al 3+ is present in the immunogenic composition in a ratio of 8:1 to 2:1 (wt / wt), and optionally, oligonucleotides and Al 3+ is present in the immunogenic composition in a ratio of 8:1 (wt / wt), and optionally, the oligonucleotide and Al 3+ The method according to any one of embodiments 13 to 19, wherein said antibody is present in the immunogenic composition in a ratio of 4:1 (wt / wt). 21. The method of any one of embodiments 1-20, wherein the human subject is 18 years of age or older. 22. The method of any one of embodiments 1-20, wherein the human subject is 50 years of age or older. 23. The method of embodiment 22, wherein the human subject is between 50 and 69 years old. 24. The method of any one of embodiments 1-23, wherein the human subject is suspected of having a latent VZV infection. 25. The method of any one of embodiments 1-24, wherein the human subject has not been previously vaccinated against chickenpox or shingles. 26. The method of any one of embodiments 1-25, wherein a human subject receives a first dose and a second dose of the immunogenic composition, the second dose being administered one month to one year after the first dose. 27. The method of embodiment 26, wherein the second dose is administered 2 to 6 months after the first dose, optionally, the second dose is administered about 2 months after the first dose. 28. The method of any one of embodiments 1 to 27, wherein the immunogenic composition is administered by intramuscular injection. 29. The method of any one of embodiments 1 to 28, wherein increasing cell-mediated immunity to VZV comprises increasing the frequency of gE antigen-reactive activated CD4+ T cells, and the activated CD4+ T cells express two or more activation markers selected from interleukin-2, interferon-gamma, tumor necrosis factor-alpha and CD40L (CD154). 30. The method of any one of embodiments 1 to 29, wherein administration of the immunogenic composition results in an increase in humoral immunity to VZV in the subject compared to humoral immunity to VZV in the subject prior to administration of the immunogenic composition. 31. The method of embodiment 30, wherein increasing humoral immunity to VZV comprises increasing the concentration of gE antigen-reactive IgG. 32. The method of embodiment 30 or embodiment 31, wherein increasing humoral immunity against VZV comprises increasing the concentration of gE and / or VZV neutralizing antibodies. 33. The method of any one of embodiments 30-32, wherein an increase in cell-mediated immunity and an increase in humoral immunity are induced in the subject by about two months after administration of the first dose of the immunogenic composition. 34. The method of any one of embodiments 30-32, wherein an increase in cell-mediated immunity and an increase in humoral immunity are induced in the subject by about one month after administration of the second dose of the immunogenic composition. 35. The method of any one of embodiments 30-34, wherein the increased cell-mediated immunity and increased humoral immunity are maintained in the subject for at least 3 months after administration of the second dose of the immunogenic composition. 36. The method of any one of embodiments 30 to 35, wherein administration of the immunogenic composition reduces the risk of the subject developing shingles compared to the risk of a control subject not receiving the immunogenic composition comprising the gE antigen. 37. The method of any one of embodiments 30-35, wherein administration of the immunogenic composition optionally prevents the subject from developing shingles for a period of at least 3 years after administration of the second dose of the immunogenic composition. 38. The method of any one of embodiments 30-35, wherein administration of the immunogenic composition optionally prevents the subject from developing postherpetic neuralgia for a period of at least 3 years following administration of the second dose of the immunogenic composition. 39. The method of any one of embodiments 30 to 38, wherein the immunogenic composition is associated with a reduced risk of occurrence of one or more reactive local reactions within 7 days (the day of vaccination and 6 days thereafter) after administration of the first dose and / or the second dose, compared to a comparative control composition comprising a gE antigen and an AS01B adjuvant. 40. The method of embodiment 39, wherein the reactive local reaction comprises pain that interferes with normal daily activities. 41. The method of any one of embodiments 30 to 40, wherein the immunogenic composition is associated with a reduced risk of occurrence of one or more solicited systemic reactions within 7 days (the day of vaccination and 6 days thereafter) following administration of the first dose and / or the second dose, compared to a comparative control composition comprising a gE antigen and an AS01B adjuvant. 42. The method of embodiment 41, wherein the solicited systemic reaction comprises fatigue, headache and / or muscle pain that interferes with normal daily activities, and / or fever above 39°C. 43. The method of any one of embodiments 30-42, wherein the immunogenic composition is associated with a reduced risk of occurrence of Guillain-Barré syndrome within 42 days (the day of vaccination and 41 days thereafter) after administration of the first dose and / or the second dose, compared to a comparative control composition comprising a gE antigen and an AS01B adjuvant. 44. An immunogenic composition for use in a method according to any one of embodiments 1 to 43. 45. An immunogenic composition comprising: i) an unmethylated cytidine-phospho-guanosine (CpG)-containing oligonucleotide comprising the sequence 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO: 1); ii) a truncated VZV glycoprotein E (gE) antigen; and iii) at least one excipient, An immunogenic composition, wherein the gE antigen is a recombinant protein produced in a mammalian cell and is present in the immunogenic composition in an amount of about 25 μg to about 150 μg, the oligonucleotide is present in the immunogenic composition in an amount of about 750 μg to about 6000 μg, and at least one excipient comprises a pharma- ceutically acceptable buffer. 46. ​​An immunogenic composition comprising: i) an unmethylated cytidine-phospho-guanosine (CpG)-containing oligonucleotide comprising the sequence 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO:1); ii) a truncated VZV glycoprotein E (gE) antigen; and iii) at least one excipient, An immunogenic composition, wherein the gE antigen is a recombinant protein produced in a mammalian cell and is present in the immunogenic composition in an amount of about 100 μg, the oligonucleotide is present in the immunogenic composition in an amount of about 3000 μg or about 6000 μg, and at least one excipient comprises a pharma- ceutically acceptable buffer. 47. The immunogenic composition further comprises an aluminum hydroxide adjuvant, wherein the immunogenic composition comprises about 0.25 mg to about 1.25 mg of Al. 3+ 47. The immunogenic composition of embodiment 45 or embodiment 46, comprising: 48. The immunogenic composition comprises about 0.75 mg of Al 3+ 48. The immunogenic composition of embodiment 47, comprising: 49. The immunogenic composition according to any one of embodiments 45 to 48, wherein the buffer is not a phosphate-containing buffer. 50. The immunogenic composition according to any one of embodiments 45 to 48, wherein the buffer is a Tris buffer. EXAMPLES

[0065] Abbreviations: CMI (cell-mediated immunity); CpG (nonmethylated cytidine-phospho-guanosine); CTRL (control); gE (glycoprotein E); ELISA (enzyme-linked immunosorbent assay); GMC (geometric mean concentration); GMFI (geometric mean fold increase); HZ (herpes zoster); IFNγ (interferon-gamma); IL-2 (interleukin-2); IM (intramuscular); mcg or μg (microgram); mcl or μl (microliter); PBMC (peripheral blood mononuclear cells); TLR9 (toll-like receptor 9); TNFα or TNF (tumor necrosis factor-alpha); VRR (vaccine response rate); and VZV (varicella zoster virus). EXAMPLES

[0066] Safety, tolerability, and immunogenicity of CpG-adjuvanted recombinant varicella zoster vaccine compared with AS01B-adjuvanted recombinant varicella zoster vaccine in human subjects This example provides a description of a Phase I clinical trial conducted in healthy adults to compare the effects of administering two doses of SHINGRIX® (a vaccine containing VZV gE adjuvanted with AS01B, GlaxoSmithKline, Research Triangle Park, NC) administered over the course of two months versus administering two doses of Z-1018 (a vaccine containing recombinant VZV gE adjuvanted with CpG 1018® adjuvant, Dynavax Technologies Corporation, Emeryville, CA) over the course of two months.

[0067] Antigen: Recombinant VZV glycoprotein E (gE) was obtained by cell culture of genetically engineered Chinese hamster ovary host cells expressing a truncated version of gE lacking the transmembrane anchor and carboxy-terminal domains, and thus secreted into the supernatant. Cell culture was supported by a medium without albumin, antibiotics or animal-derived proteins, but containing amino acids. gE was purified by chromatography and lyophilized for future use as described (Haumont et al., Virus Res, 40:199-204, 1996). For phase I clinical trials, VZV gE was obtained commercially.

[0068] Primary objective: To evaluate the safety and tolerability of Z-1018. Endpoints include evaluation of adverse events: solicited local and systemic post-injection reactions; adverse events; and evaluation of serious adverse events.

[0069] Secondary objective: to evaluate the immunogenicity of Z-1018 compared to SHINGRIX® 4 weeks after the second injection. Endpoints include: 1) the number of CD4+ T cells expressing at least two of the following activation markers: IFNγ, IL-2, TNFα and CD40L; 2) the geometric mean concentration (GMC) of IgG antibodies against VZV gE antigen; and 3) the determination of vaccine response rate (VRR). The desired endpoint is an increase in gE-reactive CD4+ T cells in PBMCs with a 2-fold or greater increase over baseline in the expression of two or more of IFNγ, IL-2, TNFα and CD40L.

[0070] Exploratory Objective: To further evaluate the immunogenicity of Z-1018. Endpoints include assessment of 1) levels of gE neutralizing antibodies; 2) T cell phenotype; 3) B cell phenotype; and 4) peripheral blood mononuclear cell (PBMC) gene expression profile.

[0071] Study Design. The study was conducted as a randomized, active-controlled, dose-escalation, multicenter study of two doses (day 1 and week 8) of an investigational varicella zoster (HZ) vaccine (Z-1018), combining varicella zoster gE antigen with an oligodeoxynucleotide, toll-like receptor 9 (TLR9) agonist adjuvant (CpG 1018® adjuvant), with and without alum, in healthy volunteers aged 50-69 years. Subjects were randomized to receive ascending dose levels of Z-1018 or SHINGRIX® as follows: i) Z-1018 dose level 1: 100 mcg gE + 3000 mcg CpG 1018® adjuvant; ii) Z-1018 dose level 1a: 100 mcg gE + 3000 mcg CpG 1018® adjuvant + alum; iii) Z-1018 dose level 2: 100 mcg gE + 6000 mcg CpG 1018® adjuvant; iv) Z-1018 dose level 2a: 100 mcg gE + 6000 mcg CpG 1018® adjuvant + alum: or v) SHINGRIX®.

[0072] All subjects received two doses of their assigned treatment and underwent safety and immunogenicity evaluations. Subjects were followed for safety for 12 weeks after the last study injection (i.e., 12 weeks after the second injection at week 8 = 20 weeks after the first injection). All subjects had pre-vaccination blood drawn and received the assigned treatment vaccine on day 1. Post-vaccination blood draws and exploratory evaluations to measure antibody levels were performed at weeks 8, 12, and 20. Subjects received the second dose of their assigned treatment at week 8 after blood draw. Safety assessments were performed at baseline (day 1), day 8, week 4, week 12, week 16, and week 20. The duration of subject participation in the screening and treatment parts of the study was approximately 24 weeks. Study population. Healthy male and female subjects aged 50-69 years were selected. Exclusion criteria included any one of the following criteria: seropositivity for human immunodeficiency virus; history of varicella zoster; previous vaccination against chickenpox or varicella zoster; receiving any non-live vaccine within 14 days of the first injection; receiving any live vaccine, systemic corticosteroids (inhaled steroids were acceptable), immunomodulators or immunosuppressants, granulocyte stimulating factors, or granulocyte-macrophage colony-stimulating factors within 28 days of the first injection; receiving or expected to receive chemotherapy; having a diagnosis of cancer within the past 5 years; and a history of autoimmune disease.

[0073] Study Treatment. The investigational product (Z-1018) or active control (SHINGRIX®) was administered as a single intramuscular (IM) injection into the deltoid muscle on Day 1 and Week 8.

[0074] A 1 ml dose of Z-1018 was combined with a 3000 mcg or 6000 mcg dose of CpG 1018® adjuvant (Dynavax Technologies Corporation. Emeryville, CA) (750 mcg of Al 3+ The adjuvant contains 100 mcg of gE (with or without a dose of aluminum hydroxide [alum]). CpG 1018® adjuvant is a single stranded 22 base phosphorothioate 2'-deoxyribo-oligonucleotide (5'-TGACTGTGAA CGTTCGAGAT GA-3', set forth in SEQ ID NO: 1) prepared by standard solid phase chemistry techniques. CpG 1018® adjuvant has a molar mass of approximately 7150 daltons. The alum in Z-1018 is ALHYDROGEL® vaccine adjuvant sold by Croda International Plc (East Yorkshire, UK).

[0075] A 0.5 ml dose of SHINGRIX® contains 50 mcg of gE combined with AS01B adjuvant (a liposomal formulation containing 1 mg dioleoylphosphatidylcholine [DOPC], 250 mcg cholesterol, 50 mcg 3-O-desacyl-4'-monophosphoryl lipid A [MPL] from Salmonella minnesota, and 50 mcg QS-21 saponin, fraction 21 from Quillaja saponaria Molina).

[0076] Immunogenicity assessment. IgG antibodies to gE (anti-gE), and T and B cell responses to gE were assessed at baseline (day 1), weeks 8, 12, and 20. Neutralizing responses to gE were assessed at baseline (day 1), weeks 8, 12, and 20.

[0077] The safety analysis population includes all subjects who received at least one dose of the study vaccine, excluding subjects without ongoing data.

[0078] The per-protocol (PP) population for immunogenicity analysis includes all subjects who received two doses of study vaccine, had no major protocol violations (as defined in the statistical analysis plan), and had CD4+ T cell counts obtained within the study visit window at week 12.

[0079] The modified intention to treat (mITT) population for immunogenicity analysis includes all eligible subjects who received at least one dose of the study vaccine and have a post-injection immunogenicity assessment.

[0080] For immunogenicity analysis, response rates for IgG and neutralizing antibodies against gE were evaluated. For cell-mediated immune (CMI) responses, the frequency of CD4+ T cells expressing ≧2 activation markers was measured. In addition, geometric mean concentrations (GMCs) were evaluated for antibodies against gE at weeks 8 and 12. Geometric mean ratios (GMRs) (e.g., week 8 vs. day 1, week 12 vs. day 1) were evaluated for antibodies against gE by dose level. T and B cell phenotypes, as well as safety data, were analyzed descriptively.

[0081] VZV Glycoprotein E (gE) ELISA. Wells of a flat-bottom microplate were coated overnight with 0.1 mcg of VZV gE antigen, washed, blocked in buffer containing 5% bovine serum albumin, and washed again. Standards, controls, and human serum samples (dilutions of 1:50 or greater) were added in duplicate to the wells of the microplate, which were incubated for 1.0 to 1.5 hours at room temperature. The standard used was an anti-VZV immunoglobulin World Health Organization standard (National Institute for Biological Standards and Control, catalog number W1044). After washing, detection antibody (dilution 1:50,000) was added to all wells of the microplate, which were incubated for 1.0 to 1.25 hours at room temperature. The detection antibody used was mouse anti-human IgG-peroxidase (Jackson ImmunoResearch, catalog number 209-035-98). After washing, 3,3',5,5'-tetramethylbenzidine substrate (Life Technologies, Cat. No. 002023) was added to all wells of the microplate and allowed to react for approximately 10 minutes before color development was stopped by the addition of 1N sulfuric acid stop solution. Absorbance was measured and concentrations were determined using a Tecan SPARK microplate reader and Magellan data analysis software.

[0082] Intracellular staining assay. PBMCs were isolated from blood samples obtained from study subjects and cryopreserved. Frozen PBMCs were gently thawed, washed in complete medium containing benzonase, and transferred to 6-well cell culture plates in complete medium without benzonase. Cells were incubated at 4°C for 24 h at 5% CO 2 The cells were then incubated at 4°C for 1 h at 37°C for 2 h and rested overnight in a 37°C incubator. The rested cells were washed, counted, and resuspended in fresh complete medium. When cell viability was at least 60-70%, the cells were distributed into wells of a 96-well cell culture plate and stimulated in complete medium containing: DMSO (0.8%), a series of peptides (15mer with 11aa overlaps) spanning the VZV gE protein (1 mcg / well). The cells were incubated at 4°C for 1 h at 37°C for 2 ... for 2 h at 37°C for 2 h at 37°C for 2 h at 37°C for 2 After stimulation for 2 hours in a 37°C incubator, an inhibitor mix containing monensin and brefeldin A was added to each well and the cell culture plate was returned to the incubator for an additional 16-18 hours. The cells were then washed, resuspended in live / dead viability dye and incubated for 20 minutes at room temperature in the dark. After washing, the cells were resuspended in cold Cytofix-Cytoperm and incubated for 20 minutes at 2-8°C in the dark. The cells were washed again and resuspended in Perm / wash buffer. The pelleted cells were resuspended in an intracellular staining panel mix containing fluorescently labeled antibodies against CD3, CD4, CD8, IFNγ, IL-2, TNFα and CD40L and incubated for 20 minutes at room temperature in the dark. The stained cells were washed, resuspended in staining buffer and stored at 2-8°C until analysis using an Attune NxT flow cytometer.

[0083] Results – Safety and Tolerability Post-injection reactions and adverse events were evaluated in the safety analysis population, which included all subjects who received at least one dose of the study vaccine. The characteristics of the safety analysis population are shown in Table 1. There were more women than men in the Z-1018 group, but the Shingrix® group was more gender-balanced. The mean age was similar across all study groups.

[0084] [Table 1]

[0085] The overall rates of reactogenicity are shown in Table 2. Local and systemic reactogenicity rates in the safety analysis population are shown in Tables 3 and 4, respectively.

[0086] [Table 2]

[0087] [Table 3]

[0088] [Table 4]

[0089] In this study, most participants in the Z-1018 group reported some reactogenicity, but most reactions were mild or moderate. Injection site pain was the most common local reaction. Fatigue was the most commonly reported systemic reaction, while nausea and diarrhea were the most frequently reported gastrointestinal symptoms. There was no clear dose response in the reported rates of local or systemic reactions in the Z-1018 groups, but more severe reactions were reported in the Z-1018 6 mg + alum group than in the other three Z-1018 groups.

[0090] The reported rate of any local and systemic reactions was higher after Shingrix® than after Z-1018. In addition, the number of reported moderate or severe reactions was higher after Shingrix® than after any formulation of Z-1018. Observations from the safety analysis population indicate that all formulations of Z-1018 have a better tolerability profile than Shingrix®.

[0091] Results - Immunogenicity VZV gE-reactive humoral and cellular immune responses were evaluated in the per-protocol population, including all subjects who received two doses of study vaccine, had no major protocol violations, and had CD4+ T cells obtained within the study visit window 4 weeks after the second injection. Characteristics of the per-protocol population are shown in Table 5. All Z-1018 groups had somewhat more females than the Shingrix® groups, but age was well balanced between all groups.

[0092] [Table 5]

[0093] Anti-gE IgG antibody levels (IU / mL) are presented as geometric mean concentrations (GMC) in Table 6, geometric mean fold increase (GMFI) in Table 7, and vaccine response rates in Table 8.

[0094] [Table 6]

[0095] [Table 7]

[0096] [Table 8]

[0097] The GMC reached at 12 weeks was comparable across all four Z-1018 groups. The GMC of the Z-1018 group was lower than that in the Shingrix® group. However, conclusions across groups are difficult to draw due to the higher baseline GMC of the Shingrix® group. At 12 weeks, the GMFI of all Z-1018 groups was comparable to or higher than Shingrix®. Among the Z-1018 groups, the highest GMFI was observed in the Z-1018 6 mg groups with and without alum. High VRR was achieved in all Z-1018 groups at 12 weeks and was comparable to Shingrix®.

[0098] The levels of gE-reactive CD4+ T cells expressing two or more activation markers (herein referred to as "CD4+(2+) T cells") were calculated using the 10-fold increase in the levels shown in Table 9 and Figures 1 to 3. 6 are shown as median frequency per CD4+ T cell, geometric mean fold increase (GMFI) in Table 10, and percentage of subjects with a 2-fold increase post-vaccination versus pre-vaccination in Table 11.

[0099] [Table 9]

[0100] [Table 10]

[0101] [Table 11]

[0102] The frequency of gE-reactive CD4+(2+) T cells expressing different combinations of activation markers (two or more of IFNγ, IL-2, TNFα, and CD40L) was similar across all groups at week 8. However, the frequency of gE-reactive CD4+(2+) T cells was higher for Shingrix® than for the four Z-1018 groups at week 12. For the Z-1018 group, similar to Shingrix®, gE-reactive CD4+ T cells expressing all four activation markers were present at the highest frequency. Robust GMFI in CD4+(2+) T cell frequency at week 12 versus baseline was observed in all four Z-1018 groups. At week 12, high vaccine response rates based on CD4+(2+) T cell frequency were seen in each Z-1018 group and were comparable to Shingrix®. A summary of the immunogenicity data for the four Z-1018 groups is shown in Table 12.

[0103] [Table 12]

[0104] Although the foregoing disclosure has been described in some detail by way of illustration and examples for purposes of clarity and understanding, it will be apparent to those skilled in the art that certain changes and modifications may be made. Accordingly, the examples should not be construed as limiting the scope of the present disclosure, which is particularly set forth in the appended claims.

Claims

1. 1. A method for increasing cell-mediated immunity to Varicella Zoster Virus (VZV) in a human subject in need thereof, comprising:

1. A method comprising administering to a human subject an immunogenic composition comprising: i) an unmethylated cytidine-phospho-guanosine (CpG)-containing oligonucleotide comprising the sequence 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO:1), present in the immunogenic composition in an amount effective to increase cell-mediated immunity to VZV compared to cell-mediated immunity to VZV in the subject prior to administration of the immunogenic composition; and ii) a truncated VZV glycoprotein E (gE) antigen, wherein a first dose and a second dose of the immunogenic composition are administered to the human subject, the second dose being administered 2 to 6 months after the first dose.

2. 10. The method of claim 1, wherein the immunogenic composition comprises about 25 μg to about 150 μg of gE antigen and about 750 μg to about 6000 μg of oligonucleotide.

3. 10. The method of claim 1, wherein the immunogenic composition comprises about 50 μg to about 100 μg of gE antigen and about 1500 μg to about 6000 μg of oligonucleotide.

4. 2. The method of claim 1, wherein the immunogenic composition comprises about 25 μg, about 50 μg, about 75 μg, about 100 μg, about 125 μg or about 150 μg of gE antigen and about 750 μg, about 1500 μg, about 3000 μg or about 6000 μg of oligonucleotide.

5. 5. The method of claim 4, wherein the immunogenic composition comprises about 100 μg of gE antigen and about 3000 μg or 6000 μg of oligonucleotide.

6. 1. A method for increasing an immune response to Varicella Zoster Virus (VZV) in a human subject in need thereof, comprising:

1. A method comprising administering to a human subject an immunogenic composition comprising: i) an unmethylated cytidine-phospho-guanosine (CpG)-containing oligonucleotide comprising the sequence 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO:1); and ii) a truncated VZV glycoprotein E (gE) antigen, wherein the immunogenic composition comprises about 100 μg of gE antigen and about 3000 μg or about 6000 μg of the oligonucleotide.

7. 7. The method of claim 5 or claim 6, wherein the immunogenic composition comprises about 100 μg of gE antigen and about 3000 μg of oligonucleotide.

8. 7. The method of claim 5 or claim 6, wherein the immunogenic composition comprises about 100 μg of gE antigen and about 6000 μg of oligonucleotide.

9. The method of any one of claims 1 to 8, wherein the oligonucleotide comprises at least one phosphorothioate linkage or all nucleotide linkages are phosphorothioate linkages.

10. The method according to any one of claims 1 to 9, wherein the oligonucleotide is a single-stranded oligodeoxynucleotide.

11. The method of any one of claims 1 to 10, wherein the gE antigen is a recombinant protein lacking the transmembrane and intravirion domains of full-length VZV gE antigen.

12. 12. The method of claim 11, wherein the gE antigen comprises the amino acid sequence of SEQ ID NO:4, or an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

4.

13. 12. The method of claim 11, wherein the gE antigen comprises the amino acid sequence of SEQ ID NO:5, or an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

5.

14. The method according to any one of claims 11 to 13, wherein the recombinant protein is produced in mammalian cells.

15. The method of any one of claims 1 to 14, wherein the immunogenic composition further comprises an aluminium salt adjuvant.

16. 16. The method of claim 15, wherein the aluminum salt adjuvant comprises one or more of the group consisting of amorphous aluminum hydroxyphosphate sulfate, aluminum hydroxide, aluminum phosphate, and potassium aluminum sulfate.

17. 16. The method of claim 15, wherein the aluminum salt adjuvant comprises aluminum hydroxide.

18. The method of any one of claims 15 to 17, wherein the immunogenic composition comprises from about 0.25 mg to about 1.25 mg of Al3+.

19. The method of any one of claims 15 to 17, wherein the immunogenic composition comprises about 0.50 mg, about 0.75 mg, or about 1.00 mg of Al3+.

20. 20. The method of claim 19, wherein the immunogenic composition comprises about 0.75 mg of Al3+.

21. The method of any one of claims 15 to 20, wherein the immunogenic composition further comprises a pharma- ceutically acceptable buffer.

22. 22. The method of claim 21, wherein the buffer is not a phosphate-containing buffer.

23. 22. The method of claim 21 , wherein the buffer is a Tris buffer.

24. The method of any one of claims 1 to 23, wherein the immunogenic composition does not comprise one or more of a saponin, a TLR4 agonist and a liposome.

25. 24. The method of any one of claims 1 to 23, wherein the immunogenic composition does not include one or more of QS21, 3-O-deacylated monophosphoryl lipid A (3D-MPL1), dioleoylphosphatidylcholine and cholesterol.

26. 26. The method of any one of claims 1 to 25, wherein the gE antigen and the oligonucleotide are present in the immunogenic composition in a ratio of 1:15 to 1:120 (wt / wt), optionally wherein the gE antigen and the oligonucleotide are present in the immunogenic composition in a ratio of 1:15 to 1:60 (wt / wt).

27. 27. The method of any one of claims 1 to 26, wherein the gE antigen and the oligonucleotide are present in the immunogenic composition in a ratio of about 1:15, about 1:20, about 1:30, about 1:40, about 1:60, about 1:80 or about 1:120 (wt / wt), optionally wherein the gE antigen and the oligonucleotide are present in the immunogenic composition in a ratio of about 1:30 (wt / wt) or about 1:60 (wt / wt).

28. 28. The method of any one of claims 21 to 27, wherein the oligonucleotide and Al3+ are present in the immunogenic composition in a ratio of 8:1 to 2:1 (wt / wt), optionally wherein the oligonucleotide and Al3+ are present in the immunogenic composition in a ratio of about 8:1 (wt / wt) or about 4:1 (wt / wt).

29. The method of any one of claims 1 to 28, wherein the human subject is 18 years of age or older.

30. The method of any one of claims 1 to 29, wherein the human subject is 50 years of age or older.

31. The method of claim 30, wherein the human subject is between 50 and 69 years old.

32. The method of any one of claims 1 to 31, wherein the human subject is suspected of having a latent VZV infection.

33. The method of any one of claims 1 to 32, wherein the human subject has not been previously vaccinated against chickenpox or shingles.

34. 34. The method of any one of claims 6 to 33, wherein a human subject receives a first dose and a second dose of the immunogenic composition, the second dose being administered one month to one year after the first dose.

35. 35. The method of claim 34, wherein the second dose is administered 2 to 6 months after the first dose.

36. 36. The method of any one of claims 1-5 and 35, wherein the second dose is administered about 2 months after the first dose.

37. The method of any one of claims 1 to 36, wherein the immunogenic composition is administered by intramuscular injection.

38. 38. The method of any one of claims 1 to 37, wherein the increased cell-mediated immunity to VZV or increased immune response to VZV comprises an increased frequency of gE antigen-reactive activated CD4+ T cells, and the activated CD4+ T cells express two or more activation markers selected from interleukin-2, interferon-gamma, tumor necrosis factor-alpha and CD40L (CD154).

39. The method of any one of claims 1 to 38, wherein administration of the immunogenic composition results in increased humoral immunity to VZV in the subject compared to humoral immunity to VZV in the subject prior to administration of the immunogenic composition.

40. 40. The method of claim 39, wherein the increased humoral immunity to VZV comprises an increased concentration of gE antigen-reactive IgG.

41. 41. The method of claim 39 or claim 40, wherein increasing humoral immunity to VZV comprises increasing the concentration of gE and / or VZV neutralizing antibodies.

42. 42. The method of any one of claims 39 to 41, wherein the increased cell-mediated immunity and increased humoral immunity are induced in the subject by about two months after administration of the first dose of the immunogenic composition.

43. 42. The method of any one of claims 39 to 41, wherein the increased cell-mediated immunity and increased humoral immunity are induced in the subject by about one month after administration of the second dose of the immunogenic composition.

44. The method of any one of claims 39 to 43, wherein the increased cell-mediated immunity and increased humoral immunity are maintained in the subject for at least three months following administration of the second dose of the immunogenic composition.

45. 45. The method of any one of claims 39 to 44, wherein administration of the immunogenic composition reduces the risk of the subject developing shingles compared to the risk of a control subject not receiving the immunogenic composition comprising the gE antigen.

46. 45. The method of any one of claims 39-44, wherein administration of the immunogenic composition optionally prevents the subject from developing shingles for a period of at least 3 years following administration of the second dose of the immunogenic composition.

47. 45. The method of any one of claims 39-44, wherein administration of the immunogenic composition optionally prevents the subject from developing postherpetic neuralgia for a period of at least 3 years following administration of the second dose of the immunogenic composition.

48. 48. The method of any one of claims 39 to 47, wherein the immunogenic composition is associated with a reduced risk of development of one or more solicited local reactions within 7 days (the day of vaccination and 6 days thereafter) following administration of the first dose and / or the second dose, compared to a comparative control composition comprising a gE antigen and an AS01B adjuvant.

49. 49. The method of claim 48, wherein the solicited local reaction comprises pain that interferes with normal daily activities.

50. 50. The method of any one of claims 39 to 49, wherein the immunogenic composition is associated with a reduced risk of developing one or more solicited systemic reactions within 7 days (the day of vaccination and 6 days thereafter) following administration of the first dose and / or the second dose, compared to a comparative control composition comprising a gE antigen and an AS01B adjuvant.

51. 51. The method of claim 50, wherein the solicited systemic reaction comprises fatigue, headache and / or muscle pain that interferes with normal daily activities, and / or fever above 39°C.

52. 52. The method of any one of claims 39 to 51, wherein the immunogenic composition is associated with a reduced risk of occurrence of Guillain-Barre Syndrome within 42 days (the day of vaccination and 41 days thereafter) following administration of the first dose and / or the second dose compared to a comparative control composition comprising a gE antigen and an AS01B adjuvant.

53. 1. An immunogenic composition for use in a method for increasing cell-mediated immunity to Varicella Zoster Virus (VZV) in a human subject in need thereof, the method comprising:

1. An immunogenic composition comprising: administering the immunogenic composition to a human subject, the immunogenic composition comprising: i) an unmethylated cytidine-phospho-guanosine (CpG)-containing oligonucleotide comprising the sequence 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO:1), present in the immunogenic composition in an amount effective to increase cell-mediated immunity to VZV compared to cell-mediated immunity to VZV in the subject prior to administration of the immunogenic composition; and ii) a truncated VZV glycoprotein E (gE) antigen, wherein a first dose and a second dose of the immunogenic composition are administered to the human subject, the second dose being administered 2 to 6 months after the first dose.

54. 1. Use of an immunogenic composition in the manufacture of a medicament for increasing cell-mediated immunity against Varicella-Zoster Virus (VZV) in a human subject in need thereof, comprising:

1. The immunogenic composition of claim 1, wherein the immunogenic composition comprises: i) an unmethylated cytidine-phospho-guanosine (CpG)-containing oligonucleotide comprising the sequence 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO:1), present in the immunogenic composition in an amount effective to increase cell-mediated immunity to VZV compared to cell-mediated immunity to VZV in the subject prior to administration of the immunogenic composition; and ii) a truncated VZV glycoprotein E (gE) antigen, wherein a first dose and a second dose of the immunogenic composition are administered to a human subject, the second dose being administered 2 to 6 months after the first dose.

55. 55. The composition or use of claim 53 or claim 54, wherein the immunogenic composition comprises from about 25 μg to about 150 μg of gE antigen and from about 750 μg to about 6000 μg of oligonucleotide.

56. 55. The composition or use of claim 53 or claim 54, wherein the immunogenic composition comprises about 50 μg to about 100 μg of gE antigen and about 1500 μg to about 6000 μg of oligonucleotide.

57. 55. The composition or use of claim 53 or claim 54, wherein the immunogenic composition comprises about 25 μg, about 50 μg, about 75 μg, about 100 μg, about 125 μg or about 150 μg of gE antigen and about 750 μg, about 1500 μg, about 3000 μg or about 6000 μg of oligonucleotide.

58. 58. The composition or use of claim 57, wherein the immunogenic composition comprises about 100 μg of gE antigen and about 3000 μg or 6000 μg of oligonucleotide.

59. 1. An immunogenic composition for use in a method of increasing an immune response to Varicella Zoster Virus (VZV) in a human subject in need thereof, the immunogenic composition comprising: i) an unmethylated cytidine-phospho-guanosine (CpG)-containing oligonucleotide comprising the sequence 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO:1); and ii) a truncated VZV glycoprotein E (gE) antigen, wherein the immunogenic composition comprises about 100 μg of gE antigen and about 3000 μg or about 6000 μg of the oligonucleotide.

60. 1. Use of an immunogenic composition in the manufacture of a medicament for increasing an immune response against Varicella-Zoster Virus (VZV) in a human subject in need thereof, comprising:

1. The immunogenic composition comprising: i) an unmethylated cytidine-phospho-guanosine (CpG)-containing oligonucleotide comprising the sequence 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO:1); and ii) a truncated VZV glycoprotein E (gE) antigen, wherein the immunogenic composition comprises about 100 μg of gE antigen and about 3000 μg or about 6000 μg of the oligonucleotide.

61. The composition or use according to any one of claims 58 to 60, wherein the immunogenic composition comprises about 100 μg of gE antigen and about 3000 μg of oligonucleotide.

62. The composition or use according to any one of claims 58 to 60, wherein the immunogenic composition comprises about 100 μg of gE antigen and about 6000 μg of oligonucleotide.

63. 63. The composition or use according to any one of claims 53 to 62, wherein the oligonucleotide comprises at least one phosphorothioate linkage or all nucleotide linkages are phosphorothioate linkages.

64. The composition or use according to any one of claims 53 to 63, wherein the oligonucleotide is a single-stranded oligodeoxynucleotide.

65. The composition or use of any one of claims 53 to 64, wherein the gE antigen is a recombinant protein lacking the transmembrane and intravirion domains of full-length VZV gE antigen.

66. 66. The composition or use of claim 65, wherein the gE antigen comprises the amino acid sequence of SEQ ID NO:4, or an amino acid sequence which is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

4.

67. 66. The composition or use of claim 65, wherein the gE antigen comprises the amino acid sequence of SEQ ID NO:5, or an amino acid sequence which is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

5.

68. The composition or use according to any one of claims 65 to 67, wherein the recombinant protein is produced in a mammalian cell.

69. The composition or use of any one of claims 53 to 68, wherein the immunogenic composition further comprises an aluminium salt adjuvant.

70. 70. The composition or use of claim 69, wherein the aluminium salt adjuvant comprises one or more of the group consisting of amorphous aluminium hydroxyphosphate sulphate, aluminium hydroxide, aluminium phosphate and potassium aluminium sulphate.

71. 70. The composition or use of claim 69, wherein the aluminium salt adjuvant comprises aluminium hydroxide.

72. 72. The composition or use of any one of claims 69 to 71, wherein the immunogenic composition comprises from about 0.25 mg to about 1.25 mg of Al3+.

73. 72. The composition or use of any one of claims 69 to 71, wherein the immunogenic composition comprises about 0.50 mg, about 0.75 mg or about 1.00 mg of Al3+.

74. 74. The composition or use of claim 73, wherein the immunogenic composition comprises about 0.75 mg of Al3+.

75. The composition or use according to any one of claims 69 to 74, wherein the immunogenic composition further comprises a pharma- ceutically acceptable buffer.

76. 76. A composition or use according to claim 75, wherein the buffer is not a phosphate-containing buffer.

77. 76. The composition or use of claim 75, wherein the buffer is a Tris buffer.

78. 78. The composition or use of any one of claims 53 to 77, wherein the immunogenic composition does not comprise one or more of a saponin, a TLR4 agonist and a liposome.

79. 78. The composition or use of any one of claims 53 to 77, wherein the immunogenic composition does not comprise one or more of QS21, 3-O-deacylated monophosphoryl lipid A (3D-MPL1), dioleoylphosphatidylcholine and cholesterol.

80. 80. The composition or use according to any one of claims 53 to 79, wherein the gE antigen and the oligonucleotide are present in the immunogenic composition in a ratio of 1:15 to 1:120 (wt / wt), optionally wherein the gE antigen and the oligonucleotide are present in the immunogenic composition in a ratio of 1:15 to 1:60 (wt / wt).

81. 81. The composition or use of any one of claims 53 to 80, wherein the gE antigen and the oligonucleotide are present in the immunogenic composition in a ratio of about 1:15, about 1:20, about 1:30, about 1:40, about 1:60, about 1:80 or about 1:120 (wt / wt), optionally wherein the gE antigen and the oligonucleotide are present in the immunogenic composition in a ratio of about 1:30 (wt / wt) or about 1:60 (wt / wt).

82. 82. The composition or use according to any one of claims 75 to 81, wherein the oligonucleotide and Al3+ are present in the immunogenic composition in a ratio of 8:1 to 2:1 (wt / wt), optionally wherein the oligonucleotide and Al3+ are present in the immunogenic composition in a ratio of about 8:1 (wt / wt) or about 4:1 (wt / wt).

83. 83. The composition or use according to any one of claims 53 to 82, wherein the human subject is aged 18 years or older.

84. 84. The composition or use according to any one of claims 53 to 83, wherein the human subject is 50 years of age or older.

85. 85. The composition or use of claim 84, wherein the human subject is aged between 50 and 69 years.

86. The composition or use according to any one of claims 53 to 85, wherein the human subject is suspected of having a latent VZV infection.

87. 87. The composition or use of any one of claims 53 to 86, wherein the human subject has not been previously vaccinated against chickenpox or shingles.

88. 88. The composition or use of any one of claims 59 to 87, wherein a human subject receives a first dose and a second dose of the immunogenic composition, the second dose being administered one month to one year after the first dose.

89. 89. The composition or use of claim 88, wherein the second dose is administered 2 to 6 months after the first dose.

90. 90. The composition or use of any one of claims 53 to 58 and 89, wherein the second dose is administered about 2 months after the first dose.

91. The composition or use according to any one of claims 53 to 90, wherein the immunogenic composition is administered by intramuscular injection.

92. 92. The composition or use of any one of claims 53 to 91, wherein the increased cell-mediated immunity to VZV or increased immune response to VZV comprises an increased frequency of gE antigen-reactive activated CD4+ T cells, wherein the activated CD4+ T cells express two or more activation markers selected from interleukin-2, interferon-gamma, tumor necrosis factor-alpha and CD40L (CD154).

93. A composition or use according to any one of claims 53 to 92, wherein administration of the immunogenic composition results in increased humoral immunity to VZV in the subject compared to humoral immunity to VZV in the subject prior to administration of the immunogenic composition.

94. 94. The composition or use of claim 93, wherein the increased humoral immunity to VZV comprises an increased concentration of gE antigen-reactive IgG.

95. 95. The composition or use of claim 93 or claim 94, wherein the increased humoral immunity to VZV comprises an increased concentration of gE and / or VZV neutralizing antibodies.

96. 96. The composition or use of any one of claims 93 to 95, wherein an increase in cell-mediated immunity and an increase in humoral immunity are induced in the subject by about 2 months after administration of the first dose of the immunogenic composition.

97. 96. The composition or use of any one of claims 93 to 95, wherein an increase in cell-mediated immunity and an increase in humoral immunity are induced in the subject by about one month after administration of the second dose of the immunogenic composition.

98. 98. The composition or use of any one of claims 93 to 97, wherein the increased cell-mediated immunity and increased humoral immunity are maintained in the subject for at least 3 months after administration of the second dose of the immunogenic composition.

99. 99. The composition or use of any one of claims 93 to 98, wherein administration of the immunogenic composition reduces the risk of the subject developing shingles compared to the risk of a control subject not receiving the immunogenic composition comprising the gE antigen.

100. 99. The composition or use of any one of claims 93 to 98, wherein administration of the immunogenic composition optionally prevents the subject from developing shingles for a period of at least 3 years following administration of the second dose of the immunogenic composition.

101. 99. The composition or use of any one of claims 93 to 98, wherein administration of the immunogenic composition optionally prevents the subject from developing post-herpetic neuralgia for a period of at least 3 years following administration of the second dose of the immunogenic composition.

102. 102. The composition or use of any one of claims 93 to 101, wherein the immunogenic composition is associated with a reduced risk of occurrence of one or more solicited local reactions within 7 days (the day of vaccination and 6 days thereafter) following administration of the first and / or second dose compared to a comparative control composition comprising gE antigen and AS01B adjuvant.

103. 103. A composition or use as described in claim 102, wherein the solicited local reaction comprises pain that interferes with normal daily activities.

104. 104. The composition or use of any one of claims 93 to 103, wherein the immunogenic composition is associated with a reduced risk of occurrence of one or more solicited systemic reactions within 7 days (the day of vaccination and 6 days thereafter) following administration of the first dose and / or the second dose compared to a comparative control composition comprising a gE antigen and an AS01B adjuvant.

105. 105. The composition or use of claim 104, wherein the solicited systemic reaction comprises fatigue, headache and / or muscle pain that interferes with normal daily activities, and / or fever above 39°C.

106. 106. The composition or use of any one of claims 93 to 105, wherein the immunogenic composition is associated with a reduced risk of occurrence of Guillain-Barre Syndrome within 42 days (the day of vaccination and 41 days thereafter) following administration of the first and / or second dose compared to a comparative control composition comprising gE antigen and AS01B adjuvant.

107. 1. An immunogenic composition comprising: i) an unmethylated cytidine-phospho-guanosine (CpG)-containing oligonucleotide comprising the sequence 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO:1); ii) a truncated VZV glycoprotein E (gE) antigen; and iii) at least one excipient, An immunogenic composition, wherein the gE antigen is a recombinant protein produced in a mammalian cell and is present in the immunogenic composition in an amount of about 25 μg to about 150 μg, the oligonucleotide is present in the immunogenic composition in an amount of about 750 μg to about 6000 μg, and the at least one excipient comprises a pharma- ceutically acceptable buffer.

108. 1. An immunogenic composition comprising: i) an unmethylated cytidine-phospho-guanosine (CpG)-containing oligonucleotide comprising the sequence 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO:1); ii) a truncated VZV glycoprotein E (gE) antigen; and iii) at least one excipient, An immunogenic composition, wherein the gE antigen is a recombinant protein produced in a mammalian cell and is present in the immunogenic composition in an amount of about 100 μg, the oligonucleotide is present in the immunogenic composition in an amount of about 3000 μg or about 6000 μg, and at least one excipient comprises a pharma- ceutically acceptable buffer.

109. 109. The immunogenic composition of claim 107 or claim 108, comprising about 100 μg of gE antigen and about 3000 μg of oligonucleotide.

110. 109. The immunogenic composition of claim 107 or claim 108, comprising about 100 μg of gE antigen and about 6000 μg of oligonucleotide.

111. The immunogenic composition of any one of claims 107 to 110, wherein the gE antigen comprises the amino acid sequence of SEQ ID NO:4, or an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

4.

112. The immunogenic composition of any one of claims 107 to 110, wherein the gE antigen comprises the amino acid sequence of SEQ ID NO:5, or an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

5.

113. The immunogenic composition of any one of claims 107 to 112, further comprising an aluminum hydroxide adjuvant, wherein the immunogenic composition comprises from about 0.25 mg to about 1.25 mg Al3+.

114. The immunogenic composition of claim 113, comprising about 0.75 mg of Al3+.

115. The immunogenic composition of any one of claims 107 to 114, wherein the buffer is not a phosphate-containing buffer.

116. The immunogenic composition of any one of claims 107 to 115, wherein the buffer is a Tris buffer.

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