Liposomal adjuvant compositions for epstein BARR virus vaccines

Abstract

The present disclosure provides a vaccine composition that comprises an Epstein Barr Virus (EBV) polypeptide and a liposomal adjuvant, and methods of inducing an immune response to an Epstein Barr Virus (EBV) or methods of preventing infection of or reducing the likelihood of infection by an Epstein Barr Virus (EBV) using the compositions, or a combination of the EBV polypeptide and the liposomal adjuvant.

Description

LIPOSOMAL ADJUVANT COMPOSITIONS FOR EPSTEIN BARR VIRUS VACCINESCROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of U. S. Provisional Patent Application Serial No. 63 / 733,174 filed December 12, 2024, the entire contents of which are incorporated by reference herein.REFERENCE TO SEQUENCE LISTING SUBMITTED ELECTRONICALLY

[0002] The contents of the electronic sequence listing (26101-WO_SL.xml; Size: 45.252 bytes; and Date of Creation: June 2, 2025) are herein incorporated by reference in their entirety.FIELD OF THE INVENTION

[0003] The invention relates generally to the prevention of Epstein Barr Virus (EBV) disease. More specifically, the invention relates to a composition comprising EBV polypeptides and a liposomal adjuvant, which can be administered as a vaccine. The liposomal adjuvant comprises a glycolipid, such as a glucopyranosyl lipid A (“GLA”) and a saponin, such as QS-21. Further provided are methods of using the disclosed compositions and formulations.BACKGROUND

[0004] Epstein Barr virus (EBV) infects about 95% of the adult population worldwide and has been known to be associated with two B-cell lymphomas, Burkitt’s and Hodgkin’s lymphomas. The virus can also infect epithelial cells and is associated with nasopharyngeal cancer.Furthermore, EBV causes most cases of infectious mononucleosis in developed countries, affecting mainly children and young adults. Infectious mononucleosis can result in a long recovery period of up to one month. There are currently no approved vaccines on the market, so there is a strong need for a preventive vaccine. Moreover, there is also a need to use adjuvants to increase the immunogenicity of the EBV vaccine.SUMMARY OF THE INVENTION

[0005] The invention provides a composition comprising: (a) an Epstein Barr Virus (EBV) polypeptide comprising one or more polypeptides selected from the group consisting of: an EBV gp42 polypeptide, an EBV gH polypeptide, and an EBV gL polypeptide; and (b) a liposomal adjuvant, wherein the liposomal adjuvant comprises a glycolipid and a saponin.

[0006] The invention further provides a composition comprising an EBV gp220 polypeptide; and a liposomal adjuvant, wherein the liposomal adjuvant comprises a glycolipid; and a saponin.

[0007] The invention further provides a pharmaceutical unit dose comprising about 5 pg to about 200 pg of EBV polypeptide consisting of any one of the amino acid sequences of SEQ ID NOs: 11-21, and about 0.1 mg to about 3 mg of a liposomal adjuvant, wherein the liposomal adjuvant comprises l,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIAacceptable salt thereof.

[0008] The invention further provides a pharmaceutical unit dose comprising about 5 pg to about 200 pg of EBV polypeptide consisting of the amino acid sequence of SEQ ID NO: 23, and about 0.1 mg to about 3 mg of a liposomal adjuvant, wherein the liposomal adjuvant comprises 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21)and the glycolipid of Formula I1IA.acceptable salt thereof.

[0009] The invention further provides a pharmaceutical unit dose comprising about 10 pg to about 100 pg of a first EBV polypeptide consisting of the amino acid sequence of SEQ ID NO: 21 and about 10 pg to about 100 pg of a second EBV polypeptide consisting of the amino acid sequence of SEQ ID NO: 23, wherein the first and the second EBV polypeptide are at a 1: 1 weight ratio; and about 0.27 mg to about 1.35 mg of a liposomal adjuvant, wherein the liposomal adjuvant comprises l,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIAacceptable salt thereof, at a ratio of 10-30:2-15: 1-5: 1-3 by weight.

[0010] The invention further provides a method of inducing an immune response to an Epstein Barr Virus (EBV) in a human patient comprising administering to the patient the composition or pharmaceutical unit dose of the invention.

[0011] The invention further provides a method of inducing an immune response to an Epstein Barr Virus (EBV) in a human patient comprising administering to the patient an EBV polypeptide described herein and the liposomal adjuvant described herein.

[0012] The disclosure further provides a method of preventing infection of or reducing the likelihood of infection of a human patient by an Epstein Barr Virus (EBV) comprising administering to the patient the composition or pharmaceutical unit dose of the invention.

[0013] The disclosure further provides a method of preventing infection of or reducing the likelihood of infection of a human patient by an Epstein Barr Virus (EBV) comprising26101administering to the patient an EBV polypeptide described herein and the liposomal adjuvant described herein.

[0014] The disclosure further provides a method of preventing cancer caused by an Epstein Barr Virus (EBV) comprising administering to the patient the composition or pharmaceutical unit dose of the invention.

[0015] The disclosure further provides a method of preventing cancer caused by an Epstein Barr Virus (EBV) comprising administering to the patient an EBV polypeptide described herein and the liposomal adjuvant described herein.

[0016] The invention also provides a kit comprising: (1) an Epstein Barr Virus (EBV) polypeptide; and (2) a liposomal adjuvant described herein.DEFINITIONS

[0017] As used throughout the specification and in the appended claims, the singular forms “a,” '“an,” and ““the” include the plural reference unless the context clearly dictates otherwise.

[0018] As used throughout the specification and appended claims, the following abbreviations and definitions apply:EBV Epstein Barr VirusID intradermalIM intramuscularMw molecular weightNMWCO nominal molecular weight cut offPS-20 polysorbate-20PS-80 polysorbate-80VLP(s) virus-like particle(s) (Protein antigens)w / v weight per volume

[0019] As used throughout the specification and appended claims, the following definitions and abbreviations apply:

[0020] AAHS: As used herein, the term “‘AAHS” refers to an amorphous aluminum hydroxyphosphate sulfate adjuvant.

[0021] About: As used herein, the term "about", when modifying the quantity (e.g., mM, or M) of a substance or composition, the percentage (v / v or w / v) of a formulation component, the pH of a solution / formulation, or the value of a parameter characterizing a step in a method, or the like refers to variation in the numerical quantity up to ± 10% rounded up where appropriate that can occur, for example, through typical measuring, handling and sampling procedures involved in thepreparation, characterization and / or use of the substance or composition; through instrumental error in these procedures; through differences in the manufacture, source, or purity of the ingredients employed to make or use the compositions or carry out the procedures; and the like. In certain embodiments, "about" can mean a variation of ± 0.1%, 0.5%. 1%, 2%, 3%, 4%, 5%, or 10%.

[0022] Adjuvant: As used herein, the term “adjuvant” refers to a composition or compound that is capable of enhancing the immune response against an antigen of interest. Adjuvants are substances or combinations of substances that are used in conjunction with a vaccine antigen to enhance (e.g., increase, accelerate, prolong and / or possibly target) the specific immune response to the vaccine antigen or modulate to a different type (e.g., switch a Thl immune response to a Th2 response, or a humoral response to a cytotoxic T cell response) in order to enhance the clinical effectiveness of the vaccine. In some embodiments, the adjuvant modifies (Thl / Th2) the immune response. In some embodiments, the adjuvant boosts the strength and longevity of the immune response. In some embodiments, the adjuvant broadens the immune response to a concomitantly administered antigen. In some embodiments, the adjuvant is capable of inducing strong antibody and T cell responses. In some embodiments, the adjuvant is capable of increasing the polyclonal ability of the induced antibodies. In some embodiments, the adjuvant is used to decrease the amount of antigen necessary to provoke the desired immune response and provide protection against the disease. In some embodiments, the adjuvant is used to decrease the number of injections needed in a clinical regimen to induce a durable immune response and provide protection against the disease. Adjuvant containing formulations described herein may demonstrate enhancements in humoral and / or cellular immunogenicity of vaccine antigens.

[0023] Administration: As used herein, the term “administration” or “administering” refers to the act of providing an active agent, composition, or formulation to a subject. Exemplary routes of administration to the human body can be through the eyes (ophthalmic), mouth (oral), skin (transdermal), nose (nasal), lungs (inhalant), oral mucosa (buccal), or ear; can be rectal or vaginal,; can be by injection (e.g., intravenously (IV). subcutaneously, intratumorally, intraperitoneally, intramuscularly (IM), or intradermally (ID) etc.) and the like.

[0024] Agent: As used herein, the term “agent” refers to a particle, compound, molecule, or entity of any chemical class including, for example, a Virus-like Particle (VLP), a small molecule, polypeptide (e.g., a protein), polynucleotide (e.g., a DNA polynucleotide or an RNA polynucleotide), saccharide, lipid, or a combination or complex thereof. In some embodiments, the term “agent” can refer to a compound, molecule, or entity that includes a polymer, or a plurality thereof.

[0025] Alkenyl: As used herein, the term "alkenyl’’ refers to a straight chain, cyclic or branched unsaturated aliphatic hydrocarbon having the specified number of carbon atoms. In one embodiment, an alkenyl group contains from 8 to 24 carbon atoms (C8-C24 alkenyl). In one embodiment, an alkenyl group is linear. In another embodiment, an alkenyl group is branched. In another embodiment the alkenyl group is unsubstituted.

[0026] Alkyl: As used herein, the term "alkyl” refers to a straight chain, cyclic or branched saturated aliphatic hydrocarbon having the specified number of carbon atoms. In one embodiment, an alkyl group contains from 8 to 24 carbon atoms (C8-C24 alkyl). In one embodiment, an alkyl group is linear. In another embodiment, an alkyd group is branched. In another embodiment the alkyl group is unsubstituted.

[0027] Antibody: As used herein, the term “antibody” (or “Ab”) refers to any form of antibody that exhibits the desired biological activity. Thus, it is used in the broadest sense and specifically covers, but is not limited to, monoclonal antibodies (including full length monoclonal antibodies), polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies), humanized, fully human antibodies, and chimeric antibodies.

[0028] Antigen: As used herein, the term “antigen” refers to any antigen that can generate one or more immune responses. The antigen may be a protein (including recombinant proteins), VLP, polypeptide, or peptide (including synthetic peptides). The antigen may be one that generates a humoral and / or CTL immune response.

[0029] API: As used herein, the term “API” refers to an active pharmaceutical ingredient, e.g., EBV protein antigens (such as the EBV polypeptide disclosed herein), which is a component of the compositions or formulations disclosed herein that is biologically active (e.g., capable of inducing an appropriate immune response) and confers a therapeutic or prophylactic benefit to a person or animal in need thereof. As used herein, an API is a vaccine active ingredient.

[0030] EBV polypeptide: As used herein, refers to a polypeptide comprising all or part of an amino acid sequence encoded by EBV, and optionally a ferritin. Similarly, gL, gH, gp42, and gp220 polypeptides refer to polypeptides comprising all or part of a gL, gH, gp42, or gp220 amino acid sequence, respectively, encoded by EBV. Polypeptides with, e.g., at least 80% identity to an EBV-encoded polypeptide will necessarily comprise part of the EBV-encoded polypeptide. The terms "gL polypeptide," "gH polypeptide," "gp42 polypeptide," and "gp220 polypeptide" are used interchangeably with " EBV gL polypeptide," " EBV gH polypeptide," " EBV gp42 polypeptide," and " EBV gp220 polypeptide," respectively. Immunization with an EBV polypeptide as part or all of an antigenic polypeptide may confer protection from infection with EBV. Unless the context dictates otherwise, any polypeptide disclosed herein comprising anEBV polypeptide can comprise all or part of multiple sequences encoded by EBV (for example, all or part of gL and gEI of EBV, or all or part of gL, gH, and gp42 of EBV).

[0031] Monomer: As used herein, a "monomer," or "monomer construct" refers to a construct expressed as a single-chain protein. A monomer, for example, may comprise gL and gH of EBV expressed in a single chain, or gL, gH. and gp42 of EBV expressed in a single chain.

[0032] Trimer: As used herein, a "trimer," or "trimer construct" refers to a construct comprising gL and / or gH of EBV together with a trimerization domain, such as a foldon trimerization domain derived from T4 phage fibritin. Other trimerization domains, such as the human collagen XVIII trimerization domain (see, e.g., Alvarez-Cienfuegos et al., Scientific Reports 2016;6:28643) and the LlORFlp trimerization domain (see, e.g., Khazina et al.. Proc Natl Acad Sci U S A 2009 Jan 12; 106(3):731-36) are also known in the art and can be used in trimeric constructs.

[0033] Ferritin: " Ferritin" or "ferritin protein," as used herein, refers to a protein with detectable sequence identity to H. pylori ferritin or another ferritin discussed herein, such as P. furiosus ferritin, Trichoplusia ni ferritin, or human ferritin, that serves to store iron, e.g., intracellularly or in tissues or to carry iron in the bloodstream. Such exemplary ferritins, including those that occur as two polypeptide chains, known as the heavy and light chains (e.g., T. ni and human ferritin), are discussed in detail below. In some embodiments, a ferritin comprises a sequence with at least 75%. 80%. 85%. 90%. 95%. 97%. 98%. 99%. or 99.5% identity to a ferritin sequence disclosed herein, e.g., in Table 2 (Sequence Table). A ferritin may be a fragment of a full-length naturally-occurring sequence.

[0034] Wild-type ferritin: " Wild-type ferritin," as used herein, refers to a ferritin whose sequence consists of a naturally-occurring sequence. Ferritins also include full-length ferritin or a fragment of ferritin with one or more differences in its amino acid sequence from a wild-type ferritin.

[0035] Ferritin monomer: As used herein, a "ferritin monomer" refers to a single ferritin molecule (or. where applicable, a single ferritin heavy or light chain) that has not assembled with other ferritin molecules. A "ferritin multimer" comprises multiple associated ferritin monomers. A "ferritin protein" includes monomeric ferritin and multimeric ferritin.

[0036] Ferritin particle: As used herein, "ferritin particle," refers to ferritin that has selfassembled into a globular form. Ferritin particles are sometimes referred to as "ferritin nanoparticles" or simply "nanoparticles". In some embodiments, a ferritin particle comprises 24 ferritin monomers (or, where applicable, 24 total heavy and light chains).

[0037] Hybrid ferritin: " Hybrid ferritin," as used herein, refers to ferritin comprising H. pylori ferritin with an amino terminal extension of bullfrog ferritin. An exemplary’ sequence used as an26101amino terminal extension of bullfrog ferritin appears as SEQ ID NO: 24, 25 or 26. In hybrid ferritin, the amino terminal extension of bullfrog ferritin can be fused to H. pylori ferritin such that immune-stimulatory moiety attachment sites are distributed evenly on the ferritin particle surface. Hybrid ferritin is also sometimes referred to as "bfpFerr" or "bfp ferritin."

[0038] Immune response: " Immune response," as used herein, refers to a response of a cell of the immune system, such as a B cell, T cell, dendritic cell, macrophage or polymorphonucleocyte, to a stimulus such as an antigen or vaccine. An immune response can include any cell of the body involved in a host defense response, including for example, an epithelial cell that secretes an interferon or a cytokine. An immune response includes, but is not limited to, an innate and / or adaptive immune response. As used herein, a "protective immune response" refers to an immune response that protects a subject from infection (e.g., prevents infection or prevents the development of disease associated with infection). Methods of measuring immune responses are well known in the art and include, for example, by measuring proliferation and / or activity of lymphocytes (such as B or T cells), secretion of cytokines or chemokines, inflammation, antibody production and the like. An "antibody response" is an immune response in which antibodies are produced.

[0039] Antigen: As used herein, an "antigen" refers to an agent that elicits an immune response, and / or an agent that is bound by a T cell receptor (e.g., when presented by an MHC molecule) or to an antibody (e.g., produced by a B cell) when exposed or administered to an organism. In some embodiments, an antigen elicits a humoral response (e.g., including production of antigenspecific antibodies) in an organism. Alternatively, or additionally, in some embodiments, an antigen elicits a cellular response (e.g., involving T-cells whose receptors specifically interact with the antigen) in an organism. A particular antigen may elicit an immune response in one or several members of a target organism (e.g., mice, rabbits, primates, humans), but not in all members of the target organism species. In some embodiments, an antigen elicits an immune response in at least about 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%. 90%. 91%. 92%. 93%. 94%. 95%. 96%. 97%. 98%. 99% of the members of a target organism species. In some embodiments, an antigen binds to an antibody and / or T cell receptor, and may or may not induce a particular physiological response in an organism. In some embodiments, for example, an antigen may bind to an antibody and / or to a T cell receptor in vitro, whether or not such an interaction occurs in vivo. In some embodiments, an antigen reacts with the products of specific humoral or cellular immunity, including those induced by heterologous immunogens. Antigens include antigenic ferritin proteins comprising ferritin (e.g., comprising one or more mutations) and a non-ferritin polypeptide as described herein.

[0040] Antigenic EBV polypeptide: An "antigenic EBV polypeptide" is used herein to refer to a polypeptide comprising all or part of an EBV amino acid sequence of sufficient length that the molecule is antigenic with respect to EBV. Antigenicity may be a feature of the EBV sequence as part of a construct further comprising a heterologous sequence, such as a ferritin or lumazine synthase protein and / or immune-stimulatory moiety. That is, if an EBV sequence is part of a construct further comprising a heterologous sequence, then it is sufficient that the construct can serve as an antigen that generates anti-EBV antibodies, regardless of whether the EBV sequence without the heterologous sequence could do so.

[0041] Antigenic ferritin polypeptide: As used herein. " Antigenic ferritin polypeptide" and "antigenic ferritin protein" are used interchangeably herein to refer to a polypeptide comprising a ferritin and anon-ferritin polypeptide (e.g., an EBV polypeptide) of sufficient length that the molecule is antigenic with respect to the non-ferritin polypeptide. Antigenicity7may be a feature of the non-ferritin sequence as part of the larger construct. That is, it is sufficient that the construct can serve as an antigen against the non-ferritin polypeptide, regardless of whether the non-ferritin polypeptide without the ferritin could do so. In some embodiments, the non-ferritin polypeptide is an EBV polypeptide, in which case the antigenic ferritin polypeptide is also an "antigenic EBV polypeptide." To be clear, however, an antigenic EBV polypeptide does not need to comprise ferritin. " Antigenic polypeptide" is used herein to refer to a polypeptide which is either or both of an antigenic ferritin polypeptide and an antigenic EBV polypeptide.

[0042] Sequence Identity7: As used herein, " Sequence identity" between two amino acid sequences indicates the percentage of amino acids that are identical between the sequences.

[0043] % Identity: The terms "% identical", "% identity" or similar terms are intended to refer, in particular, to the percentage of nucleotides or amino acids which are identical in an optimal alignment between the sequences to be compared. Said percentage is purely statistical, and the differences between the two sequences may be but are not necessarily randomly distributed over the entire length of the sequences to be compared. Comparisons of two sequences are usually earned out by comparing said sequences, after optimal alignment, with respect to a segment or "window of comparison", in order to identify local regions of corresponding sequences. The optimal alignment for a comparison may be carried out manually or with the aid of the local homology algorithm by Smith and Waterman, 1981, Ads App. Math. 2, 482, with the aid of the local homolog}7algorithm by Needleman and Wunsch, 1970, J. Mol. Biol. 48, 443. with the aid of the similarity search algorithm by Pearson and Lipman, 1988, Proc. Natl Acad. Sci. USA 88, 2444, or with the aid of computer programs using said algorithms (GAP, BESTFIT, FASTA,BLAST P, BLAST N and TFASTA in Wisconsin Genetics Software Package, Genetics Computer Group, 575 Science Drive, Madison, Wis.).

[0044] Percentage identity is obtained by determining the number of identical positions at which the sequences to be compared correspond, dividing this number by the number of positions compared (e.g., the number of positions in the reference sequence) and multiplying this result by 100.

[0045] In some embodiments, the degree of identity is given for a region which is at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90% or about 100% of the entire length of the reference sequence. For example, if the reference nucleic acid sequence consists of 200 nucleotides, the degree of identity is given for at least about 100, at least about 120, at least about 140, at least about 160, at least about 180, or about 200 nucleotides, in some embodiments in continuous nucleotides. In some embodiments, the degree of identity is given for the entire length of the reference sequence.

[0046] Nucleic acid sequences or amino acid sequences having a particular degree of identity to a given nucleic acid sequence or amino acid sequence, respectively, may have at least one functional property of said given sequence, e.g., and in some instances, are functionally equivalent to said given sequence. One important property includes the ability to act as a cytokine, in particular when administered to a subject. In some embodiments, a nucleic acid sequence or amino acid sequence having a particular degree of identity to a given nucleic acid sequence or amino acid sequence is functionally equivalent to said given sequence.

[0047] As used herein, the term "kit" refers to a packaged set of related components, such as one or more compounds or compositions and one or more related materials such as solvents, solutions, buffers, instructions, or desiccants.

[0048] Co-administration: As used herein, the term “co-administration’’ or “co-administering” in relation to the liposomal adjuvant and a pharmaceutical formulation (e.g., an EBV polypeptide) refers to administration of a liposomal adjuvant and a pharmaceutical formulation (e.g., an EBV polypeptide) concurrently, i.e., simultaneously in time, or sequentially, i.e., administration of an EBV polypeptide followed by administration of a liposomal adjuvant (or vice versa). That is, after administration of the EBV polypeptide (or liposomal adjuvant), the liposomal adjuvant (or EBV polypeptide) can be administered substantially immediately after the EBV polypeptide (or liposomal adjuvant) or the liposomal adjuvant (or the EBV polypeptide) can be administered after an effective time period after the EBV polypeptide (or liposomal adjuvant); the effective time period is the amount of time period is generally within 1, 2, 3, 5, 10, 15, 20, 25, 30, 45, or 60 minutes.

[0049] Composition: As used herein, the term “composition” refers to a formulation containing an active pharmaceutical or biological ingredient (for example, at least one protein antigen of Epstein Barr Virus (EBV) and a liposomal adjuvant, along with one or more additional components. The term “composition” is used interchangeably with “pharmaceutical composition” and “formulation.” The compositions can be liquid or solid (e.g.. lyophilized). Additional components that may be included as appropriate include pharmaceutically acceptable excipients, additives, diluents, buffers, sugars, amino acids, chelating agents, surfactants, polyols, bulking agents, stabilizers, lyo-protectants, solubilizers, emulsifiers, salts, adjuvants, tonicity enhancing agents, delivery vehicles, and anti-microbial preservatives. Compositions are nontoxic to recipients at the dosages and concentrations employed.

[0050] Consists essentially7of: As used herein, the term," consists essentially of’ or variations such as "consist essentially of' or "consisting essentially of," as used throughout the specification and claims, indicate the inclusion of any recited elements or group of elements, and the optional inclusion of other elements, of similar or different nature than the recited elements, that do not materially change the basic or novel properties of the specified dosage regimen, method, or composition. As a non-limiting example, an EBV polypeptide that consists essentially of a recited amino acid sequence may also include one or more amino acids, including substitutions of one or more amino acid residues, that do not materially affect the properties of the EBV polypeptide.

[0051] Dose: As used herein, the term “dose” means a quantity of an agent, API, formulation, or pharmaceutical composition administered or recommended to be administered at a particular time.

[0052] Immunogenic: As used herein, the term “immunogenic” or “immunogenicity” refers to the ability of an antigen to provoke an immune response in a subject. The term “immunogenic composition” refers to the ability' of an agent, API, formulation, or composition to provoke an immune response in a subject.

[0053] In need of treatment: Those “in need of treatment” include those previously exposed to or infected with Epstein Barr Virus, those who were previously vaccinated against Epstein Barr Virus, as well as those prone to have an infection or any person in which a reduction in the likelihood of infection is desired, e.g., the immunocompromised, the elderly, children, adults, or healthy individuals.

[0054] Lipid: As used herein, the term “lipid” refers to any of a group of organic compounds that are esters of fatty acids and are characterized by being insoluble in water or having low solubility' in water but may be soluble in many organic solvents. Lipids can be divided in at least26101three classes: (1) "simple lipids,” which include, e.g., fats and oils as well as waxes; (2) “compound lipids,” which include, e.g., phospholipids and glycolipids; and (3) “derived lipids,” which include, e.g., steroids.

[0055] Patient: As used herein, the term “patient” refers to any human being that is to receive the EBV vaccines, or pharmaceutical compositions, described herein. As defined herein, “patient” includes those already infected with EBV as well as those in which infection with EBV is to be prevented.

[0056] Pharmaceutically acceptable: refers to excipients (vehicles, additives) and compositions that can reasonably be administered to a subject to provide an effective dose of the active ingredient employed and that are "generally regarded as safe" e.g., that are physiologically tolerable and do not typically produce an allergic or similar untoward reaction, such as gastric upset and the like, when administered to a human. In another embodiment, this term refers to molecular entities and compositions approved by a regulatory agency of the federal or a state government or listed in the U. S. Pharmacopeia or another generally recognized pharmacopeia for use in animals, and more particularly in humans.

[0057] Pharmaceutical composition: As used herein, the term “pharmaceutical composition,” refers to a composition containing an active pharmaceutical or biological ingredient, along with one or more additional components, e.g., a composition in which an active agent is formulated together with one or more pharmaceutically acceptable carriers. As used herein, the terms “pharmaceutical formulation” and “formulation” are used interchangeably with “pharmaceutical composition.” In some embodiments, the active agent is present in a pharmaceutical unit dose amount appropriate for administration in a therapeutic regimen that shows a statistically significant probability of achieving a predetermined therapeutic effect when administered to a relevant population. The pharmaceutical compositions or formulations can be liquid or solid (e.g., lyophilized). Additional components that may be included as appropriate include pharmaceutically acceptable excipients, additives, diluents, buffers, sugars, amino acids, chelating agents, surfactants, polyols, bulking agents, stabilizers, lyo-protectants. solubilizers, emulsifiers, salts, adjuvants, tonicity enhancing agents, delivery vehicles, and anti-microbial preservatives. The pharmaceutical compositions or formulations are nontoxic to recipients at the dosages and concentrations employed. In some embodiments, a pharmaceutical composition can be specially formulated for administration in solid or liquid form, including those adapted for the following: oral administration, for example, drenches (aqueous or non-aqueous solutions or suspensions), tablets, e.g., those targeted for buccal, sublingual, and systemic absorption, boluses, powders, granules, pastes for application to the tongue; parenteral administration, for example,26101by subcutaneous, intramuscular, intravenous or epidural injection as, for example, a sterile solution or suspension, or sustained-release formulation; topical application, for example, as a cream, ointment, or a controlled-release patch or spray applied to the skin, lungs, or oral cavity; intravaginally or intrarectally, for example, as a pessary, cream, or foam; sublingually; ocularly; trans dermally; or nasally, pulmonary, and to other mucosal surfaces.

[0058] Subject: As used herein, the term “subject’’ refers to an organism, typically a mammal (e.g., a human, in some embodiments including prenatal human forms). In some embodiments, a subject is suffering from a relevant disease, disorder or condition. In some embodiments, a subject is susceptible to a disease, disorder, or condition. In some embodiments, a subject displays one or more symptoms or characteristics of a disease, disorder or condition. In some embodiments, a subject does not display any symptom or characteristic of a disease, disorder, or condition. In some embodiments, a subject is someone with one or more features characteristic of susceptibility to or risk of a disease, disorder, or condition. In some embodiments, a subject is a patient. In some embodiments, a subject is an individual to whom diagnosis and / or therapy is and / or has been administered.

[0059] Therapeutically Effective Amount: As used herein, the term “therapeutically effective amount” (or “therapeutically effective dose”) refers to an amount of the active ingredient (e.g., therapeutic protein, vaccine, or antibody) sufficient to produce the desired therapeutic effect in a human or animal, e.g., the amount necessary to elicit an immune response, treat, cure, prevent, or inhibit development and progression of a disease or the symptoms thereof and / or the amount necessary' to ameliorate symptoms or cause regression of a disease. Therapeutically effective amount may vary’ depending on the structure and potency of the active ingredient and the contemplated mode of administration. One of skill in the art can readily determine a therapeutically effective amount of a given antibody or therapeutic protein or vaccine antigen.

[0060] Vaccine: As used herein, the term “vaccine” or “vaccine composition” refers to a substance or preparation used to stimulate the production of antibodies and provide immunity against one or several diseases, prepared from the causative agent of a disease, its products, or a synthetic substitute, treated to act as an antigen without inducing the disease. A vaccine composition may include at least one antigen in a pharmaceutically acceptable vehicle useful for inducing an immune response in a subject. The vaccine composition is administered by doses and techniques known to those skilled in the pharmaceutical or veterinary fields, taking into account factors such as the age, sex, weight, species, and condition of the recipient animal and the route of administration.26101BRIEF DESCRIPTION OF THE DRAWINGS

[0061] Figure 1: Shown are the interpolated serum antibody titers against the EBV antigens gH / gL of individual animals at day 42. The horizontal bar represents the Geometric Mean Titers (GMT). Error bars represent the 95% confidence interval of the GMT.

[0062] Figure 2: Shown are the interpolated serum antibody titers against the EBV antigens gp350 of individual animals at day 42. The horizontal bar represents the GMT. Error bars represent the 95% confidence interval of the GMT.

[0063] Figure 3: Show n are the interpolated serum antibody titers against the EBV antigens gp42 of individual animals at day 42. The horizontal bar represents the GMT. Error bars represent the 95% confidence interval of the GMT.

[0064] Figure 4: Shown are the NT50 values on EBV infection of B cells and epitheli al dike cells at day 42. The horizontal bar represents the geometric mean NT50 values. Error bars represent the 95% confidence interval of the Geometric Mean NT50.

[0065] Figure 5: Rhesus monkeys (5 per group) were injected intramuscularly with 3 doses of gp350-FNP and gL-gH-gp42-FNP dual particle vaccine formulated with liposomal adjuvant B (Adjuvant B) at weeks 0, 4, and 10 (indicated by arrows). Shown are the interpolated serum antibody titers against the EBV antigens gH / gL, gp42 or gp350, through week 42. The data are presented as geometric mean titers. Error bars represent the 95% confidence interval of the GMT.

[0066] Figure 6: Rhesus monkeys (5 per group) were injected intramuscularly with 3 doses of gp350-FNP and gL-gH-gp42-FNP dual particle vaccine formulated with liposomal adjuvant B (Adjuvant B) at weeks 0, 4, and 10 (indicated by arrows). Shown are the NT50 values on EBV infection of B cells and epithelial-like cells through week 42. The data are presented as geometric mean titers. Error bars represent the 95% confidence interval of the Geometric Mean NT50.

[0067] Figure 7: CD4 T cell cytokine responses (IFN-y, IL-2 or TNF-a) to gH or gp350 stimulation determined by ICS assay. The dotted lines represent individual animals, and the solid lines represent the group median. The horizontal dotted line represents the assay detection limit.

[0068] Figure 8: CD4 T cell responses to gH peptide stimulation determined by ICS assay. Spleens from 4 immunized animals per group were harvested at day 42 and pooled. The dotted line represents the assay baseline level.DETAILED DESCRIPTION

[0069] In one aspect, the invention provides a composition comprising:26101(a) an Epstein Barr Virus (EBV) polypeptide comprising one or more polypeptides selected from the group consisting of: an EBV gp42 polypeptide, an EBV gH polypeptide, and an EBV gL polypeptide; and(b) a liposomal adjuvant, wherein the liposomal adjuvant comprises a glycolipid and a saponin.The Liposomal Adjuvant

[0070] In specific embodiments, the liposomal adjuvant of the present invention is in a multicomponent composition composed of structural lipids (such as l,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) and cholesterol) in combination with a glycolipid (e.g., Glucopyranosyl Lipid A) and saponin molecules (such as, e.g., Quillaja Saponaria 21 (QS-21).Structural Lipids

[0071] In an embodiment, a combination of lipids are included to provide structure for the liposomal adjuvant. In an embodiment, a phospholipid provides structure for the liposomal adjuvant. In an embodiment, a sterol provides structure for the liposomal adjuvant. In an embodiment, a phospholipid and a sterol provide structure for the liposomal adjuvant.

[0072] In an embodiment, the liposomal adjuvant may include a phospholipid. In an embodiment, the phospholipid is DOPC. In an embodiment, the liposomal adjuvant may include a phospholipid selected from: phospholipids, aminolipids and sphingolipids. In some embodiments, the structural lipid may include a phospholipid selected from: phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, phosphatidylinositol, phosphatidic acid, palmitoyloleryl phosphatidylcholine, lysophosphatidylcholine, lysophosphatidylethanolamine, dipalmitoylphosphatidylcholine, dioleoylphospbatidylcholme, dstearoylphosphatidylcholine and dilinoleoylphosphatidylcholine. In some embodiments, the liposomal adjuvant may include a neutral lipid selected from: sphingolipid, glycosphingolipid families, diacylglycerols and S-acyloxyacids. In some embodiments, the liposomal adjuvant may include a neutral lipid selected from: phosphatidylcholine (PC), phosphatidylethanolamine (PE), and phosphatidylglycerol (PG), phosphatidylserine (PS), phosphatidylinositol (PI), phosphatidic acid (phosphatidate) (PA), dipalmitoylphosphatidylcholine, monoacyl-phosphatidylcholine (lyso PC), l-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), N-acyl-PE, phosphoinositides, and phosphosphingolipids. In some embodiments, the structural lipid may include a neutral lipid selected from: phosphatidic acid (DMPA, DPPA, DSPA), phosphatidylcholine (DDPC, DLPC, DMPC, DPPC, DSPC, DOPC, POPC, DEPC), phosphatidylglycerol (DMPG, DPPG, DSPG, POPG), phosphatidylethanolamine (DMPE, DPPE, DSPE DOPE), and phosphatidylserine (DOPS). In26101some embodiments, the liposomal adjuvant may include a neutral lipid selected from: fatty acids (C14:0), palmitic acid (C16:0), stearic acid (C18:0), oleic acid (C 18: 1), linoleic acid (C18:2), linolenic acid (C18:3), arachidonic acid (C20:4), C20:0, C22:0 and lecithin. In some embodiments, the phospholipid may include l,2-Distearoyl-sn-glycero-3 -phosphocholine (DSPC).

[0073] In some embodiments, the liposomal adjuvant includes 40-90 mole% phospholipid. In some embodiments, the liposomal adjuvant includes 45-85 mole% phospholipid. In some embodiments, the liposomal adjuvant includes 50-85 mole% phospholipid. In some embodiments, the liposomal adjuvant includes 55-75 mole% phospholipid. In some embodiments, the liposomal adjuvant includes 60-70 mole% phospholipid. In some embodiments, the liposomal adjuvant includes 60-65 mole% phospholipid. In some embodiments, the liposomal adjuvant includes about 65 mole% phospholipid. In an embodiment, the phospholipid is DOPC.

[0074] In some embodiments, the liposomal adjuvant includes about 60 pg / mL to about 300 mg / mL phospholipid. In some embodiments, the liposomal adjuvant includes about 80 pg / mL to about 50 mg / mL phospholipid. In some embodiments, the liposomal adjuvant includes about 1 mg / mL to about 10 mg / mL phospholipid. In some embodiments, the liposomal adjuvant includes about 2.5 mg / mL to about 3.5 mg / mL phospholipid. In some embodiments, the liposomal adjuvant includes about 3.0 mg / mL phospholipid. In an embodiment, the phospholipid is DOPC.

[0075] In some embodiments, the liposomal adjuvant may include a sterol, such as cholesterol. In some embodiments, the structural lipid includes cholesterol. In some embodiments, the liposomal adjuvant includes 10-40 mole% cholesterol. In some embodiments, the liposomal adjuvant includes 15-25 mole% cholesterol. In some embodiments, the liposomal adjuvant includes 10-20 mole% cholesterol. In some embodiments, the liposomal adjuvant includes 20-30 mole% cholesterol. In some embodiments, the liposomal adjuvant includes 10-15 mole% cholesterol. In some embodiments, the liposomal adjuvant includes 25-35 mole% cholesterol. In some embodiments, the liposomal adjuvant includes 30-35 mole% cholesterol. In some embodiments, the liposomal adjuvant includes about 30 mole% cholesterol.

[0076] In some embodiments, the liposomal adjuvant includes about 0.05 pg / mL to about 10 mg / mL sterol. In some embodiments, the liposomal adjuvant includes about 0.2 pg / mL to about 3.0 mg / mL sterol. In some embodiments, the liposomal adjuvant includes about 0.25 mg / mL to about 2.0 mg / mL sterol. In some embodiments, the liposomal adjuvant includes about 0.5 mg / mL to about 1.5 mg / mL sterol. In some embodiments, the liposomal adjuvant includes about 0.75 mg / mL sterol. In an embodiment, the sterol is cholesterol.26101Glycolipid

[0077] In an embodiment, the liposomal adjuvant composition includes a glycolipid having the structure set forth in Formula I;OHFormula Iwherein Ri is P(O)(OH)2;wherein R2 is selected from H, C(O)CH2CH(OH)Cn alkyl and C(O)CH2CH(OC(O)Cn- C13 alkyljCn alkyd;wherein R3is C(O)CH2CH(OC(O)Cn-Ci3 alkyljCn alkyl;wherein R4 is selected from H. C(O)CH2CH(OH)Cn alkyl, and C(O)CH2CH(OC(O)Cn- C13 alkyljCn alkyl;wherein R5 is selected from C(O)CH2CH(OH)Cn alkyl and C(O)CH2CH(OC(O)Ci3-Cis alkyljCn alkyl; andwherein Re is H;or a pharmaceutically acceptable salt thereof.

[0078] In an embodiment, the glycolipid has the structure set forth in Formula II;Formula IIwherein Ri is H or C(O)Cn-Ci3 alkyl;wherein R2 is Cn alkyl;26101wherein R3 is C11-C13 alkyl;wherein R4 is C11 alkyl;wherein R5 is H or C(0)CH2CH(0H)Cn alkyl;wherein Re is H or C(O)Ci3-Cis alkyl; andwherein R7 is Cn alkyl;or a pharmaceutically acceptable salt thereof.

[0079] In an embodiment, the glycolipid has the structure set forth in Formula IIA;Formula IIAwherein Ri is H or C(O)Cn-Ci3 alkyl;wherein R2 is Cn alkyl;wherein R3 is C11-C13 alkyl;wherein R4 is Cn alkyd;wherein R5 is H or C(O)CH2CH(OH)Cn alkyl;wherein Re is H or C(O)Ci3-Ci5 alkyl; andwherein R7 is Cn alkyl;or a pharmaceutically acceptable salt thereof.In one embodiment under Formula II or IIA.Ri is C(O)Cn-Ci3 alkyl;R2 is Cn alkyl;R3 is C11-C13 alkyl;R+ is Cn alkyl;R5is C(O)CH2CH(OH)Cn alkyl;Re is H; andR7 is Cn alkyl.26101

[0080] In an embodiment, the glycolipid has the structure set forth in Formula III:Formula IIIor a pharmaceutically acceptable salt thereof.

[0081] In an embodiment, the glycolipid has the structure set forth in Formula IIIA:26101or a pharmaceutically acceptable salt thereof.

[0082] In an embodiment, the glycolipid has the structure set forth in Formula IV:26101Formula IV,or a pharmaceutically acceptable salt thereof.

[0083] In an embodiment, the glycolipid has the structure set forth in Formula IVA:26101or a pharmaceutically acceptable salt thereof.

[0084] In an embodiment, the glycolipid has the structure set forth in Formula V:26101Formula V,or a pharmaceutically acceptable salt thereof.

[0085] In an embodiment, the glycolipid has the structure set forth in Formula VI:26101Formula VI,or a pharmaceutically acceptable salt thereof.

[0086] In one aspect under any one of the Formulas I-VI, the pharmaceutically acceptable salt is an ammonium, sodium, potassium, calcium, or organic amine salt such as an ethylamine, diethylamine, or triethylamine salt. In one aspect under any one of Formulas I-VI, the pharmaceutically acceptable salt is an ammonium salt.

[0087] In some embodiments, the liposomal adjuvant includes 0.25-20 mole% glycolipid. In some embodiments, the liposomal adjuvant includes 0.5-10 mole% glycolipid. In some embodiments, the liposomal adjuvant includes 0.25-5 mole% glycolipid. In some embodiments, the liposomal adjuvant includes 0.25-2.5 mole% glycolipid. In some embodiments, the liposomal adjuvant includes 0.5-2.25 mole% glycolipid. In some embodiments, the liposomal adjuvant includes 0.5-2.0 mole% glycolipid. In some embodiments, the liposomal adjuvant includes 0.75-1.90 mole% glycolipid. In some embodiments, the liposomal adjuvant includes 0.95-1.85 mole% glycolipid. In some embodiments, the liposomal adjuvant includes 1.0-1.75 mole% glycolipid. In some embodiments, the liposomal adjuvant includes about 1.44 mole% glycolipid. In an embodiment, the glycolipid is GLA.

[0088] In some embodiments, the liposomal adjuvant includes about 0.01 pg / mL to about 10 mg / mL glycolipid. In some embodiments, the liposomal adjuvant includes about 0.02 pg / mL to about 3.0 mg / mL glycolipid. In some embodiments, the liposomal adjuvant includes about 0.03 mg / mL to about 2.0 mg / mL glycolipid. In some embodiments, the liposomal adjuvant includes about 0.05 mg / mL to about 1.0 mg / mL glycolipid. In some embodiments, the liposomal adjuvant includes about 0.15 mg / mL glycolipid. In an embodiment, the glycolipid is GLA.

[0089] In an embodiment, the glycolipid is selected from the structure set forth in Formula III, Formula IV, Formula V, Formula VI, a pharmaceutically acceptable salt thereof or combinations thereof. In an embodiment, the glycolipid is the structure set forth in Formula IIIA. Formula IVA. a pharmaceutically acceptable salt thereof, or combinations thereof. In an embodiment, the glycolipid is the structure set forth in Formula IIIA, a pharmaceutically acceptable salt thereof, or combinations thereof.Saponins

[0090] In an embodiment, the liposomal adjuvant includes a saponin. In an embodiment, the saponin is a derivative of Quillaja saponaria Molina quil A, suitably an immunologically active fraction of Quil A, such as QS7, QS17, QS18 or QS-21, in particular QS-21. In an embodiment, the Quil A saponin is selected from QS7, QS17. QS18, QS-21, and combinations thereof. In an embodiment, the saponin is selected from QS7, QS-21, and combinations thereof. In one embodiment, the saponin used is selected from QS-17, QS-18, QS-21, and combinations thereof. In an embodiment, the saponin is QS-21.

[0091] Commercially available QS-21 purified from Quillaja saponaria Molina tree bark can comprise one or more of QS-21-H, QS-21-Apiose and QS-21-xylose as depicted below:In one embodiment, the QS-21 comprises QS-21-H. QS-21-Apiose and QS-21-xylose. In another embodiment, the QS-21 comprises QS-21-Apiose and QS-21-xylose. See N. E. Jacobsen et al.. Carbohydr. Res., 280 (1996) 1-14.

[0092] In an embodiment, the saponin, such as Quil A and in particular QS-21, is at least 90% pure, such as at least 95% pure, especially at least 98% pure, or in particular 99% pure.

[0093] In some embodiments, the liposomal adjuvant includes 0.25-20 mole% saponin. In some embodiments, the liposomal adjuvant includes 0.5-10 mole% saponin. In some embodiments, the liposomal adjuvant includes 0.25-5 mole% saponin. In some embodiments, the liposomal adjuvant includes 0.25-2.5 mole% saponin. In some embodiments, the liposomal adjuvant includes 0.5-2.25 mole% saponin. In some embodiments, the liposomal adjuvant includes 0.5-2.0 mole% saponin. In some embodiments, the liposomal adjuvant includes 0.75-1.75 mole% saponin. In some embodiments, the liposomal adjuvant includes 0.95-1.5 mole% saponin. In some embodiments, the liposomal adjuvant includes 1.0-1.5 mole% saponin. In some embodiments, the liposomal adjuvant includes about 1.27 mole% saponin. In an embodiment, the saponin is QS-21.

[0094] In some embodiments, the liposomal adjuvant includes about 0.01 pg / mL to about 10 mg / mL saponin. In some embodiments, the liposomal adjuvant includes about 0.02 pg / mL to about 3.0 mg / mL saponin. In some embodiments, the liposomal adjuvant includes about 0.03 mg / mL to about 2.0 mg / mL saponin. In some embodiments, the liposomal adjuvant includes about 0.05 mg / mL to about 1.0 mg / mL saponin. In some embodiments, the liposomal adjuvant includes about 0.15 mg / mL saponin. In an embodiment, the saponin is QS-21.Buffer

[0095] In some embodiments, the liposomal adjuvant may include a buffer. In some embodiments, the buffer may be selected from any pharmaceutically acceptable buffer, including acetic acid, histidine, citrate, Bis-Tris, HEPES, phosphate, MES, disodium hydrogen phosphate dihydrate, potassium dihydrogen phosphate, salt, and combinations thereof. In some embodiments, the buffer may be present in an amount of 1 mM to about 100 mM.

[0096] In some embodiments, the liposomal adjuvant includes about 60 pg / mL to about 300 mg / mL buffer. In some embodiments, the liposomal adjuvant includes about 80 pg / mL to about 50 mg / mL buffer. In some embodiments, the liposomal adjuvant includes about 1 mg / mL to about 25 mg / mL buffer. In some embodiments, the liposomal adjuvant includes about 2.5 mg / mL to about 20 mg / mL buffer. In some embodiments, the liposomal adjuvant includes about 13.0 mg / mL buffer. In an embodiment, the buffer includes about 1.0 mg / mL to about 20 mg / mL disodium hydrogen phosphate dihydrate, about 1.0 mg / mL to about 20 mg / mL potassium dihydrogen phosphate, and about 1.0 mg / mL to about 20 mg / mL sodium chloride. In an embodiment, the buffer includes about 1.25 mg / mL to about 10 mg / mL disodium hydrogen26101phosphate di hydrate, about 2.5 mg / mL to about 15 mg / mL potassium dihydrogen phosphate, and about 2.5 mg / mL to about 15 mg / mL sodium chloride. In an embodiment, the buffer includes about 1.5 mg / mL to about 5 mg / mL disodium hydrogen phosphate dihydrate, about 3.5 mg / mL to about 10 mg / mL potassium dihydrogen phosphate, and about 3.5 mg / mL to about 10 mg / mL sodium chloride. In an embodiment, the buffer includes about 1.6 mg / mL disodium hydrogen phosphate dihydrate, about 5.6 mg / mL potassium dihydrogen phosphate, and about 5.8 mg / mL sodium chloride.General Methods of Making the Liposomal Adjuvant

[0097] In specific embodiments, the liposomal adjuvant of the present invention is a multicomponent composition composed of structural lipids (such as l,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) and cholesterol) in combination with a glycolipid (e.g., Glucopyranosyl Lipid A) and one or more saponin molecules (such as, e.g., Quillaja Saponaria 21 (QS-21). In an embodiment, the liposomal adjuvant is prepared by first solubilizing the structural lipids (e.g., DOPC, cholesterol) and glycolipid (e.g., GLA) at desired mass ratios (for example, see Table 1) in an organic solvent (such as ethanol, chloroform, isopropyl alcohol) to form a lipidic mixture. The organic solvent is then removed from the lipidic mixture by heat and / or a vacuum driven drying procedure (e.g.. a rotatory’ evaporation) to form a dry’ thin lipidic film. In an embodiment, the thin lipidic film is then reconstituted with an aqueous buffer (e.g., histidine, sodium or potassium phosphate buffer, and combinations thereof) to trigger the formation of a poly disperse mixture of liposomes. In an embodiment, the poly dispersity of liposome mixture is then reduced by passage of liposomes under high pressure (e.g., using an extrusion device through a defined pore size membrane) or high shear (such as, e.g., microfluidizer through a narrow chamber) to achieve a defined liposome particle size distribution, lamellarity. In an embodiment, the formed liposomes are then mixed with one or more saponin molecules (such as QS-21) dissolved in an aqueous matrix (such as, e.g., histidine buffers, or phosphate buffers, and combinations thereof), which leads to binding of the saponin molecules to the cholesterol component of the liposomes. In an embodiment, the resulting structural lipid / glycolipid / saponin liposome formulation is then sterile filtered to form the liposomal adjuvant. Several process parameters within each step, including but not limited to lipid dissolution (mixing time, pH, temperature, concentration of components), solvent removal (temperature, pressure, time), lipid film hydration (time, temperature), and liposome sizing (pressure, membrane pore size, flow rate, # of passes) may be controlled to yield a liposomal adjuvant with desired attributes.26101Table 1: Composition of a Representative Liposomal AdjuvantTarget Mass ContentConcentration ratio of Each MolecularContent of Each Lipid Component and range and Lipid and Weight(mg / mL) and range (Mole%)ranges range (g / mol)(Mass %)Phospholipid 2.25-3.75 10-30 55.56- 786.11 48.38-80.6392.5913.89- Sterol 0.56-0.94 2-15 23.15 386.65 24.59-40.98 Glycolipid 0.12-0.18 0-3 2.78-4.63 1763.46 1.08-1.80Saponin 0.12-0.18 0-5 2.78-4.63 1990.13 0.96-1.595-300mMBuffer Matrix buffer, at pH N / A5.0-8.0Lipid Stock Solution

[0098] In an embodiment, the structural lipids (for example, DOPC and cholesterol) (with or without GLA) are dissolved in an organic solvent e.g., methanol, ethanol chloroform, isopropyl alcohol (IP A)) or a mixture of solvents (e.g., 80:20 ethanol: IP A; 90:10 chloroform: methanol) to desired concentrations, ranging from 10-30 mg / mL DOPC, 2.5-10 mg / mL cholesterol and 0.5-5 mg / mL GLA. The stock solutions are then prepared separately for all 3 components or prepared as combined stock solutions containing 1-3 of the components based on ease of manufacturing. Lipid dissolution parameters such as starting mass and ratios of lipids (micrograms- grams), solvent volume mixing speeds( 100-600 rpm), temperature (40-60° C) and time (30 min- 3hrs) are ranged appropriately to achieve complete lipid dissolution towards targeted scale.

[0099] In an embodiment, 100-200 mg DOPC and 20-100 mg cholesterol are together dissolved in 1-50 mL ethanol by heating at 30-60°C for 1-25 minutes to complete dissolution; 1-50 mg GLA is then dissolved in 1-50 mL ethanol at 30-100 C for 1-60 minutes with sonication to complete dissolution. In an embodiment, each lipid is dissolved individually, i.e., 1-50 mg / mL DOPC is dissolved in chloroform, 1-50 mg / mL cholesterol is dissolved in chloroform, and 1-50 mg / mL GLA is dissolved in a 9:1 mixture of chloroform: methanol under ambient temperatures with swirl mixing. In an embodiment, all lipid components (i.e., 1-50 g DOPC, 1-25 g cholesterol, 0.25-25 g GLA), were dissolved together in 1-10 L ethanol by mixing at 100-500 RPM at temperatures (30-70° C) for 60-120 mins.

[0100] In one embodiment, 150 mg DOPC and 37.7 mg cholesterol were together dissolved in 5 mL ethanol by heating at 40° C for 5 min to complete dissolution. 11 mg GLA was then dissolved in 11 mL ethanol at 60°C for 25 min with sonication to complete dissolution. In one embodiment, each lipid was dissolved individually under ambient temperatures with swirl26101mixing. 25 mg / mL DOPC was dissolved in chloroform, lOmg / mL cholesterol was dissolved in chloroform, and 5 mg / mL GLA was dissolved in a 9:1 mixture of chloroform: methanol. In one embodiment, the lipid components (36 g DOPC, 9 g Cholesterol, 1.8 g GLA), were dissolved together in 3.6L ethanol by mixing at 350 RPM at temperatures (50°C) for 120 mins.Preparation of Thin-lipidic Film

[0101] In an embodiment, thin lipidic films are prepared via dehydration of the lipid stock solution described above. In an embodiment, the individual lipid stock solutions are combined at atarget mass ratio of DOPC: cholesterol: GLA (such as a20:5:l ratio) or prepared at the target ratios. In an embodiment, the lipid stock solution is then dehydrated using a rotatory evaporator to make a dry thin lipid film. In an embodiment, the dehydration parameters such as starting volumes of lipid mix volumes (10 mL-3.6 L), rotatory’ speed (40-280 rpm) temperature (ambient - 50°C) and time (40 min-8 hrs) are ranged appropriately to achieve complete dehydration.

[0102] In an embodiment, the rotary evaporation of 11 mL lipid mix is performed at ambient temperature (e.g., 23°C), pressure 22 mbar and rotary speed 150 rpm for 40 mins to form a thin lipid film on the inner wall of the round bottom flask. In an embodiment, the dry ing time range from (40 minutes - 16 hours). In an embodiment, the rotatory evaporation is performed at room temperature at 280 rpm and pressure is reduced from 600 mbar to 15 mbar at a rate of 100 mbar / 15-30 mins and a total drying time of approximately 1-5 hours. In an embodiment, the rotatory evaporation is performed at room temperature (e.g., about 23°C) to about 50° C at 40-110 rpm speed with -0.07 to 0. IMPa vacuum for a total of about 4-10 hours. In an embodiment, the dehydrated thin film is stored under vacuum or purged with nitrogen gas at ambient temperature for approximately 5-25 hours to ensure the complete removal of residual ethanol.Preparation of Liposomes

[0103] In an embodiment, the dried lipidic film described above is then reconstituted in an aqueous buffer (e.g.. histidine buffer, sodium buffer, or potassium phosphate buffer) to form liposomes. Rehydration parameters such as temperature, volume, mixing speeds and buffers are ranged.

[0104] In an embodiment, the lipid film is rehydrated with a buffer at ambient temperature of about 23 C to about 50°C with constant mixing to form liposomes with 1-15 mg / mL phospholipid, 0.25-2.25 mg / mL sterol and 0.05-1.25 mg / mL glycolipid concentration. In an embodiment, the lipid film is rehydrated with buffer at ambient temperature of about 23 °C to about 50°C with manual swirling to form liposomes with 1-15 mg / mL phospholipid, 0.25-2.2526101mg / mL sterol and 0.05-1.25 mg / mL glycolipid concentration. In an embodiment the lipid thin film is resuspended with a buffer at ambient temperature of about 23 °C to about 50°C over 30 minutes with 80 rpm mixing to form liposomes having the target of 1-15 mg / mL phospholipid, 0.25-2.25mg / mL sterol and 0.05-1.25mg / mL glycolipid.

[0105] In an embodiment, the lipid film is rehydrated with 30 mL of 1 mM (Na / K) phosphate. pH 6.2, 100 mM NaCl buffer at (ambient-50°C) with constant mixing to form liposomes with 5 mg / mL DOPC, 1.25mg / mL cholesterol and 0.25 mg / mL GLA concentration. In an embodiment, the lipid film is rehydrated with 20 mL of 10 mM His pH 6.2, 325 mM NaCl, 0.01% PS80 buffer at 50°C with manual swirling to form liposomes with 6 mg / mL DOPC, 1.5 mg / mL cholesterol and 0.3 mg / mL GLA concentration. In an embodiment, the lipid thin film is resuspended with a 5.4 L of 50 mM (Na / K) phosphate, pH 6.2, 100 mM NaCl buffer at 50°C over 30 minutes with 80 rpm mixing to form liposomes having the target of 6.7 mg / mL DOPC, 1.7 mg / mL cholesterol and 0.3 mg / mL GLA concentration.

[0106] In an embodiment, the poly disperse (in terms of particle size, lamellarity) mixture of liposomes are subjected to pressure or flow driven extrusion through a defined pore sized membrane or high shear (such as microfluidizer through a narrow chamber) to achieve a defined liposome particle size distribution, lamellarity. A step-wise liposome sizing procedure where extrusion starts by passing liposomes through a large (400 nm) pore size membrane and step-wise decreases to smaller pore size membrane 200 nm,100 nm and 80 nm to achieve target liposome size is also used. Process parameters such as pressure, flow rate, membrane pore size, temperature and number of passes are ranged to achieve the optimal size range and lamellarity.

[0107] In an embodiment, the rehydrated liposomes are extruded in batch mode, through an extruder device with a 100 nm pore size membrane under a pressure from 200-400 psi at ambient -50° C and passed 6 times to achieve a low- poly dispersity (poly dispersity index, PDI —0.1), lamellarity(~2) and -120 nm mean particle size. In an embodiment, the extrusion is performed under constant flow rate (3 L / min) through an extruder device with a 100 nm pore size membrane at 40 C temperature and passed 35 times through the membrane to achieve low poly dispersity (PDI -0.1), lamellarity (-2), mean particle size-100 nm. In an embodiment, the liposomes are extruded via a continuous mode of extrusion under constant flow rate at -3.5 L / min through an extruder device with a 100 nm pore size membrane at 40°C for a total of 30 minutes to achieve low polydispersity(PDI -0.1), lamellarity -2 and particle size -100 nm.

[0108] In an embodiment, a buffer including 30 mL of 50 mM (Na / K) phosphate, pH 6.2, lOOmM NaCl buffer is used to rehydrate lipid film at (ambient-50°C) with constant mixing to get 5 mg / mL DOPC, 1.25 mg / mL cholesterol and 0.25 mg / mL GLA concentration. The rehydrated26101liposomes are extruded in batch mode, through an extruder device with a 100 nm pore size membrane under a pressure from 200-400 psi at ambient -50°C and passed 6 times to achieve a low poly dispersity (poly dispersity index, PDI -0.1), lamellarity (~2) and a -120 nm mean particle size.

[0109] In an embodiment, a buffer including 20 mL of 10 mM His pH 6.2, 325 mM NaCl, 0.01% PS80 buffer is used to rehydrate lipid fdm at 50°C with manual swirling to obtain 6 mg / mL DOPC, 1.5 mg / mL Cholesterol and 0.3 mg / mL GLA concentration. Extrusion is performed under constant flow rate (3 L / min) through an extruder device with a 100 nm pore size membrane at 40°C temperature and passed 35 times through the membrane to achieve low poly dispersity (PDI -0.1), lamellarity (-2), and a mean particle size- 100 nm.

[0110] In an embodiment, the lipid thin fdm is resuspended with 5.4 L of 50 mM (Na / K) phosphate, pH 6.2, 100 mM NaCl buffer at 50°C over 30 minutes with 80 rpm mixing to the target of 6.7 mg / mL DOPC, 1.7 mg / mL cholesterol and 0.3 mg / mL GLA concentration.Continuous mode of extrusion of the mixture is performed under constant flow rate at -3.5 L / min through an extruder device with a 100 nm pore size membrane at 40°C for a total of 30 minutes to achieve low poly dispersity (PDI -0.1), lamellarity -2 and particle size -100 nm.Liposomal Adjuvant Formation

[0111] In an embodiment, the final liposomal adjuvant is formed by first dissolving saponin in an aqueous buffer and mixing the resulting solution with the liposomes described above. The liposomal adjuvant is formed by saponin binding to the cholesterol component in the liposomes. The dissolution conditions such as mass of saponin, buffers, volume and mixing time and temperature are optimized to ensure complete dissolution and stability of saponin.

[0112] In an embodiment, approximately 1-100 mL of saponin stock was prepared at ambient condition with a range of 1 to 10 mg / mL saponin concentration in a buffer solution with gentle manual swirl. In an embodiment, the buffer solution includes, e.g., approximately 50 mM (Na / K) phosphate, at pH 6.2 and approximately 100 mM NaCl buffer or approximately 20 mM histidine, pH 6.2 buffer. In an embodiment, up to IL of saponin solution are prepared with stir plate at 200-500 rpm mixing speed to assist complete dissolution.

[0113] In an embodiment, 2 g QS-21 is dissolved in 800 mL of 50 mM (Na / K) phosphate, pH 6.2, 100 mM NaCl formulation buffer at room temperature with stir mixing at 500 rpm for 50 minutes to prepare the QS-21 stock solution. In an embodiment, a 5 mg QS-21 is dissolved in 5 mL of 20 mM histidine, pH 6.2 formulation buffer at room temperature with gentle swirl. In an26101embodiment 5 mg QS-21 was dissolved in 0.5 mL of 50 mM (Na / K) phosphate, pH 6.2, 100 rnM NaCl buffer at room temperature with gentle swirl.

[0114] In an embodiment, saponin and the extruded liposomes are mixed with a targeted ratio of saponin: cholesterol at 1:5 to form a liposomal solution, the mixing parameters such as rate of addition and mixing speed, and time are ranged. In an embodiment, the mixing volume is adjusted to achieve the targeted adjuvant concentration. In an embodiment, a low volume (example < 10 mL) of saponin stock solution is added dropwise with a volumetric pipette with gentle mix, while high volume (10 mL to IL) is added with a peristaltic pump (10-20 ml / min) to mix with the extruded liposome solution at 100-500 rpm stir mixing depending on the total solution volume. In an embodiment, 0.8mL of 10 mg / mL QS-21 stock solution was added with pipette and mixed thoroughly with 30 mL of extruded liposomes and 20 mL buffer (50 mM (Na / K) phosphate, pH 6.2, 100 mM NaCl). In an embodiment, the calculated volume 630g of QS-21 stock solution and 4000 g formulation buffer is combined with the 5000 g extruded liposome at 20 mL / min with 150 rpm mixing, and then stir-mixed thoroughly at room temperature at 120 rpm for at least 30 minutes as the final adjuvant formulation (composition as described in Table 1) before filtration.

[0115] In an embodiment, after the liposomal solution is sterile filtered passed through a 0.22pm filter. In an embodiment, 0.22 pm syringe filters were used to filter ~ 60 mL of liposomal solution. In another instance, large capacity capsule filters units were connected to a peristatic pump to sterile filter approximately 10L of liposomal solution with less than 25 psi pressure throughout the filtration step.EB K Polypeptides

[0116] The EBV polypeptide used in the composition and methods of the invention are described in Section A-G as follows:A. EBV polypeptides comprising gL and gH polypeptides

[0117] EBV has three glycoproteins, glycoprotein B (gB), gH, and gL, that form the core membrane fusion machinery to allow viral penetration into a cell. gL and gH have been previously described, for example, in Matsuura et al., Proc Natl Acad Sci U S A. 2010 Dec 28; 107(52):22641-6. Monomers and trimers of gL and gH for use as vaccines have been described, for example, in Cui et al., Vaccine. 2016 Jul 25; 34(34):4050-5. The gH and gL proteins associate to form a heterodimeric complex considered necessary for efficient membrane fusion and binding to epithelial cell receptors required for viral entry'.26101

[0118] In one embodiment, the compositions or methods of the invention comprise antigenic EBV polypeptides comprising EBV gL and / or EBV gH. In some embodiments, the polypeptide exists as a single-chain. In some embodiments, the polypeptide forms atrimer, e.g., through trimerization of a trimerization domain, such as a T4 phage fibritin trimerization domain. In some embodiments, the polypeptide forms a nanoparticle (e.g., ferritin or lumazine synthase particle), e.g., through multimerization of a ferritin or lumazine synthase.

[0119] In some embodiments, an antigenic EBV polypeptide according to this disclosure comprises an EBV gL polypeptide and an EBV gH polypeptide, and a linker having a length of at least 15 amino acids separating the EBV gL polypeptide and the EBV gH polypeptide.

[0120] In some embodiments, the EBV gH and / or gL polypeptides are full-length gH and / or gL (for exemplary full-length sequences, see GenBank Accession Nos. CEQ35765.1 and YP_001129472.1, respectively). In some embodiments, the EBV gH and / or gL polypeptides are fragments of gH and / or gL. In some embodiments, the gL polypeptide is a gL(D7) construct with a 7-amino acid deletion at the end of the gL C terminus.

[0121] In some embodiments, the EBV gL polypeptide comprises an amino acid sequence with at least 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 4. In some embodiments, the EBV gH polypeptide comprises an amino acid sequence with at least 80%, 85%. 90%. 95%. 97%. 98%. 99%. or 100% identity to SEQ ID NO: 5.

[0122] In particular embodiments, the EBV polypeptide comprises an EBV polypeptide comprising an EBV gL polypeptide, and an EBV gH polypeptide, wherein the EBV gL polypeptide comprises the sequence of residues 1-115 of SEQ ID NO: 21 and the EBV gH polypeptide comprises the sequence of residues 162-823 of SEQ ID NO: 21. In such embodiments, the EBV gL polypeptide, the EBV gH polypeptide, and the ferritin are arranged in N-terminal to C-terminal order within the EBV polypeptide (i.e., the gL polypeptide is N-terminal to the gH polypeptide, which is N-terminal to the ferritin). In some such embodiments, a linker separates the EBV gL polypeptide and the EBV gH polypeptide and comprises the amino acid sequence of SEQ ID NO: 7.

[0123] Native EBV gH and / or gL sequences are shown in GenBank Accession No.NC_009334.1 (Human herpesvirus 4, complete genome, dated 26-Mar-2010). For some of the constructs disclosed herein, amino acids 23-137 of the gL amino acid sequence in NC_009334.1 was used as the gL polypeptide). For some of the constructs, amino acids 19-678 of the gH amino acid sequence in NC_009334.1 was used as the gH polypeptide. In some embodiments, the gL and gH were linked via a linker as shown in the table of sequences herein.26101

[0124] In some embodiments, gL and gH polypeptides are expressed as a single-chain monomer. A single-chain comprising gL and gH polypeptides may be referred to as "gL-gH". In some embodiments, gL and gH are provided as a trimer. In some embodiments, a trimerization domain is placed after (C -terminal to) the gH sequence. The fold on trimerization domain is exemplary, as any trimerization domain known in the art can be used, such as collagen or LI ORF Ip trimerization domains referenced herein. A gL and gH trimer has been shown to induce higher serum neutralization titers relative to a gL and gH monomer using peripheral blood human naive B cells (see, for example, Cui et al., Vaccine. 2016 Jul 25; 34(34):4050-5).

[0125] The gL-gH polypeptide can be combined with any of the ferritins. For example, in some embodiments, an antigenic EBV polypeptide comprises a monomer or trimer gL-gH polypeptide (+ / - gp42 and / or gp220) and i) a heavy or light chain ferritin (e.g., T. ni heavy or light chain ferritin); or ii) a ferritin. In one embodiment the ferritin has a sequence of SEQ ID NO: 24. In one embodiment the ferritin has a sequence of SEQ ID NO: 25. In one embodiment the ferritin has a sequence of SEQ ID NO: 26.

[0126] Additionally, in some embodiments, any antigenic EBV polypeptide comprising an EBV gL-gH polypeptide and a ferritin can be present in a composition comprising another polypeptide disclosed herein. Some such embodiments further comprise a second EBV polypeptide comprising an EBV gp220 polypeptide and ferritin, and the second EBV polypeptide comprises, consists essentially of, or consists of the amino acid sequence of SEQ ID NO: 23. In a particular embodiment, the second EBV polypeptide comprises an amino acid sequence having at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 23.B. EBV polypeptides comprising a gp42 polypeptide

[0127] In some embodiments, an antigenic EBV polypeptide comprises a gp42 polypeptide. An exemplary gp42 sequence is provided as SEQ ID NO: 1. A further exemplary gp42 sequence, suitable for inclusion in fusions e.g., with gL and gH polypeptides, is provided as SEQ ID NO: 2. Another exemplary gp42 sequence, suitable for inclusion in fusions e.g., with gL and gH polypeptides, is provided as SEQ ID NO: 3.

[0128] In some embodiments, the gp42 polypeptide comprises an amino acid sequence with at least 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1. In some embodiments, the gp42 polypeptide comprises an amino acid sequence with at least 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identity' to SEQ ID NO: 2. In some embodiments, the gp4226101polypeptide comprises an amino acid sequence with at least 80%, 85%, 90%, 95%. 97%. 98%.99%, or 100% identity to SEQ ID NO: 3.

[0129] In some embodiments, an antigenic EBV polypeptide comprising a gH and / or gL polypeptide further comprises a gp42 polypeptide. Any of the EBV polypeptides comprising a gH and / or gL polypeptide described above can further comprise a gp42 polypeptide. In some embodiments, the gp42 polypeptide is located C-terminal to the gH and / or gL polypeptide(s), as exemplified in SEQ ID NOs: 11-21. In some embodiments, the gp42 polypeptide is located N-terminal to a ferritin, also as exemplified in SEQ ID NOs: 11-21. Thus, for example, an antigenic EBV polypeptide may comprise, in N- to C-terminal order, a gL polypeptide, a gH polypeptide, a gp42 polypeptide, and optionally a ferritin. Linkers such as those described herein can separate the gp42 polypeptide from EBV polypeptides and / or ferritins located N-terminal and / or C-terminal thereto. In some embodiments, a linker separates each EBV polypeptide in an antigenic ferritin polypeptide (e.g., a gL polypeptide, a gH polypeptide, and a gp42 polypeptide), and a further linker may be present between the ferritin if present and the EBV polypeptide proximal thereto (e.g., a gp42 polypeptide).

[0130] In some embodiments, a linker having a length of at least 15 amino acids separates the EBV gH polypeptide and the EBV gp42 polypeptide. Such a linker may have a length of 15 to 60 amino acids, 20 to 60 amino acids, 30 to 60 amino acids, 40 to 60 amino acids. 30 to 50 amino acids, or 40 to 50 amino acids. In some embodiments, the linker comprises an amino acid sequence with at least 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 8.

[0131] In some embodiments, where gp42 and ferritin are present in a polypeptide, a linker separates the EBV gp42 polypeptide and the ferritin. Such a linker may have a length of at least 15 amino acids or has a length of 15 to 100 amino acids, 20 to 90 amino acids, 30 to 90 amino acids, 60 to 100 amino acids, 70 to 90 amino acids, 80 to 90 amino acids. In some embodiments, such a linker comprises an amino acid sequence with at least 80%, 85%, 90%, 95%, 97%, 98%, 99%. or 100% identity of SEQ ID NO: 9.

[0132] The gp42 polypeptide can be combined with any of the ferritins discussed herein. For example, in some embodiments, a polypeptide comprises agp42 polypeptide (+ / - gL / gH and / or gp220) and a heavy or light chain ferritin (e.g., T. ni heavy or light chain ferritin); or ii) ferritin. In one embodiment the ferritin has a sequence of SEQ ID NO: 24. In one embodiment the ferritin has a sequence of SEQ ID NO: 25. In one embodiment the ferritin has a sequence of SEQ ID NO: 26.26101

[0133] Additionally, in some embodiments, any antigenic EBV polypeptide comprising a gp42 polypeptide and a ferritin can be present in a composition comprising another polypeptide disclosed herein.C. Linkers

[0134] In some embodiments, the EBV polypeptide in Section A comprises a linker between gL and gH polypeptides. In some embodiments, the EBV polypeptide in Section B comprises a linker between gH and gp42 polypeptides. In some embodiments, the EBV polypeptide in Sections A, B, D, E or F comprises a linker between an EBV polypeptide and a ferritin of the present invention. In some embodiments, the linker is a peptide linker, which can facilitate expression of an EBV polypeptide (e.g., from a single open reading frame). In some embodiments, the linker is a glycine-serine linker. Exemplary' linkers are provided in Table 2.

[0135] In some embodiments, the linker is at least 15 amino acids in length. In some embodiments, the linker is at least 25 amino acids in length. In some embodiments, the linker is at least 30 amino acids in length. In some embodiments, the linker is at least 35 amino acids in length. In some embodiments, the linker is at least 40 amino acids in length. In some embodiments, the linker is less than or equal to 60 amino acids in length. In some embodiments, the linker is less than or equal to 50 amino acids in length. In some embodiments, the linker is about 16, 28, 40, 46, or 47 amino acids in length.

[0136] In some embodiments, the linker comprises glycine (G) and / or serine (S) amino acids. In some embodiments, the linker comprises or consists of glycine (G), serine (S), asparagine (N), and / or alanine (A) amino acids. In some embodiments, the linker comprises an amino acid sequence with at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identity to GGSGSASSGASASGSSNGSGSGSGSNSSASSGASSGGASGGSGGSG (SEQ ID NO: 27). In some embodiments, the linker comprises GGGGSGGGGSGGGGSG (SEQ ID NO: 28), GGSGSGSNSSASSGASSGGASGGSGGSG (SEQ ID NO: 29), or GS. In some embodiments, the linker comprises FR1 (GGSGSASAEAAAKEAAAKAGGSGGSG; SEQ ID NO: 30) or FR2 (GGSGSASAEAAAKEAAAKEAAAKASGGSGGSG; SEQ ID NO: 31). In some embodiments, the linker comprises or consists of SEQ ID NO: 7, 8 or 9. In some embodiments, the linker comprises an amino acid sequence with at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identity to SEQ ID NO: 7. In some embodiments, the linker comprises an amino acid sequence with at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identity' to26101SEQ ID NO: 8. In some embodiments, the linker comprises an ammo acid sequence with at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identity to SEQ ID NO: 9.

[0137] In some embodiments, a linker separates the EBV gL polypeptide and the EBV gH polypeptide and has a length of at least 15 amino acids, such as 15-100 amino acids, such as 15-90, 15-80, 15-70, 15-60, 15-50, 15-40, 15-30, or 15-20 amino acids. Such a linker may have a length of 15 to 60 amino acids, 20 to 60 amino acids, 30 to 60 amino acids, 40 to 60 amino acids, 30 to 50 amino acids, 30 to 40 amino acids, 44 to 48 amino acids, 44 amino acids, 46 amino acids, 48 amino acids, or 40 to 50 amino acids. In some embodiments, a linker separating the EBV gL polypeptide and the EBV gH polypeptide has at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identity to any one of SEQ ID Nos: 7-9. In a particular embodiment, a linker separating the EBV gL polypeptide and the EBV gH polypeptide has at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%. at least 98%, at least 99%, or 100% identity to SEQ ID NO: 7. In a particular embodiment, a linker separating the EBV gL polypeptide and the EBV gH polypeptide comprises or consists of SEQ ID NO: 7.

[0138] In some embodiments, a linker separates the EBV gH polypeptide and the EBV gp42 polypeptide and has a length of at least 15 amino acids, such as 15-100 amino acids, such as 15-90, 15-80, 15-70, 15-60, 15-50, 15-40, 15-30, or 15-20 amino acids. Such a linker may have a length of 15 to 60 amino acids, 20 to 60 amino acids, 30 to 60 amino acids, 40 to 60 amino acids, 30 to 50 amino acids, 30 to 40 amino acids, 32 amino acids, or 40 to 50 amino acids. In some embodiments, a linker separating the EBV gH polypeptide and the EBV gp42 polypeptide has at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identity to any one of SEQ ID Nos: 7-9. In a particular embodiment, a linker separating the EBV gH polypeptide and the EBV gp42 polypeptide has at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identity to SEQ ID NO: 8. In a particular embodiment, a linker separating the EBV gH polypeptide and the EBV gp42 polypeptide comprises or consists of SEQ ID NO: 8.

[0139] In some embodiments, a linker separates the EBV gp42 polypeptide and the ferritin. Such a linker may have a length of at least 15 amino acids, such as 15-100 amino acids, such as 60-100, 15-90, 15-80, 15-70, 15-60, 15-50, 15-40, 15-30, or 15-20 amino acids, or has a length of 15 to 60 amino acids, 20 to 60 amino acids, 30 to 60 amino acids, 40 to 60 amino acids, 30 to 50 amino acids, or 40 to 50 amino acids, 70 to 90 amino acids, 80 to 90 amino acids, or 88 amino acids. In some embodiments, such a linker comprises an amino acid sequence having at least2610180%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identity to any one of SEQ ID NOs: 7-9. In particular embodiments, a linker separating the EBV gp42 polypeptide and the ferritin comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identity to SEQ ID NO: 9. In particular embodiments, a linker separating the EBV gp42 polypeptide and the ferritin comprises or consists of SEQ ID NO: 9.D. EBV polypeptides comprising a gL, gH, gp42 polypeptide

[0140] In one aspect, the compositions or methods of the invention comprise antigenic EBV polypeptides comprising EBV gL, EBV gH and gp42. In some embodiments, the EBV gL, EBV gH and gp42 is in N to C-terminal order. In some embodiments, the polypeptide exists as a single-chain. In some embodiments, the polypeptide forms a trimer, e.g., through trimerization of a trimerization domain, such as a T4 phage fibritin trimerization domain. In some embodiments, the polypeptide forms a nanoparticle (e.g.. ferritin or lumazine synthase particle), e.g., through multimerization of a ferritin or lumazine synthase.

[0141] In some embodiments, the EBV polypeptide comprises an EBV gL polypeptide, an EBV gH polypeptide, an EBV gp42 polypeptide, and a ferritin, wherein the EBV gL polypeptide, the EBV gH polypeptide, the EBV gp42 polypeptide, and the ferritin are arranged in N-terminal to C-terminal order within the EBV polypeptide.

[0142] In some such embodiments, the EBV polypeptide comprises one or more, or each, of a linker between the EBV gL polypeptide and the EBV gH polypeptide, between the EBV gH polypeptide and the EBV gp42 polypeptide, and between the EBV gp42 polypeptide and the ferritin. In some embodiments, the linker has a length of at least 15 amino acids, as disclosed elsewhere herein. In such embodiments, each linker may be independently selected from any linker disclosed herein, such as a linker having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to any one of SEQ ID Nos: 7-9. In a particular embodiment, the linker between the EBV gL polypeptide and the EBV gH polypeptide is 44 to 48 amino acids in length, and the linker between the EBV gH polypeptide and the EBV gp42 polypeptide is 32 amino acids in length. In another particular embodiment, the linker between the EBV gL polypeptide and the EBV gH polypeptide is 46 amino acids in length, and the linker between the EBV gH polypeptide and the EBV gp42 polypeptide is 32 amino acids in length. In some embodiments, the EBV polypeptide lacks one or more of the linkers (such as one or more of a linker between the EBV gL polypeptide and the EBV gH polypeptide, between the EBV gH polypeptide and the EBV gp42 polypeptide, and / or26101between the EBV gp42 polypeptide and the ferritin). In particular embodiments, a linker separates the EBV gL polypeptide and the EBV gH polypeptide and comprises an amino acid sequence having at least 80% identity to SEQ ID NO: 7; and / or a linker separates the EBV gH polypeptide and the EBV gp42 polypeptide and comprises an amino acid sequence having at least 80% identity to SEQ ID NO: 8. In particular embodiments, a linker separates the EBV gL polypeptide and the EBV gH polypeptide and consists of an amino acid sequence that has at least 80% identity to SEQ ID NO: 7; and / or a linker separates the EBV gH polypeptide and the EBV gp42 polypeptide and consists of an amino acid sequence that has at least 80% identity to SEQ ID NO: 8. In some embodiments, a linker separates the EBV gp42 polypeptide and the ferritin and comprises an amino acid sequence having at least 80% identity to SEQ ID NO: 9. In some embodiments, the linker separating the EBV gp42 polypeptide and the ferritin comprises the amino acid sequence of SEQ ID NO: 9. In some embodiments, the linker separating the EBV gp42 polypeptide and the ferritin consists of the amino acid sequence of SEQ ID NO: 9.

[0143] In particular embodiments, the EBV polypeptide comprises the amino acid sequence of any one of SEQ ID Nos: 11-21. In other particular embodiments, the EBV polypeptide consists of the amino acid sequence of any one of SEQ ID Nos: 11-21. In other particular embodiments, the EBV polypeptide consists essentially of the amino acid sequence of any one of SEQ ID Nos: 11-21.

[0144] In some embodiments, the EBV polypeptide comprises a sequence with at least 80%, 85%, 90%, 95%, 98%, or 99% identity to any one of SEQ ID NOs: 11-21. In some embodiments, the EBV polypeptide comprises a sequence with at least 90% identity to SEQ ID NO: 11. In some embodiments, the EBV polypeptide comprises a sequence with at least 95% identity to SEQ ID NO: 11. In some embodiments, the EBV polypeptide comprises a sequence with at least 99% identity to SEQ ID NO: 11. In some embodiments, the EBV polypeptide comprises a sequence of SEQ ID NO: 11. In some embodiments, the EBV polypeptide consists of a sequence of SEQ ID NO: 11. In some embodiments, the EBV polypeptide consists essentially of a sequence of SEQ ID NO: 11. In some embodiments, the EBV polypeptide comprises a sequence with at least 95% identity to SEQ ID NO: 21. In some embodiments, the EBV polypeptide comprises a sequence with at least 98% identity to SEQ ID NO: 21. In some embodiments, the EBV polypeptide comprises a sequence with at least 99% identity to SEQ ID NO: 21. In some embodiments, the EBV polypeptide comprises the amino acid sequence of SEQ ID NO: 21. In some embodiments, the EBV polypeptide consists essentially of the amino acid sequence of SEQ ID NO: 21. In some embodiments, the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21.26101

[0145] In some embodiments, an EBV polypeptide that compnses gL, gH and gp42 and a ferritin comprises a linker between gL and gH, gH and gp42, and gp42 and the ferritin. In one embodiment, the linker that separates the EBV gL polypeptide and the EBV gH polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 7. In one embodiment, the linker that separates the EBV gL polypeptide and the EBV gH polypeptide has 40 to 50 amino acids. In one embodiment, the linker that separates the EBV gL polypeptide and the EBV gH polypeptide has 44, 46, or 47 amino acids. In one embodiment, the linker that separates the EBV gH polypeptide and the EBV gp42 polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 8. In one embodiment, the linker that separates the EBV gH polypeptide and the EBV gp42 polypeptide has 30 to 40 amino acids. In one embodiment, the linker that separates the EBV gH polypeptide and the EBV gp42 polypeptide has 32 amino acids. In one embodiment, the linker that separates the EBV gp42 polypeptide and the ferritin comprises or consists of the amino acid sequence of SEQ ID NO: 9. In one embodiment, the linker that separates the EBV gp42 polypeptide and the ferritin has 80 to 90 amino acids. In one embodiment, the linker that separates the EBV gp42 polypeptide and the ferritin has 88 amino acids.

[0146] In a particular embodiment, the linker between the EBV gL polypeptide and the EBV gH polypeptide is 44 to 48 amino acids, and the linker between the EBV gH polypeptide and the EBV gp42 polypeptide is 32 amino acids. In another particular embodiment, the linker between the EBV gL polypeptide and the EBV gH polypeptide is 46 amino acids, and the linker between the EBV gH polypeptide and the EBV gp42 polypeptide is 32 amino acids. In another particular embodiment, a linker separates the EBV gL polypeptide and the EBV gH polypeptide and consists of an amino acid sequence that has at least 80% identity to SEQ ID NO: 7; and / or a linker separates the EBV gH polypeptide and the EBV gp42 polypeptide and consists of an amino acid sequence that has at least 80% identity to SEQ ID NO: 8.

[0147] In some particular embodiments, an EBV polypeptide described above further comprises a further linker that separates the ferritin and the EBV gp42 polypeptide. In particular embodiments, the further linker has a length of 88 amino acids. In particular embodiments, the further linker consists of an amino acid sequence having at least 80% identity to SEQ ID NO: 9.

[0148] In other particular embodiments, the EBV polypeptide comprises an EBV gL polypeptide, an EBV gH polypeptide, and an EBV gp42 polypeptide, wherein the EBV gL polypeptide comprises the amino acid sequence of residues 1-115 of SEQ ID NO: 21, the EBV gH polypeptide comprises the amino acid sequence of residues 162-823 of SEQ ID NO: 21, and the EBV gp42 polypeptide comprises the amino acid sequence of residues 856-1038 of SEQ ID NO: 21; and a ferritin comprising SEQ ID NO: 26. In such embodiments, the EBV gL26101polypeptide, the EBV gH polypeptide, the EBV gp42 polypeptide, and the ferritin are arranged in N-terminal to C-terminal order within the EBV polypeptide. In some such embodiments, a linker separates the EBV gL polypeptide and the EBV gH polypeptide and comprises the amino acid sequence of SEQ ID NO: 7. In some such embodiments, a linker separates the EBV gH polypeptide and the EBV gp42 polypeptide and comprises the amino acid sequence of SEQ ID NO: 8. Some such embodiments further comprise a second EBV polypeptide comprising an EBV gp220 polypeptide and ferritin, and the second EBV polypeptide comprises, consists essentially of, or consists of the amino acid sequence of SEQ ID NO: 23.E. EBV polypeptides comprising a gp220 polypeptide

[0149] In some embodiments, an antigenic EBV polypeptide comprises a gp220 polypeptide. A gp220-hybrid bullfrog / H. pylori ferritin nanoparticle has been previously described in Kanekiyo Cell. 2015 Aug 27; 162(5): 1090-100.

[0150] In some embodiments, the gp220 polypeptide is an amino acid sequence with at least 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identity to gp350D123 (SEQ ID NO: 6). In some embodiments, the gp220 polypeptide comprises the amino acid sequence of SEQ ID NO: 6. In some embodiments, the gp220 polypeptide consists of the amino acid sequence of SEQ ID NO: 6. In some embodiments, the gp220 polypeptide consists essentially of the amino acid sequence of SEQ ID NO: 6.

[0151] The gp220 polypeptide can be combined with any of the ferritins discussed herein. For example, in some embodiments, an antigenic EBV polypeptide comprises a gp220 polypeptide (+ / - gL / gH and / or gp42) and i) a heavy or light chain ferritin (e g., T. ni heavy or light chain ferritin); or ii) a ferritin. In one embodiment the ferritin has a sequence of SEQ ID NO: 24. In one embodiment the ferritin has a sequence of SEQ ID NO: 25. In one embodiment the ferritin has a sequence of SEQ ID NO: 26. In some embodiments, the gp220 polypeptide with ferritin comprises the amino acid sequence of SEQ ID NO: 23. In some embodiments, the gp220 polypeptide with ferritin consists of the amino acid sequence of SEQ ID NO: 23. In some embodiments, the gp220 polypeptide with ferritin consists essentially of the amino acid sequence of SEQ ID NO: 23.

[0152] Additionally, in some embodiments, any antigenic EBV polypeptide comprising a gp220 polypeptide and a ferritin can be present in a composition comprising another polypeptide disclosed herein.26101F. EBV polypeptides comprising an EBV polypeptide and ferritin

[0153] In some embodiments, an antigenic EBV polypeptide is provided, comprising an EBV polypeptide and ferritin. The EBV polypeptide can be any of the EBV polypeptides described herein, such as a gL, gH, gL / gH. gp220, or gp42 polypeptide, or combinations thereof. The ferritin component of the polypeptide may be a ferritin from any species and may or may not have mutations.

[0154] In some embodiments, the ferritin in the polypeptide is a wild-type ferritin. In some embodiments, the ferritin is bacterial, insect, fungal, bird, or mammalian. In some embodiments, the ferritin is human. In some embodiments, the ferritin is bacterial. In one embodiment the ferritin has a sequence of SEQ ID NO: 24. In one embodiment the ferritin has a sequence of SEQ ID NO: 25. In one embodiment the ferritin has a sequence of SEQ ID NO: 26.

[0155] In some embodiments, the ferritin is a light chain and / or heavy chain ferritin. In some embodiments, the ferritin is an insect ferritin, such as Trichoplusia ni heavy chain ferritin or Trichoplusia in light chain ferritin. In some embodiments, the ferritin is a human ferritin, such as human heavy chain ferritin (GENE ID No: 2495) or human light chain ferritin (GENE ID No: 2512). In some embodiments, a ferritin nanoparticle comprises 24 total subunits of heavy chain ferritin and light chain ferritin, such as in human or Trichoplusia ni ferritin nanoparticles. T. ni ferritin nanoparticles can comprise 12 subunits of heavy chain ferritin and 12 subunits of light chain ferritin.

[0156] In some embodiments, an antigenic EBV polypeptide comprises a light chain ferritin and an EBV poly peptide. In some embodiments, an antigenic EBV polypeptide comprises a heavy chain ferritin and an EBV polypeptide. In some embodiments, an antigenic EBV polypeptide comprising a light chain ferntin and an EBV polypeptide can assemble with a heavy¬ chain ferritin that is not linked to an EBV polypeptide. In some embodiments, an antigenic EBV polypeptide comprising a heavy chain ferritin and an EBV polypeptide can assemble with a light chain ferritin that is not linked to an EBV polypeptide. A ferritin not linked to an EBV polypeptide (or, more generally, anon-ferritin polypeptide) may be referred as a “naked ferritin / ’

[0157] In some embodiments, an antigenic polypeptide comprising a heavy chain ferritin and an EBV polypeptide can assemble with an antigenic polypeptide comprising a light chain ferritin and an EBV polypeptide to allow presentation of two of the same or different non-ferritin polypeptides on a single ferritin nanoparticle.

[0158] In some embodiments, an antigenic polypeptide comprising a heavy chain ferritin and a non-ferritin polypeptide can assemble with a polypeptide comprising a light chain ferritin and a non-ferritin polypeptide to produce a bivalent composition.26101

[0159] In some embodiments, an antigenic polypeptide comprises a light chain ferritin and a gp220 and / or gp42 polypeptide. In some embodiments, an antigenic polypeptide comprises a heavy chain ferritin and a gp220 and / or gp42 polypeptide.

[0160] In some embodiments, an antigenic polypeptide comprises a light chain ferritin and a single-chain gL and gH polypeptide. In some embodiments, an antigenic polypeptide comprises a heavy chain ferritin and a single-chain gL and gH polypeptide.

[0161] In some embodiments, an antigenic polypeptide comprising a light chain ferritin and a gp220 and / or gp42 polypeptide assembles with an antigenic polypeptide comprising a heavy¬ chain ferritin and a single-chain gL and gH polypeptide.

[0162] In some embodiments, an antigenic polypeptide comprising a heavy chain ferritin and a gp220 and / or gp42 polypeptide assembles with an antigenic polypeptide comprising a light chain ferritin and a single-chain gL and gH polypeptide. In some embodiments, twelve (12) gp220 and / or gp42 polypeptides and twelve (12) single-chain gL and gH polypeptides are comprised in an assembled ferritin nanoparticle, as in the case of an assembled T. ni ferritin nanoparticle.

[0163] Any type of ferritin nanoparti cle(s) that comprises both gp220 and / or gp42 and singlechain gL and gH polypeptides may be referred to as a “bivalent” or “bivalent EBV” particle or construct. A composition comprising a gL and gH trimer together with a ferritin that comprises gp220 and / or gp42 would also be a bivalent EBV composition.

[0164] In some embodiments, the ferritin is Hybrid bullfrog-H. pylori ferritin (for example SEQ ID NO: 25), optionally with one or more mutations such as those described herein. In some embodiments, the lower sequence homology between H. pylori ferritin (or other bacterial ferritins) and human ferritin may decrease the potential for autoimmunity when used as a vaccine platform (see Kanekiyo et al., Cell 162, 1090-1100 (2015)).

[0165] In some embodiments, a nanoparticle is provided comprising an antigenic EBV polypeptide as disclosed herein comprising an EBV polypeptide and a ferritin.G. Ferritin mutations

[0166] In some embodiments, the ferritin comprises one or more mutations are disclosed herein. In some embodiments, the one or more mutations comprise changes to the amino acid sequence of a wild-type ferritin and / or an insertion, e.g., at the N- or C-terminus. In some embodiments, one. two, three, four, five, or more different amino acids are mutated in the ferritin as compared to wild-type ferritin (in some embodiments, in addition to any N-terminal insertion). The one or more mutations can change functional properties of the ferritin. In general, a mutation simply refers to a difference in the sequence (such as a substituted, added, or deleted amino acid26101residue or residues) relative to the corresponding wild-type ferritin. In one embodiment the ferritin has a sequence of SEQ ID NO: 24. In one embodiment the ferritin has a sequence of SEQ ID NO: 25. In one embodiment the ferritin has a sequence of SEQ ID NO: 26.TABLE 2 (SEQUENCE TABLE): DESCRIPTION OF THE SEQUENCES Description Sequences SEQ Key for protein sequences ID NO gL - ItalicizedLinker - double underlinedgH - BoldbfpFerr (hybrid ferritin) - wavy underlinegp220 - Italicized and boldgp42 - Italicized and underlinedGp42 DSKGSSQKGSRLLLLLVVSNLLLPQGVLAYFLPPRV 1RGGGRVAAAAITWVPKPNVEVWPVDPPPPVNFNK TAEQEYGDKEVKLPHWTPTLHTFQVPQNYTKANC TYCNTREYTFSYKGCCFYFTKKKHTWNGCFQACA ELYPCTYFYGPTPDILPVVTRNLNAIESLWVGVYRV GEGNWTSLDGGTFKVYQIFGSHCTYVSKFSTVPVS HHECSFLKPCLCVSQRSNSgp42 fusion LAYFLPPRVRGGGRVAAAAITWVPKPNVEVWPVD 2 segment PPPPVNFNKTAEQEYGDKEVKLPHWTPTLHTFQVP QNYTKANCTYCNTREYTFSYKGCCFYFTKKKHTW NGCFQACAELYPCTYFYGPTPDILPVVTRNLNAIES LWVGVYRVGEGNWTSLDGGTFKVYQIFGSHCTYV SKFSTVPVSHHECSFLKPCLCVSQRSNSgp42 fusion AITWVPKPNVEVWPVDPPPPVNFNKTAEQEYGDKE 3 segment 2 VKLPHWTPTLHTFQVPQNYTKANCTYCNTREYTFS YKGCCFYFTKKKHTWQGCFQACAELYPCTYFYGP TPDILPVVTRSLQAIESLWVGVYRVGEGNWTSLDG GTFKVYQIFGSHCTYVSKFSTVPVSHHECSFLKPCL CVSQRSNSExemplary gL NWAYPCCHVTQLRAQHLLALENISDIYLVSNQTCD 4 polypeptide GFSLASLNSPKNGSNQLV1SRCANGLNVVSFFISILK RSSSALTGHLRELLTTLETLYGSFSVEDLFGANLNR YAWHRGGExemplary gH AASLSEVKLHLDIEGHASHYTIPWTELMAKVPGLSP 5 polypeptide EALWREANVTEDLASMLNRYKLIYKTSGTLGIALA EPVDIPAVSEGSMQVDASKVHPGVISGLNSPACMLS APLEKQLFYYIGTMLPNTRPHSYVFYQLRCHLSYV ALSINGDKFQYTGAMTSKFLMGTYKRVTEKGDEH VLSLVFGKTKDLPDLRGPFSYPSLTSAQSGDYSLVI VTTFVHYANFHNYFVPNLKDMFSRAVTMTAASYA RYVLQKLVLLEMKGGCREPELDTETLTTMFEVSVA FFKVGHAVGETGNGCVDLRWLAKSFFELTVLKDII GICYGATVKGMQSYGLERLAAMLMATVKMEELG HLTTEKQEYALRLATVGYPKAGVYSGLIGGATSVLLSAYNRHPLFQPLHTVMRETLFIGSHVVLRELRLNV26101TTQGPNLALYQLLSTALCSALEIGEVLRGLALGTES GLFSPCYLSLRFDLTRDKLLSMAPQEATLDQAAVS NAVDGFLGRLSLEREDRDAWHLPAYKCVDRLDKV LMIIPLINVTFIISSDREVRGSALYEASTTYLSSSLFLS PVIMNKCSQGAVAGEPRQIPKIQNFTRTQKSCIFCGF ALLSYDEKEGLETTTYITSQEVQNSILSSNYFDFDNL HVHYLLLTTNGTVMEIAGLYEERAExemplary EAALLVCQYTIQSLIHLTGEDPGFFNVEIPEFPFYPTC 6 gp350D123 NVCTADVNVTINFDVGGKKHQLDLDFGQLTPHTK polypeptide AVYQPRGAFGGSENATNLFLLELLGAGELALTMRS KKLPINVTTGEEQQVSLESVDVYFQDVFGTMWCH HAEMQNPVYLIPETVPYIKWDNCNSTNITAVVRAQ GLDVTLPLSLPTSAQDSNFSVKTEMLGNEIDIECIME DGEISQVLPGDNKFNITCSGYESHVPSGGILTSTSPV ATPIPGTGYAYSLRLTPRPVSRFLGNNSILYVFYSGN GPKASGGDYCIQSNIVFSDEIPASQDMPTNTTDITYV GDNATYSVPMVTSEDANSPNVTVTAFWAWPNNTE TDFKCKWTLTSGTPSGCENISGAFASNRTFDITVSG LGTAPKTLIITRTATNATTTTHKVIFSKAPE46 amino acid GGSGSASSGASASGSSGGSGSGSGSGSSASSGASSG 7 linker GASGGSGGSG32 amino acid SGGGSGSASSGASASGSSGSGSGSGSSSASSG 8 linker88 amino acid GGSGSASSGASASGSSGSGSGSGSSSASSGASSGGA 9 linker SGGSGGSGGGSGSASSGASASGSSGSGSGSGSSSAS SGASSGGASGGSGGSG44 amino acid GGSGSASSGASASGSSGSGSGSGSSSASSGASSGGA 10 linker SGGSGGSG NWAYPCCHVTQLRAQHLLALENISDIYLVSNQTCDGFS 11 gL-gH-gp42 LASLNSPKNGSNQLVISRCANGLNWSFFISILKRSSSAL TGHLRELLTTLETLYGSFSVEDLFGANLNRYAWHRGG GGSGSASSGASASGSSNGSGSGSGSNSSASSGASSG GASGGSGGSGAASLSEVKLHLDIEGHASHYTIPW TELMAKVPGLSPEALWREANVTEDLASMLNRY KLIYKTSGTLGIALAEPVDIPAVSEGSMQVDASK VHPGVISGLNSPACMLSAPLEKQLFYYIGTMLPN TRPHSYVFYQLRAHLSYVALSINGDKFQYTGAM TSKFLMGTYKRVTEKGDEHVLSLVFGKTKDLPD LRGPFSYPSLTSAQSGDYSLVIVTTFVHYANFHN YFVPNLKDMFSRAVTMTAASYARYVLQKLVLLE MKGGCREPELDTETLTTMFEVSVAFFKVGHAV GETGNGCVDLRWLAKSFFELTVLKDIIGICYGAT VKGMQSYGLERLAAMLMATVKMEELGHLTTE KQEYALRLATVGYPKAGVYSGLIGGATSVLLSA YNRHPLFQPLHTVMRETLFIGSHVVLRELRLNV TTQGPNLALYQLLSTALCSALEIGEVLRGLALGT ESGLFSPCYLSLRFDLTRDKLLSMAPQEATLDQA AVSNAVDGFLGRLSLEREDRDAWHLPAYKCVDR LDKVLMIIPLINVTFIISSDREVRGSALYEASTTYLSSSLFLSPVIMNKCSQGAVAGEPRQIPKIQNFTRT26101QKSCIFCGFALLSYDEKEGLETTTYITSQEVQNSI LSSNYFDFDNLHVHYLLLTTNGTVMEIAGLYEER ASGGGSGS ASSGAS ASGSSGSGSGSGSSSASSGL4FF LPPRVRGGGRVAAAAITWVPKPNVEVWPVDPPPPVNF NKTAEOEYGDKEVKLPHWTPTLHTFQVPONYTKANC TYCNTREYTFSYKGCCFYFTKKKHTWNGCFOACAELY PCTYFYGPTPDILP WTRNLNAIESL WVGVYR VGEGNW TSLDGGTFKVYOIFGSHCTYVSKFSTVPVSHHECSFLK PCLCVSORSNSGS NWAYPCCHVTQLRAQHLLALENISDIYLVSNQTCDGFS 12 gL-gH-gp42 LASLNSPKNGSNQLVISRCANGLNWSFFISILKRSSSAL FERRITIN TGHLRELLTTLETLYGSFSVEDLFGANLNRYAWHRGG GGSGSASSGASASGSSNGSGSGSGSNSSASSGASSG GASGGSGGSGAASLSFVKEHLDTFGHASHYTIPW TELMAKVPGLSPEALWREANVTEDLASMLNRY KLIYKTSGTLGIALAEPVDIPAVSEGSMQVDASK VHPGVISGLNSPACMLSAPLEKQLFYYIGTMLPN TRPHSYVFYQLRAHLSYVALSINGDKFQYTGAM TSKFLMGTYKRVTEKGDEHVLSLVFGKTKDLPD LRGPFSYPSLTSAQSGDYSLVIVTTFVHYANFHN YFVPNLKDMFSRAVTMTAASYARYVLQKLVLLE MKGGCREPELDTETLTTMFEVSVAFFKVGHAV GETGNGCVDLRWLAKSFFELTVLKDIIGICYGAT VKGMQSYGLERLAAMLMATVKMEELGHLTTE KQEYALRLATVGYPKAGVYSGLIGGATSVLLSA YNRHPLFQPLHTVMRETLFIGSHVVLRELRLNV TTQGPNLALYQLLSTALCSALEIGEVLRGLALGT ESGLFSPCYLSLRFDLTRDKLLSMAPQEATLDQA AVSNAVDGFLGRLSLEREDRDAWHLPAYKCVDR LDKVLMTIPLINVTFIISSDREVRGSALYEASTTYL SSSLFLSPVIMNKCSQGAVAGEPRQIPKIQNFTRT QKSCIFCGFALLSYDEKEGLETTTYITSQEVQNSI LSSNYFDFDNLHVHYLLLTTNGTVMEIAGLYEER ASGGGSGS ASSGAS ASGSSGSGSGSGSSSASSGL4FF LPPRVRGGGRVAAAAITWVPKPNVEVWPVDPPPPVNF NKTAEOEYGDKEVKLPHWTPTLHTFOVPONYTKANC TYCNTREYTFSYKGCCFYFTKKKHTWNGCFOACAELY PCTYFYGPTPDILP WTRNLNAIESL WVGVYR VGEGNW TSLDGGTFKVYOIFGSHCTYVSKFSTVPVSHHECSFLK PCTCKSWSWGGSGS ASSGAS ASGSSGSGSGSGSSS ASSGASSGGASGGSGGSGGGSGSASSGASASGSSGS GSGSGSSSASSGASSGGASGGSGGSGESQVRQQFSK DIEKLLNEQVNKEMQSSNLYMSMSSWSYTHSLE)G AGLFLFDHAAEEYEHAKKrilFLNENNVPyQLTSISA PEHKFEGLTQIFQKAYEHEQHISESINNIVDHAIKCK DHAIFNI LQ\VY\ AEQHEEEX LFKDILDKIELIGXEX HGLYLADQYVKGIAKSRKS NWAYPCCHVTQLRAQHLLALENISDIYLVSNQTCDGFS 13 gL-gH-gp42- LASLNSPKNGSNQLVISRCANGLNWSFFISILKRSSSALFERRITIN C12 TGHLRELLTTLETLYGSFSVEDLFGANLNRYAWHRGG26101GGSGS ASSGAS ASGSSNGSGSGSGSNSSASSGASSG GASGGSGGSGAASESFVKEHEDTFGHASHYTIPW TELMAKVPGLSPEALWREANVTEDLASMLNRY KLIYKTSGTLGIALAEPVDIPAVSEGSMQVDASK VHPGVISGLNSPACMLSAPLEKQLFYYIGTMLPN TRPHSYVFYQLRAHLSYVALSINGDKFQYTGAM TSKFLMGTYKRVTEKGDEHVLSLVFGKTKDLPD LRGPFSYPSLTSAQSGDYSLVIVTTFVHYANFHN YFVPNLKDMFSRAVTMTAASYARYVLQKLVLLE MKGGCREPELDTETLTTMFEVSVAFFKVGHAV GETGNGCVDLRWLAKSFFELTVLKDIIGICYGAT VKGMQSYGLERLAAMLMATVKMEELGHLTTE KQEYALRLATVGYPKAGVYSGLIGGATSVLLSA YNRHPLFQPLHTVMRETLFIGSHVVLRELRLNV TTQGPNLALYQLLSTALCSALEIGEVLRGLALGT ESGLFSPCYLSLRFDLTRDKLLSMAPQEATLDQA AVSNAVDGFLGRLSLEREDRDAWHLPAYKCVDR LDKVLMIIPLINVTFIISSDREVRGSALYEASTTYL SSSLFLSPVIMNKCSQGAVAGEPRQIPKIQNFTRT QKSCIFCGFALLSYDEKEGLETTTYITSQEVQNSI LSSNYFDFDNLHVHYLLLTTNGTVMEIAGLYEER ASGGGSGS ASSGAS ASGSSGSGSGSGSSSASSGL4FF LPPRVRGGGRVAAAAITWVPKPNVEVWPVDPPPPVNF NKTAEOEYGDKEVKLPHWTPTLHTFOVPQNYTKANC TYCNTREYTFSYKGCCFYFTKKKHTWNGCFOACAELY PCTYFYGPTPDILP WTRNLNA IESL WVG VYR VGEGNW TSLDGGTFKVYQIFGSHCTYVSKFSTVPVSHHECSFLK PCLCKSmSTVSGGSGSASSGASASGSSGSGSGSGSSS ASSGASSGGASGGSGGSGESQVRQQFSKDIEKLLNE QVNKEMQSSNLYMSMSSWSYTHSLDGAGLFLFDH AAEEYEIIAKKLIIFLNENNVPVQLTSISAPEHKFEGL TQIFQKAYEHEQHISESINNIVDHAIKCKDHATFNFL o\\ Y\ AF: OHF: F: FA LFKDILI)KIF: LIG\F:\IIGLYLAD OY\ KG1AKSRKS NWAYPCCHVTQLRAQHLLALENISDIYLVSNQTCDGFS 14 gL-gH-gp42- LASLNSPKNGSNQLVISRCANGLNWSFFISILKRSSSAL FERRITIN C13 TGHLRELLTTLETLYGSFSVEDLFGANLNRYAWHRGG GGSGSASSGAS ASGSSNGSGSGSGSNSSASSGASSG GASGGSGGSGAASESFVKEHEDTFGHASHYTIPW TELMAKVPGLSPEALWREANVTEDLASMLNRY KLIYKTSGTLGIALAEPVDIPAVSEGSMQVDASK VHPGVISGLNSPACMLSAPLEKQLFYYIGTMLPN TRPHSYVFYQLRAHLSYVALSINGDKFQYTGAM TSKFLMGTYKRVTEKGDEHVLSLVFGKTKDLPD LRGPFSYPSLTSAQSGDYSLVIVTTFVHYANFHN YFVPNLKDMFSRAVTMTAASYARYVLQKLVLLE MKGGCREPELDTETLTTMFEVSVAFFKVGHAV GETGNGCVDLRWLAKSFFELTVLKDIIGICYGAT VKGMQSYGLERLAAMLMATVKMEELGHLTTEKQEYALRLATVGYPKAGVYSGLIGGATSVLLSA26101YNRHPLFQPLHTVMRETLFIGSHVVLRELRLNV TTQGPNLALYQLLSTALCSALEIGEVLRGLALGT ESGLFSPCYLSLRFDLTRDKLLSMAPQEATLDQA AVSNAVDGFLGRLSLEREDRDAWHLPAYKCVDR LDKVLMIIPLINVTFI1SSDREVRGSALYEASTTYL SSSLFLSPVIMNKCSQGAVAGEPRQIPKIQNFTRT QKSCIFCGFALLSYDEKEGLETTTYITSQEVQNSI LSSNYFDFDNLHVHYLLLTTNGTVMEIAGLYEER ASGGGSGS ASSGAS ASGSSGSGSGSGSSSASSGL4 KF LPPRVRGGGRVAAAAITWVPKPNVEVWPVDPPPPVNF NKTAEQEYGDKEVKLPHWTPTLHTFOVPQNYTKANC TYCNTREYTFSYKGCCFYFTKKKHTWNGCFOACAELY PCTYFYGPTPDILP WTRNLNAIESL WVGVYR VGEGNW TSLDGGTFKVYQIFGSHCTYVSKFSTVPVSHHECSFLK PCLCVSQRSNSEPEPEPEPEPGGVSQNWOYSKDXEK. EEXEQVXIG Y1QSSXEY YISXISSWSYTI ISEDGAGEE IJ DHAAI J AEHAKKIJIFLXEXXA 1A QLTSISAPEHK FEGLTQIFQKAYEHEQHISESINNIVDHAIKCKDHAT FNFLQWYVAEQHEEEVLFKDILDKIELIGNENHGLYI. ADQYX KGIAKSRKS NWAYPCCHVTQLRAQHLLALENISDIYLVSNQTCDGFS 15 gL-gH-gp42- LASLNSPKNGSNQLVISRCANGLNWSFFISILKRSSSAL FERRITIN C14 TGHLRELLTTLETLYGSFSVEDLFGANLNRYAWHRGG GGSGSASSGASASGSSNGSGSGSGSNSSASSGASSG GASGGSGGSGAASLSFVKLHLDTFGHASHYTIPW TELMAKVPGLSPEALWREANVTEDLASMLNRY KLIYKTSGTLGIALAEPVDIPAVSEGSMQVDASK VHPGVISGLNSPACMLSAPLEKQLFYYIGTMLPN TRPHSYVFYQLRAHLSYVALSINGDKFQYTGAM TSKFLMGTYKRVTEKGDEHVLSLVFGKTKDLPD LRGPFSYPSLTSAQSGDYSLVIVTTFVHYANFHN YFVPNLKDMFSRAVTMTAASYARYVLQKLVLLE MKGGCREPELDTETLTTMFEVSVAFFKVGHAV GETGNGCVDLRWLAKSFFELTVLKDIIGICYGAT VKGMQSYGLERLAAMLMATVKMEELGHLTTE KQEYALRLATVGYPKAGVYSGLIGGATSVLLSA YNRHPLFQPLHTVMRETLFIGSHVVLRELRLNV TTQGPNLALYQLLSTALCSALEIGEVLRGLALGT ESGLFSPCYLSLRFDLTRDKLLSMAPQEATLDQA AVSNAVDGFLGRLSLEREDRDAWHLPAYKCVDR LDKVLMIIPLINVTFIISSDREVRGSALYEASTTYL SSSLFLSPVIMNKCSQGAVAGEPRQIPKIQNFTRT QKSCIFCGFALLSYDEKEGLETTTYITSQEVQNSI LSSNYFDFDNLHVHYLLLTTNGTVMEIAGLYEER ASGGGSGSASSGASASGSSGSGSGSGSSSASSGL4KF LPPR VRGGGR VAAAAITWVPKPNVEVWP VDPPPP VNF NKTAEOEYGDKEVKLPHWTPTLHTFOVPQNYTKANC TYCNTREYTFSYKGCCFYFTKKKHTWNGCFOACAELY PCTYFYGPTPDILP WTRNLNAIESL WVG VYR VGEGNWTSLDGGTFKVYOIFGSHCTYVSKFSTVPVSHHECSFLK26101PCCCKSWWGGSGESQVRQQFSKDIEKLLNEQVN KEMQSSNLYMSMSSWSYTHSLDGAGLFLFDHAAE EYEHAKKLIIFLNENNVPVQLTSISAPEHKFEGLTQI FQKAYEHEQHISESINMVDHAIKCKDHATFNFLQW YVAEQ1 lEFIA EFlGllEDlGELlGYEYl IGEYEADOYV KGIAKSRKSgL-gH- NWAYPCCHVTQLRAQHLLALENISDIYLVSNQTCDAFS 16 gp42_FERRITIN_ LASLNSPKQGSNQLVISRCANGLNWSFFISILKRSSSAL Cl 6-1 TGHLRELLTTLETL YGSFSVEDLFGAQLNRYA WHRGG GGSGSASSGASASGSSGGSGSGSGSGSSASSGASSG GASGGSGGSGAASLSEVKLHLDIEGHASHYTIPW TELLAKVPGLSPEALWREANVTEDLASMLNRYK LIYKTSGTLGIALAEPVDIPAVSEGSMQVDASKV HPGVISGLNSPACMLSAPLEKQLFYYIGTMLPNT RPHSYVFYQLRCHLSYVALSINGDKFQYTGAMT SKFLMGTYKRVTEKGDEHVLSLVFGKTKDLPDL RGPFSYPSLTSAQSGEYSLVIVTTFVHYANFHNYF VPNLKDMFSRAVTMTAASYARYVLQKLVLLEM KGGCREPELETETLTTMFEVSVAFFKVGHAVGE TGNGCVDLRWLAKSFFELTVLKDIIGICYGATVK GMQSYGLERLAAILMATVKMEELGHLTTEKQE YALRLATVGYPKAGVYSGLIGGATSVLLSAYNR HPLFQPLHTVMRETLFIGSHVVLRELRLNVTTQ GPNLALYQLLSTALCSALEIGEVLRGLALGTESG LFSPCYLSLRFDLTRDKLLSIAPQEATLDQAAVS QAVDGFLGRLSLEREDRDAWHLPAYKCVDRLD KVLMIIPLINVTFIISSDREVRGSALYEASTTYLSS SLFLSPVILNKCSQGAVAGEPRQIPKIQNFTRTQK SCIFCGFALLSYDEKEGLETTTYITSQEVQNSILSS NYFDFDNLHVHYLLLTTNGTVMEIAGLYEERAS GGGSGSASSGASASGSSGSGSGSGSSSASSG4I7WFP KPNVEVWP VDPPPP VNFNKTAEOEYGDKEVKLPHWT PTLHTFOVPQNYTKANCTYCNTREYTFSYKGCCFYFTK KKHTWOGCFOACAELYPCTYFYGPTPDILPWTRSLO AIESL WVG VYR VGEGNWTSLDGGTFKVYQIFGSHCTY VSKFSTVPVSHHECSFLKPCLCVSORSNSGGSGS^SG ASASGSSGSGSGSGSSSASSGASSGGASGGSGGSGG GSGSASSGASASGSSGSGSGSGSSSASSGASSGGAS GGSGGSGESQVRSQFSIG^EfaLNEQVNKEMQSSN LYMSMSSWSYTHSLDGAGLFLFDHAAEEYEHAKK LI1FLNENNVPVQLTS1SAPEHKFEGLTQ1FQKAYEH EQHISESINQiyDHAIKCKDHATFNFLQWYVAEQHE EEVLFKDILDKIELIGQENHGLYLADQYVKGEYKSR KS NWAYPCCHVTQLRAQHLLALENISDIYLVSNQTCDAFS 17 gL-gH-gp42- LASLNSPKOGSNQLVISRCANGLNWSFFISILKRSSSAL FERRITIN C17-1 TGHLRELLTTLETLYGSFSVEDLFGAQLNRYAWHRGG GGSGSASSGASASGSSGGSGSGSGSGSSASSGASSG GASGGSGGSGAASLSFVKEHLDTFGHASHYTIPWTELLAKVPGLSPEALWREANVTEDLASMLNRYK26101LIYKTSGTLGIALAEPVDIPAVSEGSMQVDASKV HPGVISGLNSPACMLSAPLEKQLFYYIGTMLPNT RPHSYVFYQLRCHLSYVALSINGDKFQYTGAMT SKFLMGTYKRVTEKGDEHVLSLVFGKTKDLPDL RGPFSYPSLTSAQSGEYSLV1VTTFVHYANFHNYF VPNLKDMFSRAVTMTAASYARYVLQKLVLLEM KGGCREPELETETLTTMFEVSVAFFKVGHAVGE TGNGCVDLRWLAKSFFELTVLKDIIGICYGATVK GMQSYGLERLAAILMATVKMEELGHLTTEKQE YALRLATVGYPKAGVYSGLIGGATSVLLSAYNR HPLFQPLHTVMRETLFIGSHVVLRELRLNVTTQ GPNLALYQLLSTALCSALEIGEVLRGLALGTESG LFSPCYLSLRFDLTRDKLLSIAPQEATLDQAAVS QAVDGFLGRLSLEREDRDAWHLPAYKCVDRLD KVLMIIPLINVTFIISSDREVRGSALYEASTTYLSS SLFLSPVIMNKCSQGAVAGEPRQIPKIQNFTRTQ KSCIFCGFALLSYDEKEGLETTTYITSQEVQNSIL SSNYFDFDNLHVHYLLLTTNGTVMEIAGLYEER ASGGGSGS ASSGAS ASGSSGSGSGSGSSSASSG4 / 77F VPKPNVEVWP VDPPPP VNFNKTAEQEYGDKE VKLPH WTPTLHTFQVPQNYTKANCTYCNTREYTFSYKGCCFY FTKKKHTWOGCFOACAELYPCTYFYGPTPDILPWTR NLNAIESLWVGVYRVGEGNWTSLDGGTFKVYQIFGSH CTYVSKFSTVPVSHHECSFLKPCLCVSORSNSGGSGSA SSGASASGSSGSGSGSGSSSASSGASSGGASGGSGG SGGGSGS ASSGAS ASGSSGSGSGSGSSSASSGASSG GASGGSGGSGESOVRSOFSKDIEKLLNEQVNKEXIO S SNLYMSMS S WS Y THSIJXiACiLI Ll DHAAI JiYliHA KKLIIFLNENNVPVQLTSISAPEHKFEGLTQIFQKAY EHEQHISESINQIVDHAIKCKDHATFNFLQWYVAEQ[[EEEVLFKDILDKIELIGQENIIGLYLADQYVKGIAK SRKS NWAYPCCHVTQLRAQHLLALENISDIYLVSNQTCDAFS 18 gL-gH-gp42_ LASLNSPKQGSNQLVISRCANGLNWSFFISILKRSSSAL FERRITIN_ Cl 8-1 TGHLRELLTTLETL YGSFSVEDLFGA QLNRYA WHRGG GGSGSASSGASASGSSGGSGSGSGSGSSASSGASSG GASGGSGGSGAASLSFVKEHEDTFGHASHYTIPW TELLAKVPGLSPEALWREANVTEDLASMLNRYK LIYKTSGTLGIALAEPVDIPAVSEGSMQVDASKV HPGVISGLNSPACMLSAPLEKQLFYYIGTMLPNT RPHSYVFYQLRCHLSYVALSINGDKFQYTGAMT SKFLMGTYKRVTEKGDEHVLSLVFGKTKDLPDL RGPFSYPSLTSAQSGEYSLVIVTTFVHYANFHNYF VPNLKDMFSRAVTMTAASYARYVLQKLVLLEM KGGCREPELETETLTTMFEVSVAFFKVGHAVGE TGNGCVDLRWLAKSFFELTVLKDIIGICYGATVK GMQSYGLERLAAMLMATVKMEELGHLTTEKQ EYALRLATVGYPKAGVYSGLIGGATSVLLSAYN RHPLFQPLHTVMRETLFIGSHVVLRELRLNVTTQGPNLALYQLLSTALCSALEIGEVLRGLALGTES26101GLFSPCYLSLRFDLTRDKLLSMAPQEATLDQAA VSNAVDGFLGRLSLEREDRDAWHLPAYKCVDRL DKVLMIIPLINVTFIISSDREVRGSALYEASTTYLS SSLFLSPVIMNKCSQGAVAGEPRQIPKIQNFTRTQ KSCIFCGFALLSYDEKEGLETTTYITSQEVQNSIL SSNYFDFDNLHVHYLLLTTNGTVMEIAGLYEER ASGGGSGS ASSGAS ASGSSGSGSGSGSSSASSGA / TIF VPKPNVEVWP VDPPPP VNFNKTAEQEYGDKEVKLPH WTPTLHTFQVPQNYTKANCTYCNTREYTFSYKGCCFY FTKKKHTWNGCFQACAELYPCTYFYGPTPDILPWTR NLNAIESLWVGVYRVGEGNWTSLDGGTFKVYQIFGSH CTYVSKFSTVPVSHHECSFLKPCLCVSORSNSGGSGSA SSGASASGSSGSGSGSGSSSASSGASSGGASGGSGG SGGGSGSASSGASASGSSGSGSGSGSSSASSGASSG GASGGSGGSGESOVRSQFSKDIEKI. LXEOX XKEXIQ SSX GV XISXISSW SY I I ISCOCiAC. I I IJ OI IAAU YIA I A KKLIIFLNENNVPVQLTSISAPEHKFEGLTQIFQKAY EHEQHISESJNQIVDHAIKCKDHATFNFLQWYVAEQ HEEEYLFKDILPKIELIGQENHGLYLADQYVKGIAK SRKSgL-gH-gp42- NWAYPCCHVTQLRAQHLLALENISDIYLVSNQTCDAFS 19 FERRITIN Cl 6-2 LASLNSPKQGSNQLVISRCANGLNWSFFISILKRSSSAL TGHLRELLTTLETLYGSFSVEDLFGAQLNRYAWHRGG GGSGSASSGASASGSSGGSGSGSGSGSSASSGASSG GASGGSGGSGAASLSEVKLHLDTEGHASHYTIPW TELLAKVPGLSPEALWREANVTEDLASMLNRYK LIYKTSGTLGIALAEPVDIPAVSEGSMQVDASKV HPGVISGLNSPACMLSAPLEKQLFYYIGTMLPNT RPHSYVFYQLRCHLSYVALSINGDKFQYTGAMT SKFLMGTYKRVTEKGDEHVLSLVFGKTKDLPDL RGPFSYPSLTSAQSGEYSLVIVTTFVHYANFHNYF VPNLKDMFSRAVTMTAASYARYVLQKLVLLEM KGGCREPELETETLTTMFEVSVAFFKVGHAVGE TGNGCVDLRWLAKSFFELTVLKDIIGICYGATVK GMQSYGLERLAAILMATVKMEELGHLTTEKQE YALRLATVGYPKAGVYSGLIGGATSVLLSAYNR HPLFQPLHTVMRETLFIGSHVVLRELRLNVTTQ GPNLALYQLLSTALCSALEIGEVLRGLALGTESG LFSPCYLSLRFDLTRDKLLSIAPQEATLDQAAVS QAVDGFLGRLSLEREDRDAWHLPAYKCVDRLD KVLMIIPLINVTFIISSDREVRGSALYEASTTYLSS SLFLSPVILNKCSQGAVAGEPRQIPKIQNFTRTQK SCIFCGFALLSYDEKEGLETTTYITSQEVQNSILSS NYFDFDNLHVHYLLLTTNGTVMEIAGLYEERAS GGGSGSASSGASASGSSGSGSGSGSSSASSG4 / 7RW KPNVEVWP VDPPPP VNFNKTAEOEYGDKEVKLPHWT PTLHTFOVPONYTKANCTYCNTREYTFSYKGCCFYFTK KKHTWOGCFOACAELYPCTYFYGPTPDILPWTRSLQ AIESL WVGVYR VGEGNWTSLDGGTFKVYQIFGSHCTYVSKFSTVPVSHHECSFLKPCLCVSORSNSGGSGSASSG.26101ASASGSSGSGSGSGSSSASSGASSGGASGGSGGSGG GSGSASSGASASGSSGSGSGSGSSSASSGASSGGAS GGSGGSGI: SOVRSQI;SKDII; KLLM: OV\KIAK.)SS\ LYMSMSSWSYTHSLDGAGLFLFDHAAEEYEHAKK LIlFLNENW±yQ. LTSEW EQHISESINQIVDHAIKSKDHATFNFLQWYVAEQHE EEVLFKDILDKIELIGQENHGLYLADQYVKGIAKSR KSgL-gH-gp42- NWAYPCCHVTQLRAQHLLALENISDIYLVSNQTCDAFS 20 FERRITIN C17-2 LASLNSPKQGSNQLVISRCANGLNWSFFISILKRSSSAL TGHLRELLTTLETLYGSFSVEDLFGAQLNRYAWHRGG GGSGSASSGASASGSSGGSGSGSGSGSSASSGASSG GASGGSGGSGAASLSEVKLHLDTEGHASHYTIPW TELLAKVPGLSPEALWREANVTEDLASMLNRYK LIYKTSGTLGIALAEPVDIPAVSEGSMQVDASKV HPGVISGLNSPACMLSAPLEKQLFYYIGTMLPNT RPHSYVFYQLRCHLSYVALSINGDKFQYTGAMT SKFLMGTYKRVTEKGDEHVLSLVFGKTKDLPDL RGPFSYPSLTSAQSGEYSLVIVTTFVHYANFHNYF VPNLKDMFSRAVTMTAASYARYVLQKLVLLEM KGGCREPELETETLTTMFEVSVAFFKVGHAVGE TGNGCVDLRWLAKSFFELTVLKDIIGICYGATVK GMQSYGLERLAAILMATVKMEELGHLTTEKQE YALRLATVGYPKAGVYSGLIGGATSVLLSAYNR HPLFQPLHTVMRETLFIGSHVVLRELRLNVTTQ GPNLALYQLLSTALCSALEIGEVLRGLALGTESG LFSPCYLSLRFDLTRDKLLSIAPQEATLDQAAVS QAVDGFLGRLSLEREDRDAWHLPAYKCVDRLD KVLMIIPLINVTFIISSDREVRGSALYEASTTYLSS SLFLSPVIMNKCSQGAVAGEPRQIPKIQNFTRTQ KSCIFCGFALLSYDEKEGLETTTYITSQEVQNSIL SSNYFDFDNLHVHYLLLTTNGTVMEIAGLYEER ASGGGSGSASSGASASGSSGSGSGSGSSSASSG4 / 7W VPKPNVEVWP VDPPPP VNFNKTAEOEYGDKE VKLPH WTPTLHTFQVPQNYTKANCTYCNTREYTFSYKGCCFY FTKKKHTWQGCFOACAELYPCTYFYGPTPDILPWTR NLNAIESLWVGVYRVGEGNWTSLDGGTFKVYQIFGSH CTYVSKFSTVPVSHHECSFLKPCLCVSORSNSGGSGSk SSGASASGSSGSGSGSGSSSASSGASSGGASGGSGG SGGGSGSASSGASASGSSGSGSGSGSSSASSGASSG GASGGSGGSGESQVRSQFSKPIEKLLNEQyNKEMQ SSNLYMSMSSWSYTHSLDGAGLFLFDHAAEEYEHA KKLnFLNENNVPVQLTSISAPEHKFEGLTQIFQKAY EHEQHISESINQiy DHAIKSKDHA I I XI LQW Y\ AEQ HEEEVLFI< DILDI< IELIGQENHGLYLADQYVI< GIAI< SRKSgL-gH-gp42- NWAYPCCHVTQLRAQHLLALENISDIYLVSNQTCDAFS 21 FERRITIN Cl 8-2 LASLNSPKQGSNQLVISRCANGLNWSFFISILKRSSSALTGHLRELLTTLETLYGSFSVEDLFGAQLNRYAWHRGGGGSGSASSGASASGSSGGSGSGSGSGSSASSGASSG26101GASGGSGGSGAASESEVKEHEDTEGHASHYTIPW TELLAKVPGLSPEALWREANVTEDLASMLNRYK LIYKTSGTLGIALAEPVDIPAVSEGSMQVDASKV HPGVISGLNSPACMLSAPLEKQLFYYIGTMLPNT RPHSYVFYQLRCHLSYVALS1NGDKFQYTGAMT SKFLMGTYKRVTEKGDEHVLSLVFGKTKDLPDL RGPFSYPSLTSAQSGEYSLVIVTTFVHYANFHNYF VPNLKDMFSRAVTMTAASYARYVLQKLVLLEM KGGCREPELETETLTTMFEVSVAFFKVGHAVGE TGNGCVDLRWLAKSFFELTVLKDIIGICYGATVK GMQSYGLERLAAMLMATVKMEELGHLTTEKQ EYALRLATVGYPKAGVYSGLIGGATSVLLSAYN RHPLFQPLHTVMRETLFIGSHVVLRELRLNVTT QGPNLALYQLLSTALCSALEIGEVLRGLALGTES GLFSPCYLSLRFDLTRDKLLSMAPQEATLDQAA VSNAVDGFLGRLSLEREDRDAWHLPAYKCVDRL DKVLMIIPLINVTFIISSDREVRGSALYEASTTYLS SSLFLSPVIMNKCSQGAVAGEPRQIPKIQNFTRTQ KSCIFCGFALLSYDEKEGLETTTYITSQEVQNSIL SSNYFDFDNLHVHYLLLTTNGTVMEIAGLYEER ASGGGSGS ASSGAS ASGSSGSGSGSGSSSASSGAZ77F VPKPNVEVWP VDPPPP VNFNKTAEQEYGDKE VKLPH WTPTLHTFQVPQNYTKANCTYCNTREYTFSYKGCCFY FTKKKHTWNGCFQACAEL YPCTYFYGPTPDILP WTR NLNAIESLWVGVYRVGEGNWTSLDGGTFKVYQIFGSH CTYVSKFSTVPVSHHECSFLKPCLCVSQRSNSGC^GS\ SSGASASGSSGSGSGSGSSSASSGASSGGASGGSGG SGGGSGS ASSGAS ASGSSGSGSGSGSSSASSGASSG GASGGSGGSGESQVRSQFSKPIEKLLNEQVNKEMQ SSNLYMSMSSWSYIHSLDGAGLFLFDHAAEEYEHA KKLnFLNENNVPVQLTSISAPEI IKFEGLTQIFQKAY EHEOHISESINQIVDHAIKSKDHATFNFLQWYVAEQ[ [EEEVLFKDILDKIELIGQENI IGLYLADQYVKGIAK SJRKSgp350D123-bfpFerr EAALLVCQYTIQSLIHLTGEDPGFFNVEIPEFPFYPT 22 N19Q / C31S / S111C CNVCTADVNVTINFDVGGKKHQLDLDFGQLTPHT KAVYQPRGAFGGSENATNLFLLELLGAGELALTMR SKKLPINVTTGEEQQVSLESVDVYFQDVFGTMWCH HAEMQNPVYLIPETVPYIKWDNCNSTNITA WRAQ GLDVTLPLSLPTSAQDSNFSVKTEMLGNEIDIECIM EDGEISQVLPGDNKFNITCSGYESHVPSGGILTSTSP VA TPIPGTGYA YSLRLTPRPVSRFLGNNSIL YVFYSG NGPKASGGDYCIQSNIVFSDEIPASQDMPTNTTDIT YVGDNATYSVPMVTSEDANSPNVTVTAFWA WPNN TETDFKCKWTLTSGTPSGCENISGAFASNRTFDITV SGLGTAPKTLIITRTA TNA TTTTHKVIFSKAPE^ES QVRQQFSKDIEKLLNEQVNKEMQSSNLYMSMSSW SYTHSLIXiAGLFLFDHAAEEYEHAKKEllFLNEXXVPVQLTSISAPEHKFEGLTQIFQKAYEHEQJ JISESINNI26101VDHAIKCKDHATFNFLQWYVAEQHEEEVLFKDILD KIELIGNENHGLYLADQYyKGIAKSRKSGp350D123- EAALLVCQYTIOSLIHLTGEDPGFFNVEIPEFPFYPT 23 bfpferritin CNVCTADVNVTINFDVGGKKHQLDLDFGQLTPHT KAVYQPRGAFGGSENATNLFLLELLGAGELALTMR SKKLPINVTTGEEQQVSLESVDVYFQDVFGTMWCH HAEMQNPVYLIPETVPYIKWDNCNSTNITAWRAQ GLDVTLPLSLPTSAQDSNFSVKTEMLGNEIDIECIM EDGEISQVLPGDNKFNITCSGYESHVPSGGILTSTSP VA TPIPGTGYA YSLRLTPRPVSRFLGNNSIL YVFYSG NGPKASGGDYCIQSNIVFSDEIPASQDMPTNTTDIT YVGDNATYSVPMVTSEDANSPNVTVTAFWA WPNN TETDFKCKWTLTSGTPSGCENISGAFASNRTFDITV SGLGTAPKTLIITRTA TNA TTTTHKVIFSKAPEGSES QVRQQFSKDIEKLLXEQVXKFAIQSSXLYMSAISSW SYTHSEDGAGEFEFDI IAAEEYEH AEKEII Fl. XEXX V PVQLTSISAPEHKFEGLTQII QKAYEHEQI 1ISESIXX 1\ 1)1 lAIKCKDl [AI I XI l. OWY\ Ai:(.)l Il l l A I. I KDII. DEll: ElCiXlAl l(il. YI. Al)QY\ K(jlAKSRKSExemplary 1 ESQVRQQFSKDIEKLLNEQVNKEMQSSNLYMSMSS 24 Bfpferritin WSYTHSLDGAGLFLFDHAAEEYEHAKKLIIFLNEN(from Gp350D123- NVPVQLTSISAPEHKFEGLTQIFQKAYEHEQHISESI bfpferritin) NNIVDHAIKCKDHATFNFLQWYVAEQHEEEVLFKD ILDKIELIGNENHGLYLADQYVKGIAKSRKSExemplary 2 H. ESQVRQQFSKDIEKLLNEQVNKEMNSSNLYMSMSS 25 pylori Ferritin with WCYTHSLDGAGLFLFDHAAEEYEHAKKLIIFLNENbullfrog linker NVPVQLTSISAPEHKFEGLTQIFQKAYEHEQHISESI NNIVDHAIKSKDHATFNFLQWYVAEQHEEEVLFKD ILDKIELIGNENHGLYLADQYVKGIAKSRKSExemplary 3 ESQVRSQFSKDIEKLLNEQVNKEMQSSNLYMSMSS 26 Bfpferritin WSYTHSLDGAGLFLFDHAAEEYEHAKKLIIFLNEN(from Cl 8-2) NVPVQLTSISAPEHKFEGLTQIFQKAYEHEQHISESINQIVDHAIKSKDHATFNFLQWYVAEQHEEEVLFKDILDKIELIGQENHGLYLADQYVKGIAKSRKSCompositions of EBV polypeptide and Liposomal adjuvant

[0167] The inventions provides the following embodiments:1. A composition comprising:(a) an Epstein Barr Virus (EBV) polypeptide comprising one or more polypeptides selected from the group consisting of: an EBV gp42 polypeptide, an EBV gH polypeptide, and an EBV gL polypeptide; and(b) a liposomal adjuvant, wherein the liposomal adjuvant comprises a glycolipid and a saponin.2. The composition of embodiment 1 that comprises two or more of an EBV gp42 polypeptide, an EBV gH polypeptide, and an EBV gL polypeptide.261013. The composition of embodiment 1, wherein the EBV polypeptide comprises an EBV gp42 polypeptide, an EBV gH polypeptide, and an EBV gL polypeptide.4. The composition of embodiment 3, wherein the EBV gL polypeptide(s), the EBV gH polypeptide(s), and the EBV gp42 polypeptide(s), are arranged inN-terminal to C-terminal order.In embodiments 1-4, any of the EBV gL, EBV gH or EBV gp42 polypeptides in Sections A and B can be used.5. The composition of any one of embodiments 3-4, further comprising a linker between each EBV polypeptide.6. The composition of embodiment 5, wherein a linker having a length of at least 40 amino acids and less than or equal to 50 amino acids separates the EBV gL polypeptide and the EBV gH polypeptide.7. The composition of embodiment 6, wherein a linker having a length of 46 or 47 amino acids separates the EBV gL polypeptide and the EBV gH polypeptide.8. The composition of any one of embodiments 5 to 7, wherein a linker having a length of 15 to 60 amino acids separates the EBV gH polypeptide and the EBV gp42 polypeptide.9. The composition of any one of embodiments 5 to 7, wherein a linker having a length of 30 to 50 amino acids separates the EBV gH polypeptide and the EBV gp42 polypeptide.10. The composition of embodiment 9, wherein the linker between the EBV gL polypeptide and the EBV gH polypeptide is 44 to 48 amino acids, and the linker between the EBV gH polypeptide and the EBV gp42 polypeptide is 32 amino acids.11. The composition of embodiment 9, wherein the linker between the EBV gL polypeptide and the EBV gH polypeptide is 46 amino acids, and the linker between the EBV gH polypeptide and the EBV gp42 polypeptide is 32 amino acids.12. The composition of embodiment 5, wherein a linker separates the EBV gL polypeptide and the EBV gH polypeptide and consists of an amino acid sequence that has at least 80%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 7; and / or wherein a linker separates the EBV gH polypeptide and the EBV gp42 polypeptide and consists of an amino acid sequence that has at least 80%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 8.13. The composition of any one of embodiments 1-12, wherein the EBV polypeptide further comprises a ferritin.2610114. The composition of embodiment 13, further comprising a further linker that separates the ferritin and the EBV gp42 polypeptide.15. The composition of embodiment 14, wherein the further linker has a length of 60-100 amino acids, or in specific embodiments, about 88 amino acids or 88 amino acids.16. The composition of embodiment 15, wherein the further linker consists of an amino acid sequence having at least 80%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 9.17. The composition of any one of embodiments 6 to 16, wherein (a) the linker that separates the EBV gL polypeptide and the EBV gH polypeptide, (b) the linker that separates the EBV gH polypeptide and the EBV gp42 polypeptide, or (c) the linker of (a) and the linker of (b) comprises one or more of glycine, asparagine, serine, and alanine.

[0168] In embodiments 5-17, any of the linkers described in Sections C and D can be used.18. The composition of any one of embodiments 13-17, wherein the ferritin comprises an amino acid sequence having at least 80%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 26.19. The composition of any one of embodiments 1-18, wherein the EBV gp42 polypeptide comprises an amino acid sequence having at least 80%, 90%, 95%, 96%, 97%, 98%, or 99% identity’ to residues 856-1038 of SEQ ID NO: 21.20. The composition of any one of embodiments 1-19, wherein the EBV gp42 polypeptide consists or consists essentially of amino acid residues 856-1038 of SEQ ID NO: 21; SEQ ID NO: 1, SEQ ID NO: 2 or SEQ ID NO:3.21. The composition of embodiment 20, wherein the EBV gH polypeptide comprises an ammo acid sequence having at least 80%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to residues 162-823 of SEQ ID NO: 21.22. The composition of embodiment 21, wherein the EBV gH polypeptide consists or consists essentially of amino acid residues 162-823 of SEQ ID NO: 21 or SEQ ID NO: 5.23. The composition of any’ one of embodiments 1-22, wherein the EBV gL polypeptide comprises an amino acid sequence having at least 80%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to residues 1-115 of SEQ ID NO: 21.24. The composition of any one of embodiments 1-23, wherein the EBV gL polypeptide comprises, consists or consists essentially of amino acid residues 1-115 of any one of SEQ ID NOs: 11-21; and SEQ ID NO: 4.

[0169] In some embodiments under embodiment 24, the composition comprises an EBV polypeptide that comprises or consists of an amino acid sequence having at least 80%, at least2610185%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identity to residues 1-1038 of SEQ ID NO: 21. In some embodiments under embodiment 24, the composition comprises an EBV polypeptide that comprises or consists of an amino acid sequence having at least 99% identity to residues 1-1038 of SEQ ID NO: 21. In some embodiments under embodiment 24, the composition comprises an EBV polypeptide that comprises, consists or consists essentially of an amino acid sequence of residues 1-1038 of SEQ ID NO: 21. In some embodiments, the EBV polypeptide comprises or consists of an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identity to residues 1-1126 of SEQ ID NO: 21, and a ferritin. In one embodiment, the ferritin comprises or consists of an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 24, 25 or 26. In some embodiments, the EBV polypeptide comprises or consists of an amino acid sequence having at least 99% identity to residues 1-1126 of SEQ ID NO: 21, and a ferritin that comprises or consists of an amino acid sequence having at least 99% sequence identity to SEQ ID NO: 24, 25 or 26. In some embodiments, the EBV polypeptide comprises, consists or consists essentially of an amino acid sequence having at least 100% identity to residues 1-1126 of SEQ ID NO: 21, and a ferritin that comprises or consists of an amino acid sequence having at least 100% sequence identity to SEQ ID NO: 24, 25 or 26.25. The composition of any one of embodiments 1-24, wherein the EBV polypeptide comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to amino acid residues 1-1038 of SEQ ID NO: 21.26. The composition of embodiment 25, wherein the EBV polypeptide comprises, consists or consists essentially of a sequence of any one of SEQ ID NOs: 11-21.27. The composition of any one of embodiments 1-26, wherein the EBV polypeptide further comprises EBV gp220.28. The composition of embodiment 27, wherein the EBV gp220 polypeptide comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 6.29. A composition comprising an EBV gp220 polypeptide; and a liposomal adjuvant, wherein the liposomal adjuvant comprises a glycolipid and a saponin.30. The composition of embodiment 29, wherein the EBV gp220 polypeptide comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%,26101at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 6.31. The composition of embodiment 29 or 30, wherein the EBV gp220 polypeptide further comprises a ferritin.32. The composition of embodiment 29 or 30, wherein the ferritin comprises an amino acid sequence having at least 80%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 24, 25 or 26.33. The composition of embodiment 32, wherein the ferritin consists of the amino acid sequence of SEQ ID NO: 24, 25 or 26.34. The composition of embodiment 31, wherein the EBV gp220 polypeptide consists of SEQ ID NO: 22 or 23.

[0170] In some embodiments, a composition comprises (a) a first EBV polypeptide comprising an EBV gL polypeptide, an EBV gH polypeptide, and / or an EBV gp42 polypeptide, and optionally a ferritin, (b) a second EBV polypeptide comprising an EBV gp220 polypeptide and optionally a ferritin, and (c) a liposomal adjuvant. In one embodiment, the first EBV polypeptide comprises or consists of an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identity to residues 1-1038 of SEQ ID NO: 21. In one embodiment, the first EBV polypeptide comprises or consists of an amino acid sequence having at least 99% identity to residues 1-1038 of SEQ ID NO: 21. In one embodiment, the first EBV polypeptide comprises an EBV polypeptide that comprises, consists or consists essentially of an amino acid sequence of residues 1-1038 of SEQ ID NO: 21. In one embodiment, the first EBV polypeptide comprises or consists of an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identity to residues 1-1126 of SEQ ID NO: 21, and a ferritin. In one embodiment, the ferritin comprises or consists of an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 24, 25 or 26. In some embodiments, the first EBV polypeptide comprises or consists of an amino acid sequence having at least 99% identity to residues 1-1126 of SEQ ID NO: 21, and a ferritin that comprises or consists of an amino acid sequence having at least 99% sequence identity to SEQ ID NO: 24, 25 or 26. In some embodiments, the first EBV polypeptide comprises or consists of an amino acid sequence having at least 100% identity to residues 1-1126 of SEQ ID NO: 21, and a ferritin that comprises or consists of an amino acid sequence having at least 100% sequence identity to SEQ ID NO: 24, 25or 26. In some embodiments, the first EBV polypeptide comprises, consists or consists essentially of an amino acid sequence of SEQ ID NO: 21.

[0171] In one embodiment, the second EBV polypeptide comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%. at least 99%. or 100% sequence identity to SEQ ID NO: 6. In one embodiment, the second EBV polypeptide further comprises a ferritin. In one embodiment, the ferritin comprises an amino acid sequence having at least 80%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 24, 25 or 26. In one embodiment, the ferritin comprises, consists or consists essentially of the amino acid sequence of SEQ ID NO: 24, 25 or 26. In a preferred embodiment, the second EBV polypeptide comprises, consists or consists essentially of the amino acid sequence of SEQ ID NO: 22 or 23. In a preferred embodiment, the first EBV polypeptide comprises, consists or consists essentially of the amino acid sequence of SEQ ID NO: 21 and the second EBV polypeptide comprises, consists or consists essentially of the amino acid sequence of SEQ ID NO: 23.

[0172] In some embodiments, the EBV gp220 polypeptide of the second EBV polypeptide comprises or consists of an amino acid sequence having at least 99% sequence identity to SEQ ID NO: 6, and a ferritin. In one embodiment, the ferritin comprises or consists of an amino acid sequence having at least 99% sequence identity to SEQ ID NO: 24.

[0173] In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in a 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9, 1:10, 2:1, 3:1, 4:1, 5:1, 6:1, 7:1, 8:1, 9:1, or 10: 1 ratio by weight or wherein the second EBV polypeptide and first EBV polypeptide are present in a 1:1, 1:2, 1:3. 1:4, 1:5, 1:6, 1:7, 1:8, 1:9, 1:10, 2:1. 3:1, 4:1, 5:1, 6:1, 7:1, 8:1, 9:1, or 10: 1 molar ratio. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in about 1: 1 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in about 1:2 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in about 2:3 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in about 1:3 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in about 1:3.33 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in about 1:4 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in about 2: 1 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in about 3:2 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in about 3:1 ratio by weight. In some embodiments, the first EBV polypeptide andsecond EBV polypeptide are present in about 3.33:1 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in about 4: 1 ratio by weight.

[0174] In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in 1: 1 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in 1:2 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in 2:3 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in 1:3 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in 1:3.33 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in 1:4 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in 2: 1 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in 3:2 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in 3:1 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in 3.33: 1 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in 4: 1 ratio by weight.35. The composition of any one of embodiments 1-34, wherein the composition is a solution of the EBV polypeptide and the liposomal adjuvant.36. The composition of any one of embodiments 1-35, wherein the liposomal adjuvant comprises a glucopyranosyl lipid A (“GLA”) and saponin selected from the group consisting of QS-7, QS-21, QS-17 and QS-I8.37. The composition of any one of embodiments 1-35, wherein the glycolipid has the structure set forth in Formula 1:OHFormula Iwherein Ri is P(O)(OH)2;wherein R2 is selected from H, C(O)CH2CH(OH)Cn alkyl, and C(O)CH2CH(OC(O)Cn- C13 alkyljCn alkyl;wherein R3is C(O)CH2CH(OC(O)Cn-Ci3alkyl)Cn alkyl;wherein R4 is selected from H, C(O)CH2CH(OH)Cn alkyl, and C(O)CH2CH(OC(O)Cn- Ci3alkyl)Cn alkyd;wherein R5 is selected from C(O)CH2CH(OH)Cn alkyl and C(O)CH2CH(OC(O)CI3-CIS alkyl)Cn alkyl; andwherein Re is H;or a pharmaceutically acceptable salt thereof.38. The composition of embodiment 37, wherein the glycolipid has the structure set forth in Formula II:FormulaIIwherein Ri is H or C(O)Cn-Ci3alkyl;wherein R2is C11 alkyl;wherein R3is C11-C13 alkyl;wherein R4 is Cn alkyl;wherein R5 is H or C(O)CH2CH(OH)Cn alkyl;wherein Re is H or C(O)Ci3-Ci5 alkyl; andwherein R7 is C11 alkyl;or a pharmaceutically acceptable salt thereof.39. The composition of embodiment 38, wherein the glycolipid has the structure set forth in Formula II:26101wherein Ri is C(O)Cn-Ci3 alkyl;wherein R2 is Cn alkyd;wherein R3 is C11-C13 alkyl;wherein R4 is Cn alkyl;wherein Rs is C(O)CH2CH(OH)Cn alkyl;wherein Re is H; andwherein R7 is Cn alkyl;or a pharmaceutically acceptable salt thereof.40. The composition of embodiment 38 or 39, wherein the glycolipid has the structure set forth in Formula IIA:Formula II A, or a pharmaceutically acceptable salt thereof.41. The composition of embodiment 37, wherein the glycolipid has the structure set forth in Formula III:26101Formula III, or a pharmaceutically acceptable salt thereof.In one aspect under embodiment 37, the glycolipid is Formula IIIA, or a pharmaceutically acceptable salt thereof.26101v eFormula IIIA42. The composition of embodiment 37, wherein the glycolipid has the structure set forth in Formula IV:26101Formula IV, or a pharmaceutically acceptable salt thereof.43. The composition of embodiment 37, wherein the glycolipid has the structure set forth in Formula V:26101Formula V, or a pharmaceutically acceptable salt thereof.44. The composition of embodiment 37, wherein the glycolipid has the structure set forth in Formula VI:26101Formula VI, or a pharmaceutically acceptable salt thereof.45. The composition of any one of embodiments 1-44, wherein the liposomal adjuvant further comprises l,2-dioleoyl-sn-glycero-3 -phosphocholine (DOPC).46. The composition of any one of embodiments 1-45, wherein the liposomal adjuvant further comprises cholesterol.47. A pharmaceutical unit dose comprising about 5 pg to about 200 pg of EBV polypeptide consisting of any one of the amino acid sequences of SEQ ID NOs: 11-21. and about 0.1 mg to about 3 mg of a liposomal adjuvant, wherein the liposomal adjuvant comprises 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and26101the glycolipid of Formula IIIAv eor a pharmaceutically acceptable salt thereof.

[0175] In one aspect under embodiment 47, the pharmaceutical unit dose comprises about 5 pg to about 150 pg of EBV polypeptide comprising or consisting of any one of the amino acid sequences of SEQ ID NOs: 11-21. In another aspect under embodiment 47, the pharmaceutical unit dose comprises about 10 pg to about 1 0 pg of EBV polypeptide comprising or consisting of any one of the amino acid sequences of SEQ ID NOs: 11-21. In another aspect under embodiment 47, the pharmaceutical unit dose comprises about 10 pg to about 100 pg of EBV polypeptide comprising or consisting of any one of the amino acid sequences of SEQ ID NOs: 11-21.48. The pharmaceutical unit dose of embodiment 47, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of about 10 pg to about 100 pg, and about 0.1 mg to about 2 mg of the liposomal adjuvant, wherein the amount of26101Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula III. Formula 111A or a pharmaceutically acceptable salt thereof is each at about 5 pg to about 100 pg.

[0176] In one aspect under embodiment 47 or 48, the pharmaceutical unit dose comprises about 0.1 mg to about 0.2 mg of a liposomal adjuvant. In one aspect under embodiment 47 or 48, the pharmaceutical unit dose comprises about 0.05 mg to about 0.3 mg of a liposomal adjuvant.49. The composition or pharmaceutical unit dose of any one of embodiments 41 and 47-48, wherein the amount of Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula III, Formula IIIA or a pharmaceutically acceptable salt thereof is each at about 10 pg to about 75 pg.50. The composition or pharmaceutical unit dose of any one of embodiments 41 and 47-48, wherein the amount of Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula III, Formula IIIA or a pharmaceutically acceptable salt thereof is each at about 10 pg to about 50 pg.51. The composition of any one of embodiments 1 -46, wherein the ratio of saponin to glycolipid is 1-5: 1-3 by weight.52. The composition or pharmaceutical unit dose of any one of embodiments 46-50, wherein the ratio of l,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid is 10-30:2-15:0.1-5:0.1-3 by weight.52a. The composition or pharmaceutical unit dose of any one of embodiments 46-50, wherein the ratio of l,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria21 (QS-21) and the glycolipid is 20:5:1:1 by weight.53. The pharmaceutical unit dose of any one of embodiments 47-50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 10 pg, and the liposomal adjuvant is in the amount of 0.27 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.54. The pharmaceutical unit dose of any one of embodiments 47-50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 30 pg, and the liposomal adjuvant is in the amount of 0.27 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.55. The pharmaceutical unit dose of any one of embodiments 47-50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 100 pg, and the liposomal adjuvant is in the amount of 0.27 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.2610156. The pharmaceutical unit dose of any one of embodiments 47-50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 10 pg, and the liposomal adjuvant is in the amount of 0.68 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.57. The pharmaceutical unit dose of any one of embodiments 47-50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 30 pg, and the liposomal adjuvant is in the amount of 0.68 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.58. The pharmaceutical unit dose of any one of embodiments 47-50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 100 pg, and the liposomal adjuvant is in the amount of 0.68 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by w eight.59. The pharmaceutical unit dose of any one of embodiments 47-50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 10 pg, and the liposomal adjuvant is in the amount of 1.35 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.60. The pharmaceutical unit dose of any one of embodiments 47-50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 30 pg, and the liposomal adjuvant is in the amount of 1.35 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.61. The pharmaceutical unit dose of any one of embodiments 47-50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 100 pg, and the liposomal adjuvant is in the amount of 1.35 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by w eight.62. A pharmaceutical unit dose comprising about 5 pg to about 200 pg of EBV polypeptide consisting of the amino acid sequence of SEQ ID NO: 23, and about 0.1 mg to about 3 mg of a liposomal adjuvant, wherein the liposomal adjuvant comprises 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the26101glycolipid of Formula IIIAv e, or a pharmaceutically acceptable salt thereof.63. The pharmaceutical unit dose of embodiment 62, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of about 10 pg to about 100 pg, and about 0.1 mg to about 2 mg of the liposomal adjuvant, wherein the amount of Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is each at about 5 pg to about 100 pg.64. The pharmaceutical unit dose of any one of embodiments 62-63, wherein the ratio of l,2-dioleoyl-sn-glycero-3-phosphocholine (DOPE), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 10-30:2-15:0.1-5:0.1-3 by weight.2610165. The pharmaceutical unit dose of any one of embodiments 62-63, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 10 pg, and the liposomal adjuvant is in the amount of 0.27 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.66. The pharmaceutical unit dose of any one of embodiments 62-63, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 30 pg, and the liposomal adjuvant is in the amount of 0.27 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.67. The pharmaceutical unit dose of any one of embodiments 62-63, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 100 pg, and the liposomal adjuvant is in the amount of 0.27 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by w eight.68. The pharmaceutical unit dose of any one of embodiments 62-63, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 10 pg, and the liposomal adjuvant is in the amount of 0.68 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.69. The pharmaceutical unit dose of any one of embodiments 62-63, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 30 pg, and the liposomal adjuvant is in the amount of 0.68 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.70. The pharmaceutical unit dose of any one of embodiments 62-63, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 100 pg, and the liposomal adjuvant is in the amount of 0.68 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by w eight.71. The pharmaceutical unit dose of any one of embodiments 62-63, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 10 pg, and the liposomal adjuvant is in the amount of 1.35 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-261013-phosphocholine (DOPC), cholesterol Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.72. The pharmaceutical unit dose of any one of embodiments 62-63, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 30 pg, and the liposomal adjuvant is in the amount of 1.35 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.73. The pharmaceutical unit dose of any one of embodiments 62-63, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 100 pg, and the liposomal adjuvant is in the amount of 1.35 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.74. A pharmaceutical unit dose comprising about 10 pg to about 100 pg of a first EBV polypeptide consisting of the amino acid sequence of SEQ ID NO: 21 and about 10 pg to about 100 pg of a second EBV polypeptide consisting of the amino acid sequence of SEQ ID NO: 23, wherein the first and the second EBV polypeptide are at a 1: 1 weight ratio; and about 0.27 mg to about 1.35 mg of a liposomal adjuvant, wherein the liposomal adjuvant comprises l,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC). cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA26101salt thereof, at a ratio of 10-30:2-15: 1-5: 1-3 by weight.74a. A pharmaceutical unit dose comprising about 10 pg to about 100 pg of a first EBV polypeptide consisting of the amino acid sequence of SEQ ID NO: 21 and about 10 pg to about 100 pg of a second EBV polypeptide consisting of the amino acid sequence of SEQ ID NO: 23, wherein the first and the second EBV polypeptide are at a 1: 1 weight ratio; and about 0.54 mg to about 1.36 mg of a liposomal adjuvant, wherein the liposomal adjuvant comprises l,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA, or a pharmaceutically acceptable salt thereof, at a ratio of 10-30:2-15: 1-5: 1-3 by weight.75. The pharmaceutical unit dose of embodiment 74, wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5:1:1 by weight.26101

[0177] In one aspect under embodiment 74 or 75. the pharmaceutical unit dose comprises about 0.1 mg to about 3 mg of a liposomal adjuvant. In certain embodiments of the foregoing embodiments, the pharmaceutical dose comprises 0.27 mg of a liposomal adjuvant. In certain embodiments of the foregoing embodiments, the pharmaceutical dose comprises 0.68 mg of a liposomal adjuvant. In certain embodiments of the foregoing embodiments, the pharmaceutical dose comprises 1.35 mg of a liposomal adjuvant. In certain embodiments of the foregoing embodiments, the pharmaceutical dose comprises about 0.54 mg of a liposomal adjuvant. In certain embodiments of the foregoing embodiments, the pharmaceutical dose comprises about 1.36 mg of a liposomal adjuvant. In certain embodiments of the foregoing embodiments, the pharmaceutical dose comprises about 2.70 mg of a liposomal adjuvant. In certain embodiments of the foregoing embodiments, the pharmaceutical dose comprises about 0.4 mg to about 0.7 mg of a liposomal adjuvant. In certain embodiments of the foregoing embodiments, the pharmaceutical dose comprises about 1.20 mg to about 1.50 mg of a liposomal adjuvant. In certain embodiments of the foregoing embodiments, the pharmaceutical dose comprises about 2.5 mg to about 2.90 mg of a liposomal adjuvant. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in a 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9, 1:10, 2:1, 3:1, 4: 1, 5:1, 6:1, 7:1, 8:1, 9:1, or 10:1 ratio by weight or wherein the second EBV polypeptide and first EBV polypeptide are present in a 1:1, 1:2. 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9.1:10, 2:1, 3:1, 4:1, 5:1, 6:1, 7:1, 8:1, 9:1, or 10:1 molar ratio. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in about 1:1 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in about 1:2 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in about 2:3 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in about 1:3 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in about 1:3.33 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in about 1:4 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in about 2: 1 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in about 3:2 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in about 3:1 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in about 3.33: 1 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in about 4: 1 ratio by weight.26101

[0178] In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in 1: 1 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in 1:2 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in 2:3 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in 1:3 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in 1:3.33 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in 1:4 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in 2: 1 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in 3:2 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in 3:1 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in 3.33:1 ratio by weight. In some embodiments, the first EBV polypeptide and second EBV polypeptide are present in 4: 1 ratio by weight. In one embodiment, the first EBV polypeptide is at 10 pg. and the second EBV polypeptide is at 30 pg. In one embodiment, the first EBV polypeptide is at 10 pg, and the second EBV polypeptide is at 100 pg. In one embodiment, the first EBV polypeptide is at 30 pg, and the second EBV polypeptide is at 10 pg. In one embodiment, the first EBV polypeptide is at 30 pg, and the second EBV polypeptide is at 100 pg. In one embodiment, the first EBV polypeptide is at 100 pg, and the second EBV polypeptide is at 10 pg. In one embodiment, the first EBV polypeptide is at 100 pg, and the second EBV polypeptide is at 30 Eg- 76. The pharmaceutical unit dose of embodiment 74 or 75, wherein each of the first and second EBV polypeptide is at 10 pg.77. The pharmaceutical unit dose of embodiment 74 or 75, wherein each of the first and second EBV polypeptide is at 30 pg.78. The pharmaceutical unit dose of embodiment 74 or 75, wherein each of the first and second EBV polypeptide is at 100 pg.79. The composition or pharmaceutical unit dose of any one of embodiments 1-78, wherein the average particle size of the liposomal adjuvant is about 50-200 nm.

[0179] In one aspect under embodiment 79, the average particle size of the liposomal adjuvant is about 100 to about 200 nm. In one aspect under embodiment 79, the average particle size of the liposomal adjuvant is about 120 to about 180 nm. In one aspect under embodiment 79, the average particle size of the liposomal adjuvant is about 80 to about 120 nm. In one aspect under26101embodiment 79, the average particle size of the liposomal adjuvant is about 100 nm. In one aspect under embodiment 79, the average particle size of the liposomal adjuvant is about 120 nm.

[0180] In one aspect under any one of embodiments 37-79, the pharmaceutically acceptable salt is an ammonium, sodium, potassium, calcium, or organic amine salt such as ethylamine, diethylamine, and triethylamine salt. In one aspect under any one of the embodiments 37-79, the pharmaceutically acceptable salt is an ammonium salt.80. The composition or pharmaceutical unit dose of any one of embodiments 1-79 wherein the liposomal adjuvant further comprises a buffer selected from the group consisting of: acetic acid, histidine, citrate. Bis-Tris, HEPES, phosphate, MES, sodium chloride, succinate. Tris, and combinations thereof.81. The composition or pharmaceutical unit dose of embodiment 80, wherein the buffer is present in the amount of about ImM to about lOOmM.82. The composition or pharmaceutical unit dose of any one of embodiments 1-79, wherein the liposomal adjuvant further comprises 5 mM - 100 mM potassium dihydrogen phosphate at pH 5.1 - 7.0, 1 mM to 50 mM disodium phosphate at pH 5.1 - 7.0, and 25 mM - 300 mM NaCl at pH 5.1 - 7.0.83. The composition or pharmaceutical unit dose of any one of embodiments 1-79, wherein the liposomal adjuvant further comprises 40 mM - 50 mM potassium dihydrogen phosphate at pH 5.1 - 7.0, 5 mM to 15 mM disodium phosphate at pH 5.1 - 7.0, and 50 mM - 150 mM NaCl at pH 5.1 - 7.0.84. A method of inducing an immune response to an Epstein Barr Virus (EBV) in a human patient comprising administering to the patient a therapeutically effective amount of the composition or pharmaceutical unit dose of any one of embodiments 1-83.85. A method of preventing infection of or reducing the likelihood of infection of a human patient by an Epstein Barr Virus (EBV) comprising administering to the patient a therapeutically effective amount of the composition or pharmaceutical unit dose of any one of embodiments 1-83.86. A method of preventing cancer caused by Epstein Barr Virus (EBV) of a human patient, comprising administering to the patient a therapeutically effective amount of the composition or pharmaceutical unit dose of any one of embodiments 1-83.87. The method of embodiment 86, wherein the cancer is Burkitt’s lymphoma, Hodgkin’s lymphoma, or nasopharyngeal cancer.2610188. The composition or pharmaceutical unit dose of any one of embodiments 1-83 for use in preventing infection of or reducing the likelihood of infection of a human patient by an Epstein Barr Virus (EBV).89. The composition or pharmaceutical unit dose of any one of embodiments 1-83 for use in inducing an immune response to an Epstein Barr Virus (EBV)90. The composition or pharmaceutical unit dose of any one of embodiments 1-83 for use in preventing cancer caused by Epstein Barr Virus (EBV) of a human patient.91. A method of inducing an immune response to an Epstein Barr Virus (EBV) in a human patient comprising co-administering to the patient a therapeutically effective amount of the EBV polypeptide described in any one of embodiments 1-78 and the liposomal adjuvant described in any one of embodiments 1-83.92. A method of preventing infection of or reducing the likelihood of infection of a human patient by an Epstein Barr Virus (EBV) comprising co-administering to the patient a therapeutically effective amount of the EBV polypeptide of any one of embodiments 1-78 and a therapeutically effective amount of the liposomal adjuvant of any one of embodiments 1-83.93. A method of preventing cancer caused by Epstein Barr Virus (EBV) of a human patient, comprising co-administering to the patient a therapeutically effective amount of the EBV polypeptide of any one of embodiments 1-78 and a therapeutically effective amount of the liposomal adjuvant of any one of embodiments 1-83.94. The method of embodiment 93, wherein the cancer is Burkitt’s lymphoma, Hodgkin's lymphoma, or nasopharyngeal cancer.

[0181] In one aspect of the methods under any one of embodiments 91-94, the method comprises co-administering the EBV polypeptide that comprises or consists of the amino acid sequence of SEQ ID NO: 21 in the amount of about 10 pg to about 100 pg, and about 0.27 mg to about 1.35 mg of a liposomal adjuvant.

[0182] In one aspect of the methods under any one of embodiments 91-94, the method comprises co-administering the EBV polypeptide that comprises or consists of the amino acid sequence of SEQ ID NO: 23 in the amount of about 10 pg to about 100 pg, and about 0.27 mg to about 1.35 mg of a liposomal adjuvant.

[0183] In one embodiment of the foregoing embodiments, the EBV polypeptide is in the amount of 100 pg, and the liposomal adjuvant is in the amount of 0.27 mg. In one embodiment of the foregoing embodiments, the EBV polypeptide is in the amount of 100 pg, and the liposomal adjuvant is in the amount of 0.68 mg. In one embodiment of the foregoing embodiments, the26101EBV polypeptide is in the amount of 100 pg, and the liposomal adjuvant is in the amount of 1.35 mg.

[0184] In one embodiment of the foregoing embodiments, the EBV polypeptide is in the amount of 30 pg, and the liposomal adjuvant is in the amount of 0.27 mg. In one embodiment of the foregoing embodiments, the EBV polypeptide is in the amount of 30 pg, and the liposomal adjuvant is in the amount of 0.68 mg. In one embodiment of the foregoing embodiments, the EBV polypeptide is in the amount of 30 pg, and the liposomal adjuvant is in the amount of 1.35 mg. In one embodiment of the foregoing embodiments, the EBV polypeptide is in the amount of 10 pg, and the liposomal adjuvant is in the amount of 0.27 mg. In one embodiment of the foregoing embodiments, the EBV polypeptide is in the amount of 10 pg, and the liposomal adjuvant is in the amount of 0.68 mg. In one embodiment of the foregoing embodiments, the EBV polypeptide is in the amount of 10 pg, and the liposomal adjuvant is in the amount of 1.35 mg. In one embodiment, the ratio of l,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is at a ratio of 10-30:2-15: 1-5: 1-3 by weight. In one embodiment, the ratio of l,2-dioleoyl-sn-glycero-3 -phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is at a ratio of 20: 5: 1: 1 by weight.

[0185] In one aspect under the methods and use of embodiments 84 to 94, the composition or pharmaceutical unit dose, or EBV polypeptide and liposomal adjuvant is administered in a 1 dose, 2 dose, 3 dose or 4 dose regimen. In one embodiment, a dose is administered, and then a second dose is administered about 50 to 65 days from the first dose, and then a third dose is administered 160 to 190 days from the first dose. In another embodiment, two doses are administered. In one embodiment, one dose is administered followed by a second dose after about 2 months. In another embodiment, three doses are administered, one dose is administered followed by a second dose after about 2 months, and followed by a third dose at about 6 months.

[0186] Any of the EBV polypeptides described in Sections A-G and Table 2 are suitable for use in the compositions or pharmaceutical compositions and methods of the invention. The EBV polypeptide can be field mixed with a liposomal adjuvant of the invention (e.g., mixed by the clinician just prior to administration to a patient, lyophilized EBV polypeptide reconstituted with liposomal adjuvant solution) or can be formulated together with the liposomal adjuvant.

[0187] Compositions of the invention may be administered subcutaneously, topically, orally, on the mucosa, intravenously, or intramuscularly. In a preferred embodiment, the compositions are administered intramuscularly. The compositions are administered in an amount sufficient to elicit26101a protective response. Compositions can be administered by various routes, for example, orally, parenterally, subcutaneously, on the mucosa, or intramuscularly. The dose administered may vary depending on the general condition, sex, weight and age of the patient, and the route of administration.

[0188] Compositions of the invention, as highlighted in the various embodiments above, may be referred to as immunogenic compositions. Compositions of the invention, as highlighted in the various embodiments above may also be referred to as vaccines or vaccine compositions.

[0189] In some embodiments, a vaccine composition is provided that comprises about 5 to about 200 pg of EBV polypeptide of SEQ ID NO: 21 or 23. and about 0.1 mg to about 3 mg liposomal adjuvant.

[0190] The vaccines of the invention comprise protein antigens containing the antigenic determinants required to induce the generation of neutralizing antibodies in the subject. The vaccines are expected to be sufficiently safe to be administered without the risk of clinical infection, have no toxic side effects, are stable, compatible with conventional carriers and can be administered effectively.

[0191] Pharmaceutical compositions, formulations, and vaccines of the invention may be administered subcutaneously, topically, orally, on the mucosa, intravenously, or intramuscularly. The pharmaceutical compositions, formulations, and vaccines are administered in an amount sufficient to elicit a protective response. Vaccines, pharmaceutical compositions and formulations can be administered by various routes, for example, orally, parenterally, subcutaneously, on the mucosa, or intramuscularly. The dose administered may vary depending on the general condition, sex, weight and age of the patient, the route of administration and the ty pe of EBV polypeptide in the vaccine. The vaccine, pharmaceutical composition, or formulation may be in the form of a capsule, suspension, elixir or solution. Such vaccine, pharmaceutical compositions or formulations may be formulated with an immunologically acceptable carrier.Kits of the Invention

[0192] Also provided herein are kits including any of the compositions or pharmaceutical compositions as described above and instructions for use.

[0193] Also provided herein are kits including (1) a pharmaceutical composition comprising any of the EBV polypeptides described in Sections A-G and (2) a liposomal adjuvant described in the Liposomal Adjuvant Section.26101

[0194] In some embodiments, the kit includes a buffer. In some embodiments, the kit includes a tonicity modifier. In some embodiments, the kit includes a detergent.

[0195] In some embodiments of the kits, the kit includes a label or packaging insert that includes a description of the components and / or instructions for use in vivo of the components therein. In some embodiments, the kits include instructions for co-administering (or vaccinating) (1) the pharmaceutical composition comprising the EBV polypeptide and (2) the liposomal adjuvant. In some embodiments, the kits include instructions for admixing (1) the pharmaceutical composition comprising the EBV polypeptide and (2) the liposomal adjuvant and subsequentially administering (or vaccinating) the admixture to a patient. The kit may further comprise a diluent for reconstituting the EBV polypeptide or adjuvant.Methods of Treatment of the Invention

[0196] Also provided herein is a method of inducing an immune response to an Epstein Barr Virus (EBV) in a human patient comprising administering to the patient a therapeutically effective amount of any one of the EBV polypeptides described in Sections A to G and Table 2 and a therapeutically effective amount of a liposomal adjuvant described in the Liposomal Adjuvant Section.

[0197] Also provided herein is a method of inducing an immune response to an Epstein Barr Virus (EBV) in a human patient comprising administering to the patient a therapeutically effective amount of the compositions or pharmaceutical unit dose described in the Section Compositions of EBV polypeptide and Liposomal adjuvant.

[0198] Also provided herein is a method of preventing infection of or reducing the likelihood of infection of a human patient to an Epstein Barr Virus (EBV) in a human patient comprising administering to the patient a therapeutically effective amount of any one of the EBV polypeptides described in Sections A to G and Table 2 and a therapeutically effective amount of a liposomal adjuvant described in the Liposomal Adjuvant Section.

[0199] Also provided herein is a method of preventing infection of or reducing the likelihood of infection of a human patient to an Epstein Barr Virus (EBV) in a human patient comprising administering to the patient a therapeutically effective amount of the compositions or pharmaceutical unit dose described in the Section Compositions of EBV polypeptide and Liposomal adjuvant.

[0200] In some embodiments, the liposomal adjuvant is formulated separately from the EBV polypeptide. In some embodiments, the liposomal adjuvant is formulated with the EBV polypeptide. In some embodiments, the liposomal adjuvant and EBV polypeptide are field-mixed26101to form a pharmaceutical composition prior to administration to the patient. In some embodiments, the liposomal adjuvant and EBV polypeptide are administered sequentially to a patient.

[0201] Also provided herein is a method of delivering the pharmaceutical unit dose or compositions of the invention to a subject that comprises administering to the subject the compositions or pharmaceutical unit dose described in the Section Compositions of EBV polypeptide and Liposomal adjuvant, whereby the administration of the pharmaceutical unit dose or composition induces a neutralizing titer against the EBV antigen in the subject, and wherein the pharmaceutical unit dose or composition provides enhanced or comparable neutralizing titers when compared to the same composition that is formulated without a liposomal adjuvant.

[0202] Also provided herein is a method for preventing cancer caused by Epstein Barr Virus (EBV) of a human patient, comprising administering to the patient a therapeutically effective amount of any one of the EBV polypeptides described in Sections A to G and Table 2 and a therapeutically effective amount of a liposomal adjuvant described in the Liposomal Adjuvant Section, wherein the cancer is Burkitt’s lymphoma, Hodgkin’s lymphoma, or nasopharyngeal cancer.

[0203] Also provided herein is a method for preventing cancer caused by Epstein Barr Virus (EBV) of a human patient, comprising administering to the patient a therapeutically effective amount of the compositions or pharmaceutical unit dose described in the Section Compositions of EBV polypeptide and Liposomal adjuvant, wherein the cancer is Burkitt’s lymphoma, Hodgkin's ly mphoma, or nasopharyngeal cancer.

[0204] Embodiments of the invention also include one or more of the pharmaceutical compositions or compositions described herein (1) for use in, (2) for use as a medicament or composition for, or (3) for use in the preparation of a medicament for: (a) therapy (e.g., of the human body); (b) medicine; (c) induction of an immune response against the EBV polypeptide in the vaccine (d) decreasing the likelihood of EBV infection in a patient; (e) prevention of infection with EBV, (f) prevention or reduction of the likelihood of Burkitt's lymphoma, (g) prevention or reduction of the likelihood of Hodgkin’s lymphoma, (h) prevention or reduction of the likelihood of nasopharyngeal cancer.

[0205] In embodiment 1, a composition comprises an Epstein Barr Virus (EBV) poly peptide comprising one or more polypeptides selected from the group consisting of: an EBV gp42 polypeptide, an EBV gH polypeptide, and an EBV gL polypeptide; and a liposomal adjuvant described herein. In embodiment 2, the composition of embodimentl is provided, wherein the composition is made by mixing an EBV vaccine and a liposomal adjuvant described herein.26101

[0206] All publications mentioned herein are incorporated by-reference for the purpose of describing and disclosing methodologies and materials that might be used in connection with the invention.

[0207] Having described embodiments of the invention with reference to the accompanying drawings, it is to be understood that the invention is not limited to those precise embodiments, and that various changes and modifications may be used by one skilled in the art without departing from the scope or spirit of the invention as defined in the appended claims.

[0208] The following examples illustrate, but do not limit the invention.EXAMPLESExample 1: Preparation of Formulation Buffer

[0209] A buffer formulation was prepared by dispensing water for injection (WFI) into a buffer preparation vessel. 15.54 grams of powdered di-sodium hydrogen phosphate dibasic. 54.31 grams of potassium dihydrogen phosphate, and 56.75 grams sodium chloride were then added to the vessel. The resulting formulation was stirred using a stir bar until dissolution of ingredients was adjusted to pH 6.2±0.1. WFI was added to bring the buffer to the final target volume of 10L. The buffer was then filtered through a 0.22 pg filter to form the Formulation Buffer.Example 2: Preparation of a Liposomal Adjuvant

[0210] The liposomal adjuvant formulation was prepared by mixing 2mL of 10.0 mg / mL of cholesterol / chloroform stock solution (Avanti’s Cat# 700000 dissolved in chloroform), 3.2 mL of 25mg / mL DOPC / chloroform stock solution (Avanti’s Cat# 850375C), and 4mL of chloroform, and 2mL of glycolipid that is the ammonium salt of Formula IIIA above (2mg / mL) in chloroform: methanol (9:1 v / v ratio) solution. The individual lipid solutions were then pooled together to make a lipid stock solution in a round bottom flask.

[0211] The round bottom flask containing the lipid stock solution was attached to a rotatory evaporator. The rotation rate was set at 280 rpm and a water bath at 20°C, the pressure set point was incrementally reduced to prevent boiling from lOOOmbar dow n to 15mbar gradually and the rotatory evaporation process was continued until the lipid stock solution formed a thin lipidic film. The round bottom flask was purged with nitrogen gas to complete the drying process.

[0212] The thin lipidic film in the round bottom flask was then reconstituted with 14.0 mL of the Formulation Buffer described in Example 1 (50 mM Na / K Phosphate, 100 mM NaCl pH 6.2). The combination was then heated at 50°C until a poly disperse mixture of liposomes was formed.26101

[0213] The resulting poly disperse liposome mixture was then passed through a high-pressure extruder device fitted with a lOOnm pore size polycarbonate membrane. The pressure was adjusted manually between 200psi - 380psi to drive the liposomes through the polycarbonate membrane. The extruder temperature was controlled at 50°C during the extrusion. The liposomes were extruded 6 times (6 passes) through the membrane to get the particle size ~100nm. The resulting liposomes were placed in a glass bottle.Example 3: Preparation of Liposomal Adjuvant Formulation (Liposomal adjuvant B)

[0214] QS-21 (Desert King International Adjuvant QS-21, Fisher Scientific cat # NC0949192) was dissolved in lOmg / mL of the Formulation Buffer described in Example 1 (50 mM Na / K Phosphate, 100 mM NaCl pH 6.2). 0.4mL of lOmg / mL QS-21 solution was added to the glass bottle containing the extruded liposomes and mixed well. An additional 7.2 rnL of the Formulation Buffer described in Example 1 (50 mM Na / K Phosphate, 100 mM NaCl pH 6.2) was added to the glass bottle to arrive at the final liposomal formulation having a Formula III A concentration of 0.2mg / mL and QS-21 concentration of 0.2mg / mL. The mass ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the ammonium salt of Formula IIIA is 20:5: 1: 1 by weight.Example 4: EBV Antigen-Specific Antibody Titers in Mice Immunized With 2 Doses of gp350-FNP and gL-gH-gp42-FNP dual particle vaccine non-adjuvanted or adj uv anted with aluminum hydroxide, aluminum phosphate or liposomal adjuvant B

[0215] Preclinical studies were performed in mice to evaluate the immunogenicity of gp350-FNP and gL-gH-gp42-FNP dual particle vaccine. Immunogenicity was assessed by measuring serum antibody titers against gp350, gH / gL, and gp42 individually by ELISA.

[0216] To investigate whether the immune responses against gp350-FNP and gL-gH-gp42-FNP can be enhanced by adjuvants, the dual particle vaccine formulated with 25 pg aluminum hydroxide, 25 pg aluminum phosphate or 10 mg liposomal adjuvant B (adjuvant B) was first evaluated in mice for its potential to induce EBV specific antibody responses. Female BALB / c mice (7 weeks old, 10 per group) were injected intramuscularly with 2 doses (0.25 pg or 1 pg) of gp350-FNP and gL-gH-gp42-FNP dual particles alone or adjuvanted with aluminum phosphate, aluminum hydroxide or liposomal adjuvant B at days 0 and 28. Sera were collected at 42 (2 weeks post dose 2). Sera were collected at day 42 and the total IgG responses were determined by gH / gL, gp42, gp350 ELISA titers.26101

[0217] An ELISA was used to determine the titers of mouse serum antibodies that bind to EBV gp350, gH / gL, and gp42. Maxisorp black 384-well plates were coated with 25 ng per well of one of the EBV antigens in DPBS for overnight incubation at 4 °C. After coating, plates were blocked with PBST with 3% NFDM for 30 minutes. Serum samples starting at 1:50 were diluted 4-fold across 10 concentrations in blocking buffer, transferred to pre-coated assay plate, and incubated for 2 hours. EBV antigen-bound antibodies were detected by addition of 1:10000 dilution of goat anti -mouse IgG Fc HRP-conjugated antibody for 1 hour. Plates were washed 6 times with PBST after each step. Plates were developed with West Pico PLUS Chemiluminescent HRP Substrate that was prepared according to the manufacturer's instructions. The luminescent assay signal generated by HRP cleavage of the chemiluminescent substrate was read on an EnVision plate reader (PerkinElmer, Waltham, Massachusetts, USA) after a 15-minute incubation.

[0218] An interpolated end point titer was calculated for each serum sample using the luminescence relative light unit (RLU) values and the following formula:(starting fold dilution of the sample / series dilution factor) x (series dilution factorAt) where t = x - [(cut-off - L) / (H - L)].

[0219] The cut-off value was designated as 550000. H is the high well RLU value (the RLU value of the first titration point above 50000), L is the low well RLU value (the RLU of the first titration point below 50000), and x is the low well number (the number in the titration series of L where the first dilution in the titration series was 1 and the highest serum dilution of the titration series was 10). Samples that did not cross the cut-off value were given a placeholder titer of 25, or one-half the initial starting serum dilution.

[0220] At two weeks post dose 2, antigen specific antibodies were detected in all dose groups. Greater titer variability was observed in 0.25 pg dose groups. In the animals that received 1 pg EBV-FNPs, all adjuvants tested were able to boost the gH / gL, gp42 and gp350 antibody titers. See Figures 1-3. Liposomal adjuvant B group showed the largest titer increase compared to the nonadjuvanted formulation.Example 5: EBV -Neutralizing Antibody Titers in Mice Immunized With 2 Doses of gp350-FNP and gL-gH-gp42-FNP dual particle vaccine non- adj uv anted or adj uv anted with aluminum hydroxide, aluminum phosphate or liposomal adjuvant B

[0221] Preclinical studies were performed in mice to evaluate the immunogenicity of gp350-FNP and gL-gH-gp42-FNP dual particle vaccine. Immunogenicity was assessed by measuring26101EBV-neutralizing serum antibody titers in a neutralization assay using EBV-GFP, B cells (4E3), and epithelial-like cells (293T).

[0222] Female BALB / c mice (10 per group) were injected intramuscularly with 2 doses of gp350-FNP and gL-gH-gp42-FNP dual particles alone or adjuvanted with aluminum phosphate, aluminum hydroxide or liposomal adjuvant B at days 0 and 28. Sera were collected at day 42 (2 weeks post dose 2). A neutralization assay using EBV-GFP, B cells, and epithelial-like cells was used to determine the titers of mouse serum antibodies that neutralize EBV.

[0223] Human B lymphoma cells (4E3) and human embry onic kidney cells (293T) cells were seeded in a 96-well black wall transparent plate at 2.5 x io6cells and 2.0 x io6cells. respectively, in 50 pL per well of assay medium (RPMI-1640 with no phenol red complete medium) for 4 hours at 37 °C, 5% CO2 for attachment. Serum starting at 1: 10 dilution or control antibody samples were 2-fold serially diluted (60:60 pL) across 11 concentrations in assay medium in a separate U-bottomed 96-well sample plate. Concentrated cell-free Akata EBV-GFP at 1.5 x 105ffu / rnL in assay medium was added to each well at 60 pL, for a total volume of 120 pL / well. The antibody and virus were mixed for 5 minutes on a plate shaker, and mixtures were maintained at ambient temperature for 1 hour. The antibody and virus mixtures were then added to the seeded 4E3 and 293T cell plates at 50 pL for a final volume of 100 pL / well, and the resulting antibody dilution at this stage was the final dilution recorded. The plates were cultured in a 36 °C, 5% CO2 incubator for 3 days, then scanned by an Acumen Cellista (SPT Labtech, Covina, California, USA) instrument to count the number of GFP-expressing cells which reflects the EBV-GFP infections. The number of EBV-GFP-infected B cells in control wells after two days is usually around 1000. The neutralization percentage (NT) of each well was calculated using the formula:NT = ((control well green cell count - antibody well green cell count) / control well green cell count) x 100%NT50 antibody titer values were determined using a 4-parameter logistic curve fitting algorithm (y = d+(a-d) / (l+(x / c)Ab)) in Excel software (Microsoft, Redmond, Washington, USA).

[0224] The antibodies induced by all formulations were capable of neutralizing EBV infection of B cells and epithelial cells. The trend of neutralizing antibody response is similar to the total antibody response, with liposomal adjuvant B producing the highest titers. See Figure 4.Collectively, the data demonstrated that gp350-FNP and gL-gH-gp42-FNP dual particle vaccine was immunogenic in BALB / c mice and that its immunogenicity was enhanced by liposomal adjuvant B.26101Example 6: Cellular Responses Against gH From Mice Immunized With 2 Doses of gp350-FNP and gL-gH-gp42-FNP dual particle vaccine non-adjuv anted or adjuvanted with aluminum hydroxide, aluminum phosphate or liposomal adjuvant B

[0225] Following the preclinical studies in Example 4, at two weeks post dose 2, spleens from four immunized animals were also harvested and pooled. The cell mediated responses were determined by an ICS assay. Among the adjuvant formulations evaluated, only liposomal adjuvant B produced measurable anti-gH CD4 T cell cytokine responses above baseline (dotted line), suggesting that liposomal adjuvant B might be more potent in eliciting cell mediated immune response compared to aluminum phosphate or aluminum hydroxide adjuvants [Figure 8], Minimal to no CD4 T cell responses against gp42, gL or gp350 were detected.

[0226] Collectively, the data demonstrated that the gp350-FNP and gL-gH-gp42-FNP dual particle vaccine was immunogenic in BALB / c mice and that its immunogenicity was enhanced by liposomal adjuvant B.Example 7: EBV Antigen-Specific Antibody Titers in Rhesus Monkeys Immunized With 3 Doses of gp350-FNP and gL-gH-gp42-FNP dual particle vaccine formulated with liposomal adjuvant B

[0227] The immunogenicity of the dual particle vaccine formulated with liposomal adjuvant B was further assessed in rhesus monkeys. Animals (5 per group) were immunized intramuscularly with 15 pg gp350-FNP and 15 pg gL-gH-gp42-FNP formulated with 10 mg adjuvanted at weeks 0, 4, and 10. Sera were collected at 2-week intervals from w eek 0 (prior to study start) to w eek 14, week 18, and then at 6-week intervals to week 42. Immunogenicity was assessed by measuring serum antibody titers against gp350, gH / gL, and gp42 individually by ELISA using methods according to Example 4 except goat anti-human IgG Fc HRP -conjugated antibody was used as the detection antibody.

[0228] Antibody titers were observed to gp350, gH / gL and gp42 at week 0 due to pre-existing cross-reactive immunity between EBV and RhLCV, a herpesvirus closely related to EBV that infects most rhesus monkeys early in life, and a significant increase in titers was observed after the 1stdose of the vaccine. One dose of vaccine maximized the boost of gH / gL and gp42 titers while 2 doses were needed to achieve peak response against gp350. The antibody titers gradually declined after peak levels and remained above the basal titers (week 0) up to 42 weeks after the 1stdose for both vaccines. See Figure 5.26101Example 8: EBV-Neutralizing Antibody Titers in Rhesus Monkeys Immunized With 3 Doses of gp350-FNP and gL-gH-gp42-FNP dual particle vaccine formulated with liposomal adjuvant B

[0229] Rhesus monkeys (5 per group) were injected intramuscularly with 3 doses of gp350-FNP and gL-gH-gp42-FNP dual particle vaccine formulated with liposomal adjuvant B at weeks 0, 4, and 10. Sera were collected at 2-week intervals from weeks 0 to 14, week 18, and then at 6-week intervals to week 42. Vaccination of rhesus monkeys with gp350-FNP and gH / gL / gp42-FNP dual particle vaccine formulated with liposomal adjuvant B boosted EBV-neutralizing antibody titers in a neutralization assay using EBV-GFP, B cells (4E3), and epithelial-like cells (293T) according to Example 5. Neutralization titers remained above basal titers (week 0) up to 42 weeks after the 1st dose. See Figure 6.Example 9: Cellular Responses Against gH and gp350 From Rhesus Monkeys Immunized With 3 Doses of gp350-FNP and gL-gH-gp42-FNP dual particle vaccine formulated with liposomal adjuvant B

[0230] PBMCs were collected at weeks 0, 6, 12, and 42, and analyzed in the intracellular cytokine staining (ICS) assay to determine the cellular responses induced by the EBV antigens gp350, gH / gL, or gp42.

[0231] Rhesus monkey PBMCs were prepared and subjected to stimulation by EBV-antigens followed by ICS. Cells were plated at 1 x 106cells per 100 pL per well in 96-well plates. Ninety-five (95) pL of stimulation cocktail consisting of CD28 / BV785 antibody and CD49d antibody at a final concentration of 1.25 pg / mL, and gp350, gH, gL, or gp42 peptide pools were added to a final concentration of 2 pg / ml. Plates were incubated at 37 °C for 1 hour. To prevent cytokines from being secreted, 5 pL of BFA was added to a final concentration of 2 pg / mL, and plates were incubated for an additional 5 hours at 37 °C, 5% CO2. Twenty (20) pL of 20 mM EDTA were added to halt cell stimulation 6 hours after peptide addition, and cells were stored at 4 °C until FACS staining. Cells were washed with PBS prior to the addition of 125 pL of Live / Dead Aqua at 1:4000 in PBS and incubated for 15 minutes at ambient temperature for cell viability staining. Cells were then washed in FACS buffer (PBS with 1% FBS, 0.01% sodium azide) by centrifugation of plates at 500 xg. All subsequent steps were performed at 2 to 8 °C with cell washes by centrifugation of plates at 500 xg for 5 minutes, unless otherwise indicated.

[0232] Cells were then stained with 150 pL of a surface antibody cocktail consisting of CD4 / BV605, CD8a / BUV395, CD95 / PE-Cy5, CD14 / BV711, CD20 / BV711, andCD3 / APC-Cy7 antibodies with BD Brilliant stain buffer for 30 minutes. Cells were washed once and then were permeabilized with 200 pL of BD Cytofix / Cy toperm solution for 20 to 2526101minutes. After an additional wash, cells were stained with 150 pL of an intracellular antibody cocktail consisting of TNF-a / PE-Cy7, IL-2 / PE, and IFN-y / FITC antibodies in BD Permeabilization wash buffer and BD Brilliant stain buffer for 60 minutes. Cells were washed twice in permeabilization wash buffer, fixed in 200 pL of BD Stabilizing Fixative, and analyzed on the LSRFortessa X-50 flow cytometer (Becton Dickinson. Franklin Lakes, New Jersey. USA) within 24 hours.

[0233] T cell cytokine responses are reported as % of non-naive CD4 or CD8 lymphocytes. Antigen-specific responses are calculated as follows:% T cellantigen-specific = % T cellpeptide-stimulated - % T cellmediaNegative values are imputed as 0 (no response), and values above 0.1% (baseline) are considered as true antigen specific response.

[0234] CD4 responses (IFN-y, IL -2, and TNF-a) to gH or gp350 peptide stimulation were observed in some animals that received the vaccine (Figure 7). No CD4 response to other antigens was detected (data not shown), and no CD8 response to any antigen was detected (data not show n). Collectively, the data demonstrated that gp350-FNP and gL-gH-gp42-FNP dual particle vaccine formulated with liposomal adjuvant B was immunogenic in Rhesus monkeys, and provided boost to both humoral and cellular responses.Example 10: A Phase 1, Randomized, Double-Blind, Placebo-Controlled, Dose Escalation Trial to Evaluate the Safety, Tolerability, and Immunogenicity of EBV polypeptide and Liposomal adjuvant B (LP or LP adjuvant B) in Healthy Participants

[0235] This is a randomized placebo-controlled, double-blind, multiple panel, multi-site study to evaluate the safety, tolerability, and immunogenicity of EBV gp350-FNP (SEQ ID NO: 23) and EBV gL-gH-gp42-FNP (SEQ ID NO: 21) dual particle vaccine and liposomal adjuvant B (EBV-LP) in healthy young adult participants (>18 to 30 years of age).

[0236] Part 1 will evaluate progressively higher antigen and / or adjuvant doses of each EBV polypeptide and LP adjuvant in seropositive participants. The scope of Part 2 will depend on enrollment of EBV-naive (seronegative) participants, and will evaluate a subset of expansion panels in seronegative participants at antigen and / or adjuvant dose levels equal to or low er than those being evaluated in Part 1. These panels selected for Part 2 may be initiated at any point following safety review of the relevant sentinel cohorts in Part 1. The number of cohorts and the selection of doses for Part 2 will be kept flexible to optimize immunogenicity information in seronegatives, pending safety and tolerability in Part 1, feasibility / seronegative recruitment numbers, and any available immunogenicity' data. Finally, Part 3 will further explore dose26101ranging in seropositive participants, at antigen and / or adjuvant doses equal to or lower than the maximum well-tolerated dose being evaluated in dose escalation. Panels selected for Part 3 may be initiated at any point following safety review of the relevant sentinel cohorts in Part 1. The dose escalation panels for the EBV vaccine will follow a similar approach to increase antigen and adjuvant dose. Based on available safety and tolerability data through 7 days postvaccination (described further below), the dose of either antigen or adjuvant will be increased in a step-wise manner until reaching the highest antigen and adjuvant dose (100 pg of each FNP and 0.68 mg of LP with an option escalate to 1.35 mg of LP.

[0237] Each EBV-LP in Part 1 will have a Lead Cohort of 4 participants. The first two participants in each of these panels will be dosed at least 2 hours apart and then, assuming sufficient safety and tolerability for the first two participants, the next two participants will be dosed approximately 24 hours later (or longer) and at least 2 hours apart from each other. All Lead Cohort participants in each panel will be dosed at the same clinical site. The Lead Cohort approach will be repeated after Dose 2 and Dose 3. This will provide the opportunity to identify potential safety' issues before subsequent participants are dosed.Part 2 (flexible): Expansion in seronegative participants

[0238] Part 2 will be implemented if 1) safety and tolerability of EBV-LP from Part 1 are supportive of further study in seronegative participants and 2) the prevalence of seronegative individuals at the FIH clinical sites is high enough to enable enrollment. Therefore, Part 2 is included as flexible to explore the safety, tolerability', and immunogenicity of EBV-LP in a seronegative population and will be based on the feasibility of enrolling seronegative participants. Up to 4 panels for EBV-LP may be included in Part 2. In each panel, 12 participants will be randomized in a 5: 1 ratio to receive either the experimental vaccine or placebo.Part 3 (flexible): Additional exploration in seropositive participants

[0239] Part 3 is included to further explore the safety, tolerability, and immunogenicity of EBV-LP in a seropositive population. Up to 2 additional panels for EBV-LP may be included in Part 3 to explore doses equal to or lower than the maximum well-tolerated dose in Part 1. In each panel, approximately 15 participants will be randomized in a 4: 1 ratio to receive either the experimental vaccine or placebo. Part 3 will be implemented if safety and tolerability of EBV-LP from Part 1 are supportive of further study in seropositive participants.26101Table 3 Sample Allocation TableEBV-LP Escalation Phase Seropositive Participants (Part l)aSubjects Panel E Panel F Panel G (N=15)per (N=15) (N=15)Paneln= 12 30 pg of each FNP, 100 pg of each FNP, 100 pg of each FNP, 1.35 mg 0.68 mg LP 0.68 mg LP LPn= 3 Placebo Placebo PlaceboEBV-LP Expansion Phase Seronegative Participants (Part 2 flexible)15Subjects Panel K Panel L Panel Mper (5:1) (5:1) (5:1)Paneln= 10 30 pg of each FNP. 10 pg of each FNP. 10 pg of each FNP, 1.35 mg 0.68 mg LP 0.68mg LP LPn= 2 Placebo Placebo PlaceboEBV-LP Expansion Phase Seronegative Participants Continued (Part 2 flexible)3Subjects Panel O Panel P Panel Qper Panel (5:1) (5:1) (5:1)n= 10 30 pg of each 10 pg of each 30 pg of each FNP, 0.27 mg LP FNP, 1.35 mg LP FNP, 0.68 mg LPn= 2 Placebo Placebo PlaceboEBV-LP Additional Exploration Phase Seropositive Participants (Part 3 flexible)aSubjects Panel S Panel T Panel Uper Panel (N=15) (N=15) (N=15)n= 12 30 pg of each 10 pg of each 30 pg of each FNP, 0.27 mg LP FNP, 1.35 mg LP FNP. 0.68 mg LPn= 3 Placebo Placebo PlaceboFNP=ferritin nanoparticleWithin each panel, participants will be randomized to receive EBV-LP or matching placebo in a 4: 1 ratio (Part 1 and Part 3) according to a computer-generated, permuted-block allocation schedule.bThe exact number of subjects per seronegative panel (Part 2) will depend on enrollment of seronegatives, but randomization is planned in a 5:1 ratio (active: placebo) with a target of n=12.Placebo: Diluent: (Sodium Phosphate Sucrose Buffer)26101Table 4: Intervention Groups and Duration:Unit Dose Dosage Level(s) Route of Regimen / Treatment Period / Strength(s) Administration Vaccination Regimengp350-FNP: 10 pg / each FNP Intramuscular Days 1, 57, and0.8 mg / mL 30 pg / each FNP 169gL / gH / gp42- 100 pg / each FNPFNP:0.8 mg / mLLP adjuvant B: LP adjuvant B4.05 mg / mL 0.27 mg0.68 mg1.35 mgDiluent:Sterile Diluentfor Reconstitution(SodiumPhosphateSucrose Buffer)0 pg / 0 mLTable 5Primary Objective Primary EndpointTo assess the safety and tolerability of EBV- Solicited injection site adverse events 7 LP compared with placebo in healthy young days after each vaccination.adult participants. Solicited systemic adverse events 7 days after each vaccination.Immediate reactions occurring within 30 minutes after each vaccination.Unsolicited adverse events 28 days after each vaccination.Serious adverse events Day 1 through 12 months after the final vaccination.Medically attended adverse events (MAAEs) Day 1 through 12 months after the final vaccination.Events of clinical interest (including potential immune-mediated disease) Day 1 through 12 months after the final vaccination.Secondary Objectives Secondary Endpoints26101To assess the immunogenicity of the EBV- Serum IgG and neutralizing (cLBA) LP using assays to measure antibody antibody responses (geometric mean titers responses to gp350 and gp42 / gH / gL in and / or geometric mean fold rise from healthy young adult participants. baseline postvaccination) at all evaluable timepoints for gp350, gp42. and gH / gL.

[0240] The dual particle vaccine was generally well tolerated across a range of dose levels of EBV gp350-FNP and EBV gL-gH-gp42-FNP antigens and the liposomal adjuvant B. Clear immunogenicity boosts were observed in seropositive and negative participants.

Claims

1. WHAT IS CLAIMED IS:

1. A composition comprising:3.(a) an Epstein Barr Virus (EBV) polypeptide comprising one or more polypeptides selected from the group consisting of: an EBV gp42 polypeptide, an EBV gH polypeptide, and an EBV gL polypeptide; and4.(b) a liposomal adjuvant, wherein the liposomal adjuvant comprises a glycolipid and a saponin.

2. The composition of claim 1 that comprises two or more of an EBV gp42 polypeptide, an EBV gH polypeptide, and an EBV gL polypeptide.

3. The composition of claim 1, wherein the EBV polypeptide comprises an EBV gp42 polypeptide, an EBV gH polypeptide, and an EBV gL polypeptide.

4. The composition of claim 3, wherein the EBV gL polypeptide(s), the EBV gH polypeptide(s), and the EBV gp42 polypeptide(s), are arranged in N-terminal to C-terminal order.

5. The composition of any one of claims 3-4, further comprising a linker between each EBV polypeptide.

6. The composition of claim 5, wherein a linker having a length of at least 40 amino acids and less than or equal to 50 amino acids separates the EBV gL polypeptide and the EBV gH polypeptide.

7. The composition of claim 6, wherein a linker having a length of 46 or 47 amino acids separates the EBV gL polypeptide and the EBV gH polypeptide.

8. The composition of any one of claims 5 to 7, wherein a linker having a length of 15 to 60 amino acids separates the EBV gH polypeptide and the EBV gp42 polypeptide.

9. The composition of any one of claims 5 to 7, wherein a linker having a length of 30 to 50 amino acids separates the EBV gH polypeptide and the EBV gp42 polypeptide.

10. The composition of claim 9, wherein the linker between the EBV gL polypeptide and the EBV gH polypeptide is 44 to 48 amino acids, and the linker between the EBV gH polypeptide and the EBV gp42 polypeptide is 32 amino acids.

11. The composition of claim 9, wherein the linker between the EBV gL polypeptide and the EBV gH polypeptide is 46 amino acids, and the linker between the EBV gH polypeptide and the EBV gp42 polypeptide is 32 amino acids.

12. The composition of claim 5, wherein a linker separates the EBV gL polypeptide and the EBV gH polypeptide and consists of an amino acid sequence that has at least 80%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 7; and / or wherein a linker separates the EBV gH polypeptide and the EBV gp42 polypeptide and consists of an amino acid sequence that has at least 80%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 8.

13. The composition of any one of claims 1-12, wherein the EBV polypeptide further comprises a ferritin.

14. The composition of claim 13, further comprising a further linker that separates the ferritin and the EBV gp42 polypeptide.

15. The composition of claim 14, wherein the further linker has a length of 60 to 100 amino acids.

16. The composition of claim 15, wherein the further linker consists of an amino acid sequence having at least 80%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 9.

17. The composition of any one of claims 6 to 16, wherein (a) the linker that separates the EBV gL polypeptide and the EBV gH polypeptide, (b) the linker that separates the EBV gH polypeptide and the EBV gp42 polypeptide, or (c) the linker of (a) and the linker of (b) comprises one or more of glycine, asparagine, serine, and alanine.

18. The composition of any one of claims 13-17, wherein the ferritin comprises an amino acid sequence having at least 80%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 26.

19. The composition of any one of claims 1-18, wherein the EBV gp42 polypeptide comprises an amino acid sequence having at least 80%, 90%, 95%, 96%, 97%, 98%, or 99% identity to residues 856-1038 of SEQ ID NO: 21.

20. The composition of any one of claims 1-19, wherein the EBV gp42 polypeptide consists of amino acid residues 856-1038 of SEQ ID NO: 21; SEQ ID NO: 1, SEQ ID NO: 2 or SEQ ID NO:3.

21. The composition of claim 20, wherein the EBV gEI polypeptide comprises an amino acid sequence having at least 80%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to residues 162-823 of SEQ ID NO: 21.

22. The composition of claim 21, wherein the EBV gEI polypeptide consists of amino acid residues 162-823 of SEQ ID NO: 21 or SEQ ID NO: 5.

23. The composition of any one of claims 1-22, wherein the EBV gL polypeptide comprises an amino acid sequence having at least 80%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to residues 1-115 of SEQ ID NO: 21.

24. The composition of any one of claims 1-23, wherein the EBV gL polypeptide comprises amino acid residues 1-115 of any one of SEQ IDNOs: 11-21; or the amino acid sequence set forth in SEQ ID NO: 4.

25. The composition of any one of claims 1-24, wherein the EBV polypeptide comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to amino acid residues 1-1038 of SEQ ID NO: 21.

26. The composition of claim 25, wherein the EBV polypeptide comprises a sequence of any one of SEQ ID NOs: 11-21.2610127. The composition of any one of claims 1-26, wherein the EBV polypeptide further comprises EBV gp220.

28. The composition of claim 27, wherein the EBV gp220 polypeptide comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 6.

29. A composition comprising an EBV gp220 polypeptide; and a liposomal adjuvant, wherein the liposomal adjuvant comprises a glycolipid and a saponin.

30. The composition of claim 29, wherein the EBV gp220 polypeptide comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 6.

31. The composition of claim 29 or 30, wherein the EBV gp220 polypeptide further comprises a ferritin.

32. The composition of claim 29 or 30, wherein the ferritin comprises an amino acid sequence having at least 80%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 24, 25 or 26.

33. The composition of claim 32, wherein the ferritin consists of the amino acid sequence of SEQ ID NO: 24, 25 or 26.

34. The composition of claim 31, wherein the EBV gp220 polypeptide consists of SEQ ID NO: 22 or 23.

35. The composition of any one of claims 1-34, wherein the composition is a solution of the EBV polypeptide and the liposomal adjuvant.

36. The composition of any one of claims 1-35, wherein the liposomal adjuvant comprises a glucopyranosyl lipid A (“GLA’’) and saponin selected from the group consisting of QS-21, QS-17 and QS-18.2610137. The composition of any one of claims 1-35, wherein the glycolipid has the structure set forth in Formula I:39.OH41. 43.Formula I,44.wherein Ri is P(O)(OH)2;45.wherein R2 is selected from H, C(O)CH2CH(OH)Cn alkyl, and C(O)CH2CH(OC(O)Cn- C13 alkyl)Cn alkyl;46.wherein R3is C(O)CH2CH(OC(O)Cn-Ci3 alkyl)Cn alkyl;47.wherein R4 is selected from H, C(O)CH2CH(OH)Cn alkyl, and C(O)CH2CH(OC(O)Cn- C13 alkyl)Cn alkyl;48.wherein R5 is selected from C(O)CH2CH(OH)Cn alkyl and C(O)CH2CH(OC(O)Ci3-Ci5 alkyl)Cn alkyl; and49.wherein Re is H;50.or a pharmaceutically acceptable salt thereof.

38. The composition of claim 37, wherein the glycolipid has the structure set forth in Formula II:

53. 55.wherein Ri is H or C(O)Cn-Ci3 alkyl; wherein R2 is Cn alkyl;56.wherein Rs is C11-C13 alkyl;57.wherein R4 is Cn alkyl;58.wherein R5 is H or C(0)CH2CH(0H)Cn alkyl;59.wherein Rs is H or C(O)Ci3-Cis alkyl; and60.wherein R7 is Cn alkyl;61.or a pharmaceutically acceptable salt thereof.

39. The composition of claim 38, wherein the glycolipid has the structure set forth in Formula II:63.Formula64.

65. II66.wherein Ri is C(O)Cn-Ci3 alkyl;67.wherein R2 is Cn alkyl;68.wherein R3 is C11-C13 alkyl;69.wherein R4 is Cn alkyd;70.wherein R5 is C(O)CH2CH(OH)C alkyl;71.wherein Rs is H; and72.wherein R7 is Cn alkyl;73.or a pharmaceutically acceptable salt thereof.

40. The composition of claim 38 or 39, wherein the glycolipid has the structure set forth in Formula I1A:

75. 77.Formula 11 A,78.or a pharmaceutically acceptable salt thereof.

41. The composition of claim 37, wherein the glycolipid has the structure set forth in Formula III:

81. 83.Formula III, or a pharmaceutically acceptable salt thereof.

42. The composition of claim 37, wherein the glycolipid has the structure set forth in Formula IV:

86. 88.Formula IV,89.or a pharmaceutically acceptable salt thereof.

43. The composition of claim 37, wherein the glycolipid has the structure set forth in Formula V:2610192. 94.Formula V,95.or a pharmaceutically acceptable salt thereof.

44. The composition of claim 37, wherein the glycolipid has the structure set forth in Formula VI:

97. 99.or a pharmaceutically acceptable salt thereof.

45. The composition of any one of claims 1-44, wherein the liposomal adjuvant further comprises l,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC).

46. The composition of any one of claims 1-45, wherein the liposomal adjuvant further comprises cholesterol.

47. A pharmaceutical unit dose comprising about 5 pg to about 200 pg of EBV polypeptide consisting of any one of the amino acid sequences of SEQ ID NOs: 11-21, and about 0.1 mg to about 3 mg of a liposomal adjuvant, wherein the liposomal adjuvantcomprises l,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol Quillaja Saponaria21 (QS-21) and the glycolipid of Formula IIIA103.OH104.?. OH105.HO**’106.O HO O H O O HO HO107.e e108.e110.

111. or a pharmaceutically acceptable salt thereof.

48. The pharmaceutical unit dose of claim 47, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of about 10 pg to about 100 pg, and about 0.1 mg to about 2 mg of the liposomal adjuvant, wherein the amount of Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is each at about 5 pg to about 100 pg.2610149. The composition or pharmaceutical unit dose of any one of claims 41 and 47-48, wherein the amount of Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula III, Formula IIIA or a pharmaceutically acceptable salt thereof is each at about 10 pg to about 75 pg.

50. The composition or pharmaceutical unit dose of any one of claims 41 and 47-48, wherein the amount of Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula III, Formula IIIA or a pharmaceutically acceptable salt thereof is each at about 10 pg to about 50 pg.

51. The composition of any one of claims 1-46, wherein the ratio of saponin to glycolipid is 1-5: 1-3 by weight.

52. The composition or pharmaceutical unit dose of any one of claims 46-50, wherein the ratio of l,2-dioleoyl-sn-glycero-3 -phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid is 10-30:2-15:0.1-5:0.1-3 by weight.

53. The pharmaceutical unit dose of any one of claims 47-50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 10 pg, and the liposomal adjuvant is in the amount of 0.27 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.

54. The pharmaceutical unit dose of any one of claims 47-50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 30 pg, and the liposomal adjuvant is in the amount of 0.27 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.

55. The pharmaceutical unit dose of any one of claims 47-50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 100 pg, and the liposomal adjuvant is in the amount of 0.27 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.2610156. The pharmaceutical unit dose of any one of claims 47-50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 10 pg, and the liposomal adjuvant is in the amount of 0.68 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.

57. The pharmaceutical unit dose of any one of claims 47-50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 30 pg, and the liposomal adjuvant is in the amount of 0.68 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.

58. The pharmaceutical unit dose of any one of claims 47-50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 100 pg, and the liposomal adjuvant is in the amount of 0.68 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.

59. The pharmaceutical unit dose of any one of claims 47-50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 10 pg, and the liposomal adjuvant is in the amount of 1.35 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.

60. The pharmaceutical unit dose of any one of claims 47-50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 30 pg, and the liposomal adjuvant is in the amount of 1.35 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.

61. The pharmaceutical unit dose of any one of claims 47-50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 100 pg, and the liposomal adjuvant is in the amount of 1.35 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-26101126.3-phosphocholine (DOPC), cholesterol Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.

62. A pharmaceutical unit dose comprising about 5 pg to about 200 pg of EBV polypeptide consisting of the amino acid sequence of SEQ ID NO: 23, and about 0.1 mg to about 3 mg of a liposomal adjuvant, wherein the liposomal adjuvant comprises 1,2-dioleoyl-sn-glycero- 3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of128.Formula II129.

130. IA,131.or a pharmaceutically acceptable salt thereof.

63. The pharmaceutical unit dose of claim 62, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of about 10 pg to about 100 pg, and about 0.1 mg to about 2 mg of the liposomal adjuvant, wherein the amount of Quillaja Saponaria26101133.21 (QS-21) and the glycolipid of Formula I1IA or a pharmaceutically acceptable salt thereof is each at about 5 pg to about 100 pg.

64. The pharmaceutical unit dose of any one of claims 62-63, wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC). cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 10-30:2-15:0.1- 5: 0.1 -3 by weight.

65. The pharmaceutical unit dose of any one of claims 62-63, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 10 pg, and the liposomal adjuvant is in the amount of 0.27 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.

66. The pharmaceutical unit dose of any one of claims 62-63, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 30 pg, and the liposomal adjuvant is in the amount of 0.27 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.

67. The pharmaceutical unit dose of any one of claims 62-63, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 100 pg, and the liposomal adjuvant is in the amount of 0.27 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.

68. The pharmaceutical unit dose of any one of claims 62-63, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 10 pg, and the liposomal adjuvant is in the amount of 0.68 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.

69. The pharmaceutical unit dose of any one of claims 62-63, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 30 pg, and26101140.the liposomal adjuvant is in the amount of 0.68 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.

70. The pharmaceutical unit dose of any one of claims 62-63, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 100 pg, and the liposomal adjuvant is in the amount of 0.68 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.

71. The pharmaceutical unit dose of any one of claims 62-63, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 10 pg, and the liposomal adjuvant is in the amount of 1.35 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by w eight.

72. The pharmaceutical unit dose of any one of claims 62-63, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 30 pg, and the liposomal adjuvant is in the amount of 1.35 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by w eight.

73. The pharmaceutical unit dose of any one of claims 62-63, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 100 pg, and the liposomal adjuvant is in the amount of 1.35 mg; wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5: 1: 1 by weight.

74. A pharmaceutical unit dose comprising about 10 pg to about 100 pg of a first EBV polypeptide consisting of the amino acid sequence of SEQ ID NO: 21 and about 10 pg to about 100 pg of a second EBV polypeptide consisting of the amino acid sequence of SEQ ID NO: 23, wherein the first and the second EBV polypeptide are at a 1: 1 weight ratio; and about 0.54 mg to about 1.36 mg of a liposomal adjuvant, wherein the liposomal adjuvant comprises l,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21)146.- Ill - 26101147.and the glycolipid of Formula I1IA148.n nu150. 151.v e, or a pharmaceutically acceptable salt thereof, at a ratio of 10-30:2-15: 1-5: 1-3 by weight.

75. The pharmaceutical unit dose of claim 74, wherein the ratio of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), cholesterol, Quillaja Saponaria 21 (QS-21) and the glycolipid of Formula IIIA or a pharmaceutically acceptable salt thereof is 20:5:1:1 by weight.

76. The pharmaceutical unit dose of claim 74 or 75, wherein each of the first and second EBV polypeptide is at 10 pg.

77. The pharmaceutical unit dose of claim 74 or 75, wherein each of the first and second EBV polypeptide is at 30 pg.2610178. The pharmaceutical unit dose of claim 74 or 75, wherein each of the first and second EBV polypeptide is at 100 pg.

79. The composition or pharmaceutical unit dose of any one of claims 1-78, wherein the average particle size of the liposomal adjuvant is about 50-200 nm.

80. The composition or pharmaceutical unit dose of any one of claims 1-79 wherein the liposomal adjuvant further comprises a buffer selected from the group consisting of: acetic acid, histidine, citrate, Bis-Tns, HEPES, phosphate, MES, sodium chloride, succinate. Tris, and combinations thereof.

81. The composition or pharmaceutical unit dose of claim 80, wherein the buffer is present in the amount of about ImM to about lOOmM.

82. The composition or pharmaceutical unit dose of any one of claims 1-79, wherein the liposomal adjuvant further comprises 5 mM - 100 mM potassium dihydrogen phosphate at pH 5.1 - 7.0, 1 mM to 50 rnM disodium phosphate at pH 5.1 - 7.0, and 25 mM - 300 mM NaCl at pH 5.1 - 7.0.

83. The composition or pharmaceutical unit dose of any one of claims 1-79, wherein the liposomal adjuvant further comprises 40 mM - 50 mM potassium dihydrogen phosphate at pH 5.1 - 7.0, 5 mM to 15 mM disodium phosphate at pH 5.1 - 7.0, and 50 mM - 150 mM NaCl at pH 5.1 - 7.0.

84. The composition or pharmaceutical unit dose of any one of claims 1-83, wherein the QS-21 comprises one or more of QS-21-H, QS-21-Apiose and QS-21-xylose as depicted below:26101163.

85. The composition or pharmaceutical unit dose of any one of claims 1-84, wherein the QS-21 comprises QS-21-Apiose and QS-21 -xylose.2610186. A method of inducing an immune response to an Epstein Barr Virus (EBV) in a human patient comprising administering to the patient a therapeutically effective amount of the composition or pharmaceutical unit dose of any one of claims 1-85.

87. A method of preventing infection of or reducing the likelihood of infection of a human patient by an Epstein Barr Virus (EBV) comprising administering to the patient a therapeutically effective amount of the composition or pharmaceutical unit dose of any one of claims 1-85.

88. A method of preventing cancer caused by Epstein Barr Virus (EBV) of a human patient, comprising administering to the patient a therapeutically effective amount of the composition or pharmaceutical unit dose of any one of claims 1-85.

89. The method of claim 88, wherein the cancer is Burkitt’s lymphoma. Hodgkin’s lymphoma, or nasopharyngeal cancer.

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