Nanoemulsion 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 squalene nanoemulsion (SNE) 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 SNE adjuvant.

Description

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

[0001] This application claims the benefit of U.S. Provisional Patent Application Serial Nos. 63 / 725,928 filed November 27, 2024, and 63 / 733,595 filed December 13, 2024 the entire contents of which are incorporated by references herein.REFERENCE TO SEQUENCE LISTING SUBMITTED ELECTRONICALLY

[0002] The contents of the electronic sequence listing (26100-WO-PCT-SEQLIST- 08SEPT2025.xml; Size: 45,249 bytes; and Date of Creation: June 11, 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 squalene nanoemulsion (SNE) adjuvant, which can be administered as a vaccine. The SNE adjuvant comprises surfactants, terpenes, terpanoid-based oils, cationic lipids, or mixtures thereof. 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 EBVgp42 polypeptide, an EBV gH polypeptide, and an EBV gL polypeptide; and (b) a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN- 85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80); and squalene.

[0006] The invention further provides a composition comprising an EBV gp220 polypeptide; and a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80); and squalene.

[0007] The invention further provides a pharmaceutical unit dose comprising about 5 pg to about 200 pg of an EBV polypeptide disclosed herein, and about 2 mg to about 30 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80); and about 2 mg to about 25 mg of squalene.

[0008] The invention further provides a pharmaceutical unit dose comprising about 5 pg to about 200 pg of EBV polypeptide comprising, consisting of, or consisting essentially of any one of the amino acid sequences of SEQ ID Nos: 11-21, and about 2 mg to about 30 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN- 85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80); and about 2 mg to about 25 mg of squalene.

[0009] The invention further provides a pharmaceutical unit dose comprising about 5 pg to about 200 pg of EBV polypeptide comprising, consisting of or consisting essentially of the amino acid sequence of SEQ ID NO: 23, and about 2 mg to about 30 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN- 85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80); and about 2 mg to about 25 mg of squalene.

[0010] The invention further provides a pharmaceutical unit dose comprising about 10 pg to about 100 pg of a first EBV polypeptide comprising, consisting of or consisting essentially 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; and about 6 mg to about 18 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80); and squalene; wherein the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10:1:1 by weight.

[0011] The invention further provides a method of inducing an immune response to a Epstein Barr Virus (EBV) in a human patient comprising administering to the patient the composition or pharmaceutical unit dose of the invention.

[0012] 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 SNE adjuvant described herein.

[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) comprising administering to the patient the composition or pharmaceutical unit dose of the invention.

[0014] 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 an EBV polypeptide described herein and the SNE adjuvant described herein.

[0015] 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.

[0016] 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 SNE adjuvant described herein.

[0017] The invention also provides a kit comprising: (1) an Epstein Barr Virus (EBV) polypeptide; and (2) a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80); and squalene.DEFINITIONS

[0018] 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.

[0019] As used throughout the specification and appended claims, the following abbreviations and definitions apply:CLA ((13Z,16Z)-N,N-dimethyl-3-nonyldocosa-I3,16-dien-l-amine)EBV Epstein Barr VirusID intradermalIM intramuscularME microemulsionMNS microfluidic nanoemulsion self-assembly26100Mw molecular weightNE nanoemulsionNMWCO nominal molecular weight cut offPHE pre-homogenized emulsionPS-20 polysorbate-20PS-80 polysorbate-80SNE squalene nanoemulsionSPAN-85 sorbitan-trioleateVLP(s) virus-like particle(s) (Protein antigens) w / v weight per volume

[0020] As used throughout the specification and appended claims, the following definitions and abbreviations apply:

[0021] AAHS: As used herein, the term “AAHS” refers to an amorphous aluminum hydroxyphosphate sulfate adjuvant.

[0022] About: As used herein, the term "about", when modifying the quantify (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 quantify up to ± 10% rounded up where appropriate that can occur, for example, through typical measuring, handling and sampling procedures involved in the preparation, characterization and / or use of the substance or composition; through instrumental error in these procedures; through differences in the manufacture, source, or purify 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%.

[0023] 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., sw itch 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 a26100 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.

[0024] 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.

[0025] 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.

[0026] 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.

[0027] 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 alkyd group contains from 8 to 24 carbon atoms (C8-C24 alkyl). In one embodiment, an alky 1 group is linear. In another embodiment, an alkyl group is branched. In another embodiment the alky 1 group is unsubstituted.

[0028] 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 monoclonalantibodies), polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies), humanized, fully human antibodies, and chimeric antibodies.

[0029] 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.

[0030] 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.

[0031] EBV poly peptide: 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 an EBV polypeptide can comprise all or part of multiple sequences encoded by EBV (for example, all or part of gL and gH of EBV. or all or part of gL, gH, and gp42 of EBV).

[0032] 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.

[0033] 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.

[0034] 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 1 (Sequence Table). A ferritin may be a fragment of a full-length naturally- occurring sequence.

[0035] Wild-type ferritin: "Wild-ri pe 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 w ith one or more differences in its amino acid sequence from a wild-type ferritin.

[0036] 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.

[0037] 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).

[0038] 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 an amino 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."

[0039] 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 immune26100 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.

[0040] 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.

[0041] 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.26100

[0042] 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 a non-ferritin polypeptide (e.g., an EBV polypeptide) of sufficient length that the molecule is antigenic with respect to the non-ferritin polypeptide. Antigenicity may 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.

[0043] Sequence Identity : As used herein, "Sequence identity" between two amino acid sequences indicates the percentage of amino acids that are identical between the sequences.

[0044] % 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 carried 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}' algorithm 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.).

[0045] 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.

[0046] 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 about100% 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.

[0047] 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.

[0048] 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.

[0049] Cationic lipid: As used herein, the term “cationic lipid” refers to a lipid species that carries a net positive charge at a selected pH, such as physiological pH. A cationic lipid may be utilized as an ingredient in a multi-component SNE adjuvant formulation. Those of skill in the art will appreciate that a cationic lipids can include, but are not limited to, those disclosed in US Patent Application Publication Nos. US2008 / 0085870, US2008 / 0057080, US2009 / 0263407, US2009 / 0285881, US2010 / 0055168, US2010 / 0055169, US2010 / 0063135, US2010 / 0076055, US2010 / 0099738, US2010 / 0104629, US2013 / 0017239, and US2016 / 0361411, International Application Publication Nos. WO2011 / 022460. WO2012 / 040184, WO2011 / 076807.W02010 / 021865, WO 2009 / 132131, WO2010 / 042877, W02010 / 146740, and WO2010 / 105209, and US Patent Nos. US5,208,036, US5,264,618, US5,279,833, US5,283,185, US6,890,557, and US9, 669.097.

[0050] Co-administration: As used herein, the term “co-administration” or “co-administering” in relation to the SNE adjuvant and a pharmaceutical formulation (e g., an EBV polypeptide) refers to administration of an SNE 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 an SNE adjuvant (or vice versa). That is, after administration of the EBV polypeptide (or SNE adjuvant), the SNE adjuvant (or EBV polypeptide) can be administered substantially immediately after the EBV polypeptide (or SNEadjuvant) or the SNE adjuvant (or the EBV polypeptide) can be administered after an effective time period after the EBV polypeptide (or SNE 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.

[0051] 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 SNE 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.

[0052] Consists essentially of: 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.

[0053] 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.

[0054] 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.

[0055] 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 the26100 likelihood of infection is desired, e.g., the immunocompromised, the elderly, children, adults, or healthy individuals.

[0056] 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 least three 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.

[0057] 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.

[0058] 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.

[0059] 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-microbial26100 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, by 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; transdermally; or nasally, pulmonary', and to other mucosal surfaces.

[0060] Squalene nanoemulsion: As used herein, the terms “squalene nanoemulsion” or “SNE” refer to a formulation of one or more of an emulsifier, a solubilizer, a surfactant, and a lipid that have adjuvant properties in an EBV vaccine. In one embodiment, SNE refers to a SNE adjuvant formulation comprising (1) sorbitan trioleate (SPAN-85); (2) polysorbate-20 (PS-20); (3) squalene; and an optional (4) cationic lipid. The formulation may comprise one or more of a buffer, surfactant and salt.

[0061] 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 susceptibility7to 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.

[0062] Surfactants: As used herein, the term “surfactants” refers to stabilizing ingredients in a multi-component SNE adjuvant formulation and include the polyoxyethylene sorbitan esters surfactants (commonly referred to as the Tweens, especially PS-20 and PS-80), copolymers of ethylene oxide (EO). propylene oxide (PO), and / or butylene oxide (BO), sold under the DOWFAX™ tradename, such as linear EO / PO block copolymers (poloxamers); octoxynols, which can vary' in the number of repeating ethoxy' (oxy-1, 2-ethanediyl) groups, with octoxynol-926100(Triton X-100, or t-octylphenoxypolyethoxyethanol) being of particular interest; (octylphenoxy)poly ethoxyethanol (IGEPAL CA-630 / NP-40); nonylphenol ethoxylates, such as the Tergitol™ NP series; polyoxyethylene fatty ethers derived from laur l. cetyl, slear l and oleyl alcohols (known as Brij surfactants), such as tri ethyleneglycol monolauryl ether (Brij 30); and sorbitan esters (commonly known as the SPANs). such as sorbitan trioleate (Span-85, Tween-85 or [2-[(2R,3S,4R)-4-hydroxy-3-[(Z)-octadec-9-enoyl]oxyoxolan-2-yl]-2-[(Z)-octadec- 9-enoyl] oxy ethyl] (Z)-octadec-9-enoate) and sorbitan monolaurate. In an embodiment, surfactants are selected from sorbitan esters and poloxamers. In an embodiment, surfactants are selected from polysorbate-20 (PS-20) and polysorbate-80 (PS-80).

[0063] Terpenes: As used herein, the term '‘terpenes” refers to stabilizing ingredients in a multicomponent SNE adjuvant formulation and include, but are not limited to: monoterpenes including geraniol, terpineol, limonene, myrcene, linalool and pinene; sesquiterpenes including humulene, famesenes and famesol; diterpenes including cafestol, kahweol, cembrene and taxadiene; triterpenes including squalene and squalane; tetraterpenes including acyclic lycopene, monocyclic gamma-carotene, bicyclic alpha- and beta-carotenes; polyterpines and norisopredoids. In an embodiment, the terpene is an oxidative degradant of terpene. In an embodiment, a terpene is squalene.

[0064] 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.

[0065] 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 account26100 factors such as the age, sex, weight, species, and condition of the recipient animal and the route of administration.BRIEF DESCRIPTION OF THE DRAWINGS

[0066] Figure 1: Select structures of cationic lipids: (13Z.16Z)-N. N-dimethyl-3-nonyldocosa- 13,16-dien-l-amine (CLA) ; (6Z,9Z,26Z,29Z)-N,N-dimethylpentatriaconta-6,9,26,29-tetraen-18- amine (CLX) ; and N,N-dimethyl-l -(( 1 S,2R)-2-octylcyclopropyl) heptadecan- 8 -amine (CLY). See Example 1.

[0067] Figure 2: CLA-SNE components: (13Z,16Z)-N, N-dimethyl-3-nonyldocosa-13.16-dien- 1 -amine (CLA), SPAN-85, PS-20 and squalene. See Example 1.

[0068] Figure 3: Characterization of CLA-SNE adjuvant A bulk preparation utilizing static light scattering (SLS). See Example 2.

[0069] Figure 4: Impact of the formulation process on the incorporation of CLA into an SNE. See Example 2.

[0070] Figures 5A-5D: Nanotracking analysis (NT A) of CLA-SNE and SNE formulations stored at 4°C and 37°C for 1 month (Figure 5A: CLA-SNE [6mg / mL CLA and 30mg / mL squalene]; Figure 5B: SNE [40mg / mL squalene]; Figure 5C: CLA-SNE [4mg / mL CLA and 4mg / mL squalene]; and Figure 5D: SNE [8mg / mL squalene]). See Example 3.

[0071] Figures 6A-6D: Dynamic light scattering (DLS) of CLA-SNE and SNE formulations stored at 4°C, 25°C and 37°C for 1 month (Figure 6A: CLA-SNE [6mg / mL CLA and 30mg / mL squalene]; Figure 6B: CLA-SNE [4mg / mL CLA and 4mg / mL squalene]; Figure 6C: SNE [40mg / mL squalene]; and Figure 6D: SNE [8mg / mL squalene]). See Example 3.

[0072] Figure 7A: CLA concentration (mg / mL) as measured by UPLC-CAD for CLA-SNE and SNE formulations stored at 4°C, 25°C and 37°C for 1 month. See Example 4.

[0073] Figure 7B: Squalene concentration (mg / mL) as measured by UPLC-CAD for CLA-SNE and SNE formulations stored at 4°C. 25 °C and 37°C for 1 month. See Example 4.

[0074] Figure 8A: The CLA / squalene (w / w) % after dialysis is plotted versus the “target” (w / w) % before self-assembly. The CLA / squalene w / w % ratios (X) were measured by reverse phase UPLC-CAD before and after self-assembly and nanoemulsion dialysis. See Example 5.

[0075] Figure 8B: The measured intensity weighted Z-average DLS diameters of CLA-SNE nanoparticles after dialysis (X) is plotted versus the measured CLA / squalene (w / w) % after dialysis for each of MNS formulation. See Example 5.26100

[0076] Figure 8C : The measured Zeta Potential of CLA-SNE squalene nanoparticles (X) after dialysis at pH 5.5 is plotted versus the measured CLA / squalene (w / w) % after dialysis for each of MNS prepared formulations. See Example 6.

[0077] Figure 9: DLS Z-averages diameters of CLA-SNE samples formed and processed with aqueous phase (20 mM L-Histidine) of increasing pH values. See Example 6.

[0078] Figure 10: Final [CLA] (mg / mL) of CLA-SNE samples formed and processed with aqueous phase (20 mM L-Histidine) of increasing pH values. See Example 6.

[0079] Figure 11 : Show n 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.

[0080] Figure 12: Shown 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.

[0081] Figure 13: 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.

[0082] Figure 14: Shown are the NTso values on EBV infection of B cells and epithelial-like cells at day 42. The horizontal bar represents the geometric mean NTso values. Error bars represent the 95% confidence interval of the geometric mean NTso values.

[0083] Figure 15: Rhesus monkeys (5 per group) were injected intramuscularly with 3 doses of gp350-FNP and gL-gH-gp42-FNP dual particle vaccine formulated with SNE adjuvant A at weeks 0, 4. and 10 (indicated by arrows). Shown are the interpolated serum antibody titers against the EBV antigens gp350, gH / gL, or gp42 through week 42. The data are presented as geometric mean titers. Error bars represent the 95% confidence interval of the GMT.

[0084] Figure 16: Rhesus monkeys (5 per group) were injected intramuscularly with 3 doses of gp350-FNP and gL-gH-gp42-FNP dual particle vaccine formulated with SNE adjuvant A at weeks 0, 4. and 10 (indicated by arrows). Shown are the NTsovalues 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 GMT.

[0085] Figure 17: CD4 T cell cytokine responses (IFN-y) to gH 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.26100

[0086] Figure 18: CD4 T cell cytokine responses (IL-2) to gH 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.

[0087] Figure 19: CD4 T cell cytokine responses (TNF-a) to gH 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.

[0088] Figure 20: CD4 T cell cytokine responses (IFN-y) to 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.

[0089] Figure 21 : CD4 T cell cytokine responses (IL-2) to 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.

[0090] Figure 22: CD4 T cell cytokine responses (TNF-a) to 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.DETAILED DESCRIPTION

[0091] In one aspect, 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 squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80); and squalene.Squalene Nanoemulsion

[0092] Squalene nanoemulsions (“SNE’") of the invention refer to a formulation of emulsifiers, solubilizers, surfactants, lipids, or mixtures of the foregoing. In one embodiment, the disclosure provides, among other things, a composition that comprises three SNE components: (1) sorbitan trioleate (SPAN-85); (2) polysorbate-20 (PS-20) or polysorbate-80 (PS-80), and (3) squalene. In one embodiment, the disclosure provides, among other things, a composition that comprises four SNE components: (1) a cationic lipid; (2) sorbitan trioleate (SPAN-85); (3) polysorbate-20 (PS- 20) or polysorbate-80 (PS-80), and (4) squalene. In one embodiment, the SNE composition comprises the cationic lipid (13Z,16Z)-N, N-dimethyl-3-nonyldocosa-13,16-dien-l-amine (‘ CLA” or, when the cationic lipid is included in the SNE, “CLA-SNE”).26100

[0093] Cationic lipids and methods of making cationic lipids are well know n in the art.

[0094] In some embodiments, the cationic lipid includes any cationic lipid mentioned in U.S. Patent Application Publication Nos. US 2008 / 0085870, US 2008 / 0057080, US 2009 / 0263407, US 2009 / 0285881, US 2010 / 0055168, US 2010 / 0055169, US 2010 / 0063135, US 2010 / 0076055, US 2010 / 0099738. US 2010 / 0104629. US 2013 / 0017239, and US 2016 / 0361411, International Patent Application Publication No. WO2011 / 022460; WO2012 / 040184, WO2011 / 076807, W02010 / 021865, WO 2009 / 132131, WO2010 / 042877, WO2010 / 146740, WO2010 / 105209, and in U.S. Pat. Nos. 5.208,036, 5,264,618, 5,279,833, 5,283,185, 6,890,557, and 9,669,097.

[0095] In some embodiments, cationic lipids useful in the compositions of the invention have the following structure, illustrated by Formula 1:Formula 1 wherein:R1and R2are each methyl;R3is H; n is 1 or 2;Li is selected from C8-C24 alkyd and C8-C24 alkenyl; andL2 is selected from C4-C9 alkyl and C4-C9 alkenyl; or any pharmaceutically acceptable salt or stereoisomer thereof.

[0096] In some embodiments, the cationic lipid is an aminoalkyl lipid. In some embodiments, the cationic lipid is an asymmetric aminoalkyl lipid. In an embodiment of the invention, the cationic lipid is (13Z,16Z)-N,N-dimethyl-3-nonyldocosa-13,16-dien-l-amine (CLA); or (6Z,9Z,26Z,29Z)-N,N-dimethylpentatriaconta-6,9,26,29-tetraen-l 8-amine (CLX); or N,N- dimethyl-l-((lS,2R)-2-octylcyclopropyl)heptadecan-8-amine (CLY).

[0097] In another embodiment of the invention, the cationic lipid is selected from: DLinDMA;DLinKC2DMA; DLin-MC3-DMA; CLinDMA; S-Octyl CLinDMA; (2S)-l-{7-[(3P)-cholest-5- en-3-yloxy]heptyloxy}-3-[(4Z)-dec-4-en-l- yloxy]-N,N-dimethylpropan-2-amine; (2R)-l-{4- [(3P)-cholest-5-en-3-yloxy]butoxy}-3-[(4Z)-dec-4-en-l-yloxy]-N,N-dimethylpropan-2-amine; 1- [(2R)-l-{4-[(3 )-cholest-5-en-3-yloxy]butoxy}-3-(octyloxy)propan-2- yl]guanidine; l-[(2R)-l-{7- [(3|3)-cholest-5-en-3-yloxy]heptyloxy}-N,N-dimethyl-3-[(9Z,12Z)-octadeca-9,12-dien-l-26100 yloxy]propan-2-amine; l-[(2R)-l-{4-[(3P)-cholest-5-en-3-yloxy]butoxy}-N,N-dimethyl-3- [(9Z,12Z)-octadeca-9,12-dien-l-yloxy]propan-2-amine; (2S)-l-({6-[(3P))-cholest-5-en-3- yloxy]hexyl}oxy)-N,N-dimethyl-3-[(9Z)-octadec-9-en-l-yloxy]propan-2-amine; (3P)-3-[6- {[(2S)-3-[(9Z)-octadec-9-en-l-yloxyl]-2-(pyrrolidin-l-yl)propyl]oxy}hexyl)oxy] cholest-5-ene; (2R)-l-{4-[(3P)-cholest-5-en-3-yloxy]butoxy}-3-(octyloxy)propan-2 -amine; (2R)-l-({8-[(3P)- cholest-5-en-3-yloxy]octyl}oxy)-N,N-dimethyl-3-(pentyloxy)propan-2-amine; (2R)-l-({8-[(3P)- cholest-5-en-3-yloxy]octyl}oxy)-3-(heptyloxy)-N,N- dimethylpropan-2-amine; (2R)-l-({8-[(3P)- cholest-5-en-3-yloxy]octyl}oxy)-N,N-dimethyl-3-[(2Z)-pent- 2-en-l-yloxy]propan-2-amine; (2S)-l-butoxy-3-({8-[(3P)-cholest-5-en-3-yloxy]octyl}oxy)-N,N-dimethylpropan-2-amine; (2S-1- ({8-[(3P)-cholest-5-en-3-yloxy]octyl}oxy)-3-[2,2, 3, 3, 4, 4, 5, 5,6, 6,7, 7, 8, 8,9, 9- hexadecafluorononyl)oxy]-N,N-dimethylpropan-2-amine; 2-amino-2-{[(9Z, 12Z)-octadeca-9, 12-dien-l-yloxy]methyl} propane-1, 3-diol; 2-amino-3-({9-[(3p,8^,9^,14^,17c,,20£,)-cholest-5-en- 3-yloxy]nonyl}oxy)-2-{[(9Z,12Z)-octadeca-9,12-dien-l-yloxy]melhyl}propan-l-ol; 2-amino-3- ({6-[(3P,8^,9^,14^.17 20^)-cholest-5-en-3-yloxy]nonyl}oxy)-2-{[(9Z)-octadec-9-en- 1- yloxy]methyl}propan-l-ol; (20Z,23Z)-N,N-dimethylnonacosa-20,23-dien-10-amine; (17Z.20Z)- N,N-dimethylhexacosa-17,20-dien-9-amine; (16Z,19Z)-N,N-dimethylpentacosa-16,19-dien-8- amine; ( 13Z, 16Z)-N,N-dimethyldocosa- 13,16-dien-5-amine; (12Z, 15Z)-N,N-dimethy Ihenicosa- 12,15 -dien-4-amine; (14Z,17Z)-N,N-dimethyltricosa-14.17-dien-6-amine; (15Z.18Z)-N,N- dimethyltetracosa- 15,18-dien-7-amine; (18Z,21 Z)-N,N-dimethylheptacosa- 18,21 -di en- 10- amine; (15Z, 18Z)-N,N-dimethyltetracosa-l 5, 18-dien-5 -amine; (14Z, 17Z)-N,N-dimethyltricosa- 14,17-dien-4-amine; (19Z,22Z)-N,N-dimethyloctacosa-19,22-dien-9-amine; (18Z,21Z)-N,N- dimethylheptacosa- 18,21 -dien-8-amine; ( 17Z.20Z)-N,N-dimethy lhexacosa- 17.20-dien-7 -amine; (16Z,19Z)-N,N-dimethylpentacosa-16,19-dien-6-amine; (22Z,25Z)-N,N-dimethylhentriaconta- 22,25-dien-10-amine; (21 Z,24Z)-N,N-dimethyltriaconta-21,24-dien-9-amine; (18Z)-N,N- dimethylheptacos- 18-en- 10-amine; (17Z)-N,N-dimethylhexacos- 17-en-9-amine; (19Z,22Z)-N,N- dimethyloctacosa-19,22-dien-7-amine; N,N-dimethylheptacosan-l 0-amine; (20Z,23Z)-N-ethyl- N-methylnonacosa-20.23-dien-l 0-amine; 1-[(1 lZ,14Z)-l-nonylicosa-l 1,14-dien-l-yl] pyrrolidine; (20Z)-N,N-dimethylheptacos-20-en-l 0-amine; (15Z)-N,N-dimethylheptacos-15-en-10-amine; (14Z)-N,N-dimethylnonacos-14-en-l 0-amine; (17Z)-N,N-dimethylnonacos-17-en-10-amine; (24Z)-N,N-dimethyltritriacont-24-en-l 0-amine; (20Z)-N,N-dimethylnonacos-20-en-l 0-amine; (22Z)-N,N-dimethylhentriacont-22-en- 10-amine; ( 16Z)-N,N-dimethylpentacos- 16-en-8-amine; (12Z,15Z)-N,N-dimethyl-2-nonylhenicosa-12,15-dien-l-amine; (13Z,16Z)-N,N-dimethyl-3- nonyldocosa-13,16-dien-l-amine; N,N-dimethyl-l-[(lS,2R)-2-octylcyclopropyl]heptadecan-8-26100 amine; 1-[(1 S,2R)-2-hexylcyclopropyl]-N,N-dimethylnonadecan-10-amine; N,N-dimethyl-l- [(lS,2R)-2-octylcyclopropyl]nonadecan-10-amine; N,N-dimethyl-21 -[(lS,2R)-2- octylcyclopropyl]henicosan-10-amine; N,N-dimethyl-l-[(lS,2S)-2-{[(lR,2R)-2- pentylcyclopropyl]methyl}cyclopropyl]nonadecan-10-amine; N,N-dimethyl-l-[(lS,2R)-2- octylcyclopropyl]hexadecan-8-amine; N,N-dimethyl-l-[(lR.2S)-2- undecylcyclopropyl]tetradecan-5-amine; N,N-dimethyl-3-{7-[(l S,2R)-2- octylcyclopropyl]heptyl}dodecan-l-amine; l-[(lR,2S)-2-heptylcyclopropyl]-N,N- dimethyloctadecan-9-amine; l-[(lS,2R)-2-decylcyclopropyl]-N,N-dimethylpentadecan-6-amine; N,N-dimethyl-l-[(lS,2R)-2-octylcyclopropyl]pentadecan-8-amine; and (11E,2OZ,23Z)-N.N- dimethylnonacosa-1 l,20,23-trien-10-amine; or a pharmaceutically acceptable salt thereof, or a stereoisomer of any of the foregoing.

[0098] In another embodiment of the invention, the cationic lipid is (13Z,16Z)-N,N-dimethyl- 3-nonyldocosa-13,16-dien-l-amine, or a pharmaceutically acceptable salt or stereoisomer thereof.

[0099] In another embodiment of the invention, the cationic lipid is (13Z,16Z)-N,N-dimethyl- 3-nonyldocosa- 13,16-dien- 1 -amine (CLA).

[0100] In some embodiments, the disclosure provides, among other things, a composition that comprises three SNE components: (1) sorbitan trioleate (SPAN-85), (2) polysorbate-20 (PS-20) or polysorbate-80 (PS-80), and (3) squalene. In embodiments of this aspect of the invention, the SNE does not comprise a cationic lipid.

[0101] In some embodiments, the SNE comprises 32-97 mole % squalene, 1-34 mole % SPAN- 85 and 1-34 mole % of PS-20 or PS-80.

[0102] In some embodiments, the SNE comprises 86-98 mole % squalene, 1-7 mole % SPAN- 85 and 1-7 mole % of PS-20 or PS-80.

[0103] In some embodiments, the SNE comprises 92-94 mole % squalene, 3-4 mole % SPAN- 85 and 3-4 mole % of PS-20 or PS-80.

[0104] In one embodiment of the invention, the SNE comprises 92.91 mole % squalene, 3.98 mole % SPAN-85 and 3. 11 mole % of PS-20 or PS-80.

[0105] The disclosure also provides, among other things, a composition that comprises four SNE components: (1) a cationic lipid; (2) sorbitan trioleate (SPAN-85); (3) polysorbate-20 (PS- 20) or polysorbate-80 (PS-80), and (4) squalene. A particular SNE composition comprises the cationic lipid (13Z,16Z)-N, N-dimethyl-3-nonyldocosa-13,16-dien-l-amine ('‘CLA” or, when the cationic lipid is included in the SNE, “CLA-SNE”).26100

[0106] In some embodiments, the SNE comprises 1-60 mole % cationic lipid, 32-97 mole % squalene, 1-4 mole % SPAN-85 and 1-4 mole % of PS-20 or PS-80.

[0107] In some embodiments, the SNE comprises 10-14 mole % cationic lipid, 78-84 mole % squalene, 1-6 mole % SPAN-85 and 1-6 mole % of PS-20 or PS-80.

[0108] In some embodiments, the SNE comprises 40-46 mole % cationic lipid. 44-52 mole % squalene, 1-8 mole % SPAN-85 and 1-8 mole % of PS-20 or PS-80.

[0109] In one embodiment of the invention, the SNE comprises 13.82 mole % cationic lipid, 80.07 mole % squalene, 3.43 mole % SPAN-85 and 2.68 mole % of PS-20 or PS-80.

[0110] In one embodiment of the invention, the SNE comprises 44.5 mole % cationic lipid, 51.56 mole % squalene, 2.21 mole % SPAN-85 and 1.72 mole % of PS-20 or PS-80.[OHl] In one embodiment of the invention, the ratio of Squalene: SPAN-85:polysorbate 20 or 80 in the SNE is about 11-7: 1-3: 1-3 by weight.

[0112] In one embodiment of the invention, the ratio of Squalene: SPAN-85:polysorbate 20 or 80 in the SNE is about 10-7: 1-3: 1-3 by weight.

[0113] In one embodiment of the invention, the ratio of Squalene: SPAN-85:polysorbate 20 or 80 in the SNE is about 10-8:1-3:1-3 by weight.

[0114] In one embodiment of the invention, the ratio of Squalene: SPAN-85:polysorbate 20 or 80 in the SNE is about 10: 1-3: 1-3 by weight.

[0115] In one embodiment of the invention, the ratio of Squalene: SPAN-85:polysorbate 20 or 80 in the SNE is about 9: 1-3: 1 -3 by weight.

[0116] In one embodiment of the invention, the ratio of Squalene: SPAN-85:polysorbate 20 or 80 in the SNE is about 8: 1-3: 1-3 by weight.

[0117] In one embodiment of the invention, the ratio of Squalene: SPAN-85:polysorbate 20 or 80 in the SNE is about 7: 1 -3: 1 -3 by weight.

[0118] In one embodiment of the invention, the ratio of Squalene: SPAN-85:polysorbate 20 or 80 in the SNE is about 10: 1 : 1 by weight.

[0119] In some embodiments, the SNE comprises PS-20.

[0120] In some embodiments, the SNE comprises PS-80.

[0121] In some embodiments, the disclosure provides, among other things, a composition that comprises one or more non-cationic lipids which can be selected from a surfactant, a mixture of surfactants, a phospholipid, a terpene, a terpenoid, a triterpene; or a combination of the foregoing.26100

[0122] In some embodiments, the surfactant is selected from the group consisting of the polyoxyethylene sorbitan esters surfactants (commonly referred to as the Tweens), especially PS- 20 and PS-80; copolymers of ethylene oxide (EO), propylene oxide (PO), and / or butylene oxide (BO), sold under the DOWFAX™ tradename, such as linear EO / PO block copolymers; octoxynols, which can vary in the number of repeating ethoxy (oxy-1.2-ethanediyl) groups, with octoxynol-9 (Triton X-100, or t-octylphenoxypoly ethoxy ethanol) being of particular interest (octylphenoxy)poly ethoxyethanol (IGEPAL CA-630 / NP-40); nonylphenol ethoxylates, such as the Tergitol™ NP series; polyoxyethylene fatty ethers derived from lauryl, cetyl, st ear l and oleyl alcohols (known as Brij surfactants), such as triethyleneglycol monolauryl ether (Brij 30); and sorbitan esters (commonly known as the SPANs), such as sorbitan trioleate (SPAN-85, Tween-85 or [2-[(27?,3S',4J?)-4-hydroxy-3-[(Z)-octadec-9-enoyl]oxyoxolan-2-yl]-2-[(Z)-octadec- 9-enoyl] oxy ethyl] (Z)-octadec-9-enoate) and sorbitan monolaurate.

[0123] In some embodiments, mixtures of surfactants are used, e.g., PS-20 / SPAN 85 or PS- 80 / SPAN 85 mixtures. A combination of a polyoxyethylene sorbitan ester such as polyoxyethylene sorbitan monooleate (PS-80) and an octoxynol such as t- octylphenoxypoly ethoxy ethanol (Triton X-100) are also suitable. Another useful combination comprises laureth 9 plus a polyoxyethylene sorbitan ester and / or an octoxynol.

[0124] In some embodiments, the amounts of surfactants or emulsifiers are: polyoxyethylene sorbitan esters (such as PS-20 or PS-80) 0.01 to 10 mole %, in particular about 1 to 4 mole %; octyl- or nonylphenoxy poly oxyethanols (such as Triton X-100, or other detergents in the Triton series) 0.001 to 10 mole %, in particular about 1 to 4 mole %; w / v, in particular 0.01 to 0.1% w / v; polyoxyethylene ethers (such as laureth 9) 0.1 to 20 mole %, preferably 0.5 to 10 mole % and in particular 1 to 4% mole % or about 10 % by mass.

[0125] In some embodiments, the phospholipid is selected from natural phospholipids including 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. Phospholipid derivatives include phosphatidic acid (DMPA, DPP A, DSPA), phosphatidylcholine (DDPC, DLPC, DMPC, DPPC, DSPC, DOPC, POPC, DEPC), phosphatidylglycerol (DMPG, DPPG, DSPG, POPG), phosphatidylethanolamine (DMPE, DPPE, DSPE DOPE), phosphatidylserine (DOPS). Fatty acids include C 14:0, palmitic acid (C16:0), stearic acid (C18:0), oleic acid (C18: 1), linoleic acid (C18:2), linolenic acid (C18:3), and arachidonic acid26100(C20:4), C20:0, C22:0 and lethicin. In certain embodiments of the invention, the phospholipid is phosphatidyl serine, l,2-distearoyl-sn-glycero-3- phosphocholine (DSPC), 1,2-dipalmitoleoyl-sn- glycero-3-phosphocholine, 1 ,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC), dilauroylphosphatidylcholine (DLPC), l,2-dieicosenoyl-sn-glycero-3-phosphocholine, or 1,2- dioleoyl-sn-glycero-3-phosphocholine (DOPC).

[0126] In some embodiments, the terpine is selected from monoterpenes including geraniol, terpineol, limonene, myrcene, linalool or pinene; sesquiterpenes including humulene, famesenes, famesol; diterpenes including cafestol, kahweol, cembrene and taxadiene, triterpenes including squalene and squalane; tetraterpenes including acyclic lycopene, the monocyclic gammacarotene, and the bicyclic alpha- and beta-carotenes; polyterpenes and norisoprenoids. In some embodiments, the terpine is squalene.

[0127] In one embodiment of the invention, the SNE comprises 50-85 mole % squalene, and 1- 10 mole % non-ionic surfactants. In one aspect of this embodiment, the non-ionic surfactant comprises a mixture of PS-20 and SPAN-85 or a mixture of PS-80 and SPAN-85.

[0128] In one embodiment of the invention, the SNE comprises 0-45 mole % cationic lipid, 50- 85 mole % squalene, and 1-10 mole % non-ionic surfactants. In one aspect of this embodiment, the non-ionic surfactant comprises a mixture of PS-20 and SPAN-85 or a mixture of PS-80 and SPAN-85.

[0129] In one embodiment of the invention, the SNE comprises one or more cationic lipids, one or more terpenes (e.g., squalene), and / or one or more sorbitan-based surfactants (e.g., PS-20 or PS-80; SPAN-85) at specific molar ratios.General Methods of Making SNEs (with and without a cationic lipid)

[0130] Generally, SNEs may be formed, for example, by initially combining and mixing the lipid components together, or initially utilizing a single lipid, such as a cationic lipid. Once mixed and blended (when combining and mixing lipid components together), an aqueous buffer is added and mixed with the initial lipid or lipid components to form a blended emulsion mixture. The blended emulsion components are first subjected to course homogenization followed by fine homogenization. Then, the resulting formulation is subjected to a final filtration step and stored at 4°C. A lipid solution may include one or more cationic lipids, one or more terpenes (e g., squalene), one or more sorbitan-based surfactants (e.g., PS-20 or PS-80; SPAN-85) at specific molar ratios.26100EBV Polypeptides

[0131] 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

[0132] 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.

[0133] 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 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.

[0134] 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.

[0135] 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.

[0136] 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.

[0137] 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 such26100 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.

[0138] 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.

[0139] In some embodiments, gL and gH poly peptides 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 tnmerization 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).

[0140] 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.

[0141] 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%, atleast 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

[0142] 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.

[0143] 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 gp42 polypeptide comprises an ammo acid sequence with at least 80%, 85%, 90%, 95%. 97%. 98%, 99%, or 100% identity to SEQ ID NO: 3.

[0144] 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).

[0145] 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 acidsequence with at least 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 8.

[0146] 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.

[0147] The gp42 polypeptide can be combined with any of the ferritins discussed herein. For example, in some embodiments, a polypeptide comprises a gp42 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.

[0148] 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

[0149] 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 ferntin 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 1.

[0150] 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.26100

[0151] 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 to SEQ ID NO: 8. 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: 9.

[0152] 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.

[0153] 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 15-2610090, 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.

[0154] 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 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 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% identity7to 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

[0155] 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.

[0156] 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,26100 the EBV gH polypeptide, the EBV gp42 polypeptide, and the ferritin are arranged in N-terminal to C-terminal order within the EBV polypeptide.

[0157] 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 / or between 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.

[0158] 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,26100 the EBV polypeptide consists essentially of the amino acid sequence of any one of SEQ ID Nos: 11-21.

[0159] 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.

[0160] In some embodiments, an EBV polypeptide that comprises 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 ammo 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.26100

[0161] 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.

[0162] 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.

[0163] 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 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. 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: 23.26100E. EBV polypeptides comprising a gp220 polypeptide

[0164] 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.

[0165] 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.

[0166] 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.

[0167] 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.F. EBV polypeptides comprising an EBV polypeptide and ferritin

[0168] 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.

[0169] 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 the26100 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.

[0170] 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.

[0171] 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 ferritin and an EBV polypeptide can assemble with a heavychain 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, a non-ferritin polypeptide) may be referred as a “naked ferritin."

[0172] 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.

[0173] In some embodiments, an antigenic polypeptide comprising a heavy chain ferntin 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.

[0174] 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.

[0175] 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.26100

[0176] 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.

[0177] 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.

[0178] Any type of ferritin nanoparticle(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.

[0179] 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)).

[0180] 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

[0181] 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 acid residue 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 1 (SEQUENCE TABLE): DESCRIPTION OF THE SEQUENCES26100261002610026100261002610026100261002610026100261002610026100261002610026100Compositions ofEBV polypeptide and SNE adjuvant

[0182] The inventions provides the following embodiments:1. A composition comprising: (a) an Epstein Ban 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 squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN- 85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80); and squalene.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.3. 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 in N-terminal to C-terminal order.26100In 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.14. 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.2610016. 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.

[0183] 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 ammo 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 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 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.

[0184] 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 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 some embodiments under embodiment 24, the composition comprises an EBV polypeptide that comprises or consists of an amino acid sequence26100 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 squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN- 85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80); and squalene.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%,26100 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 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.

[0185] 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) an SNE 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, 2526100 or 26. In some embodiments, the first EBV polypeptide comprises, consists or consists essentially of an amino acid sequence of SEQ ID NO: 21.

[0186] 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.

[0187] 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 at least 99% sequence identity to SEQ ID NO: 24.

[0188] 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 polypeptide26100 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.

[0189] 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 SNE adjuvant.36. A pharmaceutical unit dose comprising about 5 pg to about 200 pg of EBV polypeptide comprising or consisting of any one of the amino acid sequences of SEQ ID Nos: 11- 21, and about 2 mg to about 30 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80); and about 2 mg to about 25 mg of squalene.

[0190] In one aspect under embodiment 36, 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 36, the pharmaceutical unit dose comprises about 10 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 36, 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: 11-21.2610037. The pharmaceutical unit dose of embodiment 36, wherein the EBV polypeptide 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 6 mg to about 18 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80); and about 5 mg to about 15 mg of squalene.

[0191] In one aspect under embodiment 37, the pharmaceutical unit dose comprises about 1 mg to about 4 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80) and squalene. In one aspect under embodiment 37, the pharmaceutical unit dose comprises about 2 mg to about 5 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80) and squalene.38. The composition or pharmaceutical unit dose of any one of embodiments 1-37, wherein the ratio of Squalene: SPAN-85 :poly sorbate 20 or 80 is about 11-7: 1-3: 1-3 by w eight.

[0192] In one aspect under embodiment 38, the composition or pharmaceutical unit dose of any one of embodiments 1-37, the ratio of Squalene: SPAN-85 :polysorbate 20 or 80 is 10-7: 1 : 1 by weight. In one aspect under embodiment 38, the composition or pharmaceutical unit dose of any one of embodiments 1-37, the ratio of Squalene: SPAN-85 :polysorbate 20 or 80 is 10-8: 1 : 1 by weight.39. The composition or pharmaceutical unit dose of any one of embodiments 1-37, wherein the ratio of Squalene: SPAN-85 :polysorbate 20 or 80 is about 10: 1 : 1 by weight.

[0193] In one aspect under embodiment 39, the composition or pharmaceutical unit dose of any one of embodiments 1-37, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20); and squalene, wherein the ratio of Squalene: SPAN-85:polysorbate 20 is 10: 1: 1 by weight.40. The pharmaceutical unit dose of embodiment 36, wherein the EBV polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 10 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 6 mg; wherein the ratio of Squalene: SPAN-85:PS-20 or PS-80 is 10: 1 : 1 by weight.41. The pharmaceutical unit dose of embodiment 36, wherein the EBV polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 30 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 6 mg; wherein the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10: 1: 1 by weight.2610042. The pharmaceutical unit dose of embodiment 36, wherein the EBV polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 100 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 6 mg; wherein the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10: 1 : 1 by weight.43. The pharmaceutical unit dose of embodiment 36, wherein the EBV polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 10 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 12 mg; wherein the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10: 1 : 1 by weight.44. The pharmaceutical unit dose of embodiment 36, wherein the EBV polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 30 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 12 mg; wherein the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10: 1 : 1 by weight.45. The pharmaceutical unit dose of embodiment 36, wherein the EBV polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 100 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 12 mg; wherein the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10: 1 : 1 by weight.46. The pharmaceutical unit dose of embodiment 36, wherein the EBV polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 10 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 18 mg; wherein the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10: 1 : 1 by weight.47. The pharmaceutical unit dose of embodiment 36, wherein the EBV polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 30 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 18 mg; wherein the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10: 1 : 1 by weight.48. The pharmaceutical unit dose of embodiment 36, wherein the EBV polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 100 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 18 mg; wherein the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10: 1 : 1 by weight.49. A pharmaceutical unit dose comprising about 5 pg to about 200 pg of EBV polypeptide comprising or consisting of the amino acid sequence of SEQ ID NO: 23, and about 2 mg to about 30 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80); and about 2 mg to about 25 mg of squalene.2610050. The pharmaceutical unit dose of embodiment 49, wherein the EBV polypeptide 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 6 mg to about 18 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80); and about 5 mg to about 15 mg of squalene.

[0194] In one aspect under embodiment 50, the pharmaceutical unit dose comprises about 1 mg to about 4 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80) and squalene. In one aspect under embodiment 50, the pharmaceutical unit dose comprises about 2 mg to about 5 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80) and squalene.51. The pharmaceutical unit dose of any one of embodiments 49-50, wherein the ratio of Squalene: SPAN-85:polysorbate 20 or 80 is about 11-7: 1-3: 1-3 by weight.

[0195] In one aspect under embodiment 51, the ratio of Squalene: SPAN-85 :polysorbate 20 or 80 is 10-7: 1: 1 by weight. In one aspect under embodiment 51, the ratio of Squalene: SPAN- 85:polysorbate 20 or 80 is 10-8: 1: 1 by weight.52. The pharmaceutical unit dose of any one of embodiments 49-50, wherein the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10: 1 : 1 by weight.53. The pharmaceutical unit dose of embodiment 49, wherein the EBV polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 10 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 6 mg; wherein the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10: 1 : 1 by weight.54. The pharmaceutical unit dose of embodiment 50, wherein the EBV polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 30 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 6 mg; wherein the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10: 1 : 1 by weight.55. The pharmaceutical unit dose of embodiment 50, wherein the EBV polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 100 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 6 mg; wherein the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10: 1 : 1 by weight.56. The pharmaceutical unit dose of embodiment 50, wherein the EBV polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 10 pg, andthe squalene nanoemulsion (SNE) adjuvant is in the amount of 12 mg; wherein the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10: 1 : 1 by weight.57. The pharmaceutical unit dose of embodiment 50, wherein the EBV polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 30 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 12 mg; wherein the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10: 1 : 1 by weight.58. The pharmaceutical unit dose of embodiment 50, wherein the EBV polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 100 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 12 mg; wherein the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10: 1 : 1 by weight.59. The pharmaceutical unit dose of embodiment 50, wherein the EBV polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 10 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 18 mg; wherein the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10: 1 : 1 by weight.60. The pharmaceutical unit dose of embodiment 50, wherein the EBV polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 30 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 18 mg; wherein the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10: 1 : 1 by weight.61. The pharmaceutical unit dose of embodiment 50, wherein the EBV polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 100 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 18 mg; wherein the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10: 1 : 1 by weight.62. A pharmaceutical unit dose comprising about 1 pg to about 100 pg of a first EBV polypeptide comprising or consisting of the amino acid sequence of SEQ ID NO: 21 and about 10 pg to about 100 pg of a second EBV polypeptide comprising or consisting of the amino acid sequence of SEQ ID NO: 23; and about 6 mg to about 36 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate- 20 (PS-20) or polysorbate-80 (PS-80); and squalene; wherein the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 11-7: 1-3: 1-3 by weight.

[0196] In one aspect under embodiment 62, the pharmaceutical unit dose comprises about 1 mg to about 4 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80) and squalene. In one aspect under embodiment 62, the pharmaceutical unit dose comprises about 2 mg to about 5mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80) and squalene. In one aspect under embodiment 62, the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10-8:1-2: 1-2 by weight. In one aspect under embodiment 62, the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10-7: 1-2: 1-2 by weight. In one aspect under embodiment 62, the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10: 1: 1 by weight. In certain embodiments of the foregoing embodiments, the pharmaceutical dose comprises 12 mg of a squalene nanoemulsion (SNE) adjuvant. In certain embodiments of the foregoing embodiments, the pharmaceutical dose comprises 24 mg of a squalene nanoemulsion (SNE) adjuvant. In certain embodiments of the foregoing embodiments, the pharmaceutical dose comprises 36 mg of a squalene nanoemulsion (SNE) 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.

[0197] 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 some26100 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.62a. A pharmaceutical unit dose comprising about 10 pg to about 100 pg of a first EBV polypeptide comprising or consisting of the amino acid sequence of SEQ ID NO: 21 and about 10 pg to about 100 pg of a second EBV polypeptide comprising or consisting of the amino acid sequence of SEQ ID NO: 23; and about 6 mg to about 36 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate- 20 (PS-20) or polysorbate-80 (PS-80); and squalene; wherein the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10: 1 : 1 by weight.

[0198] In one aspect under embodiment 62a, the pharmaceutical unit dose comprises about 1 mg to about 4 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80) and squalene. In one aspect under embodiment 62a, the pharmaceutical unit dose comprises about 2 mg to about 5 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80) and squalene. In certain embodiments of the foregoing embodiments, the pharmaceutical dose comprises 12 mg of a squalene nanoemulsion (SNE) adjuvant. In certain embodiments of the foregoing embodiments, the pharmaceutical dose comprises 24 mg of a squalene nanoemulsion (SNE) adjuvant. In certain embodiments of the foregoing embodiments, the pharmaceutical dose comprises 36 mg of a squalene nanoemulsion (SNE) adjuvant.

[0199] In one aspect under embodiment 62 or 62a, 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 one26100 embodiment the first EBV polypeptide is at 100 pg, and the second EBV polypeptide is at 30 fig-63. The pharmaceutical unit dose of embodiment 62 or 62a, wherein each of the first and second EBV polypeptide is at 10 pg.64. The pharmaceutical unit dose of embodiment 62 or 62a. wherein each of the first and second EBV polypeptide is at 30 pg.65. The pharmaceutical unit dose of embodiment 62 or 62a, wherein each of the first and second EBV polypeptide is at 100 pg.66. The pharmaceutical unit dose of embodiment 62a, wherein the first EBV polypeptide is at 10 pg, and the second EBV polypeptide is at 30 pg.67. The pharmaceutical unit dose of embodiment 62 or 62a, wherein the first EBV polypeptide is at 10 pg, and the second EBV polypeptide is at 100 pg.68. The pharmaceutical unit dose of embodiment 62 or 62a, wherein the first EBV polypeptide is at 30 pg, and the second EBV polypeptide is at 10 pg.69. The pharmaceutical unit dose of embodiment 62 or 62a, wherein the first EBV polypeptide is at 30 pg, and the second EBV polypeptide is at 100 pg.70. The pharmaceutical unit dose of embodiment 62 or 62a, wherein the first EBV polypeptide is at 100 pg. and the second EBV polypeptide is at 10 pg.71. The pharmaceutical unit dose of embodiment 62 or 62a, wherein the first EBV polypeptide is at 100 pg, and the second EBV polypeptide is at 30 pg.72. The composition or pharmaceutical unit dose of any one of embodiments 1-71, wherein the SNE adjuvant further comprises a cationic lipid.73. The composition or pharmaceutical unit dose of embodiment 72, wherein the cationic lipid is (13Z,16Z)-N,N-dimethyl-3-nonyldocosa-13,16-dien-l-amine.74. The composition or pharmaceutical unit dose of any one of embodiments 1-73, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) and squalene.75. The composition or pharmaceutical unit dose of any one of embodiments 1-74, wherein the SNE adjuvant comprises emulsion particles of about 10 to about 1000 nm.75 a. The composition or pharmaceutical unit dose of any one of embodiments 1-74, wherein the Z-av erage of the SNE adjuvant is about 20 to about 500 nm.

[0200] In one aspect under embodiment 75a, the Z-average of the SNE adjuvant is about 30 to about 300 nm. In one aspect under embodiment 75a, the Z-average of the SNE adjuvant is about2610050 to about 200 nm. In one aspect under embodiment 75a, the Z-average of the SNE adjuvant is about 100 to about 300 nm. In one aspect under embodiment 75a, the Z-average of the SNE adjuvant is about 100 to about 200 nm. In one aspect under embodiment 75a, the Z-average of the SNE adjuvant is about 120 to about 180 nm. In one aspect under embodiment 75a, the Z- average of the SNE adjuvant is about 80 to about 180 nm. In one aspect under embodiment 75a, the Z-average of the SNE adjuvant is about 130 to about 170 nm. In one aspect under embodiment 75a, the Z-average of the SNE adjuvant is about 150 to about 170 nm. In one aspect under embodiment 75a, the Z-average of the SNE adjuvant is about 160 nm. In one aspect of the foregoing embodiments, the Z-average is measured with dynamic light scattering.76. The composition or pharmaceutical unit dose of any one of embodiments 1-75 wherein the SNE adjuvant further comprises a buffer.77. The composition or pharmaceutical unit dose of embodiment 76, wherein the buffer is selected from the group consisting of: acetic acid, histidine, citrate, Bis-Tris, HEPES, phosphate, MES. sodium chloride, succinate, Tris, and combinations thereof.78. The composition or pharmaceutical unit dose of any one of embodiments 76 and 77, wherein the buffer is present in the amount of about ImM to about lOOmM.79. The composition or pharmaceutical unit dose of any one of embodiments 1-78, wherein the SNE adjuvant further comprises a salt.80. The composition or pharmaceutical unit dose of embodiment 79, wherein the salt is NaCl.81. The composition or pharmaceutical unit dose of embodiment 79, wherein the SNE adjuvant comprises 5 mM - 40 mM histidine at pH 5. 1 - 7.0 and 25 mM - 300 mM NaCl.82. The composition or pharmaceutical unit dose of embodiment 79, wherein the SNE adjuvant comprises about 20 mM histidine at about pH 5.8 and about 75 mM NaCl.83. The composition or pharmaceutical unit dose of any one of embodiments 1-82, wherein the SNE adjuvant comprises about 4 mg / ml sorbitan trioleate (SPAN-85); about 0.05% polysorbate-20 (PS-20); about 40 mg / ml squalene, about 20 mM histidine at about pH 5.8 and about 75 mM NaCl.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 1-83.2610085. 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 of any one of embodiments 1-82.86. 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 1-82.87. The method of embodiment 86, wherein the cancer is Burkitt’s or Hodgkin’s lymphoma, or nasopharyngeal cancer.88. The composition or pharmaceutical unit dose of any one of embodiments 1-82 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 1-82 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 1-82 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 administering to the patient a therapeutically effective amount of the EBV polypeptide described in any one of embodiments 1-71 and the SNE 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 administering to the patient a therapeutically effective amount of the EBV polypeptide of any one of embodiments 1-71 and a therapeutically effective amount of the SNE 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 administering to the patient a therapeutically effective amount of the EBV polypeptide of any one of embodiments 1-71 and a therapeutically effective amount of the SNE adjuvant of any one of embodiments 1-83.94. The method of embodiment 93, wherein the cancer is Burkitt’s or Hodgkin’s lymphomas, or nasopharyngeal cancer.

[0201] In one aspect of the methods under any one of embodiments 90-94, 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 6 mg to about 18 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises26100 sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80); and about 5 mg to about 15 mg of squalene.

[0202] In one aspect of the methods under any one of embodiments 90-94, 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 6 mg to about 18 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80); and about 5 mg to about 15 mg of squalene.

[0203] In one embodiment of the foregoing embodiments, the EBV polypeptide is in the amount of 100 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 6 mg. In one embodiment of the foregoing embodiments, the EBV polypeptide is in the amount of 100 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 12 mg. In one embodiment of the foregoing embodiments, the EBV polypeptide is in the amount of 100 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 18 mg.

[0204] In one embodiment of the foregoing embodiments, the EBV polypeptide is in the amount of 30 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 6 mg. In one embodiment of the foregoing embodiments, the EBV polypeptide is in the amount of 30 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 12 mg. In one embodiment of the foregoing embodiments, the EBV polypeptide is in the amount of 30 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 18 mg. In one embodiment of the foregoing embodiments, the EBV polypeptide is in the amount of 10 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 6 mg. In one embodiment of the foregoing embodiments, the EBV polypeptide is in the amount of 10 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 12 mg. In one embodiment of the foregoing embodiments, the EBV polypeptide is in the amount of 10 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 18 mg. In one embodiment, the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10: 1: 1 by weight.

[0205] In one aspect under the methods and use of the above embodiments, the composition or pharmaceutical unit dose, or EBV polypeptide and SNE 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. In26100 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.

[0206] Any of the EBV polypeptides described in Sections A-G and Table 1 are suitable for use in the compositions or pharmaceutical compositions and methods of the invention. The EBV polypeptide can be field mixed with an SNE adjuvant of the invention (e.g., mixed by the clinician just prior to administration to a patient, lyophilized EBV polypeptide reconstituted with SNE adjuvant solution) or can be formulated together with the SNE adjuvant.

[0207] 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 elicit a 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.

[0208] 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.

[0209] In an embodiment, a composition is provided, wherein the CLA-SNE comprises PS-20, sorbitan trioleate, squalene and (13Z, 16Z) - N, N-dimethyl-3-nonyldocosa 13, 16-dien-l -amine.

[0210] In an embodiment, a composition is provided, wherein the CLA-SNE comprises 5-15 mol% sorbitan trioleate, 25-35 mole% PS-20 or PS-80, 1-2.5 mol% squalene, and 55-65 mol% (13Z. 16Z) - N, N-dimethyl-3-nonyldocosa 13, 16-dien-l -amine.

[0211] In an embodiment, a composition is provided, wherein the SNE, including the cationic lipid, comprises up to 75 mol% of cationic lipid, up to 30 mol% of sorbitan trioleate, up to 30 mol% of polysorbate-20 or polysorbate-80 and 25-85 mol% of squalene.

[0212] In an embodiment, a composition is provided, wherein the SNE, including the cationic lipid, comprises up to 50 mol% of cationic lipid, up to 10 mol% of sorbitan trioleate, up to 10 mol% of polysorbate-20 or polysorbate-80 and 50-80 mol% of squalene.

[0213] In an embodiment, a composition is provided, wherein the SNE, including the cationic lipid, comprises up to 24 mol% of cationic lipid, 1-8 mol% of sorbitan trioleate, 1-8 mol% of polysorbate-20 or polysorbate-80 and 60-75 mol% of squalene.26100

[0214] In an embodiment, a composition is provided, wherein the SNE, including the cationic lipid, comprises about 10-14 mol% of cationic lipid, 1-4 mol% of sorbitan trioleate, 1-4 mol% of polysorbate-20 or polysorbate-80 and 50-80 mol% of squalene.

[0215] In an embodiment, a composition is provided, wherein the SNE, including the cationic lipid, comprises 30-65 mol % cationic lipid. 5-30 mol % sorbitan trioleate, 10-40 mol% squalene, and 0.5-4 mol% PS-20 or PS-80.

[0216] In an embodiment, a composition is provided, wherein the SNE, including the cationic lipid, comprises 55-65 mol % cationic lipid, 5-15 mol% sorbitan trioleate, 25-35 mol% squalene, and 1-2.5 mol % PS-20 or PS-80.

[0217] In an embodiment, a composition is provided, wherein the SNE, including the cationic lipid, comprises 13-45 mol % cationic lipid, 2-4 mol % sorbitan trioleate, 50-82 mol% squalene, and 1.5-3 mol% PS-20 or PS-80.

[0218] In an embodiment, a composition is provided, wherein the SNE. including the cationic lipid, comprises 13-14 mol % cationic lipid, 1-2 mol % sorbitan trioleate, 79-81 mol% squalene, and 1-2 mol% PS-20 or PS-80.

[0219] In an embodiment, a composition is provided, wherein the SNE, including the cationic lipid, comprises 20 mol % cationic lipid, 30 mol% sorbitan trioleate, 20 mol% squalene, and 30 mol % PS-20 or PS-80.

[0220] In an embodiment, a composition is provided, wherein the SNE, including the cationic lipid, comprises about 2 mol % cationic lipid, about 8 mol% sorbitan trioleate, about 82 mol% squalene, and about 8 mol % PS-20 or PS-80.

[0221] In an embodiment, a composition is provided, wherein the SNE. including the cationic lipid, comprises 2 mol % cationic lipid, 8 mol% sorbitan trioleate, 82 mol% squalene, and 8 mol % PS-20 or PS-80.

[0222] In an embodiment, a composition is provided, wherein the SNE, including the cationic lipid, comprises about 13.82 mol % cationic lipid, about 3.43 mol% sorbitan trioleate, about 80.07 mol% squalene, and about 2.68 mol % PS-20 or PS-80.

[0223] In an embodiment, a composition is provided, wherein the SNE, including the cationic lipid, comprises 13.82 mol % cationic lipid, 3.43 mol% sorbitan trioleate, 80.07 mol% squalene, and 2.68 mol % PS-20 or PS-80.

[0224] In an embodiment, a composition is provided, wherein the SNE. including the cationic lipid, comprises about 44.5 mol % cationic lipid, about 2.21 mol% sorbitan trioleate, about 51.56 mol% squalene, and about 1.72 mol % PS-20 or PS-80.26100

[0225] In an embodiment, a composition is provided, wherein the SNE, including the cationic lipid, comprises 44.5 mol % cationic lipid, 2.21 mol% sorbitan trioleate, 51.56 mol% squalene, and 1.72 mol % PS-20 or PS-80.

[0226] In an embodiment, a composition is provided, wherein the SNE comprises 32 mole % squalene. 34 mole % SPAN-85 and 34 mole % of PS-20 or PS-80.

[0227] In an embodiment, a composition is provided, wherein the SNE comprises 98 mole % squalene, 1 mole % SPAN-85 and 1 mole % of PS-20 or PS-80.

[0228] In an embodiment, a composition is provided, wherein the SNE comprises 86 mole % squalene. 7 mole % SPAN-85 and 7 mole % of PS-20 or PS-80.

[0229] In an embodiment, a composition is provided, wherein the SNE comprises 92 mole % squalene, 4 mole % SPAN-85 and 4 mole % of PS-20 or PS-80.

[0230] In an embodiment, a composition is provided, wherein the SNE comprises 94 mole % squalene. 3 mole % SPAN-85 and 3 mole % of PS-20 or PS-80.

[0231] In an embodiment, a composition is provided, wherein the SNE comprises 92.91 mole % squalene, 3.98 mole % SPAN-85 and 3.11 mole % of PS-20 or PS-80.

[0232] In an embodiment, a composition is provided, wherein the SNE comprises 62 mole % squalene, 17 mole % SPAN-85 and 17 mole % of PS-20 or PS-80.

[0233] 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 1 mg to about 30 mg SNE adjuvant.

[0234] 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.

[0235] 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 type of EBV polypeptide in the vaccine. The vaccine, pharmaceutical composition, or26100 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

[0236] Also provided herein are kits including any of the compositions or pharmaceutical compositions as described above and instructions for use.

[0237] Also provided herein are kits including (1) a pharmaceutical composition comprising any of the EBV polypeptides described in Sections A-G and (2) an SNE adjuvant.

[0238] In an embodiment, a kit is provided comprising: (a) an Epstein Barr Virus (EBV) vaccine; and (b) a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80); and squalene.

[0239] In some embodiments of the kits, the SNE adjuvant is any of the SNE adjuvants described herein above. In some embodiments, the kit includes about 1 mg to about 30 mg of an SNE adjuvant comprising SPAN-85, PS-20 or PS-80 and squalene. In some embodiments, the kit includes about 5 mg to about 30 mg of an SNE adjuvant comprising SPAN-85, PS-20 or PS-80 and squalene. In some embodiments, the kit includes about 6 mg to about 18 mg of an SNE adjuvant comprising SPAN-85, PS-20 or PS-80 and squalene. In some embodiments, the kit includes about 6 mg of an SNE adjuvant comprising SPAN-85, PS-20 or PS-80 and squalene. In some embodiments, the kit includes about 12 mg of an SNE adjuvant comprising SPAN-85, PS- 20 or PS-80 and squalene. In some embodiments, the kit includes about 18 mg of an SNE adjuvant comprising SPAN-85, PS-20 or PS-80 and squalene. In the foregoing embodiments, in one embodiment, the weight ratio of Squalene: SPAN-25 :PS-20 is 7-11 : 1: 1. In the foregoing embodiments, in one embodiment, the weight ratio of Squalene: SPAN-25 :PS-20 is 8: 1 : 1. In the foregoing embodiments, in one embodiment, the weight ratio of Squalene: SPAN-25:PS-20 is 9: 1 : 1. In the foregoing embodiments, in one embodiment, the weight ratio of Squalene: SPAN- 25:PS-20 is 10: 1 : 1. In the foregoing embodiments, in one embodiment, the weight ratio of Squalene: SPAN-25:PS-80 is 7-11: 1: 1. In the foregoing embodiments, in one embodiment, the weight ratio of Squalene: SPAN-25 :PS-80 is 8: 1 : 1. In the foregoing embodiments, in one embodiment, the weight ratio of Squalene: SPAN-25: PS-80 is 9: 1 : 1. In the foregoing embodiments, in one embodiment, the weight ratio of Squalene: SPAN-25 : PS-80 is 10: 1 : 1. In some embodiments, the kit further comprises a cationic lipid. In another embodiment, the26100 cationic lipid is CLA. In another embodiment, the cationic lipid is CLX. In another embodiment, the cationic lipid is CLY.

[0240] In some embodiments, the kit includes about 6 pg / mL - 24 mg / rnL SPAN-85, 6 pg / mL - 24 mg / mL PS-20 or PS-80 and 60 pg / mL - 240 mg / mL of squalene. In some embodiments, the kit includes about 2 pg / mL - 24 mg / mL SPAN-85, 2 pg / mL - 2.4 mg / mL PS-20 or PS-80 and 20 pg / mL - 24 mg / mL of squalene. In another embodiment, the kit includes 6 pg / mL - 2.4 mg / mL SPAN-85, 6 pg / mL - 2.4 mg / mL PS-20 or PS-80 and 60 pg / mL - 24 mg / mL of squalene.

[0241] In some embodiments, the kit includes 30 pg / mL to about 2.4 mg / mL cationic lipid and further includes 6 pg / mL - 14 mg / mL SPAN-85, 6 pg / mL - 14 mg / mL PS-20 or PS-80 and 60 pg / mL - 34 mg / mL of squalene. In another embodiment, the cationic lipid is CLA. In another embodiment, the cationic lipid is CLX. In another embodiment, the cationic lipid is CLY.

[0242] In an embodiment, the kit includes a CLA-SNE adjuvant, wherein the SNE comprises PS-20, sorbitan trioleate, squalene and (13Z, 16Z) - N, N-dimethyl-3-nonyldocosa 13, 16-dien-l- amine.

[0243] In an embodiment, the kit includes an SNE adjuvant, wherein the SNE, comprises 32 mole % squalene, 34 mole % SPAN-85 and 34 mole % of PS-20 or PS-80.

[0244] In an embodiment, the kit includes an SNE adjuvant, wherein the SNE, comprises 98 mole % squalene, 1 mole % SPAN-85 and 1 mole % of PS-20 or PS-80.

[0245] In an embodiment, the kit includes an SNE adjuvant, wherein the SNE, comprises 86 mole % squalene, 7 mole % SPAN-85 and 7 mole % of PS-20 or PS-80.

[0246] In an embodiment, the kit includes an SNE adjuvant, wherein the SNE, comprises 92 mole % squalene, 4 mole % SPAN-85 and 4 mole % of PS-20 or PS-80.

[0247] In an embodiment, the kit includes an SNE adjuvant, wherein the SNE, comprises 94 mole % squalene, 3 mole % SPAN-85 and 3 mole % of PS-20 or PS-80.

[0248] In an embodiment, the kit includes an SNE adjuvant, wherein the SNE, comprises 92.91 mole % squalene, 3.98 mole % SPAN-85 and 3.11 mole % of PS-20 or PS-80.

[0249] In an embodiment, the kit includes an SNE adjuvant, wherein the SNE, comprises 62 mole % squalene, 17 mole % SPAN-85 and 17 mole % of PS-20 or PS-80.

[0250] 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.

[0251] 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)26100(1) the pharmaceutical composition comprising the EBV polypeptide and (2) the SNE adjuvant. In some embodiments, the kits include instructions for admixing (1) the pharmaceutical composition comprising the EBV polypeptide and (2) the SNE 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

[0252] Also provided herein is a method of inducing an immune response to an Epstein Ban- 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 1 and a therapeutically effective amount of an SNE adjuvant described in the Squalene Nanoemulsion Adjuvant Section.

[0253] Also provided herein is a method of inducing an immune response to an Epstein Ban- 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 SNE adjuvant.

[0254] Also provided herein is a method of preventing infection of or reducing the likelihood of infection of a human patient to a 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 1 and a therapeutically effective amount of an SNE adjuvant described in the Squalene Nanoemulsion Adjuvant Section.

[0255] 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 SNE adjuvant.

[0256] In some embodiments, the SNE adjuvant is formulated separately from the EBV polypeptide. In some embodiments, the SNE adjuvant is formulated with the EBV polypeptide. In some embodiments, the SNE adjuvant and EBV polypeptide are field-mixed to form a pharmaceutical composition prior to administration to the patient. In some embodiments, the SNE adjuvant and EBV polypeptide are administered sequentially to a patient.

[0257] 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 the26100 compositions or pharmaceutical unit dose described in the Section Compositions of EBV polypeptide and SNE 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 an SNE adjuvant.

[0258] 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 1 and a therapeutically effective amount of an SNE adjuvant described in the Squalene Nanoemulsion Adjuvant Section, wherein the cancer is Burkit’s or Hodgkin’s lymphomas, or nasopharyngeal cancer.

[0259] 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 SNE adjuvant, wherein the cancer is Burkit’s or Hodgkin’s lymphomas, or nasopharyngeal cancer.

[0260] 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 Burkit’s lymphomas, (g) prevention or reduction of the likelihood of Hodgkin’s lymphomas, (h) prevention or reduction of the likelihood of nasopharyngeal cancer.

[0261] In embodiment 1, a composition comprises 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 a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate- 20 (PS-20) or polysorbate-80 (PS-80); and squalene.

[0262] In embodiment 2, the composition of embodiment! is provided, wherein the composition is made by mixing an EBV vaccine and a squalene nanoemulsion (SNE) adjuvant; wherein the EBV vaccine comprises the EBV polypeptide and a pharmaceutically acceptable26100 carrier and the squalene nanoemulsion (SNE) adjuvant comprises sorbitan trioleate (SPAN-85), polysorbate-20 (PS-20) or polysorbate-80 (PS-80), and squalene.

[0263] 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.

[0264] 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.

[0265] The following examples illustrate, but do not limit the invention.EXAMPLESExample 1: Preparation of a Squalene Nanoemulsion (SNE) Adjuvant System with and without the Cationic Lipid, (13Z.16Z)-N, N-dimethyl-3-nonyldocosa-13,16-dien-l-amine)

[0266] The SNE adjuvant can be prepared with and without cationic lipids, (13Z,16Z)-N,N- dimethyl-3-nonyldocosa-13,16-dien-l-amine) also referred to as CLA, or (6Z,9Z,26Z,29Z)-N,N- dimethylpentatriaconta-6,9,26.29-tetraen-18-amine, also referred to as CLX; or N,N-dimethyl-l- ((lS,2R)-2-octylcyclopropyl)heptadecan-8-amine, also referred to as CLY (Figure 1). The SNE adjuvant A (‘'Adjuvant A” or “SNE’’ hereafter) is a multi-component emulsion formulation consisting of 3 stabilizing ingredients; squalene, sorbitan trioleate (SPAN-85), and polysorbate- 20 (PS-20) (see Table 3); CLA-SNE is SNE Adjuvant A with a cationic lipid CLA (see Table 2). This formulation was prepared by combining and mixing the cationic lipid (if used), squalene, SPAN-85 and PS-20 or similar (e.g., surfactants, oils, and solubilizers) components together (Table 2 and Figure 2). Once mixed and blended, a histidine buffer was added and mixed with the initial emulsion components. Blended emulsion components were first subjected to course homogenization followed by fine homogenization, as described below. The resulting formulation was subjected to a final 0.2 mm filtration step. Several process parameters within each step, such as order of addition, mixing time, pH, temperature, concentration of components, homogenization, microfluidization were controlled to yield an emulsion system with desired attributes.Table 2: Composition of a Representative CLA-SNE Adjuvant26100Table 3: Composition of a Representative SNE Adjuvant AFormulation Preparation

[0267] The squalene and solubilizer formulation (referred to as the oil phase) of the emulsion was prepared by addition of squalene, SPAN-85, PS-20 and CLA to a vessel. The oil phase was then mixed using magnetic stirring at 100-1000 RPM for 10 to 120 minutes. After mixing of these components, an aqueous phase comprised of 20 mM Histidine pH 5.8 was slowly added to the oil phase while being mixed using a magnetic stir bar. This formulation was then mixed again for 1 hour.Coarse Homogenization

[0268] The oil and aqueous phase mixture (referred to as the pre-homogenized emulsion or PHE) was then homogenized and size reduced to form a rough emulsion using a rotor stator homogenizer at ambient temperature. The homogenizer arm tip was submerged into the PHE and held in place near the bottom of the formulation vessel and was operated at 6 to 10 kRPM for 5- 15 minutes. This process resulted in a homogenous micro-emulsion (ME) suspension of squalene emulsion particles in the 4 to 20 pm diameter range which were suitable for additional size26100 reduction by microfluidization in a high-pressure homogenizer to create a squalene nanoemulsion (SNE).Fine Homogenization to Produce the Squalene Nanoemulsion (SNE)

[0269] After coarse homogenization, the emulsion was further processed using a high-pressure homogenizer / microfluidizer to produce nanometer-sized emulsion particles. The ME was introduced to a high-pressure homogenizer such as the Microfluidics low volume Microfluidizer®, the GEA Group PandaPlus 2000 or Bee International. NanoDeBEE and a recirculation loop is established. A counter-flow heat exchanger, fed by a Controlled Temperature Unit with a set point of 5°C, is included in the recirculation loop to neutralize the heat generated through high pressure homogenization. For the production of emulsion particles of desired size and processability', 20 kPSI was selected as the operating set point for this process step. The high-pressure homogenizer operates at a constant and unalterable flow rate through the established recirculating loop. Using this measured flow rate and the volume of ME to be processed, the theoretical time required for the entirety of the formulation to make a single pass through the recirculation loop was calculated. Given this calculated single pass time, the high- pressure homogenizer was usually operated until the desired pass count of at least 10 was reached, yielding either the SNE or CLA-SNE.Filtration

[0270] After formulation, the SNE or CLA-SNE was passed through a 0.8 / 0.2 pm PES filter. A flux of 42 LMH through the filter was selected given its optimal mass yield and particle stability through filtration.

[0271] A laser diffraction or static light scattering (SLS) technique using a Malvern Panalytical Ltd. MS3000 instrument was utilized to measure the volume-weighed size distribution of a nanoemulsion during preparation. This data was then analyzed to calculate the size of the particles that created the scattering pattern. Sample fractions of pre-homogenized emulsion (PHE), micro-emulsion (ME), and squalene nanoemulsion (SNE) were obtained. These emulsion formulations were diluted to target an obscuration of 3% into 5 rnM Histidine pH 5.8 and 2.5 mM NaCl buffer and SLS was performed and collected under recirculation of 1200 RPM.Sample data sets were collected with a scan of 30 seconds per data set. Three data sets from each step in the CLA-SNE formulation process are summarized in Figure 3. Although 20 mM Histidine pH 5.8 buffer was a perfectly suitable formulation for the stability of the bulk during26100 process and when stored in polymeric containers (e.g., plastic), upon storage in glass, nonspecific absorption of the CLA-SNE or SNE to the surface of the glass was observed. A screen evaluating surfactants / solubilizers, buffers and salts was evaluated, and multiple formulations show success in eliminating this stability- issue with the selected formulation of 20 mM Histidine 0.05% PS-20 and 75 mM NaCl being selected as the stabilizing formulation (data not shown).Example 2: Preparation of a Squalene Nanoemulsion (SNE) Adjuvant System and Addition of the Cationic Lipid, (13Z,16Z)-N, N-dimethyl-3-nonyldocosa-13,16-dien-l-amine) or CLA as Free Base Directly After Microfluidization of the SNE

[0272] Two formulation processes were evaluated for incorporating CLA into the nanoemulsion particle which includes PS-20, sorbitan trioleate (SPAN-85) and squalene formulated in Histidine pH 5.8 buffer. In the first process (referred to as Process 1), the SNE was prepared using the process described in Example 1. In the second process, (referred to as Process 2), only PS-20, sorbitan trioleate (SPAN-85) and squalene were combined and mixed together. Once mixed and blended, a histidine buffer was added and mixed with the initial emulsion components (PS-20, sorbitan trioleate and squalene). Blended emulsion components were first subjected to course homogenization to produce the microemulsion ME followed by microfluidization to produce the nanoemulsion (NE), as described in Example 1. In a separate glass vessel, 0.25 mg / ruL CLA was dissolved in 100% ethanol at room temperature. A sufficient volume of this CLA ethanol solution, to produce the desired final CLA concentration, was then added to the SNE containing PS-20, sorbitan trioleate and squalene in Histidine buffer pH 5.8 and then mixed for 60 minutes at room temperature. After incubation, the formulation was then dialyzed against 5 mM histidine 2.5 mM NaCl pH 5.8 at 10 mL sample to 500 mL buffer over night at 4°C, with two buffer changes. The two processed emulsions (Process 1 and 2) were then examined for CLA incorporation into the SNE using UPLC-CAD. The results indicate that no significant quantity- of CLA was incorporated as compared to the individual formulation (w7w)% target incorporation using Process 2 which indicates that Process 1 is preferred for the successful incorporation and stability of CLA in the nanoemulsion (Figure 4).Example 3: Impact of Time and Temperature on Nanoemulsion Formulation Physical Stability Using NTA and DLS

[0273] As shown in Figures 5A-5D, to assess stability of the nanoemulsion systems (CLA-SNE or SNE), prepared as described in Examples, supra, nanoparticle tracking analysis (NTA) was26100 utilized. The technique collects videos of directly tracked nanoparticle populations as they move by Brownian motion to extrapolate particle size and concentration. A class 1, 635 nm laser focuses an 80 mm red laser beam through the liquid sample, illuminating particles as rapidly diffusing points of light. A CCD camera records a 30 frame per second video to track the movement of each individual illuminated particle over time. The system software identifies the center of each individual particle from the video and tracks the distance independently traversed to determine the mean square displacement. This tracking was performed simultaneously for every particle within the sample population in each frame until the raw data collected from the entire video was analyzed. By simultaneously measuring the mean square displacement of every individual particle tracked, its diffusion coefficient (Dt) and the spherical equivalent hydrodynamic radius (rh) were determined by applying the Stokes-Einstein equation. The software then represents this accumulated data as a particle size and concentration distribution. Raw data information on not only particle size and concentration, but also intensity, or brightness of the individual particle were gathered. Taken together the data were fitted and plotted individually as particle intensity relative to particle size, and particle concentration relative to particle size, and then on three-dimensional contour plots comparing particle size, concentration, and intensity of all particle populations.

[0274] Upon exposure of the nanoemulsion formulations (CLA-SNE or SNE) to 37°C for up to 1 month, no significant change in particle concentration or size distribution of the nanoemulsions was observed as evaluated using NTA (Figure 5).

[0275] A nanoemulsion may be susceptible to aggregation within the 10-1000 nm particle size range, thereby making DLS a suitable stabi 1 i ty indicating technique for assessing and quantitating aggregation phenomena. To assess stability of the nanoemulsion systems, prepared as described in examples, supra, dynamic light scattering (DLS) was utilized to measure the average particle size distribution. DLS instruments use a laser to illuminate particles in a solution and then examine the changes in intensity of the scattered light over time as a result of Brownian motion. The correlation of the scattered light intensity over time to the intensity at time zero results in an exponential decay curve or correlation function. The rate of decay in the correlation function with respect to time is much faster for smaller particles than larger particles and this forms the basis for calculation of the particle sizes. Upon exposure of the nanoemulsion to 4°C, 25°C or 37°C for up to 1 month, no change in the size distribution of the nanoemulsions was observed by DLS (Figures 6A-6D). The Z-av erage remained around 110 nm to 180 nm for the CLA-SNE or SNE.26100Example 4: Impact of Time and Temperature on Nanoemulsion Formulation Chemical Stability Using UPLC-CAD

[0276] To assess the chemical stability of the nanoemulsion systems, prepared as described in the Examples, supra, ultra-performance liquid chromatography coupled with a charge aerosol detector (UPLC-CAD) was utilized to measure the stability of the CLA (CLA-SNE only) and squalene concentration upon storage at 4°C, 25°C and 37°C for 1 month. Upon exposure of the nanoemulsion to 4°C, 25°C and 37°C for up to 1 month, the concentration of CLA (Figure 7A) or squalene (Figure 7B) in the SNE was not impacted, demonstrating excellent chemical stability in both CLA-SNE and SNE adjuvants.

[0277] Moreover, UPLC-CAD can quantitate the production of degradation products because of chemical breakdown of either squalene or CLA. Upon exposure of the SNE or CLA-SNE adjuvant systems to elevated temperature, no detectable degradation peaks were observed indicating that the squalene and CLA components of CLA-SNE and SNE adjuvant systems have excellent thermal stability (data not shown).Example 5: Preparation of a Squalene Nanoemulsion (SNE) Adjuvant System with and without the Cationic Lipid. (13Z.16Z)-N, N-dimethyl-3-nonyldocosa-13, 16-dien-l -amine) by Microfluidic Nanoemulsion Self-assembly (MNS)

[0278] The squalene nanoemulsion adjuvant is prepared with and without the ionizable cationic lipid (13Z,16Z)-N, N-dimethyl-3-nonyldocosa-13, 16-dien-l-amine), also referred to as CLA (Figure 1). The microfluidic nanoemulsion self-assembly (MNS) process used to prepare an SNE. The SNE is multi-component emulsion formulation consisting of 3 stabilizing ingredients; squalene, sorbitan trioleate (SPAN-85), and polysorbate-20 (PS-20) with CLA (referred to as CLA-SNE, Table 4) or without a CLA (referred to as SNE, Table 5). The biophysical characteristics (e.g., particle size, chemical compositions) and stability of MNS-prepared-SNE / CLA-SNE formulations are very similar to that of the high pressure fine homogenization process for preparation of SNE / CLA-SNE formulations described in Example 3. Essentially, the microfluidic nanoemulsion self-assembly (MNS) process described in this example is an alternative process for preparing a squalene nanoemulsion (SNE) adjuvant system. The nanoparticle self-assembly process described in this example was conducted using a ■‘microfluidics’’ ethanol / aqueous mixing instrument. However, the ethanol / aqueous stream nanoparticle self-assembly process described in this invention is not limited by “microfluidic”26100 mixing. Mixing larger volume streams of hydrophobic solvents with aqueous solutions can be accomplished using a Tee-mixing process outlined in Example 4.

[0279] SNE MNS formulations can generally be prepared by dissolving the cationic lipid, squalene, SPAN-85, and PS-20 at the targeted concentrations into an appropriate non-aqueous solvent such as ethanol. The self-assembly procedure involves combining a stream of the ethanol dissolved hydrophobic emulsion components with a stream of the aqueous emulsion solution. As the two solvent streams combine, the hydrophobic molecules (i.e., the cationic lipid, squalene, SPAN-85, and PS-20) interact with the aqueous solvent. The molecules then assemble themselves into an emulsion of nanosized particles, as described below. Following the formation of the self-assembled emulsion of nanoparticles, the residual ethanol can be removed from the squalene emulsion by several suitable means. In this example, the ethanol was reduced to less than 0.1% (w / v) by overnight dialysis with the aqueous buffer. The resulting SNE formulation was sterilized by fdtration through a 0.2 pm pore size sterilization fdter. Several process parameters within each step, such as order or addition, mixing times, temperature, concentration of non-aqueous components, concentrations of aqueous buffer components, aqueous pH, nonaqueous to aqueous solution mixing ratio, total flow rate, and waste discard volumes were controlled to yield SNE adjuvant systems with the desired attributes.Table 4 Composition of CLA-SNE Adjuvant Prepared by MNSTable 5: Composition of SNE Adjuvant Prepared by MNS26100Formulation Preparation using Microfluidic Nanoemulsion Self-assembly

[0280] In this example, one SNE and four CLA-SNE formulations were prepared by the microfluidic nanoemulsion self-assembly procedure in 20 mM histidine pH 5.8 for biophysical characterization. The self-assembled nanoemulsion process starts with 15 mg / mL squalene, 1.5 mg / mL SPAN-85 and 1.5 mg / mL PS-20 completely dissolved in ethanol. In addition, each of the ethanol solutions described above also contained CLA at either 0.75, 1.5, 5.0 or 15.0 mg CLA Z mL. Thus, the initial “target” CLA / squalene (w / w) % for all five formulations would be 0.0, 5.0. 10.0, 33.3. and 100 (w / w) % CLA / squalene. The aqueous buffer for all the formulations was 20 mM histidine at pH 5.8. A benchtop NanoAssemblr™ instrument from Precision NanoSystems, Inc. (Vancouver, BC, Canada) was used to self-assemble the one SNE and four CLA-SNE adjuvant nanoemulsions

[0281] The self-assembled squalene nanoparticle formulations were prepared in the following manner. A 1 mL syringe was filled with a little over 0.7 mL of the hydrophobic compound mixture dissolved in ethanol, while a 3 mL syringe was filled with a little over 1.4 mL of the aqueous 20 mM histidine pH 5.8 buffer. After both syringes were loaded with the appropriate amount of solution, the syringes were attached to the NanoAssemblr™ instrument. The following microfluidic mixing parameters were programed into the NanoAssemblr™: a) Total volume = 2 mL, b) Flow rate ratio =2: 1 (aqueous to ethanol), c) Total Flow Rate = 12 mL / min, d) Start waste volume = 0.25 mL, and e) End waste volume = 0.05 mL. The instrument w as activated to start the ethanol and aqueous solution mixing process in as little as a few seconds. Approximately 2.0 mL of post-mixing nanoparticle emulsion in approximately 30 % ethanol was collected in a 15 mL Falcon tube for each of the 5 formulations. A fresh NanoAssemblrTM mixing cartridge w as used for each of the 5 different SNE and CLA-SNE formulations described above. The ethanol concentration was reduced in each formulation by overnight dialysis. After dialysis, all of the samples were stored at 4 °C prior to analytical characterization.Analytical Characterization

[0282] The cationic lipid, CLA, and squalene were equally incorporated into the squalene CLA-SNE nanoparticles prepared by MNS as shown in Figure 8A. The CLA / squalene (w / w) %26100 ratio after dialysis (i.e., the y axis) to remove the process ethanol was compared to the CLA / squalene (w / w) % before self-assembly (i.e., the x axis) while in the ethanol solution for all the formulation samples described in this example. The “measured"’ CLA / squalene (w / w) % after MNS and dialysis was equal to the “target” (w / w) % up to at least 35 (w / w) %. Even at a 100 % “target” CLA / squalene (w / w) % prior to self-assembly over 70 % of the available CLA was incorporated into the CLA-SNE nanoparticles relative to the squalene content in the MNS prepared nanoparticle emulsion. The CLA / squalene (w / w) % ratios were measured by reverse phase UPLC-CAD. CLA was clearly incorporated into CLA-SNE by prepared by microfluidic nanoemulsion self-assembly (MNS) process.

[0283] The intensity weighted Z-average DLS diameters of the CLA-SNE formulations prepared the MNS process were measured using a Malvern ZetaSizer Ultra. Aliquots of postdialyzed CLA-SNE samples from each formulation were diluted at either 50- or 100-fold in 2.0 rnL 20 mM histidine pH 5.8 buffer. Average DLS diameter and standard deviation was plotted versus the measured post-dialysis CLA / squalene (w / w) % for each formulation and is shown in Figure 8B. Three DLS measurements were made at room temperature for each formulation. The standard deviation bars are show unless the standard deviation is less that data point image. The intensity weight Z-average DLS diameters of MNS prepared CLA-SNE ranged from approximately 150 to 280 nm which is similar to CLA-SNE nanoparticles prepared by high- pressure homogenization. Varying MNS process parameters such as those described above in this example were controlled to yield CLA-SNE adjuvant systems with the desired diameters.

[0284] The measured Zeta Potential of CLA-SNE squalene nanoparticle formulations at pH 5.5 prepared the MNS process are shown in Figure 8C. The Zeta Potential was measured using a Malvern ZetaSizer Ultra. Aliquots of post-dialyzed CLA-SNE samples from each formulation were diluted at either 50 or 100 X in 2.0 mL of 20 mM citrate BIS TRIS propane buffer at pH 5.5. Three Zeta potential measurements were made at room temperature for each formulation. The standard deviation bars are show unless the standard deviation is less that data point image. The Zeta Potential of the 0 (w / w) % CLA CLA-SNE formulation, i.e., no CLA, was around- 5 mV. As illustrated in Figure 8C, addition of CLA significantly increased the nanoparticle Zeta Potential to around +10 mV.Example 6: Optimization of CLA-SNE Preparation by Alteration of the Aqueous Phase pH.

[0285] The CLA-SNE process involves the use of a reversible cationic CLA molecule with an observed pKa of 6.4. Addition of CLA to the SNE preparation process and final matrix with a pH26100 of 5.8 results in the protonation of CLA, which functions to give an overall net positive charge to CLA-SNE particles as well as any intermediates of the preparation process. CLA-SNE preparation culminates in a 0.8 / 0.2 pm filtration event, a process step which had proved difficult to perform, with significant filter fouling and low product yield consistently obser ed. However, the filtration of SNE did not demonstrate the same magnitude of these filtration challenges and was a more efficient nanoemulsion filtration step. Importantly, without CLA present SNE does not carry the strong positive charge observed with CLA-SNE. In an effort to prepare an uncharged CLA-SNE for a higher efficiency CLA-SNE filtration step, a series of experiments were performed in which the pH of the aqueous phase (20 mM L-Histidine) was adjusted prior to use in the preparation of CLA-SNE. 20 mM L-Histidine was prepared with pH targets of 5.0, 5.7, 5.8, 6.0, 6.2, 7.0, and 7.7. Each buffer was then used as the aqueous phase during the CLA-SNE preparation process with a target formulation target of 15 mg / mL CLA, CLA-SNE preparation proceeded exactly as described in Example 3. Upon completion of homogenization process step, the particle size of the CLA-SNE intermediate was measured by DLS using a Malvern Panalytical Nano ZS. Filtration with 0.8 / 0.2pm PES filter was then performed. Particle size distribution was measured post-filtration by DLS and [CLA] was quantified by UPLC-CAD. For CLA-SNE samples prepared with 20 mM L-histidine with pH 7.0 or 7.7, complete filter fouling was observed immediately upon application of material to the filter and no collection of filtered material was possible making quantification by DLS or UPLC-CAD impossible, values of 0 are reported for illustration purposes. In pre-filtered samples, a trend w as observed of increasing particle size by DLS as the pH of the aqueous phase buffer was increased (Figure 9). This relationship was maintained in post-filter samples with each CLA-SNE sample demonstrating a modest reduction in particle size post-filtration, with the obvious exception of pH 7.0 and 7.7 samples which again demonstrated complete filter fouling immediately and no material recovery was possible. For post-filtered samples, it was observed that [CLA] decreases in the final CLA- SNE material as the pH of the aqueous phase is increased (Figure 10). This relationship demonstrates that CLA-SNE preparation using an aqueous phase at a lower pH results in an increase in process yield in the terminal filtration and a more efficient manufacturing process.Example 7: EBV Antigen-Specific Antibody Titers in 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 SNE adjuvant A26100

[0286] Preclinical studies were performed in mice to evaluate the immunogenicity of gp350- FNP (SEQ ID NO: 23) and gL-gH-gp42-FNP (SEQ ID NO: 21) dual particle vaccine. Immunogenicity was assessed by measuring serum antibody titers against gp350, gH / gL, and gp42 individually by ELISA.

[0287] 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 SNE adjuvant A 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 SNE adjuvant A 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.

[0288] 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.

[0289] 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)] .

[0290] The cut-off value was designated as 50000. 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 titration26100 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.

[0291] 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 11-13. SNE Adjuvant A group showed the largest titer increase compared to the nonadjuvanted formulation.Example 8: EBV-Neutralizing Antibody Titers in 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 SNE adjuvant A

[0292] Preclinical studies were performed in mice to evaluate the immunogenicity of gp350- FNP (SEQ ID NO: 23) and gL-gH-gp42-FNP (SEQ ID NO: 21) dual particle vaccine.Immunogenicity was assessed by measuring EBVneutralizing serum antibody titers in a neutralization assay using EBV-GFP, B cells (4E3). and epitheliallike cells (293T).

[0293] 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 SNE adjuvant A at days 0 and 28. Sera were collected at day 42 (2 w eeks 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.

[0294] Human B lymphoma cells (4E3) and human embryonic kidney cells (293T) cells were seeded in a 96-well black wall transparent plate at 2.5 x io6cells and 2.0 * 106cells, 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 1 1 concentrations in assay medium in a separate U-bottomed 96-w ell sample plate. Concentrated cell-free Akata EBV-GFP at 1.5 x 105ffu / rnL in assay medium w as added to each w ell 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 w as 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,26100Covina, 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).

[0295] 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 SNE adjuvant A producing the highest titers. See Figure 14.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 SNE adjuvant A.Example 9: 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 SNE adjuvant A.

[0296] The immunogenicity of the dual particle vaccine formulated with SNE adjuvant A was further assessed in rhesus monkeys. Animals (5 per group) were immunized intramuscularly with 15 pg gp350-FNP and 1 pg gL-gH-gp42-FNP formulated with 10 mg adjuvanted at weeks 0, 4, and 10. Sera were collected at 2-week intervals from week 0 (prior to study start) to week 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 7.

[0297] 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 1st dose 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 1st dose for both vaccines. See Figure 15.26100Example 10: EBV-Neutrahzing Antibody Titers in Rhesus Monkeys Immunized With 3 Doses of gp350-FNP and gL-gH-gp42-FNP dual particle vaccine formulated with SNE adjuvant A

[0298] Rhesus monkeys (5 per group) were injected intramuscularly with 3 doses of gp350- FNP (SEQ ID NO: 23) and gL-gH-gp42-FNP (SEQ ID NO: 21) dual particle vaccine formulated with SNE adjuvant A 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 SNE adjuvant A boosted EBVneutralizing antibody titers in a neutralization assay using EBV-GFP, B cells (4E3), and epitheliallike cells (293T) according to Example 8. Neutralization titers remained above basal titers (week 0) up to 42 weeks after the 1st dose. See Figure 16.Example 11: 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 SNE adjuvant A.

[0299] 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.

[0300] 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. Tw enty (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 w ere performed at 2 to 8 °C with cell washes by centrifugation of plates at 500 xg for 5 minutes, unless otherwise indicated.

[0301] 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, and26100CD3 / 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 / Cytoperm solution for 20 to 25 minutes. 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.

[0302] 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 cellmedia

[0303] Negative values are imputed as 0 (no response), and values above 0.1% (baseline) are considered as true antigen specific response.

[0304] CD4 responses (IFN-y, IL-2, and TNF-a) to gH peptide stimulation were observed in some animals that received the vaccine (Figures 17-19). No CD4 response to other antigens was detected (Figures 20-22), and no CD8 response to any antigen was detected (data not shown). Collectively, the data demonstrated that gp350-FNP and gL-gH-gp42-FNP dual particle vaccine formulated with SNE adjuvant A was immunogenic in Rhesus monkeys, and provided boost to both humoral and cellular responses.Example 12: A Phase 1, Randomized, Double-Blind, Placebo-Controlled, Dose Escalation Trial to Evaluate the Safety, Tolerability, and Immunogenicity of EBV polypeptide and SNE adjuvant A in Healthy Participants

[0305] 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 SNE adjuvant A (EBV- SNE) in healthy young adult participants (>18 to 30 years of age).

[0306] Part 1 will evaluate progressively higher antigen and / or adjuvant doses of each EBV polypeptide and SNE adjuvant A 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 lower 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 the26100 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 dose ranging 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 safety7review 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 12 mg of SNE Adjuvant A with an option escalate to 18 mg of SNE Adjuvant A.

[0307] Each EBV-SNE 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

[0308] Part 2 will be implemented if 1) safety and tolerability of EBV-SNE from Part 1 are supportive of further study in seronegative participants and 2) the prevalence of seronegative individuals at the clinical sites is high enough to enable enrollment. Therefore, Part 2 is included as flexible to explore the safety, tolerability, and immunogenicity of EBV-SNE in a seronegative population and will be based on the feasibility of enrolling seronegative participants. Up to 4 panels for EBV-SNE 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

[0309] Part 3 is included to further explore the safety7, tolerability, and immunogenicity of EBV-SNE in a seropositive population. Up to 2 additional panels for EBV-SNE 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 the26100 experimental vaccine or placebo. Part 3 will be implemented if safety and tolerability of EBV- SNE from Part 1 are supportive of further study in seropositive participants.Table 6 Sample Allocation TableFNP=ferritin nanoparticle a Within each panel, participants will be randomized to receive EBV-SNE or matching placebo in a 4: 1 ratio (Part 1 and Part 3) according to a computer-generated, permuted-block allocation schedule. b The 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)26100Table 7: Intervention Groups and Duration:26100Table 8

[0310] 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 SNE adjuvant A. Clear immunogenicity boosts were observed in seropositive participants.

Claims

1. WHAT IS CLAIMED IS:

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 squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80); and squalene.

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 gHpolypeptide 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 ID NOs: 11-21; or the amino acid sequence of 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: 21.2610026. The composition of claim 25, wherein the EBV polypeptide comprises a sequence of any one of SEQ ID NOs: 11-21.

27. 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 squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN- 85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80); and squalene.

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 23.2610035. The composition of any one of claims 1-34, wherein the composition is a solution of the EBV polypeptide and the SNE adjuvant.

36. 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 2 mg to about 30 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80); and about 2 mg to about 25 mg of squalene.

37. The pharmaceutical unit dose of claim 36, 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 6 mg to about 18 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS- 80); and about 5 mg to about 15 mg of squalene.

38. The composition or pharmaceutical unit dose of any one of claims 1-37, wherein the ratio of Squalene: SPAN-85 :polysorbate 20 or 80 is 10-8: 1 : 1 by weight.

39. The composition or pharmaceutical unit dose of any one of claims 1-37, wherein the ratio of Squalene: SPAN-85 :polysorbate 20 is 10: 1 : 1 by weight.

40. The pharmaceutical unit dose of claim 36, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 10 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 6 mg; wherein the ratio of Squalene: SPAN- 85 :PS-20 or PS-80 is 10:1 :1 by weight.

41. The pharmaceutical unit dose of claim 36, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 30 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 6 mg; wherein the ratio of Squalene: SPAN- 85: PS-20 or PS-80 is 10: 1 : 1 by weight.

42. The pharmaceutical unit dose of claim 36, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 100 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 6 mg; wherein the ratio of Squalene: SPAN- 85 : PS-20 or PS-80 is 10: 1 : 1 by weight.

43. The pharmaceutical unit dose of claim 36, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 10 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 12 mg; wherein the ratio of Squalene: SPAN- 85: PS20 or PS-80 is 10:1 : 1 by weight.

44. The pharmaceutical unit dose of claim 36, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 30 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 12 mg; wherein the ratio of Squalene: SPAN- 85: PS-20 or PS-80 is 10: 1 : 1 by weight.

45. The pharmaceutical unit dose of claim 36, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 100 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 12 mg; wherein the ratio of Squalene: SPAN- 85 : PS-20 or PS-80 is 10: 1 : 1 by weight.

46. The pharmaceutical unit dose of claim 36, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 10 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 18 mg; wherein the ratio of Squalene: SPAN- 85 : PS-20 or PS-80 is 10: 1 : 1 by weight.

47. The pharmaceutical unit dose of claim 36, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 30 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 18 mg; wherein the ratio of Squalene: SPAN- 85: PS-20 or PS-80 is 10: 1 : 1 by weight.

48. The pharmaceutical unit dose of claim 36, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 21 in the amount of 100 pg, and the squalenenanoemulsion (SNE) adjuvant is in the amount of 18 mg; wherein the ratio of Squalene: SPAN- 85 : PS-20 or PS-80 is 10: 1 : 1 by weight.

49. 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 2 mg to about 30 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80); and about 2 mg to about 25 mg of squalene.

50. The pharmaceutical unit dose of claim 49, 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 6 mg to about 18 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS- 80); and about 5 mg to about 15 mg of squalene.

51. The pharmaceutical unit dose of any one of claims 49-50, wherein the ratio of Squalene: SPAN-85 :polysorbate 20 or 80 is 10-8: 1 : 1 by weight.

52. The pharmaceutical unit dose of any one of claims 49-50, wherein the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10: 1: 1: by weight.

53. The pharmaceutical unit dose of claim 49, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 10 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 6 mg; wherein the ratio of Squalene: SPAN- 85: PS-20 or PS-80 is 10: 1 : 1 by weight.

54. The pharmaceutical unit dose of claim 50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 30 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 6 mg; wherein the ratio of Squalene: SPAN- 85: PS-20 or PS-80 is 10: 1: 1 by weight.

55. The pharmaceutical unit dose of claim 50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 100 pg, and the squalene- 100 -26100 nanoemulsion (SNE) adjuvant is in the amount of 6 mg; wherein the ratio of Squalene: SPAN- 85 : PS-20 or PS-80 is 10: 1 : 1 by weight.

56. The pharmaceutical unit dose of claim 50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 10 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 12 mg; wherein the ratio of Squalene: SPAN- 85: PS-20 or PS-80 is 10: 1 : 1 by weight.

57. The pharmaceutical unit dose of claim 50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 30 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 12 mg; wherein the ratio of Squalene: SPAN- 85: PS-20 or PS-80 is 10: 1: 1 by weight.

58. The pharmaceutical unit dose of claim 50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 100 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 12 mg; wherein the ratio of Squalene: SPAN- 85: PS-20 or PS-80 is 10: 1: 1 by weight.

59. The pharmaceutical unit dose of claim 50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 10 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 18 mg; wherein the ratio of Squalene: SPAN- 85: PS-20 or PS-80 is 10: 1: 1 by weight.

60. The pharmaceutical unit dose of claim 50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 30 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 18 mg; wherein the ratio of Squalene: SPAN- 85 : PS-20 or PS-80 is 10: 1 : 1 by weight.

61. The pharmaceutical unit dose of claim 50, wherein the EBV polypeptide consists of the amino acid sequence of SEQ ID NO: 23 in the amount of 100 pg, and the squalene nanoemulsion (SNE) adjuvant is in the amount of 18 mg; wherein the ratio of Squalene: SPAN- 85 : PS-20 or PS-80 is 10: 1 : 1 by weight.2610062. 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 6 mg to about 36 mg of a squalene nanoemulsion (SNE) adjuvant, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) or polysorbate-80 (PS-80); and squalene; wherein the ratio of Squalene: SPAN-85: PS-20 or PS-80 is 10: 1 : 1 by weight.

63. The pharmaceutical unit dose of claim 62, wherein each of the first and second EBV polypeptide is at 10 pg.

64. The pharmaceutical unit dose of claim 62, wherein each of the first and second EBV polypeptide is at 30 pg.

65. The pharmaceutical unit dose of claim 62, wherein each of the first and second EBV polypeptide is at 100 pg.

66. The composition or pharmaceutical unit dose of any one of claims 1-65, wherein the SNE adjuvant further comprises a cationic lipid.

67. The composition or pharmaceutical unit dose of claim 66, wherein the cationic lipid is (13Z, 16Z)-N,N-dimethy 1-3-nonyldocosa- 13,16-dien-l -amine.

68. The composition or pharmaceutical unit dose of any one of claims 1-67, wherein the SNE adjuvant comprises sorbitan trioleate (SPAN-85); polysorbate-20 (PS-20) and squalene.

69. The composition or pharmaceutical unit dose of any one of claims 1-68, wherein the Z-average of the SNE adjuvant is about 50 to about 200 nm.

70. The composition of any one of claims 1-69 wherein the SNE adjuvant further comprises a buffer.

71. The composition or pharmaceutical unit dose of claim 70. wherein the buffer is selected from the group consisting of: acetic acid, histidine, citrate, Bis-Tris, HEPES, phosphate, MES, sodium chloride, succinate, Tris, and combinations thereof.

72. The composition or pharmaceutical unit dose of any one of claims 70 and 71. wherein the buffer is present in the amount of about ImM to about lOOmM.

73. The composition or pharmaceutical unit dose of any one of claims 1-72, wherein the SNE adjuvant further comprises a salt.

74. The composition or pharmaceutical unit dose of claim 73, w herein the salt is NaCl.

75. The composition or pharmaceutical unit dose of claim 74, wherein the SNE adjuvant comprises 5 mM - 40 mM histidine at pEI 5.1 - 7.0 and 25 mM - 300 mM NaCl.

76. The composition or pharmaceutical unit dose of claim 74, wherein the SNE adjuvant comprises about 20 mM histidine at about pH 5.8 and about 75 mM NaCl.

77. The composition or pharmaceutical unit dose of any one of claims 1-76, wherein the SNE adjuvant comprises about 4 mg / ml sorbitan trioleate (SPAN-85); about 0.05% polysorbate-20 (PS-20); about 40 mg / ml squalene, about 20 mM histidine at about pH 5.8 and about 75 mM NaCl.78 The composition or pharmaceutical unit dose of any one of claims 1-77, w herein the SNE adjuvant comprises emulsion particles in the range of about 10 to about 1000 nm.

79. The composition or pharmaceutical unit dose of any one of claims 1-78 wherein the Z-average of the SNE adjuvant is about 100 to about 300 nm.

80. 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 1-79.

81. 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 1-79.

82. 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 1-79.

83. The method of claim 82, wherein the cancer is Burkitt’s or Hodgkin’s lymphomas, or nasopharyngeal cancer.

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