Vaccine composition comprising recombinant nipah virus protein

Abstract

The present invention relates to a vaccine composition comprising a recombinant Nipah virus protein. The vaccine composition according to the present invention not only exhibits excellent immunogenicity against both the G protein and the F protein of Nipah virus, but also exhibits immunogenicity against the G protein and the F protein of Hendra virus, and thus can be used as a universal vaccine against Henipavirus. In addition, the vaccine composition has the characteristic of inducing cell-mediated immune responses, and therefore can be provided as a formulation for enhancing specific immune responses to an administered drug such as an adjuvant or a vaccine.

Description

Vaccine composition containing Nipah virus recombinant protein

[0001] The present invention relates to a vaccine composition comprising a Nipah virus recombinant protein.

[0002] Nipahvirus and Hendravirus are two zoonotic pathogens that originated in the Asia-Pacific region. Both are RNA viruses belonging to the paramyxoviruses family and classified under the genus Henipavirus. They share antigenic, serological, and microstructural features and are distinguished from other paramyxoviruses.

[0003] Both Nipah and Hendra viruses express G and F proteins anchored to the viral membrane, which are essential for fusion with host cells. The G protein is a type 2 membrane protein that does not possess erythrocyte agglutination or neuraminidase activity; as an adhesion glycoprotein, it has an amino (N) terminus facing the cytoplasm and a carboxyl (C) terminus facing outward. The F protein is a type 1 membrane protein in which the molecule is a trimer containing two heptad repeat (HR) regions and a single hydrophobic fusion peptide. After binding to cell receptors, Nipah and Hendra viruses infect cells through a pH-dependent membrane fusion process facilitated by the action of the G and F proteins. As such, surface structural proteins essential for the development of vaccines for henipah viruses, such as Nipah and Hendra viruses, are known to be Attachment glycoprotein (G) and Fusion glycoprotein (F), which are fixed to the viral membrane and are essential for fusion with host cells; however, there are currently no effective vaccines or treatments for the prevention and treatment of Nipah or Hendra virus infections. Therefore, the development of serological diagnostic methods and vaccines is required.

[0004] The present invention aims to provide a vaccine composition for preventing or treating henipharon infection.

[0005] The present invention aims to provide an immune-enhancing composition.

[0006] The present invention aims to provide a composition for diagnosing henipharon infection or antibody possession.

[0007] The present invention aims to provide a method for preventing or treating henipharon infection.

[0008] 1. A vaccine composition for the prevention or treatment of heniphavirus infection, comprising as an active ingredient a recombinant protein comprising at least one of the polypeptide represented by SEQ ID NO. 6 and the polypeptide represented by SEQ ID NO. 8.

[0009] 2. A vaccine composition for the prevention or treatment of henpavirus infection, wherein the henpavirus in 1 above is a Nipah virus or a Hendra virus.

[0010] 3. A vaccine composition for the prevention or treatment of henipahvirus infection, wherein the polypeptide represented by SEQ ID NO. 6 is derived from the G protein of the Bangladesh Nipahvirus strain and the Malaysia Nipahvirus strain.

[0011] 4. A vaccine composition for the prevention or treatment of henipharvirus infection, wherein, in the above 1, the recombinant protein comprising the polypeptide represented by SEQ ID NO. 6 is encoded by the polynucleotide represented by SEQ ID NO. 5.

[0012] 5. A vaccine composition for the prevention or treatment of henipahvirus infection, wherein the polypeptide represented by SEQ ID NO 8 is derived from the F protein of the Bangladesh Nipahvirus strain and the Malaysia Nipahvirus strain.

[0013] 6. A vaccine composition for the prevention or treatment of henipahvirus infection, wherein, in the above 1, the recombinant protein comprising the polypeptide represented by SEQ ID NO. 8 is encoded by the polynucleotide represented by SEQ ID NO. 7.

[0014] 7. A vaccine composition for the prevention or treatment of henipahvirus infection, wherein the above 1 further comprises an adjuvant.

[0015] 8. A vaccine composition for the prevention or treatment of henipharon infection, wherein the immunoadjuvant of 1 above is at least one selected from the group consisting of Alum, Adju-Phos, MPLA-SM VacciGrade, AddaS03, and AddaVax.

[0016] 9. A vaccine composition for the prevention or treatment of henipahvirus infection, wherein, in addition to the above 1, the recombinant protein has activity inducing a cell-mediated immune response.

[0017] 10. An immune-enhancing composition comprising, as an active ingredient, a recombinant protein comprising at least one of the polypeptide represented by SEQ ID NO. 6 and the polypeptide represented by SEQ ID NO. 8.

[0018] 11. A composition for diagnosing heniphavirus infection or antibody possession, comprising at least one of the polypeptide represented by SEQ ID NO. 6 and the polypeptide represented by SEQ ID NO. 8 as an active ingredient.

[0019] 12. A composition for diagnosing henpavirus infection or antibody possession, wherein the henpavirus in 11 above is a Nipah virus or a Hendra virus.

[0020] 13. A method for preventing or treating heniphavirus infection, comprising the step of administering a recombinant protein, excluding humans, to an animal comprising at least one of the polypeptide represented by SEQ ID NO. 6 and the polypeptide represented by SEQ ID NO. 8.

[0021] The vaccine composition according to the present invention has excellent immunogenicity against the G protein and F protein of Nipah virus.

[0022] The vaccine composition according to the present invention has excellent immunogenicity against the G protein and F protein of Hendra virus as well as Nipah virus.

[0023] The vaccine composition according to the present invention has an excellent effect of inducing a cell-mediated immune response.

[0024] Figure 1 is a figure showing the design of the Nipah virus recombinant G protein antigen.

[0025] Figure 2 is a figure showing the amino acid sequence of the Nipah virus recombinant G protein.

[0026] Figure 3 is a figure showing the design of the Nipah virus recombinant F protein antigen.

[0027] Figure 4 is a figure showing the amino acid sequence of the Nipah virus recombinant F protein.

[0028] Figure 5 shows the SDS-PAGE and Western blot results confirming the expression of Nipah virus recombinant G protein.

[0029] Figure 6 shows the SDS-PAGE and Western blot results confirming the expression of Nipah virus recombinant F protein.

[0030] Figure 7 is a schematic diagram of a mouse experiment to evaluate the immunogenicity of Nipah virus recombinant protein.

[0031] Figure 8 shows the antibody titers against Nipah Virus G protein according to the inoculation dose of the recombinant protein in mouse serum. ELISA coated antigen: Nipah Virus Glycoprotein G (Human Fc-Tag (HEK293), REC31633, Native Antigen).

[0032] Figure 9 shows the antibody titers against Nipah Virus F protein according to the inoculation dose of the recombinant protein in mouse serum. ELISA coated antigen: Nipah Virus F protein (Nipah Virus Glycoprotein F, Human Fc-Tag, REC31637, Native Antigen).

[0033] Figure 10 shows the antibody titers against Hendra virus G protein according to the inoculation dose of the recombinant protein in mouse serum. ELISA coated antigen: Hendra virus G protein (Hendra virus Glycoprotein, His Tag (MALS verified), GLN-H52H3, Acro).

[0034] Figure 11 shows the antibody titers against Hendra virus F protein according to the inoculation dose of the recombinant protein in mouse serum. ELISA coated antigen: Hendra virus F protein (Hendra virus Glycoprotein / GP Protein (HEK293, Fc), FUN-H52H4, Acro).

[0035] Figure 12 shows the neutralizing antibody titer in mouse serum treated with a recombinant VSV vector-based Nifas Pseudovirus. It was confirmed in mouse serum 3 weeks after the first inoculation with the recombinant protein.

[0036] Figure 13 shows the neutralizing antibody titers in mouse serum treated with a recombinant VSV vector-based Nifas pseudovirus. These titers were confirmed in mouse serum 3 weeks after the second inoculation with the recombinant protein.

[0037] Figure 14 shows the pooling results of the neutralizing ability in mouse serum treated with a recombinant VSV vector-based Nifas Pseudovirus by inoculation test group. The results were confirmed in mouse serum 3 weeks after the first inoculation of the recombinant protein.

[0038] Figure 15 shows the pooling results of the neutralizing ability in mouse serum treated with a recombinant VSV vector-based Nifas Pseudovirus by inoculation test group. The results were confirmed in mouse serum 3 weeks after the second inoculation of the recombinant protein.

[0039] Figure 16 is a schematic diagram of a hamster experiment to evaluate the immunogenicity of Nipah virus recombinant protein.

[0040] Figure 17 shows the antibody titers against Nipah Virus G protein in hamster serum according to recombinant protein and adjuvant. ELISA coated antigen: Nipah Virus G protein (Nipah Virus Glycoprotein G, Human Fc-Tag (HEK293), REC31633, Native Antigen).

[0041] Figure 18 shows the antibody titers against Nipah Virus F protein in hamster serum according to recombinant protein and adjuvant. ELISA coated antigen: Nipah Virus F protein (Nipah Virus Glycoprotein F, Human Fc-Tag, REC31637, Native Antigen).

[0042] Figure 19 shows the antibody titers against Hendra virus G protein in hamster serum according to recombinant protein and adjuvant. ELISA coated antigen: Hendra virus G protein (Hendra virus Glycoprotein, His Tag (MALS verified), GLN-H52H3, Acro).

[0043] Figure 20 shows the antibody titers against Hendra virus F protein in hamster serum according to recombinant protein and adjuvant. ELISA coated antigen: Hendra virus F protein (Hendra virus Glycoprotein / GP Protein (HEK293, Fc), FUN-H52H4, Acro).

[0044] Figures 21a and 21b show the neutralizing antibody titers in hamster serum treated with a recombinant VSV vector-based Nifas pseudodovirus. These titers were confirmed in mouse serum 3 weeks after the first inoculation with the recombinant protein (*p<0.05).

[0045] Figures 22a-22b show the neutralizing antibody titers in hamster serum treated with a recombinant VSV vector-based Nifas pseudodovirus. These titers were confirmed in mouse serum 3 weeks after the second inoculation with the recombinant protein (*p<0.05).

[0046] Figure 23 shows the pooling results of neutralizing ability by inoculation test group in hamster serum treated with recombinant VSV vector-based Nipash pseudovirus 3 weeks after the first inoculation and 3 weeks after the second inoculation.

[0047] Figures 24a-24b are the ELISpot results confirming INF-γ cytokine expression by recombinant G protein in hamster splenocytes.

[0048] Figures 25a-25b are the ELISpot results confirming INF-γ cytokine expression by recombinant F protein in hamster splenocytes.

[0049] The present invention relates to a universal hendravirus recombinant protein for the serological diagnosis of Nipah virus or Hendravirus infection, prevention of infection, treatment, and vaccine development.

[0050] In one aspect, the present invention provides a vaccine composition for the prevention or treatment of heniphavirus infection, comprising as an active ingredient a recombinant protein comprising at least one of the polypeptide represented by SEQ ID NO. 6 and the polypeptide represented by SEQ ID NO. 8.

[0051] In the present invention, the henpah virus may be a Nipah virus or a Hendra virus.

[0052] Both Hendra and Nipah viruses express G and F proteins anchored to the viral membrane, which are essential for fusion with host cells. The G protein is a type 2 membrane protein that does not possess erythrocyte agglutination or neuraminidase activity; as an adhesion glycoprotein, it has an amino (N) terminus facing the cytoplasm and a carboxyl (C) terminus facing outward. The F protein is a type 1 membrane protein in which the molecule is a trimer containing two heptad repeat (HR) regions and a single hydrophobic fusion peptide. After binding to cell receptors, Hendra and Nipah viruses infect cells through a pH-dependent membrane fusion process facilitated by the action of the G and F proteins.

[0053] The recombinant protein of the present invention was produced by extracting representative reference sequences for each cluster from the sequences encoding the F protein and the sequence encoding the G protein of Bangladesh Nipah virus strains (BD1, BD2, India) and Malaysia Nipah virus strains (MY) for the development of a universal vaccine, and by obtaining representative consensus sequences of the F protein and the G protein covering each reference.

[0054] In the present invention, the polypeptide represented by SEQ ID NO. 6 may be derived from the G protein of the Bangladesh Nipah virus strain and the Malaysia Nipah virus strain. More specifically, it may be a consensus sequence capable of covering the entire sequence of the G protein of the Bangladesh Nipah virus strain and the Malaysia Nipah virus strain. Preferably, it may be the amino acid sequence represented by SEQ ID NO. 6 or a functional equivalent thereof.

[0055] In the present invention, the polypeptide represented by SEQ ID NO. 8 may be derived from the F protein of the Bangladeshi Nipah virus strain and the Malaysian Nipah virus strain. More specifically, it may be a consensus sequence capable of covering the entire sequence of the F protein of the Bangladeshi Nipah virus strain and the F protein of the Malaysian Nipah virus strain. Preferably, it may be an amino acid sequence represented by SEQ ID NO. 8 or a functional equivalent thereof.

[0056] In the present invention, 'origin' means that it occurs due to a source. The source may be a producer or starting point of a substance or phenomenon. Derivation of the G protein or F protein of the Nipah virus strain may mean that it is generated or made from the G protein or F protein contained in the Nipah virus strain. Derivation of the G protein or F protein of the Hendra virus strain may mean that it is generated or made from the G protein or F protein contained in the Hendra virus strain.

[0057] In the present invention, the 'consensus sequence' is also referred to as the canonical sequence and means a generalized DNA, RNA, or protein sequence representing the nucleotide or amino acid most frequently found at each position of an aligned sequence group. This reveals conserved regions important for gene regulation (e.g., splice sites, transcription factor binding, etc.) or the function of protein structure.

[0058] In the present invention, 'functional equivalent' means having at least 80%, or 85%, or 90%, or 95%, or 97%, or 99% sequence homology with the amino acid sequence represented by SEQ ID NO. 6 or SEQ ID NO. 8 as a result of the addition, substitution, or deletion of amino acids, and exhibiting substantially the same physiological activity as the polypeptide represented by SEQ ID NO. 6 or the polypeptide represented by SEQ ID NO. 8.

[0059] In the present invention, 'recombinant protein' may be used interchangeably with 'recombinant G protein', 'recombinant F protein', or 'recombinant G protein and recombinant F protein'.

[0060] The recombinant protein of the present invention may include a polypeptide represented by SEQ ID NO. 6. The recombinant protein including the polypeptide represented by SEQ ID NO. 6 may be a recombinant G protein.

[0061] The recombinant protein of the present invention may include a polypeptide represented by SEQ ID NO. 8. The recombinant protein including the polypeptide represented by SEQ ID NO. 8 may be a recombinant F protein.

[0062] The recombinant protein of the present invention may include both the polypeptide represented by SEQ ID NO. 6 and the polypeptide represented by SEQ ID NO. 8. The recombinant protein may be a recombinant G protein and a recombinant F protein.

[0063] In the present invention, the recombinant protein may be produced by any suitable procedure known to those skilled in the art, namely, genetic engineering methods, such as recombinant techniques. Additionally, the polypeptide may additionally include a targeting sequence, a signal sequence, a tag, or an amino acid sequence prepared for a specific purpose for a labeled residue, according to conventional methods. The additional sequence may be included at the N-terminus, C-terminus, or both ends of the polypeptide or its functional equivalent as needed.

[0064] In addition, to obtain better chemical stability, enhanced pharmacological properties (half-life, absorption, potency, efficacy, etc.), modified specificity (e.g., broad spectrum of biological activity), and reduced antigenicity, a protecting group may be attached to the N-terminus or C-terminus of the polypeptide. The protecting group may be, for example, an acetyl group, a fluorenyl methoxycarbonyl group, a formyl group, a palmitoyl group, a myristyl group, a stearyl group, or polyethylene glycol (PEG), but may be included without limitation as long as it is a substance capable of modifying the protein, particularly enhancing the stability of the protein.

[0065] In one embodiment, the recombinant protein may include the signal peptide of SEQ ID NO. 9. Specifically, the signal peptide of SEQ ID NO. 9 may be bound to the N-terminus of the polypeptide represented by SEQ ID NO. 6 or the polypeptide represented by SEQ ID NO. 8.

[0066] In one embodiment, when the recombinant protein includes a polypeptide represented by SEQ ID NO. 8, a GCNt trimerization motif may be attached to the C-terminus of the polypeptide for structural stability.

[0067] In the present invention, the recombinant protein may be obtained by culturing a host cell (transformer) transformed with a recombinant expression vector in a medium suitable for protein expression using a known technique.

[0068] The recombinant expression vector of the present invention may include at least one of the polynucleotide represented by SEQ ID NO. 5 and the polynucleotide represented by SEQ ID NO. 7.

[0069] In the present invention, the recombinant protein comprising the polypeptide represented by SEQ ID NO. 6 may be encoded by the polynucleotide represented by SEQ ID NO. 5.

[0070] In the present invention, the recombinant protein comprising the polypeptide represented by SEQ ID NO. 8 may be encoded by the polynucleotide represented by SEQ ID NO. 7.

[0071] The recombinant protein according to the present invention may be a tetramer or a trimer, but is not limited thereto.

[0072] The recombinant protein containing the polypeptide represented by SEQ ID NO. 6 may be a tetramer, but is not limited thereto.

[0073] The recombinant protein containing the polypeptide represented by SEQ ID NO 8 may be a trimer, but is not limited thereto.

[0074] The vaccine composition according to the present invention may further comprise a recombinant protein comprising at least one of the polypeptide represented by SEQ ID NO. 6 and the polypeptide represented by SEQ ID NO. 8, or a functional equivalent thereof, and optionally one or more pharmaceutically acceptable carriers, excipients, and / or diluents. The carrier may further contain other pharmaceutically acceptable therapeutic / prophylactic drugs, vectors, or diluents suitable for administration for the treatment or prevention of henipharon. Pharmaceutically acceptable carriers are known in the art and may be aqueous or non-aqueous solutions, suspensions, and emulsions. Additionally, they may further comprise antimicrobial agents, antioxidants, chelating agents, inert gases, etc., as preservatives and other additives.

[0075] In the present invention, the vaccine composition may be administered by methods such as sublingual administration, transdermal administration, rectal administration, transmucosal administration, local administration, oral administration, intrapleural administration, intravenous administration, intra-arterial administration, intraperitoneal administration, subcutaneous administration, intramuscular administration, nasal administration, spinal administration, and intra-articular administration, but is not limited thereto.

[0076] The dosage of the vaccine composition of the present invention is selected by taking into consideration the age, weight, sex, physical condition, etc. of the individual. The amount required to induce an immunoprotective response in the individual without significant side effects may vary depending on the presence of the recombinant protein used as an immunogen and any excipient. Generally, each dose contains 0.1 to 1000 μg of protein, preferably 0.1 to 100 μg, per ml of sterile solution of the recombinant protein of the present invention. In the case of the vaccine composition, antigen stimulation may be performed arbitrarily repeatedly following the initial dose as needed.

[0077] Alternatively, the vaccine composition of the present invention may be administered at an immunologically effective amount. An "immunologically effective amount" refers to a sufficient amount to produce a preventive effect against disease and not cause side effects or severe or excessive immune responses, and may vary depending on the type or age of the animal to be vaccinated. The effective bacterial count content in the vaccine composition of the present invention is 10 4 to 10 9 cfu, preferably 10 5 to 10 8 It could be cfu.

[0078] The vaccine composition according to the present invention may further include an adjuvant. The adjuvant or adjuvant component refers to a broad spectrum of substances capable of increasing the immunogenicity of an antigen incorporated into or co-administered with an adjuvant, as a substance that enhances the efficacy of other agents, such as drugs or vaccines, and is, for example, a preparation or composition that can be pharmacologically or immunologically modified.

[0079] In the present invention, the adjuvant can enhance the henipahvirus-specific immunogenic effect of the recombinant protein according to the present invention. The adjuvant can allow for a lower dose of the immuno-interacting agent to increase the efficacy or safety of a specific dose of the immuno-interacting agent. For example, the adjuvant can increase the efficacy or safety of a specific immuno-interacting agent by preventing T cell depletion. The adjuvant includes, but is not limited to, mineral substances, aluminum hydroxide, aluminum phosphate, bacterial extracts (e.g., bacterial lipoglycans, Freund adjuvants, and / or MDP), oily emulsions, saponins, squalene, potassium aluminum sulfate, calcium hydroxide, TLR agonists, etc.

[0080] In one embodiment, the adjuvant may be at least one selected from the group consisting of Alum, Adju-Phos, MPLA-SM VacciGrade, AddaS03, and AddaVax, but is not limited thereto.

[0081] In another aspect, the present invention provides an immune-enhancing composition comprising, as an active ingredient, a recombinant protein comprising at least one of the polypeptide represented by SEQ ID NO. 6 and the polypeptide represented by SEQ ID NO. 8.

[0082] The recombinant protein of the present invention is characterized by having activity that induces a cell-mediated immune response. Therefore, the recombinant protein of the present invention can not only induce an adaptive immune response that triggers antibody production but also stimulate T cells to induce an innate immune response.

[0083] In one embodiment, it was confirmed that the recombinant protein can induce T cell activation by inducing the secretion of INF-γ cytokine in splenocytes stimulated with Nipah virus G peptide or Nipah virus F peptide.

[0084] The immunoadjuvant composition of the present invention may be a drug that enhances specific immune responses to a vaccine or inoculated drug. The immunoadjuvant composition may be provided in the form of an adjuvant as any substance or preparation that, when incorporated into a vaccine or inoculated drug, acts to accelerate, prolong, or enhance the quality of specific immune reactivity to immunogenic substances.

[0085] The immunostimulating composition of the present invention may include additional components known to those skilled in the art and may further include one or more pharmaceutically acceptable carriers, excipients, and / or diluents.

[0086] In another aspect, the present invention provides a composition for diagnosing heniphavirus infection or antibody possession, comprising as an active ingredient a recombinant protein comprising at least one of the polypeptide represented by SEQ ID NO. 6 and the polypeptide represented by SEQ ID NO. 8.

[0087] The recombinant protein of the present invention may be provided in a form coated on the surface of a plate. In this case, a sample is treated on the plate and reacted under appropriate conditions, and then the binding of the recombinant protein (antigen) of the present invention on the surface of the plate to the antibody in the sample is observed to diagnose whether there is a heniphavirus infection or whether antibodies against heniphavirus are present.

[0088] In the present invention, diagnosing (detecting) serum of henpavirus infection includes confirming the presence or / and amount of the anti-Nipah virus antibody or anti-Hendra virus antibody present in a sample of an individual by binding to the recombinant protein of the present invention (forming an antigen-antibody complex). If the presence of an anti-Nipah virus antibody is detected or / and confirmed in a sample of an individual, the individual may be determined to be infected with Nipah virus. If the presence of an anti-Hendra virus antibody is detected or / and confirmed in a sample of an individual, the individual may be determined to be infected with Henra virus. The sample may be a solid sample or a fluid sample, and preferably may be serum, plasma, whole blood, lymph fluid, or a homogenate thereof.

[0089] Since the diagnostic composition according to the present invention can diagnose whether there is a heniphavirus infection or whether there is an antibody against heniphavirus, the present invention can further provide a kit for diagnosing heniphavirus infection or a kit for diagnosing whether there is an antibody against heniphavirus comprising the diagnostic composition.

[0090] In the present invention, the diagnostic kit may be performed using an immunoassay method provided in the art, preferably an ELISA method, more preferably a sandwich-type ELISA, but is not limited thereto. Kit components according to each method are well known in the art. In addition, in addition to the recombinant protein of the present invention, a suitable buffer solution or medium for the binding reaction between the recombinant protein and an anti-Nipah virus antibody or an anti-Hendra virus may be additionally included. Furthermore, if the recombinant protein of the present invention is provided without being directly labeled, other detectable labeling means for labeling the recombinant protein may be additionally included in the kit.

[0091] In another aspect, the present invention provides a method for preventing or treating heniphavirus infection, comprising the step of administering to an animal a recombinant protein comprising at least one of the polypeptide represented by SEQ ID NO. 6 and the polypeptide represented by SEQ ID NO. 8.

[0092] The above animal may be an animal including humans or an animal excluding humans.

[0093] The above animal may be an animal that requires administration of the recombinant protein according to the present invention. The recombinant protein may be a recombinant protein comprising an effective amount of the polypeptide represented by SEQ ID NO. 6, the polypeptide represented by SEQ ID NO. 8, and any one of combinations thereof.

[0094] The above animals may be mammals, but are not limited thereto.

[0095] The animals other than the human mentioned above may be mammals such as cows, monkeys, birds, cats, mice, rats, hamsters, pigs, goats, dogs, rabbits, sheep, horses, etc.

[0096] In the present invention, 'prevention' refers to all effects that suppress or block heniphavirus infection or transmission in advance, and further includes all effects that delay the growth, proliferation, invasiveness, or infectivity of heniphavirus.

[0097] In the present invention, "administration" means introducing an active ingredient to the animal by an appropriate method ordinarily accepted in the art.

[0098] In the present invention, the method of administration may be administered through any general route as long as it can reach the target tissue. Examples may include, but are not limited to, sublingual administration, transdermal administration, rectal administration, transmucosal administration, local administration, oral administration, intrapleural administration, intravenous administration, intra-arterial administration, intraperitoneal administration, subcutaneous administration, intramuscular administration, intranasal administration, intravertebral administration, and intra-articular administration.

[0099] In the present invention, the dosage of the recombinant protein refers to a sufficient amount with a reasonable benefit / risk ratio applicable to treatment or improvement, and the effective dose level may be determined according to factors including the type of wound, application site, type and severity of the target, age, gender, drug activity, sensitivity to the drug, time of administration, route of administration and elimination rate, duration of treatment, concurrently used drugs, and other factors well known in the medical field.

[0100] Since the above method of administration is determined in light of the various relevant factors mentioned above, the dosage, route of administration, frequency of administration, etc., should not be understood as limiting the scope of the present invention in any aspect.

[0101]

[0102] The present invention will be described in detail below through examples. However, these are presented as preferred examples of the invention and should not be interpreted as limiting the invention. Details not described herein can be sufficiently technically inferred by those skilled in the art, so such descriptions are omitted.

[0103]

[0104] Example 1: Derivation of consensus sequences for the entire Bangladeshi and Malaysian Nipah virus strains

[0105] To develop a universal vaccine, complete sequences of Nipah viruses from Bangladesh, India, and Malaysia registered on NCBI were obtained, and fusion glycoprotein sequences (F gene) and glycoprotein sequences (G gene) were collected. A phylogenetic tree of complete genome / F gene / G gene ectodomain sequences was analyzed using Bayesian MCMC analysis. Subsequently, BD1, BD2, India, and MY clusters of the Bangladeshi and Malaysian types were distinguished using inter-genotypic or intra-genotypic p-distance analysis. Then, a rootless phylogenetic tree of complete genome / F gene / G gene ectodomain sequences was constructed using Maximum Likelihood analysis, and representative reference sequences for each cluster were obtained. The consensus sequence of the F gene (hereinafter referred to as NiV_conF) and the consensus sequence of the G gene (hereinafter referred to as NiV_conG) covering the references were extracted using the DNAMAN program and obtained using the Seaview program (Tables 1 and 2).

[0106] 서열번호 1NiV_conG_BPseqAGAAAGGGAGGTATGATAAAGTTATGCCATATGGACCCTCAGGCATCAAACAAGGTGACACCCTGTACTTTCCTGCTGTAGGATTTTTGGTCAGGACAGAGTTCAAATACAATGATTCAAATTGTCCCATCGCAAAGTGTCAATACAGCAAACCTGAAAACTGCAGGCTATCTATGGGGATTAGACCAAACAGTCATTATATCCTTCGATCTGGACTACTAAAATACAATCTATCAGATGAGGAGAACTCTAAAATTGTATTCATTGAAATATCTGATCAAAGACTATCTATTGGATCTCCTAGCAAAATCTATGATTCTTTGGGTCAACCTGTTTTCTACCAAGCGTCTTTTTCATGGGACACTATGATTAAATTTGGAGATGTTCAAACAGTTAACCCTTTGGTTGTAAATTGGCGTGACAACACGGTAATCTCAAGACCTGGGCAATCACAATGCCCTAGATTCAACACGTGCCCAGAGGTTTGCTGGGAAGGGGTTTATAATGATGCCTTCCTGATTGACAGAATCAATTGGATAAGCGCGGGTGTATTCCTTGACAGCAACCAGACCGCAGAGAATCCTGTTTTTACTGTATTCAAAGATAATGAAGTACTTTACAGAGCACAACTAGCTTCCGAGGACACCAATGCACAAAAAACAATAACTAATTGCTTCCTTTTGAAGAATAAGATCTGGTGCATATCACTGGTTGAGATATACGACACAGGAGACAATGTTATAAGACCTAAACTATTCGCAGTTAAGATACCAGAGCAATGTACATAA서열번호2NiV_conG_AAseqMPTESKKVRFENTASDKGKNPSKVIKSYYGTMDIKKINEGLLDSKILSAFNTVIALLGSIVIIVMNIMIIQNYTRSTDNQAMIKDALQSIQQQIKGLADKIGTEIGPKVSLIDTSSTITIPANIGLLGSKISQSTASINENVNEKCKFTLPPLKIHECNISCPNPLPFREYKPQTEGVSNLVGLPNNICLQKTSNQILKPKLISYTLPVVGQSGTCITDPLLAMDEGYFAYSHLEKIGSCSRGVSKQRIIGVGEVLDRGDEVPSLFMTNVWTPSNPNTVYHCSAVYNNEFYYVLCAVSVVGDPILNSTYWSGSLMMTRLAVKPKNNGESYNQHQFALRNIEKGRYDKVMPYGPSGIKQGDTLYFPAVGFLVRTEFKYNDSNCPIAKCQYSKPENCRLSMGIRPNSHYILRSGLLKYNLSDEENSKIVFIEISDQRLSIGSPSKIYDSLGQPVFYQASFSWDTMIKFGDVQTVNPLVVNWRDNTVISRPGQSQCPRFNTCPEVCWEGVYNDAFLIDRINWISAGVFLDSNQTAENPVFTVFKDNEVLYRAQLASEDTNAQKTITNCFLLKNKIWCISLVEIYDTGDNVIRPKLFAVKIPEQCT*

[0107] 서열번호 3NiV_conF_BPseqACCAAGATTATGCAACACCCATGACAAACAATATGAGGGAATGTTTGACGGGGTCGACTGAGAAGTGTCCTCGAGAGCTGGTTGTTTCATCACACGTTCCCAGATTTGCACTATCTAACGGGGTTTTGTTTGCTAATTGCATAAGTGTCACATGCCAGTGTCAAACAACAGGTAGGGCAATCTCACAGTCAGGAGAACAAACTCTGCTGATGATTGATAACACCACCTGTCCTACAGCTGTACTCGGTAATGTGATCATCAGCTTAGGAAAATATCTTGGGTCAGTAAATTATAACTCTGAAGGCATTGCTATTGGTCCTCCTGTCTTTACTGATAAAGTTGACATATCAAGTCAAATATCTAGCATGAATCAGTCCTTACAACAATCTAAGGACTATATCAAAGAGGCTCAACGACTCCTTGATACTGTTAACCCGTCATTAATAAGCATGTTGTCTATGATCATACTGTATGTACTATCAATTGCATCATTGTGTATAGGGTTGATTACATTTATCAGTTTTATCATTGTTGAGAAAAAAAGAAACACCTATAGCAGATTAGAGGACAGGAGAGTCAGACCTACAAGCAGTGGGGATCTCTATTACATTGGGACATAG서열번호4NiV_conF_*

[0108] (1) Verification of the representativeness of the NiV_conG sequence

[0109] The derived NiV_conG sequence showed a higher minimum similarity (MIN) value than any sequence of the collected G gene and showed a higher average similarity for all sequences within the group than the overall average similarity, confirming that this sequence is representative enough to cover the entire sequences of the Nipah virus Bangladesh type (BD1, BD2, India) and Malaysia type (MY) (Table 3).

[0110] Similarity (%) Average similarity of all sequences within the collected G gene 96.453 Average similarity between the consensus sequence and the entire G gene sequence 97.4493 Average minimum similarity (MIN) of the entire G gene sequence 92.5465 Maximum minimum similarity of the entire G gene sequence 92.9 Minimum similarity of the entire G gene sequence to the consensus sequence 93.5

[0111] In addition, it was confirmed that it is located exactly in the center of the group in a phylogenetic tree constructed by bootstrapping 1,000 times using the neighbor joining method with the whole G gene sequence and consensus sequence.

[0112] (2) Verification of the representativeness of the NiV_conF sequence

[0113] The derived NiV_conF sequence showed a higher minimum similarity (MIN) value than any sequence of the collected F gene and showed a higher average similarity for all sequences within the group than the overall average similarity, confirming that this sequence is representative enough to cover the entire sequences of the Nipah virus Bangladesh type (BD1, BD2, India) and Malaysia type (MY) (Table 4).

[0114] Similarity (%) Average similarity of all sequences within the collected F gene 96.95 23 Average similarity between the consensus sequence and the entire F gene sequence 97.88 33 Average minimum similarity (MIN) of the entire F gene sequence 93.13 08 Maximum minimum similarity of the entire F gene sequence 93.3 Minimum similarity of the entire F gene sequence to the consensus sequence 93.9

[0115] In addition, it was confirmed that it is located exactly in the center of the group in a phylogenetic tree constructed by bootstrapping 1,000 times using the neighbor joining method with the whole F gene sequence and consensus sequence.

[0116]

[0117] Example 2: Recombinant Protein Design and Expression

[0118] NiV_conG and NiV_conF, derived from the whole consensus sequences of Bangladeshi and Malaysian Nipah virus strains, were optimized for protein expression. A recombinant G protein was designed by sequentially placing a restriction enzyme recognition site, a Kozac sequence, and a signal peptide (MGWSCIILFLVATATGVHS; SEQ ID NO. 9) at the N-terminus of the optimized NiV_conG sequence, and placing a stop codon containing a 6XHis tag and a restriction enzyme recognition site at the C-terminus (Figs. 1-2). The coding sequence of the recombinant G protein is indicated as SEQ ID NO. 5 in Table 5.

[0119] 서열번호 5NiV_G_polynucleotidetctagaGCCACCAAAATTGTAGACTGTCTATGGGCATCAGGCCTAACAGCCACTACATCCTGCGCAGCGGCCTGCTGAAGTACAATCTGTCTGATGAGGAAAACAGCAAAATCGTGTTTATCGAAATCAGCGACCAGCGGCTGAGCATCGGCAGCCCCAGCAAGATCTATGATAGCCTGGGCCAGCCCGTGTTCTACCAGGCTAGCTTCAGCTGGGACACAATGATCAAGTTCGGCGACGTGCAGACAGTGAACCCTCTGGTGGTCAACTGGCGGGACAACACCGTGATCAGCAGACCAGGCCAGTCTCAGTGTCCTAGATTCAACACCTGTCCTGAGGTGTGCTGGGAGGGTGTCTACAACGACGCCTTCCTGATCGACAGAATCAACTGGATCTCTGCTGGCGTGTTTCTGGACAGCAATCAGACCGCCGAGAATCCGGTGTTCACAGTGTTTAAGGACAACGAGGTCCTGTACAGAGCTCAACTGGCCAGTGAAGATACCAACGCCCAGAAGACCATCACAAATTGCTTCCTGCTGAAAAACAAGATCTGGTGCATCAGCCTGGTCGAGATCTACGACACAGGCGACAACGTGATCCGGCCCAAGCTGTTTGCTGTCAAGATCCCTGAACAGTGCACCCACCATCACCACCACCATTGAgatatc서열번호6NiV_conG_

[0120] Next, a recombinant F protein was designed by sequentially positioning a restriction enzyme recognition site, a Kozac sequence, and a signal peptide (MGWSCIILFLVATATGVHS; SEQ ID NO. 9) at the N-terminus of the optimized NiV_conF sequence, inserting a GCNt trimerization motif at the C-terminus, and then positioning a stop codon containing a 6XHis tag and a restriction enzyme recognition site (Figs. 3-4). The coding sequence of the recombinant F protein is indicated as SEQ ID NO. 7 in Table 6.

[0121] 서열번호 7NiV_F_polynucleotideGAATTCCCGCCGCCACCTGACCAACAACATGAGAGAGTGCTTGACAGGCAGTACAGAGAAGTGCCCCCGGGAACTGGTCGTGAGCTCCCACGTGCCAAGATTCGCCCTGTCTAATGGAGTTCTGTTCGCCAACTGCATCAGCGTGACCTGCCAGTGTCAGACAACCGGCAGAGCCATCTCTCAGAGCGGCGAGCAGACCCTGCTGATGATCGATAATACCACCTGTCCTACCGCCGTGCTCGGCAATGTGATCATTAGCCTGGGCAAGTACCTGGGTAGCGTGAACTACAACAGCGAGGGCATTGCTATCGGCCCTCCTGTGTTTACAGACAAGGTGGACATCAGCAGCCAGATCTCCAGCATGAACCAGTCTCTGCAACAGAGCAAGGACTACATCAAGGAAGCTCAGAGACTGCTGGACACCGTCAACCCCAGCCTGATGAAGCAGATCGAAGATAAGATCGAGGAAATCCTGTCCAAGATCTATCACATCGAGAACGAGATTGCCAGAATCAAAAAGCTGATCGGCGAAGGCGGCGGAGGCTCTCACCACCACCACCATCACTGATAAGCTT서열번호8NiV_conF_polypeptideILHYEKLSKIGLVKGITRKYKIKSNPLTKDIVIKMIPNVSNMSQCTGSVMENYKTRLNGILTPIKGALEIYKNNTHDCVGDVRLAGVCMAGVAIGIATAAQITAGVAL YEAMKNADNINKLKSSIESTNEAVVKLQETAEKTVYVFTALQDYINTNLVPTIDKIPCKQTELSLDLALSKYLSDLLFVFGPNLQDPVSNSMTIQAISQAFGGNYETLLRTLGYATEDFDDLLESDSITG QIIYVDLSGYYIIVRVYFPILTEIQQAYIQELLPVSFNNDNSEWISIVPNFILVRNTLISNIEIGFCLITKRSVICNQDYATPMTNNMRECLTGSTEKCPRELVVSSHVPRFALSNGVLFANCISVTCQC QTTGRAISQSGEQTLLMIDNTTCPTAVLGNVIISLGKYLGSVNYNSEGIAIGPPVFTDKVDISSQISSMNQSLQQSKDYIKEAQRLLDTVNPSLMKQIEDKIEEILSKIYHIENEIARIKKLIGEGGGGS

[0122] After expressing the G protein using a plasmid constructed by inserting the 1,689 bp NiV_G_polynucleotide sequence into the pcDNA3.4 (6011 kb) vector, the expression of the recombinant G protein was confirmed by SDS-PAGE and Western blot (reducing, non-reducing conditions) using a 6XHis tag (Fig. 5).

[0123] After expressing the F protein using a plasmid constructed by inserting the 1,596 bp NiV_F_polynucleotide sequence into the pcDNA3.4 (6011 kb) vector, the expression of the recombinant F protein was confirmed by SDS-PAGE and Western blot (reducing, non-reducing conditions) using a 6XHis tag (Fig. 6).

[0124]

[0125] Example 3: Production and Purification of Recombinant Protein

[0126] To produce recombinant proteins of Nipah and Hendra viruses, the plasmid constructed in Example 2 was transformed into E. coli via heat shock, and the plasmid amount was increased. Protein expression was carried out by transfecting the plasmid into the Expi293F cell line. Specifically, the cells were set at 2.5–3 x 10⁶ one day prior. 6 Subcultured at cells / ml, and on D-day, the cells were 3 x 10 6 When the cell count was 95% or higher and the viability was 95% or higher, the cells were transferred to a new medium, and infection was carried out by slowly adding ExpiFectamine 293 reagent and plasmid in the specified ratio. The required number of cells and the amounts of plasmid and reagent according to the passage volume are as follows (Table 7).

[0127] Cell System Large Volume Plasmid DNA Concentration Plasmid DNA Amount Opti-MEM I Reduced Serum Medium Expi Fectamine 293 Regent Opti-MEM I Reduced Serum Medium Transfection Enhancer 1 Transfection Enhancer 2 Final Culture Volume 2 x 10 6 800ul1.0ug / ml0.8ul50ul2.5ul50ul5ul50ul~1ml7.5X10 6 2.5ml2.5ul150ul8ul140ul15ul150ul~3ml45X10 6 15ml15ul900ul50ul850ul90ul900ul~20ml75X10 6 25ml25ul1.5ml80ul1.4ml150ul1.5ml~30ml150X10 6 50ml50ul3ml160ul2.8ml300ul3ml~60ml

[0128] On Day 1, Transfection Enhancer 1 and Transfection Enhancer 2 were added 18–22 hours after transfection. After 5–7 days, recombinant proteins were harvested by filtering through a 0.22 µm filter at 3000–5000 g for 20–30 minutes. The harvested recombinant proteins were purified using a prepared Ni-NTA column and the 6XHis-tag embedded in the protein. The prepared Ni-NTA resin was added to the samples to be purified and bound by rotating at 4°C for 1 hour. Subsequently, the samples were washed 5 times with 5 mL of buffer and divided into 6 portions with 1 mL of elution buffer. The concentration of each eluted sample was measured using a nanodrop, and samples with high concentration and quality were selected.

[0129] To prepare the produced protein for animal inoculation, a buffer exchange step with PBS was added. The cap on the PD-10 desalted column was opened, the column storage solution was drained, and the bottom section was cut to allow the solution to pass through easily. To equilibrate the column, the PBS intended for buffer exchange was flowed through until the column was completely filled; this process was repeated at least four times to ensure a final flow of over 25 ml. 2.5 ml of the high-concentration and high-quality sample (total 3 ml) was added to the column and flowed through. A tube was placed under the column, 3.5 ml of PBS was added for elution, and the final concentration was checked using a NanoDrop.

[0130] It was confirmed that more than 1 mg of protein was produced for each of the recombinant G protein and recombinant F protein based on a 60 ml culture of Expi293F cells. Recombinant F protein is characterized by concentration reduction and clumping due to aggregation, resulting in variations in production volume between production batches; however, the decrease in protein expression efficiency was improved by adding an immediate PBS buffer exchange step after purification to minimize the influence of imidazole in the buffer used during the purification step.

[0131]

[0132] Example 4: Evaluation of Immunogenicity of Recombinant Protein in Mice (balb / c)

[0133] (1) Evaluation of binding antibodies

[0134] To confirm the immunogenicity of the recombinant G protein (hereinafter referred to as G(ct)) and recombinant F protein (hereinafter referred to as F(ct)) obtained in Example 3 as universal vaccine candidates, 6-week-old male mice (BALB / cAnNCrlOri, SPF) were vaccinated with a 3-week interval between the first and second vaccinations, and blood was collected 3 weeks after the first vaccination and necropsy was performed 3 weeks after the second vaccination (Fig. 7).

[0135] The vaccination test group was composed as shown in Table 8. The adjuvant was standardized to observe the effect according to the vaccination dose. A bivalent vaccine composition (G1-G2 group) of Malaysia G protein (hereinafter referred to as G(Mt)) and Bangladesh G protein (hereinafter referred to as G(Bt)), which showed high binding antibody titers and neutralizing antibody titers in previous studies, and a mixed composition (G3-G4 group) of G(Mt), G(Bt) and F protein (hereinafter referred to as F(t)) were used as positive controls, and the G13 group, which received PBS, was used as a negative control.

[0136] Group Candidate Substance Number of Immunostimulants Dose G1 NiV Recombinant Protein G(Mt) + G(Bt)7Alum 10 µg + 10 µg / dose G2 NiV Recombinant Protein G(Mt) + G(Bt)7Alum 20 µg + 20 µg / dose G3 NiV Recombinant Protein G(Mt) + G(Bt) + F(t)7Alum 10 µg + 10 µg + 10 µg / dose G4 NiV Recombinant Protein G(Mt) + G(Bt) + F(t)7Alum 20 µg + 20 µg + 20 µg / dose G5 NiV Recombinant Protein G(ct)7Alum 10 µg / dose G6 NiV Recombinant Protein G(ct)7Alum 20 µg / dose G7 NiV Recombinant Protein G(ct)7Alum 40ug / dose G8NiV recombinant protein F(ct)7Alum 10ug / dose G9NiV recombinant protein F(ct)7Alum 20ug / dose G10NiV recombinant protein F(ct)7Alum 40ug / dose G11NiV recombinant protein G(ct) + F(ct)7Alum 10ug+10ug / dose G12NiV recombinant protein G(ct) + F(ct)7Alum 20ug+20ug / dose G13PBS4--Total90

[0137] As a universal vaccine candidate, binding antibodies against Nipah virus G protein and F protein, as well as Hendra virus G protein and F protein, were also confirmed.

[0138] 1st 3w1st 3wMean2nd 3w2nd 3wMean 1st3w1st3wMean2nd3w2nd3wMeanNiV GO.DO.DO.DO.DO.DO.DNiV GO.DO.DO.DO.DO.DO.DG13.373.163.273.453.303.38G63.413.203.303.583.333.463.203.203.203.433.403.413.503.383.443.503.463.483.073.073.073.433.403.413.403.553.473.443.423.433.273.163.223.493.463.473.473.513.493.463.523.493.313.173.243.493.403.453.493.363.433.553.463.503.373.413.393.533.483.513.463.523.493.513.553.533.093.033.063.393.363.383.513.483.50- - - G23.063.113.083.513.313.41G73.223.213.213.263.303.283.173.073.123.313.293.303.233.153.193.363.313.333.103.053.083.463.463.463.593.533.563.443.463.453.353.303.333.523.493.503.573.473.523.343.343.343.443.273.363.483.463.473.313.253.283.563.473.513.323.213.263.433.353.393.503.473.483.543.413.483.133.153.14- - - 3.423.433.42 -- - G33.073.183.123.353.343.34G113.283.293.293.123.143.132.993.053.023.153.223.183.263.253.253.263.283.273.073.043.053.383.393.383.423.393.413.223.283.253.213.203.213.413.313.363.423.353.383.323.253.283.143.163.153.363.323.343.363.373.363.173.213.193.293.313.303.273.203.243.303.343.323.353.363.363.143.113.123.253.313.283.263 .343.303.313.423.37G43.143.043.093.283.343.31G123.303.273.283.103.123.113.193.283.243.383.26 3.323.223.203.213.313.273.293.083.143.113.353.313.333.393.313.353.403.383.393.213.193.203.433.353.393.413.303.363.253.263.253.253.323.293.473.463.463.453.483.473.353.313.333.133.073.10 -- - 3.283.283.283.313.323.323.203.233.21- - - 3.323.383.353.423.483.45G53.193.123.153.253.193.22G130.180.250.210.540.280.413.073.093.08 3.233.323.280.240.260.250.670.360.523.233.143.193.263.303.280.260.300.280.590.320.453.333. 183.253.323.323.320.230.230.230.450.310.383.353.253.303.463.453.46 Positive control 3.043.333.193.193.393.293.263.313.293.263.353.30 Negative control 0.310.430.370.310.550.433.213.193.203.253.333.29-------.

[0139] As shown in Table 9, high levels of binding antibodies against the Nipah virus G protein were confirmed in the serum of the G5-G7 groups administered the G(ct) vaccine candidate 3 weeks after both the first and second doses. Furthermore, there was no difference in binding antibody titers when compared to the positive control group G1-G2 of the bivalent vaccine, confirming that the recombinant G protein alone possesses excellent immunogenicity. The highest binding antibody titer was observed at a dose of 20 µg / dose (Fig. 9).

[0140] 1st3w1st3wMean2nd3w2nd3wMean 1st3w1st3wMean2nd3w2nd3wMeanNiV FO.DO.DO.DO.DO.DO.DNiV FO.DO.DO.DO.DO.DO.DG30.310.270.291.071.031.05G100.370.380.382.362.412.390.300.320.310.870.940.910.460.490.482.212.262.230.250.240.250.850.900.870.380.390. 392,032,112,070,240,250,251,091,201,140,590,580,592,702,662,680,340,320,331,291,421,360,700,740,722,872,862,870,270,250,261,191,261,230,480,470,482,642,672 .660.290.280.281.161.161.160.320.330.321.821.861.84G40.210.220.211.441.371.40 G110.570.540.550.740.670.700.300.290.291.171.241.200.360.360.360.800.900.850. 270.290.280.800.780.790.410.440.420.930.970.950.630.680.661.301.281.290.380.410.390.760.880.820.300.370.331.441.411.430.270.260.260.920.930.930.260.260.260.260.26- - - 0.230.230.231.101.171.130.260.280.27- - - 0.480.480.481.751.801.77G80.490.520.511.301.311.31G120.520.500 .511.131.191.160.470.510.492.312.402.350.450.450.450.750.780.77 0.330.340.341.751.741.740.400.400.400.930.920.920.290.300.291.711.731.720.280.270.271.321.271.290.230.250.241.881.901.890.510.500.501.671.721.690.260.250.252.082.022.050.300.290.301.751.771.760.380.360.371.261.301.280.250.270.261.511.511. 51G90.330.330.331.291.321.31G130.210.200.200.250.270.260.310.310.312.552.522.540.240.240.240.240.220.230.360.360 .362.112.062.090.260.270.260.220.230.220.340.370.360.910.910.910.220.230.230.210.210.210.230.230.231.221.241.23 Positive control 3.333.543.443.573.433.500.330.340.340.140.140.14 Negative control 0.430.300.370.400.330.360.250.260.262.642.642.64-------.

[0141] As shown in Table 10, the G8-G10 groups vaccinated with the F(ct) vaccine candidate showed higher levels of binding antibodies against the Nipah virus F protein in the serum 3 weeks after the second dose compared to the serum 3 weeks after the first dose. The highest binding antibody levels were observed at a dose of 40 µg / dose.

[0142] 1st 3w1st 3wMean2nd3w2nd3wMean 1st 3w1st 3wMean2nd3w2nd3wMeanHeV GO.DO.DO.DO.DO.DO.DHeV GO.DO.DO.DO.DO.DO.DG12.812.472.642.892.732.81G62.742.572.653.233.143.192.832.792.812.652.922.782.962.952.953.133.173.152.822.772.792.562.722.642.942.882.912.892.822.852.812.8 62.842.862.972.912.812.742.783.113.093.102.932.782.862.832.852.842.712.602.652.933.002.972.802.672.742.762.782.772.922.912.913.052.993.022.642.582.612.862.842.852.822.712.77- -- G22.592.552.572.712.632.67G72.562.602.583.062.953.012.702.592.652.912.972.94 2.612.572.592.972.942.952.963.002.982.932.802.862.892.912.902.872.982.932.87 2.782.833.042.902.972.792.772.782.912.882.902.892.912.902.802.672.742.802.722.763.053.133.092.742.712.732.892.822.862.892.842.872.862.862.882.792.83- - - 2.882.842.86- - - G32.582.542.562.822.872.85G112.432.452.443.053.003.032.412.492.452.612.782.692.622.652.642.993.073.032.502.502.502. 792.932.862.642.662.652.932.882.902.712.782.752.732.822.77 2.562.632.592.812.722.762.192.162.182.772.692.732.502.572.542.742.702.721.151.121.132.772.732.752.552.652.602.892.852.872.192.212.202.772.812.792.702. 652.682.972.972.97G42.722.632.672.632.762.69G122.572.512.542.812.842.822.892.822.852.712.892 .802.932.942.932.812.912.862.832.712.772.913.022.972.762.742.752.942.952.952.832.782.812.873.052.962.562.522.542.732.802.772.822.822.822.682.712.692.472.502.482.932.932.932.492.472.48- - - 2.562.562.562.782.802.792.672.652.66- - - 2.492.502.492.932.982.96G52.752.672.712.732.762.75G130.080.120.100.080.120.102.892.682.79 2.972.842.900.100.110.110.060.060.062.912.802.852.792.742.770.090.160.120.060.070.072.682. 592.642.852.842.850.060.060.060.060.060.062.972.952.962.792.802.79Positive control 3.052.963.002.883.143.012.502.522.512.612.622.62Negative control 0.100.060.080.060.070.072.402.352.382.802.752.77-------.

[0143] As shown in Table 11, the G8-G10 groups administered the G(ct) vaccine candidate showed a constant level of binding antibody titers against the Hendra virus G protein in both serum samples 3 weeks after the first and second doses, and there was no difference in binding antibody titers compared to the positive control group, demonstrating immunogenicity against the Hendra virus with the recombinant G protein alone (Fig. 10).

[0144] 1st 3w1st 3wMean2nd3w2nd3wMean 1st3w1st 3wMean2nd3w2nd3wMeanHeV FO.DO.DO.DO.DO.DO.DHeV FO.DO.DO.DO.DO.DO.DG30.930.900.912.662.392.52G101.041.061.052.362.322.340.590.600.602.792.762.781.711.741.732.552.552.550.460.420.442.442.432.432.632.582. 612.622.652.630.500.440.472.622.462.540.920.950.942.782.832.810.280.260.272.902.802.851.381.341.362.892.812.850.140.130.142.902.792.840.981.000.992.942.96 2.950.310.280.302.802.682.741.141.121.132.702.762.73G40.300.300.302.952.952.9 5G112.662.572.622.962.802.881.251.191.222.742.642.691.291.191.241.951.881.910 .750.770.762.021.992.010.330.320.322.742.662.702.052.012.032.872.792.830.540.520.532.902.852.880.360.360.362.912.932.920.180.160.171.951.921.930.380.390.39 0.390.370.382.682.802.741.071.081.08- - - 0.860.830.843.153.153.15G81.171.171.172.492.412.45G122.332.322 .332.832.832.831.791.851.822.352.402.372.172.242.212.792.742.77 1.171.181.172.662.532.600.790.750.772.492.452.471.531.561.542.722.692.700.480.450.473.013.013.010.420.380.402.352.372.361.851.821.832.362.382.370.350.330.342.772.672.720.500.480.492.952.932.941.801.791.791.931.941.930.540.510.532.852.872. 86G91.171.191.182.652.672.66G130.050.050.050.080.080.081.301.341.322.712.632.670.090.080.080.070.070.071.301.291 .292.482.462.470.080.080.080.070.070.071.521.521.521.481.471.480.060.070.070.070.060.071.161.181.172.762.692.73Positive control 3.333.543.443.573.433.500.620.640.632.542.522.53Negative control 0.080.070.070.080.040.060.710.700.712.432.472.45-------.

[0145] As shown in Table 12, the G8-G10 groups administered the F(ct) vaccine candidate showed higher levels of binding antibodies against the Hendra virus F protein in the serum 3 weeks after the second dose compared to the serum 3 weeks after the first dose. In particular, levels higher than those against the Nipah virus F protein were observed. Regarding the dosage, the highest binding antibody levels were observed in the serum 3 weeks after the first dose at 40 µg / dose, and 3 weeks after the second dose, binding antibody levels were detected in all groups except the group administered PBS (Fig. 11).

[0146]

[0147] (2) Evaluation of neutralizing antibody activity using pseudovirus

[0148] In the above experiment, each mouse serum was treated with a recombinant VSV vector-based Nifas Pseudovirus, and the IC against neutralizing antibodies generated in the serum 50The values ​​were calculated. The neutralizing ability 3 weeks after the first vaccination and the neutralizing ability 3 weeks after the second vaccination are shown in Table 13 and Figure 12, and Table 14 and Figure 13, respectively.

[0149] Group IC 50 Group IC 50 Group IC 50 G116146.85G511875.58G9189.812658.521681.472162.131923.83107 0.07391.934788.554668.364111.952273.75513.555261.9661449.726 7288.116339.197425.537924.667361.7G211339.17G61734.45G10164. 5821194.9821135.992227.163523.5332046.46361.567874534.624206 5.11432.8183254359.315506.95555.3813463328.2161109.33690.213 4971264.4671334.83771.01927G313212.04G712376.62G111493.44322 3384.1922164.012406.06443645.9831137.683749.872841295.634124 1.714842.89785705.345593.965686.21546776.56273.796143.20987- 71197.4671696.701G414419.64G81124.93G1211051.89125161.662235.542267.294933089.773152.673248.1292428 82.724310.214375.230451254.056596.335211.01356372.3145637.326220.44937782.28697161.527320.0952-G131 -203040

[0150] Group IC 50 Group IC 50 Group IC 50G11159781.4G6122220.55G11119494.53270643.21262346.04233542.63337414.99353576.94338774.942 24653.44205248.3473281.175451422.6571146.665116608.96130695.96199073.86200928.1743288.447 -731100.46G21467076.7G7119505.63G12154118.452193725.22149207.42226768.8377440.4138222.363 95212.03431762.48440722.25485287.365136503.6530962.125181722.7651621.1639446.766115625.47 -7- 7415541.9G3181970.91G8113573.8G1313.42276218.125121.97203204148.937922.18304529631.144141.98405211181.45690.2611149 9.5619532.727293612.179271.7G411238735G911936.012513814.52 4256.653688690.9317706.164177284642363.652034055510529.696 -65954.287 -70G511241124G1011763.582100300122712.933462950.133822.3643221 1.9842564.985144065.251299.566127691.661021.43721241.6471204.88

[0151] The group pooling results of serum neutralizing ability 3 weeks after the first vaccination against Nipah pseudovirus are shown in Table 15 and Figure 14, and the group pooling results of serum neutralizing ability 3 weeks after the second vaccination are shown in Table 16 and Figure 15.

[0152] No. Vaccine Candidate IC 50G1NiV Recombinant Protein G(Mt) + G(Bt) / 10ug+10ug / dose 2298.08 G2NiV Recombinant Protein G(Mt) + G(Bt) / 20ug+20ug / dose 991.61 G3NiV Recombinant Protein G(Mt) + G(Bt) + F(t) / 10ug+10ug+10ug / dose 619.3 G4NiV Recombinant Protein G(Mt) + G(Bt) + F(t) / 20ug+20ug+20ug / dose 1073.86 G5NiV Recombinant Protein G(ct) / 10ug / dose 1175.16 G6NiV Recombinant Protein G(ct) / 20ug / dose 1946.9 G7NiV Recombinant Protein G(ct) / 40ug / dose 832.95G8NiV Recombinant Protein F(ct) / 10ug / dose 117.91G9NiV Recombinant Protein F(ct) / 20ug / dose 97.22G10NiV Recombinant Protein F(ct) / 40ug / dose 108.21G11NiV Recombinant Protein G(ct) + F(ct) / 10ug+10ug / dose 907.29G12NiV Recombinant Protein G(ct) + F(ct) / 20ug+20ug / dose 540.63G13PBS0

[0153] No. Vaccine Candidate IC 50G1NiV Recombinant Protein G(Mt) + G(Bt) / 10ug+10ug / dose 159700 G2NiV Recombinant Protein G(Mt) + G(Bt) / 20ug+20ug / dose 159688.3 G3NiV Recombinant Protein G(Mt) + G(Bt) + F(t) / 10ug+10ug+10ug / dose 215466 G4NiV Recombinant Protein G(Mt) + G(Bt) + F(t) / 20ug+20ug+20ug / dose 1249628 G5NiV Recombinant Protein G(ct) / 10ug / dose 433183.7 G6NiV Recombinant Protein G(ct) / 20ug / dose 102268.7 G7NiV Recombinant Protein G(ct) / 40ug / dose 48011.08G8NiV Recombinant Protein F(ct) / 10ug / dose 8607.793G9NiV Recombinant Protein F(ct) / 20ug / dose 6106.627G10NiV Recombinant Protein F(ct) / 40ug / dose 2055.674G11NiV Recombinant Protein G(ct) + F(ct) / 10ug+10ug / dose 73390.1G12NiV Recombinant Protein G(ct) + F(ct) / 20ug+20ug / dose 167753.8G13PBS0

[0154] Upon examination, a certain level of neutralizing antibody titers against Nipah pseudovirus was confirmed in the serum 3 weeks after the first dose of the G(ct) vaccine candidate; excluding the positive control group with the bivalent vaccine composition, the highest neutralizing antibody titers against Nipah pseudovirus were observed in the G6 group, which received 20 µg / dose of the G(ct) vaccine candidate. Furthermore, high levels of neutralizing antibody titers were confirmed in the serum 3 weeks after the second dose, with the highest titers observed in the G5 group, which received 10 µg / dose of the G(ct) vaccine candidate. This indicates that the recombinant G protein demonstrated excellent immunogenicity against Nipah virus even at low doses when administered alone.

[0155]

[0156] Example 5: Evaluation of Immunogenicity of Recombinant Protein in Hamsters (Syrian Golden Hamster)

[0157] (1) Binding antibodies against Nipah virus and Hendra virus

[0158] The binding antibody titers and neutralizing antibody titers of the G(ct) vaccine candidate and the F(ct) vaccine candidate were evaluated using Syrian Golden Hamsters, which are used as animal models for Nipah virus infection. The first and second doses were administered with an interval of 3 weeks, and blood was collected 3 weeks later, and blood collection and autopsy were performed 3 weeks after the second dose (Fig. 16).

[0159] The vaccination test group was composed as shown in Table 17 to observe the effects of the adjuvant. As in mouse experiments, the bivalent vaccine composition of G(Mt) and G(Bt) (G1 group), which showed high binding antibody and neutralizing antibody titers against Nipah virus in previous studies, and the mixed composition of G(Mt), G(Bt), and F(t) (G2 group) were used as positive controls, and the G10 group, which was vaccinated with PBS, was used as a negative control.

[0160] No. Vaccine Candidate Number of Agents Adjuvant dose G1 NiV Recombinant Protein G(Mt) + G(Bt)5 Alum 20ug + 20ug / dose G2 NiV Recombinant Protein G(Mt) + G(Bt) + F(t)5 Alum 20ug + 20ug + 20ug / dose G3 NiV Recombinant Protein G(ct)5 Alum 20ug / dose G4 NiV Recombinant Protein G(ct)5 Adju-Phos 20ug / dose G5 NiV Recombinant Protein G(ct)5 MPLA-SM VacciGrade 20ug / dose G6 NiV Recombinant Protein G(ct)5 AddaS03 20ug / dose G7 NiV Recombinant Protein G(ct)5 AddaVax 20ug / dose G8 NiV Recombinant Protein F(ct)5Alum20ug / doseG9NiVrecombinant protein G(ct) + F(ct)5Alum20ug+20ug / doseG10PBS5Alum- Total50

[0161] As a universal vaccine candidate, binding antibodies against Nipah virus G protein and F protein, as well as Hendra virus G protein and F protein, were also identified.

[0162] 1st3w1st3wMean2nd3w2nd3wMean 1st3w1st3wMean2nd3w2nd3wMeanNiV GO.DO.DO.DO.DO.DO.DNiV GO.DO.DO.DO.DO.DO.DG12.422.102.263.062.802.93G62.192.082.133.082.882.982.322.212.272.962.912.932.021.931.972.832.662.742.132.032.082.782.682.731.821.781.802.592.452.522.262.182.222.572.532.550.830.830.832.622.522.572.392.222.313.083.023.050.620.580.602.722.692.71G22.012.052.032.742.652.70G71.001.021.012.492.322.412.082.092.082.732.702.710.340.340.342.712.692.701.741.811.782.842.822.830.720.700.712.432.402.421.861.891.882.762.742.750.420.430.432.262.292.271.681.711.702.942.972.950.130.130.13- -- G31.831.711.772.602.602.60G91.962.052.012.732.772.751.851.841.842.522.512.511.731.731.731.971.941.951.731.741.732.882.932.901.461.411.442.672.662.670.690.670.681.891.891.891.981.981.982.502.612.552.082.012.042.422.392.402.222.122.172.272.222.24G40.390.390.392.292.252.27G100.120.120.120.150.150.151.661.721.692.292.182.240.120.120.120.140.140.140.680.660.672.602.632.610.120.120.120.130.140.131.311.411.362.052.092.070.120.130.120.140.140.140.590.620.602.222.222.220.120.130.130.130.140.13G50.120.120.120.540.530.54PositiveControl3.143.003.073.603.603.6 00.130.120.120.930.930.93 Negative control group 0.290.210.250.250.200.230.100.110.111.441.481.46 - 0.200.190.200.330.330.330.130.130.130.340.330.34.

[0163] As shown in Table 18, the results of confirming the binding antibody titers against Nipah virus G protein showed that in all groups except the G10 group vaccinated with PBS, the binding antibody titers were higher 3 weeks after the second vaccination than 3 weeks after the first vaccination. Regarding differences according to the adjuvant, among the groups excluding the positive control groups (G1, G2), the G6 group using AddaSO3 had the highest binding antibody titer, followed by the group using AddaVax, the group using Alum, the group using Adju-Phos, and the group using MPLA-SM VacciGrade (Fig. 17).

[0164] 1st3w1st3wMean2nd3w2nd3wMeanNiV FO.DO.DO.DO.DO.DO.DG20.210.180.190.770.750.760.250.240.250.280.290.280 .130.130.130.360.320.340.160.230.190.610.650.630.200.270.230.310.310.31 G80.150.160.160.510.520.520.230.190.210.310.340.320.210.220.220.350.280 .310.210.200.200.390.440.420.230.180.200.270.280.28G90.130.130.130.450. 460.460.220.190.210.310.250.280.230.160.190.360.340.350.150.130.140.40 0.400.400.190.140.160.450.450.45G100.130.120.130.190.200.200.130.160.14 0.190.190.190.130.150.140.200.210.210.120.120.120.180.200.190.230.230.230.200.190.19 Positive control 3.40 3.45 3.42 3.59 3.67 3.63 Negative control 0.18 0.09 0.140.28 0.270.28

[0165] As shown in Table 19, when confirming the binding antibody titer against Nipah virus F protein, unlike the results in mice, no binding antibody titers were observed in the serum 3 weeks after the first and second vaccinations (Fig. 18).

[0166] 1st 3w1st 3wMean2nd 3w2nd 3wMean 1st 3w1st 3wMean2nd 3w2nd 3wMeanHeV GO.DO.DO.DO.DO.DO.DHeV GO.DO.DO.DO.DO.DO.DG11.861.661.762.902.712.80G62.612.332.473.0 12.742.882.322.142.232.822.712.760.800.730.762.722.662.692.562 .412.492.852.702.771.481.411.452.722.642.682.142.062.102.712.7 02.710.480.430.462.982.982.981.651.591.622.632.472.550.290.260 .282.582.502.54G21.411.351.382.032.052.04G70.320.320.322.522.4 92.511.661.681.672.702.642.670.160.150.152.592.672.631.811.861 .842.112.012.060.110.110.112.682.572.621.241.231.242.282.082.180.190.190.191.091.051.071.241.301.272.672.572.620.050.040.05- -- G32.032.022.022.232.322.28G90.690.680.691.451.451.450.981.000.992.722.762.740.520.530.522.102.132.120 .550.540.552.422.392.411.891.931.912.612.602.610.410.430.422.222.192.210.981.011.002.352.352.352.332.3 42.332.832.792.811.911.981.951.531.511.52G40.190.190.191.091.071.08G100.040.050.040.070.080.080.440.44 0.441.531.511.520.040.050.050.070.080.070.200.200.202.342.292.310.040.040.040.060.060.060.360.360.360.880.890.880.040.040.040.060.060.060.310.300.311.911.931.920.040.040.040.050.060.06G50.050.050.050.110.110.11 Positive control 3.453.413.433.583.5 83.580.050.100.080.680.730.70 Negative Control Group 0.090.270.180.120.090.110.050.050.050.510.500.51-0.060.060.060.530.540.540.060.060.060.120.100.11.

[0167] As shown in Table 20, the results of confirming the binding antibody titers against Hendra virus G protein showed that in all groups except the G10 group vaccinated with PBS, the binding antibody titers increased 3 weeks after the second vaccination compared to 3 weeks after the first vaccination. Regarding the differences according to the adjuvant, similar to the results for the binding antibody titers against Nipah virus G protein, the G6 group using AddaSO3 had the highest binding antibody titer among the groups excluding the positive controls (G1, G2); however, slightly different from the results for the Nipah virus G protein, the binding antibody titers were confirmed in the order of the group using Alum, the group using AddaVax, the group using Adju-Phos, and the group using MPLA-SM VacciGrade (Fig. 19).

[0168] 1st3w1st3wMean2nd3w2nd3wMeanHeV FO.DO.DO.DO.DO.DO.DG20.810.720.771.771.671.720.130.100.110.600.560.580 .200.170.190.300.280.290.130.110.121.441.351.400.540.320.431.211.131.17 G80.390.410.401.461.431.450.250.240.241.741.791.770.340.330.340.960.970 .970.070.080.080.770.780.780.300.300.300.560.550.56G90.360.390.371.291. 291.290.530.500.510.650.680.670.130.140.141.341.311.320.520.500.511.49 1.741.610.190.200.201.731.751.74G100.050.060.060.100.100.100.050.050.05 0.180.150.170.050.050.050.170.170.170.050.050.050.180.120.150.050.050.050.140.100.12 Positive control 3.573.423.493.433.503.47 Negative control 0.100.130.120.150.150.15

[0169] As shown in Table 21, the results of confirming the binding antibody titers for the Hendra virus F protein showed that the binding antibody titers in the serum were higher 3 weeks after the second vaccination than 3 weeks after the first vaccination, but this was not significant. However, similar to the results in mice, it was confirmed that the binding antibody titers for the Hendra virus F protein were higher than those for the Nipah virus F protein (Fig. 20).

[0170]

[0171] (2) Neutralizing antibodies against Nipah virus and Hendra virus

[0172] In the above experiment, each hamster serum was treated with a recombinant VSV vector-based Nipapsedovirus, and the IC against neutralizing antibodies generated in the serum50 The values ​​were calculated and shown in Table 22. The neutralizing ability 3 weeks after the first vaccination and 3 weeks after the second vaccination for each group are shown in Figures 21a-21b and Figures 22a-22b, respectively.

[0173] 1st 3w 2nd 3w 1st 3w 2nd 3w Group IC 50 IC 50 Group IC 50 IC 50 G13906.682898.33G61616.084163.232282.272535.79354.444940.515 64.69014.39431.4510916.19779.053465.3810.5511773.34742.175061 .1907711.31G2826.692778.32G701618.411611.581226.380310.28207 3.251029.770.41202.481095.81718.6211.491202.45176.122142.190- G399.91993.49G8114.073394.94169.093520.3402852.11795.34811.46105. 24307.891.77011.581053.19338.031110.790.867.06G40138.58G9602.8426 70.7245.25150.99190.5178.862.57959.93104.175524.9712.722.57870.46 9332.440372.752649.792623.09G500G102.29000000279.8001.51000000.850

[0174] The group pooling results of serum neutralizing ability 3 weeks after the first vaccination against Nipah pseudovirus are shown in Table 15 and Figure 14, and the group pooling results of serum neutralizing ability 3 weeks after the second vaccination are shown in Table 23 and Figure 23.

[0175] 1st 3w 2nd 3w group vaccine candidate adjuvant IC 50 MeanIC 50MeanG1NiV recombinant protein G(Mt) + G(Bt)Alum1854.9544595.016G2NiV recombinant protein G(Mt) + G(Bt) + F(t)Alum1156.6881779.056G3NiV recombinant protein G(ct)Alum280.8281287.216G4NiV recombinant protein G(ct)Adju-Phos52.108324.964G5NiV recombinant protein G(ct)MPLA-SM VacciGrade0.30255.96G6NiV recombinant protein G(ct)AddaS03482.5047900.914G7NiV recombinant protein G(ct)AddaVax 2.378 108 3.405G8NiVRecombinant Protein F(ct)Alum 46.338 1535.038G9NiVRecombinant Protein G(ct) + F(ct)Alum 88 3.55 240 66.012G10PBSAlum 0.6280

[0176] Upon examination, high levels of neutralizing antibodies were found in the serum 3 weeks after the second dose in all groups except the G10 group that received PBS, and the highest neutralizing ability was confirmed in the G6 group that received AddaSO3, even when compared to the positive control group (G1, G2, G9).

[0177]

[0178] Example 6: Confirmation of cell-mediated immune response to recombinant protein in hamsters (Syrian golden hamster)

[0179] To confirm the cell-mediated immune response of the G(ct) vaccine candidate and the F(ct) vaccine candidate in the hamster of Example 5, the spleen was removed 3 weeks after the second vaccination and an ELISpot was performed.

[0180] Splenocytes were stimulated with Nipah virus G peptide, and as a result of confirming cell-mediated immunogenicity, INF-γ cytokine levels were detected in all groups vaccinated with the G(ct) vaccine candidate and the F(ct) vaccine candidate. Additionally, similar to humoral immune responses, INF-γ cytokine levels were highest in the G6 group, which used the AddaSO3 adjuvant. In contrast, no INF-γ cytokine levels were detected in the positive control groups G1 and G2 (Figs. 24a–24b). Next, splenocytes were stimulated with Nipah virus F peptide, and as a result of confirming cell-mediated immunogenicity, high INF-γ cytokine levels were detected in the G9 group, which was vaccinated with a mixture of the G(ct) vaccine candidate and the F(ct) vaccine candidate (Figs. 25a–25b).

[0181] These results are novel findings confirming the cell-mediated immunogenicity of the recombinant G protein and recombinant F protein according to the present invention.

Claims

1. A vaccine composition for the prevention or treatment of heniphavirus infection, comprising as an active ingredient a recombinant protein comprising at least one of the polypeptide represented by SEQ ID NO. 6 and the polypeptide represented by SEQ ID NO.

8.

2. A vaccine composition for the prevention or treatment of henipah virus infection according to claim 1, wherein the henipah virus is a Nipah virus or a Hendra virus.

3. A vaccine composition for the prevention or treatment of henipahvirus infection according to Claim 1, wherein the polypeptide represented by SEQ ID NO. 6 is derived from the G protein of the Bangladesh Nipahvirus strain and the Malaysia Nipahvirus strain.

4. A vaccine composition for the prevention or treatment of henipahvirus infection according to Claim 1, wherein the recombinant protein comprising the polypeptide represented by SEQ ID NO. 6 is encoded by the polynucleotide represented by SEQ ID NO.

5.

5. A vaccine composition for the prevention or treatment of henipahvirus infection according to Claim 1, wherein the polypeptide represented by SEQ ID NO. 8 is derived from the F protein of the Bangladesh Nipahvirus strain and the Malaysia Nipahvirus strain.

6. A vaccine composition for the prevention or treatment of henipahvirus infection according to Claim 1, wherein the recombinant protein comprising the polypeptide represented by SEQ ID NO. 8 is encoded by the polynucleotide represented by SEQ ID NO.

7.

7. A vaccine composition for the prevention or treatment of henipahvirus infection according to claim 1, further comprising an adjuvant.

8. A vaccine composition for the prevention or treatment of henipharon infection according to claim 1, wherein the adjuvant is at least one selected from the group consisting of Alum, Adju-Phos, MPLA-SM VacciGrade, AddaS03, and AddaVax.

9. A vaccine composition for the prevention or treatment of henipahvirus infection according to claim 1, wherein the recombinant protein has activity to induce a cell-mediated immune response.

10. An immune-enhancing composition comprising, as an active ingredient, a recombinant protein comprising at least one of the polypeptide represented by SEQ ID NO. 6 and the polypeptide represented by SEQ ID NO.

8.

11. A composition for diagnosing heniphavirus infection or antibody possession, comprising as an active ingredient a recombinant protein comprising at least one of the polypeptide represented by SEQ ID NO. 6 and the polypeptide represented by SEQ ID NO.

8.

12. A composition for diagnosing henpavirus infection or antibody possession according to claim 11, wherein the henpavirus is a Nipah virus or a Hendra virus.

13. A method for preventing or treating heniphavirus infection, comprising the step of administering a recombinant protein, excluding humans, to an animal comprising at least one of the polypeptide represented by SEQ ID NO. 6 and the polypeptide represented by SEQ ID NO. 8.

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