Construction and Application of a Fusion Protein Vaccine Platform

Abstract

The present invention relates to the construction and application of a fusion protein vaccine platform. The present invention provides a vaccine comprising a fusion protein containing an interferon, a target antigen, an immunoglobulin Fc region (or antibody), and a Th cell helper epitope. The present invention also relates to the use of a fusion protein containing an interferon, a target antigen, an immunoglobulin Fc region (or antibody), and a Th cell helper epitope for preparing a prophylactic or therapeutic composition. The vaccines of the present invention are produced using a eukaryotic cell expression system, and wild-type and various mutant antigen vaccines can be prepared. These vaccines can induce a strong immune response in the body by inoculation via immunization routes such as subcutaneous / muscle or intranasal. The vaccines of the present invention can be used as prophylactic or therapeutic vaccines.

Description

Technical Field

[0001] The present invention belongs to the fields of genetic engineering and biomedical technology, and specifically relates to a vaccine containing a fusion protein having interferon-target antigen-immunoglobulin Fc region (antibody) as a main backbone as a vaccine. The vaccine of the present invention is used as a vaccine platform for the prevention of hepatitis B virus (HBV) infection, the prevention of HPV, EBV, HIV, SARA-COV2, influenza virus infection, and the prevention of the occurrence of HPV, EBV-related tumors, as well as the treatment of chronic hepatitis B (CHB) infection and the treatment of HBV, HP, EBV-related tumors.

Background Art

[0002] Worldwide, approximately 257 million people are chronically infected with the virus, and approximately 88,700 people die each year from end-stage liver diseases caused by HBV such as liver failure, cirrhosis, and hepatocellular carcinoma [1-3] . Approximately 30% of cirrhotic patients are caused by HBV, and approximately 40% of hepatocellular carcinoma (HCC) is caused by HBV. Hepatitis B virus infection remains a major public health problem worldwide [4] . However, there is still no effective treatment for chronic hepatitis B. Existing HBV treatments mainly include antiviral drugs (nucleoside / nucleotide analogs) and interferon. Although these have a certain therapeutic effect, they usually cannot induce an effective immune response, so HBV infection cannot be completely removed. And the side effects caused by long-term medication are relatively large, and antiviral drugs may also cause drug resistance. Chronic HBV infection is one of the major diseases threatening human health, and it is urgent to explore an effective immunotherapy strategy for chronic hepatitis B. The development of a therapeutic vaccine for chronic hepatitis B is of great social and economic importance.

[0003] Seasonal influenza causes serious illness in 1 million to 4 million people every year, and 200,000 to 500,000 people die [5]Vaccines are the best way to prevent and control influenza. Vaccines can reduce the incidence of disease and mitigate the severity of infections, especially in young children and the elderly who are at risk of complications from influenza. Currently approved influenza vaccines provide sufficient protection for healthy young adults, but there are still some issues that need to be addressed. For example, the production pathways of some vaccines, such as inactivated influenza vaccines and live attenuated influenza vaccines, rely on chicken embryos. The drawback of these vaccines is that when the prevalent strains are avian-derived, an influenza epidemic may increase the demand for both the vaccine and chicken embryos, potentially causing problems in the supply of chicken embryos. [6] Another drawback is that the production of these vaccines takes a long time. The elderly are prone to severe syndromes caused by the influenza virus. At the same time, standard vaccines are generally less effective in the elderly, and the immune systems of the elderly gradually weaken with age. [7] In view of the problems with current influenza vaccines, in response to the influenza virus epidemic, there is an urgent need to develop influenza vaccines with high immunogenicity, independent of chicken embryos, and can be manufactured quickly.

[0004] Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the pathogen that caused the COVID-19 pandemic in 2019. The clinical symptoms caused by SARS-CoV-2 are mainly asymptomatic infection, mild influenza-like symptoms, pneumonia, and severe acute respiratory distress syndrome. In severe cases, infected patients may die. [8] Currently, there is no specific drug for the novel coronavirus, and vaccines are the basic countermeasure to control and end the novel coronavirus pandemic. [9] In addition, the emergence of novel coronavirus variants poses new challenges to existing candidate vaccines and epidemic suppression.

[10] Therefore, under the current epidemic situation, there is an urgent need for vaccines that are powerful and can also act against novel coronavirus variants.

[0005] When an antigen is linked to the Fc region of an immunoglobulin, the half-life of the antigen is significantly extended, and since the Fc region of the immunoglobulin can bind to the Fc receptor on the surface of antigen-presenting cells, it becomes easier for antigen-presenting cells to process and present the antigen. [11-13] Type I interferons have many biological activities as antiviral cytokines, one of which is the stimulatory effect on immune cells.

[14] IFNα can strongly induce the differentiation and activation of human DC cells.

[15] After type I interferons act on immature DCs, they promote the expression of MHC molecules and co-stimulatory molecules on the surface of DCs such as MHC class I, CD80, and CD86, thereby enhancing the ability of DCs to activate T cells. [16-18] It has been reported that type I interferons can promote the antigen-presenting ability of DCs after infection with vaccinia virus and lymphocytic choriomeningitis virus (LCMV). [19-21] In addition, after acting on DCs, type I interferons can promote the migration of DCs to lymph nodes by upregulating the expression of chemokine receptors, thereby promoting the activation of T cells. [22、23] Recently, more and more studies have shown that type I interferons can be used as immune adjuvants. In the study by Le Bon et al., when mice were immunized with a weak immunogen, type I interferons showed a strong immune adjuvant effect in mice, inducing long-term antibodies and immune memory.

[24] Also, it was found that the main cell population on which type I interferons act is DC cells. At the same time, using antibodies, vaccines can be targeted and delivered to DCs to stimulate the activation and cross-presentation function of DCs, further enhancing the activity and efficacy of the vaccines.

[0006] There is a need to provide a vaccine platform that enhances the body's response to viral, bacterial, or tumor antigens in the present invention.

Summary of the Invention

[0007] Vaccines are an effective way to prevent and control the spread of infectious diseases. There are various types of vaccines, one of which is the protein subunit vaccine. Generally, simple protein subunit vaccines often have low immunogenicity, which frequently limits their use. Therefore, a general-purpose protein subunit vaccine platform is urgently needed. Based on the effects of immunoglobulin Fc region and type I interferon on the immune system, the inventors specifically proposed an interferon α-virus antigen, bacteria or tumor-immunoglobulin Fc region fusion protein vaccine platform to enhance the body's response to viral, bacterial or tumor antigens. The present invention provides a type I interferon-protein antigen-immunoglobulin Fc vaccine platform. Type I interferon acts on antigen-presenting cells to mature and migrate them, enabling them to more appropriately play the role of antigen presentation and T cell activation. On the other hand, the Fc portion of the vaccine platform binds to Fc receptors on the surface of antigen-presenting cells, promoting the uptake of antigens by antigen-presenting cells and further assisting the function of antigen-presenting cells. The inventors proposed the fusion of Th cell helper epitopes, which can further improve the immune response effect of the type I interferon-protein antigen-immunoglobulin Fc vaccine and is an important component of the vaccine. The inventors proposed replacing Fc with antibodies such as anti-PD-L1 to target and deliver the vaccine to DCs, stimulating the activation and cross-presentation function of DCs and further enhancing the activity and efficacy of the vaccine. The present invention can be used as a novel vaccine platform for the prevention and treatment of diseases such as viral infections, bacterial infections or tumors.

[0008] In some embodiments, the present invention provides a vaccine comprising a fusion protein (with Th epitope addition) containing interferon-target antigen-immunoglobulin Fc region (or antibody). In some embodiments, the present invention further provides the use of a fusion protein (with Th epitope addition) containing interferon-target antigen-immunoglobulin Fc region (or antibody) for preparing a prophylactic or therapeutic composition or kit (e.g., a drug or vaccine composition or kit). The vaccine of the present invention is produced by a eukaryotic cell expression system and can be inoculated by an immunization route such as subcutaneous / muscular or nasal. In the fusion polypeptide of the present invention, the antibody (abbreviated as Ab) as one of its structural units is not particularly limited and may include, for example, a complete antibody, an antibody fragment such as a heavy chain or a light chain of an antibody, or a single-chain antibody, or may be a DC-target activating antibody containing an antibody such as anti-PD-L1, anti-DEC205, anti-CD80 / 86, etc.

[0009] In some embodiments, the target antigen described herein is not particularly limited and can be any suitable antigen. In some embodiments, the target antigen described herein can be, for example, a tumor antigen and / or a pathogen antigen (e.g., a viral or bacterial antigen). In some embodiments, the target antigen described herein can be, for example, a tumor antigen, such as a tumor cell highly expressed protein molecule, such as human epidermal growth factor receptor 2 (HER2 / neu), epidermal growth factor receptor (EGFR).

[0010] In some embodiments, the target antigen used in the vaccines provided herein can be, for example, a mutant target antigen that is different from the wild type. In some embodiments, the target antigens described herein can be, for example, variants of tumor antigens and / or pathogen antigens (e.g., viral or bacterial antigens). In some embodiments, the target antigen can be, for example, the full-length SARS-COV-2 virus S protein or the S1 region, and for example, the target antigen can be the antigen represented by SEQ ID NO: 76 or SEQ ID NO: 77. As used herein, the wild-type target antigen refers to a protein having immunogenicity expressed by a virus or other infectious agent or tumor encoded by a wild-type gene (the wild-type gene refers to the dominant allele in nature and is often used as a standard control gene in biological experiments), for example, the Spike protein (S protein) derived from the original wild-type strain of SARS-CoV-2. As used herein, the mutant target antigen (variant) refers to a mutant viral protein expressed by a mutant viral strain encoded by a mutant gene mutated from a wild-type gene. For example, the point mutations found in different mutant SARS-CoV-2 S proteins include deletions of 69-70 in the NTD region, deletion of Y144, deletions of 242-244, L18F, D80A, D215, R246I mutations, mutations such as K417, E484, N501Y in the RBD region, L452R mutation, T478K mutation, D614G, H655Y mutation. For example, these point mutations are present in various combinations in mutant SARS-CoV-2s derived from, for example, the B.1.1.7 (Alpha) mutant strain in the UK, the B.1.351 (Beta) mutant strain in South Africa, the P1 (Gamma) mutant strain in Brazil, B.1.617, B.1.617.1 (Kappa), B.1.617.2 (Delta), B.1.617.3 mutant strains in India, the California B.1.429 mutant strain, and the like.In some embodiments, the mutant target antigen may include, for example, natural point mutations / deletion mutations / insertion mutations / truncations, artificial point mutations / deletion mutations / insertion mutations / truncations, any combination of natural or artificial mutations, and subtypes generated after mutation, where the target antigen can be a tumor antigen, a pathogen antigen, such as a viral antigen (e.g., SARS-COV-2) or a bacterial antigen. In some embodiments, the target antigen used in the vaccine provided by the present invention is a mutant viral antigen. For example, the mutant viral antigen may be a variant of SARS-COV-2. For example, the mutant viral antigen may include natural point mutations / deletion mutations / insertion mutations / truncations of at least one selected from the group consisting of SARS-COV-2 proteins (e.g., S protein, N protein, M protein, E protein), artificial point mutations / deletion mutations / insertion mutations / truncations, any combination of natural or artificial mutations, and subtypes generated after mutation. For example, the mutant viral antigen may be a variant of the full-length S protein, the S1 region, or the RBD region. For example, the mutant viral antigen may include at least one mutation selected from the group consisting of NTD region 69-70 deletion, Y144 deletion, 242-244 deletion, L18F, D80A, D215, R246I mutations, K417, E484, N501Y mutations, L452R mutation, T478K mutation, D614G, H655Y mutations in the RBD region of the S protein of SARS-COV-2. For example, the mutant viral antigen may include mutations derived from the B.1.1.7 (Alpha) variant strain in the UK, the B.1.351 (Beta) variant strain in South Africa, the P1 (Gamma) variant strain in Brazil, B.1.617, B.1.617.1 (Kappa), B.1.617.2 (Delta), B.1.617.3 variant strains in India, and the B.1.429 variant strain in California. For example, the mutant viral antigen may include a variant of the mutation represented by any one of SEQ ID NO: 79, SEQ ID NO: 80, and SEQ ID NO: 81. For example, the mutant viral antigen may include a variant of the sequence represented by any one of SEQ ID NO: 79, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 82, SEQ ID NO: 83, and SEQ ID NO: 84.Unless otherwise specified in this specification or clearly limited by the context, the target antigens referred to in this specification generally include wild-type target antigens and mutant target antigens.

[0011] The present invention aims to provide a (Th epitope-added) vaccine platform consisting of interferon (IFN) and a tumor, bacterial or viral antigen (hepatitis B virus Pres1 antigen, SARS-COV2 RBD antigen, influenza HA antigen, human papillomavirus HPV E7 antigen, hepatitis B virus surface antigen (HBsAg) antigen or peptide, varicella-zoster virus (VZV) gE antigen, Epstein-Barr virus (EBV) EBNA1 / LMP2 / gp350, herpes simplex virus 2 (HSV-2) gD antigen, human immunodeficiency virus (HIV) gp120 antigen) and an immunoglobulin Fc region (or antibody). The fusion protein may be a homodimeric protein or a heterodimeric protein. When the fusion protein is a dimer, interferon, the target antigen, and the immunoglobulin Fc region (or antibody Ab) as structural units may be present in the first polypeptide chain and / or the second polypeptide chain. The existence form of each structural unit is not particularly limited. For example, they may be present simultaneously in one chain, or any one or more structural units may be present in one chain, and another one or more structural units may be present in another chain.

[0012] The interferon described in the present invention can be selected from the group consisting of type I interferon, type II interferon, type III interferon, for example, IFN-α, IFN-β, IFN-γ, IFN-λ1 (IL-29), IFN-λ2 (IL-28a), IFN-λ (IL-28b), and IFN-ω. The IFN may be of human origin or murine origin. The interferon described in the present invention is preferably IFN-α (SEQ ID NO: 1, SEQ ID NO: 21, SEQ ID NO: 22) as type I interferon.

[0013] The Fc region of the immunoglobulin described in the present invention can be selected from the group consisting of the constant region amino acid sequences of IgG1, IgG2, IgG3, IgG4 and / or IgM, and preferably is IgG1 (SEQ ID NO: 2, SEQ ID NO: 23, SEQ ID NO: 24).

[0014] In addition, the fusion polypeptide of the present invention may optionally contain one or more Th cell helper epitopes and / or a linking fragment (linker). For example, when the fusion protein is a dimer, optionally, the fusion protein may also contain one or more Th cell helper epitopes and / or a linking fragment in any one or two chains of the homodimer or heterodimer (i.e., the first polypeptide chain and / or the second polypeptide chain). As known to those skilled in the art, between each structural unit of the fusion protein, it can be linked by an appropriate linking fragment (linker). The linking fragment that can be used in the vaccine of the present invention is not particularly limited and can be any appropriate peptide fragment known in the art. The linking fragment of each structural unit described in the present invention may be a flexible polypeptide sequence, and may be those represented by the amino acid sequences of linking fragments 1 and 2, for example, SEQ ID NO: 4, SEQ ID NO: 25.

[0015] At the N-terminus of the polypeptide sequence composed of each structural unit described in the present invention, there is included a corresponding signal peptide that can promote protein secretion, for example, that represented by the amino acid sequence of SEQ ID NO: 5.

[0016] Preferred antigens according to the present invention include hepatitis B Pres1 antigens including ad subtype (SEQ ID NO: 6), ay subtype (SEQ ID NO: 26), adr subtype (SEQ ID NO: 7), adw subtype (SEQ ID NO: 27), ayw subtype (SEQ ID NO: 28) (each subtype and peptide), SARA-COV2 RBD antigen (SEQ ID NO: 8), influenza virus HA antigen (SEQ ID NO: 9), HPV E7 antigen (SEQ ID NO: 10), herpesvirus VZV-gE antigen (SEQ ID NO: 91), EBV-gp350 antigen (SEQ ID NO: 92), HSV-2-gD antigen (SEQ ID NO: 93).

[0017] The homodimeric protein according to the present invention includes a first polypeptide and a second polypeptide, and the first polypeptide and the second polypeptide are completely identical. The first polypeptide and the second polypeptide are, in order from the N-terminus to the C-terminus, IFN-tumor or viral antigen (hepatitis B Pres1 antigen, SARS-COV2 RBD antigen, influenza HA antigen, HPV E7 antigen, HBsAg antigen, VZV-gE antigen, EBV EBNA1 / LMP2 / gp350, HSV-2-gD antigen, HIV gp120 antigen)-immunoglobulin Fc region, or a Pan epitope-containing polypeptide, and include the amino acid sequences shown in SEQ ID NO: 11, 12, 13, 14, 29, 30, 31, 32, 38, 39, 40, 47, 48, 49, 50, 51, 56, 57, 59, 58, 65, 66, 67, 68.

[0018] The heterodimer described in the present invention comprises a first polypeptide and a second polypeptide, wherein the first polypeptide and the second polypeptide are not the same polypeptide. The first polypeptide is, in order from the C-terminus to the N-terminus, an IFN-immunoglobulin Fc region respectively, and comprises the amino acid sequences shown by SEQ ID NO: 15, 33, 42, 51, 60, 69. The second polypeptide is, in order from the C-terminus to the N-terminus, a tumor or viral antigen (Hepatitis B Pres1 antigen, SARS-COV2 RBD antigen, Influenza HA antigen, HPV E7 antigen, VZV-gE antigen, EBV EBNA1 / LMP2 / gp350, HSV-2-gD antigen, HIV gp120 antigen)-immunoglobulin Fc region respectively, and comprises the amino acid sequences shown by SEQ ID NO: 16, 17, 18, 19, 34, 35, 36, 37, 43, 44, 45, 46, 52, 53, 54, 55, 61, 62, 63, 64, 70, 71, 72, 73.

[0019] Furthermore, the present invention provides an amino acid sequence encoding the above IFN-tumor or viral antigen (Hepatitis B Pres1 antigen, HBsAg antigen or peptide, SARS-COV2 RBD antigen, Influenza HA antigen, HPV E7 antigen, VZV-gE antigen, EBV EBNA1 / LMP2 / gp350, HSV-2-gD antigen, HIV gp120 antigen)-immunoglobulin Fc vaccine platform.

[0020] Furthermore, the present invention relates to a nucleotide segment encoding the above vaccine platform and fusion protein.

[0021] Furthermore, the present invention relates to a method for preparing the above fusion protein or vaccine platform. For example, the preparation method (1) constructing an expression vector containing a coding gene encoding the fusion protein or vaccine platform, preferably, the expression vector is a pEE12.4 expression vector; (2) Constructing a host cell containing the expression vector by transiently transfecting a host cell, preferably, the host cell is a 293F cell; (3) Culturing the host cell and collecting the cell supernatant; (4) Purifying the fusion protein or vaccine platform by purifying the protein with a Protein A / G affinity chromatography column; The method includes the above steps.

[0022] In addition, the present invention includes the application of the vaccine platform, and the vaccine platform can be used as a hepatitis B preventive vaccine, the vaccine platform can be used as a hepatitis B therapeutic vaccine, the vaccine platform can be used as an influenza preventive vaccine, the vaccine platform can be used as a SARA-COV2, influenza, HPV, VZV, EBV, HSV-2, HIV preventive vaccine, and the vaccine platform can be used as an HPV, EBV-related tumor preventive vaccine.

[0023] The present invention includes an adjuvant used by the vaccine platform, and the adjuvant includes an aluminum adjuvant (Alum), a Toll-like receptor 4 activator ligand MPLA, a Toll-like receptor 9 ligand, M59, an oligodeoxynucleotide (CpG-ODN), and a Freund's adjuvant.

[0024] The present invention includes the clinical use of the combination of the vaccine platform as an HBV therapeutic vaccine and the hepatitis B virus envelope protein HBsAg vaccine in the treatment of chronic hepatitis B virus infection.

[0025] The present invention includes the clinical use of the combination of the vaccine platform as an HBV therapeutic vaccine and a nucleoside or nucleotide analog in the treatment of chronic hepatitis B virus infection.

[0026] The present invention includes the combined use of the vaccine platform as an HBV, influenza, SARA-COV2, HPV, VZV, EBV, HSV-2, HIV preventive or therapeutic vaccine, etc., and antiviral drugs and other therapies, and also includes the combined use of the vaccine platform as an HBV, HPV, EBV-related tumor preventive or therapeutic vaccine and antiviral, tumor drugs and therapies.

[0027] The present invention includes a multivalent mixed vaccine composed of the vaccine platform as a component of a vaccine and other viral or pathogen or tumor vaccines.

[0028] The present invention includes any fusion protein vaccine of the vaccine platform immunized by sequential or simultaneous immunization procedures with an adenovirus vaccine or mRNA vaccine or inactivated vaccine or DNA vaccine of the same virus, pathogen, or tumor.

[0029] The present invention includes the full-length sequences and any truncated sequences of the vaccine platform antigen, such as SEQ ID NO: 76, SEQ ID NO: 77, SEQ ID NO: 78.

[0030] The present invention includes any possible variants of the fusion protein vaccine antigen, including natural point mutations / deletion mutations / truncates, any combination of natural point mutations, subtypes generated after mutation, and mutant sequences such as artificial point mutations / deletion mutations / truncates constructed by the inventor to enhance the vaccine effect, such as SEQ ID NO: 79, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 82, SEQ ID NO: 83, SEQ ID NO: 84.

[0031] The present invention includes a multivalent mixed vaccine composed of any vaccine of the present invention as a component of a vaccine and another vaccine of the present invention or another vaccine different from the vaccine of the present invention, for example, another virus or pathogen or tumor vaccine. For example, a multivalent vaccine in which the SARS-CoV-2 fusion protein vaccine of the present invention is combined with an influenza vaccine or another vaccine. For example, any vaccine of the present invention is immunized with an adenovirus vaccine or mRNA vaccine or inactivated vaccine or DNA vaccine of the same virus, pathogen, or tumor in a sequential or simultaneous immunization procedure. For example, the SARS-CoV-2 fusion protein vaccine is immunized with an adenovirus vaccine or mRNA vaccine or inactivated vaccine or DNA vaccine of SARS-CoV-2 in a sequential or simultaneous immunization procedure. For example, the order of immunization can be: 1) First immunize with the SARS-CoV-2 fusion protein vaccine of the present invention, and then immunize with an adenovirus vaccine or mRNA vaccine or inactivated vaccine or DNA vaccine of SARS-CoV-2; 2) First immunize with an adenovirus vaccine or mRNA vaccine or inactivated vaccine or DNA vaccine of SARS-CoV-2, and then immunize with the SARS-CoV-2 fusion protein vaccine; 3) Immunize with the SARS-CoV-2 fusion protein vaccine and an adenovirus vaccine or mRNA vaccine or inactivated vaccine or DNA vaccine of SARS-CoV-2 simultaneously. As is known in the art, when used in combination, the vaccines for combined use can be prepared as a convenient kit.

[0032] Compared with the prior art, the present invention includes, but is not limited to, the following beneficial effects.

[0033] 1. In the IFN-tumor or virus antigen-immunoglobulin Fc (or antibody) vaccine platform provided by the present invention, the antigen can vary in multiple components, and can be a tumor-associated antigen or a virus-specific antigen, thereby improving the flexibility of the use of the vaccine platform and also expanding the scope of use of this vaccine platform.

[0034] 2. In the IFN-tumor or virus antigen-immunoglobulin Fc (or antibody) vaccine platform provided by the present invention, among them, interferon (IFN) promotes the migration and maturation of antigen-presenting cells, increases the expression of co-stimulatory molecules, thereby being beneficial for antigen presentation to T cells. At the same time, the Fc region (or antibody) of the vaccine platform, on the one hand, increases the molecular weight of the antigen and its half-life, and on the other hand, the Fc region (or antibody) binds to the Fc receptor on the surface of antigen-presenting cells, promoting the processing and presentation of antigens by antigen-presenting cells, thereby being beneficial for the generation of immune responses.

[0035] 3. The IFN-tumor or virus antigen-immunoglobulin Fc (or antibody) vaccine platform provided by the present invention is expressed by a eukaryotic HEK293 cell expression system, and the protein expressed by HEK293 cells is close to natural protein molecules in terms of molecular structure, physical and chemical properties, protein modification, and protein biological functions.

[0036] 4. The IFN-tumor or virus antigen-immunoglobulin Fc (or antibody) vaccine platform provided by the present invention has two structures, homodimer or heterodimer, and has better selectivity for different antigens.

[0037] 5. The IFN-tumor or virus antigen-immunoglobulin Fc vaccine platform provided by the present invention fuses Th cell helper epitopes, such as Pan epitopes, utilizes DC-targeting antibodies such as anti-PD-L1, and adds various adjuvants that stimulate the immune response, thereby activating DCs and enhancing DC cross-presentation, and can produce a strong B cell and T cell immune response.

[0038] 6. The IFN-tumor or virus antigen-immunoglobulin Fc (or antibody) vaccine platform provided by the present invention has a wide range of applications and can be used not only as a preventive vaccine but also as a therapeutic vaccine.

[0039] 7. The IFN-tumor or virus antigen-immunoglobulin Fc (or antibody) vaccine platform provided by the present invention can not only be used alone but also be used as a therapeutic vaccine in combination with existing commercially available HBsAg vaccines and nucleoside / nucleotide analogs.

[0040] 8. The vaccine provided by the present invention can form a multivalent mixed vaccine with other viruses or pathogens or tumor vaccines as one component of the vaccine.

[0041] 9. Any fusion protein vaccine in the vaccine platform provided by the present invention can be immunized with an adenovirus vaccine or mRNA vaccine or inactivated vaccine or DNA vaccine of the same virus, pathogen, or tumor in a sequential or simultaneous immunization procedure.

[0042] 10. The present invention provides the full-length sequence and any truncated sequences of the vaccine platform antigen.

[0043] 11. The present invention provides any possible mutants of a vaccine platform antigen, including natural point mutations / deletion mutations / insertion mutations / truncations, any combination of natural point mutations, subtypes generated after mutation, and mutant sequences such as artificial point mutations / deletion mutations / insertion mutations / truncations constructed by the inventor to enhance the vaccine effect.

[0044] The information on the sequences related to the present invention is shown below. 1. Unit constituent sequence: SEQ ID NO: 1: Mouse mIFNα4 amino acid sequence (mIFNα) CDLPHTYNLGNKRALTVLEEMRRLPPLSCLKDRKDFGFPLEKVDNQQIQKAQAILVLRDLTQQILNLFTSKDLSATWNATLLDSFCNDLHQQLNDLKACVMQEPPLTQEDSLLAVRTYFHRITVYLRKKKHSLCAWEVIRAEVWRALSSSTNLLARLSEEKE SEQ ID NO: 21: Human IFNα2 amino acid sequence (hIFNα) CDLPQTHSLGSRRTLMLLAQMRRISLFSCLKDRHDFGFPQEEFGNQFQKAETIPVLHEMIQQIFNLFSTKDSSAAWDETLLDKFYTELYQQLNDLEACVIQGVGVTETPLMKEDSILAVRKYFQRITLYLKEKKYSPCAWEVVRAEIMRSFSLSTNLQESLRSKE SEQ ID.NO.22: Human mutant IFNα2 (Q124R) amino acid sequence (hmIFNα) CDLPQTHSLGSRRTLMLLAQMRRISLFSCLKDRHDFGFPQEEFGNQFQKAETIPVLHEMIQQIFNLFSTKDSSAAWDETLLDKFYTELYQQLNDLEACVIQGVGVTETPLMKEDSILAVRKYFRRITLYLKEKKYSPCAWEVVRAEIMRSFSLSTNLQESLRSKE SEQ ID NO: 2: Human IgG1-Fc amino acid sequence EPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGKHV SEQ ID NO: 23: Heterodimer Fc-hole DKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 24: Heterodimer Fc-knob DKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 3: Th Helper Epitope Pan HLA DR-binding epitope (PADER) Amino Acid Sequence AKFVAAWTLKAAA SEQ ID NO: 4: Linker 1 Amino Acid Sequence: GGGGSGGGGSGGGGS SEQ ID NO: 25: Amino acid sequence of Linker 2 GSGSGS SEQ ID NO: 5: Amino acid sequence of signal peptide MARLCAFLMILVMMSYYWSACSLG SEQ ID NO: 6: Amino acid sequence of HBV Pres1 (ad subtype) MGGWSSKPRKGMGTNLSVPNPLGFFPDHQLDPAFGANSNNPDWDFNPIKDHWPAANQVGVGAFGPGLTPPHGGILGWSPQAQGILTTVSTIPPPASTNRQSGRQPTPISPPLRDSHPQA SEQ ID NO: 26: Amino acid sequence of HBV Pres1 (ay subtype) MGQNLSTSNPLGFFPDHQLDPAFRANTANPDWDFNPNKDTWPDANKVGAGAFGLGFTPPHGGLLGWSPQAQGILQTLPANPPPASTNRQTGRQPTPLSPPLRNTHPQA SEQ ID NO: 7: Amino acid sequence of HBV HBsAg (adr subtype) MENTTSGFLGPLLVLQAGFFLLTRILTIPQSLDSWWTSLNFLGGAPTCPGQNSQSPTSNHSPTSCPPICPGYRWMCLRRFIIFLFILLLCLIFLLVLLDYQGMLPVCPLLPGTSTTSTGPCKTCTIPAQGTSMFPSCCCTKPSDGNCTCIPIPSSWAFARFLWEWASVRFSWLSLLVPFVQWFVGLSPTVWLSVIWMMWYWGPSLYNILSPFLPLLPIFFCLWVYI SEQ ID NO: 27: Amino acid sequence of HBV HBsAg (adw subtype) MENITSGLLGPLLVLQAGFFLLTRILTIPQSLDSWWTSLSFLGEAPVCLGQNSQSPTRNHSPTSCPPICPGYRWMCLRRFIIFLFILLLCLIFLLVLLDYQGMLPVCPLIPGSTTTSTGPCKTCTTPAQGNSMFPSCCCTKPTDGNCTCIPIPSSWAFAKYLWEWASVRFSWLSLLVPFVQWFVGLSPTVWLSAIWMIWYWGPSLYSIVCPFTPLLQIFCCLWVFI SEQ ID NO: 28: Amino acid sequence of HBV HBsAg (ayw subtype) MENITSGFLGPLLVLQAGFFLLTRILTIPQSLDSWWTSLNFLGGTTVCLGQSSQSPTSNHSPTSCPPTCPGYRWMCLRRFIIFLFILLLCLIFLLVLLDYQGMLPVCPLIPGSSTTSTGPCRTCMTTAQGTSMYPSCCCTKPSDGNCTCIPIPSSWAFGKFLWEWASARFSWLSLLVPFVQWFVGLSPTVWLSVIWMMWYWGPSLYSILSPFLPLLPIFFCLWVYI SEQ ID NO: 8: Amino acid sequence of SARS-CoV-2 RBD RVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNF SEQ ID NO: 9: Amino acid sequence of influenza virus HA DTICIGYHANNSTDTVDTVLEKNVTVTHSVNLLEDSHNGKLCRLKGIAPLQLGKCNIAGWLLGNPECDPLLPVRSWSYIVETPNSENGICYPGDFIDYEELREQLSSVSSFERFEIFPKESSWPNHNTNGVTAACSHEGKSSFYRNLLWLTEKEGSYPKLKNSYVNKKGKEVLVLWGIHHPPNSKEQQNLYQNENAYVSVVTSNYNRRFTPEIAERPKVRDQAGRMNYYWTLLKPGDTIIFEANGNLIAPMYAFALSRGFGSGIITSNASMHECNTKCQTPLGAINSSLPYQNIHPVTIGECPKYVRSAKLRMVTGLRNNPSIQSR SEQ ID NO: 10: Amino acid sequence of HPV-E7 antigen MHGDTPTLHEYMLDLQPETTDLYCYEQLNDSSEEEDEIDGPAGQAEPDRAHYNIVTFCCKCDSTLRLCVQSTHVDIRTLEDLLMGTLGIVCPICSQKP 2. Mouse-derived IFN vaccine mIFNα-antigen-Fc sequence: SEQ ID NO: 11: Amino acid sequence of mIFNα-Pres1-Fc in homodimer CDLPHTYNLGNKRALTVLEEMRRLPPLSCLKDRKDFGFPLEKVDNQQIQKAQAILVLRDLTQQILNLFTSKDLSATWNATLLDSFCNDLHQQLNDLKACVMQEPPLTQEDSLLAVRTYFHRITVYLRKKKHSLCAWEVIRAEVWRALSSSTNLLARLSEEKESGGGGSGGGGSGGGGSGGGGRTMGQNLSTSNPLGFFPDHQLDPAFRANTANPDWDFNPNKDTWPDANKVGAGAFGLGFTPPHGGLLGWSPQAQGILQTLPANPPPASTNRQTGRQPTPLSPPLRNTHPQAFEEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGKHV SEQ ID NO: 12: Amino acid sequence of mIFNα-RBD(SARS-CoV-2)-Fc in homodimer CDLPHTYNLGNKRALTVLEEMRRLPPLSCLKDRKDFGFPLEKVDNQQIQKAQAILVLRDLTQQILNLFTSKDLSATWNATLLDSFCNDLHQQLNDLKACVMQEPPLTQEDSLLAVRTYFHRITVYLRKKKHSLCAWEVIRAEVWRALSSSTNLLARLSEEKESGGGGSGGGGSGGGGSGGGGRTRVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFFEEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGKHV SEQ ID NO: 13: Amino acid sequence of mIFNα-HA-Fc in a homodimer CDLPHTYNLGNKRALTVLEEMRRLPPLSCLKDRKDFGFPLEKVDNQQIQKAQAILVLRDLTQQILNLFTSKDLSATWNATLLDSFCNDLHQQLNDLKACVMQEPPLTQEDSLLAVRTYFHRITVYLRKKKHSLCAWEVIRAEVWRALSSSTNLLARLSEEKESGGGGSGGGGSGGGGSGGGGRTDTICIGYHANNSTDTVDTVLEKNVTVTHSVNLLEDSHNGKLCRLKGIAPLQLGKCNIAGWLLGNPECDPLLPVRSWSYIVETPNSENGICYPGDFIDYEELREQLSSVSSFERFEIFPKESSWPNHNTNGVTAACSHEGKSSFYRNLLWLTEKEGSYPKLKNSYVNKKGKEVLVLWGIHHPPNSKEQQNLYQNENAYVSVVTSNYNRRFTPEIAERPKVRDQAGRMNYYWTLLKPGDTIIFEANGNLIAPMYAFALSRGFGSGIITSNASMHECNTKCQTPLGAINSSLPYQNIHPVTIGECPKYVRSAKLRMVTGLRNNPSIQSRFEEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGKHV SEQ ID NO: 14: mIFNα-E7(HPV)-Fc amino acid sequence in homodimer CDLPHTYNLGNKRALTVLEEMRRLPPLSCLKDRKDFGFPLEKVDNQQIQKAQAILVLRDLTQQILNLFTSKDLSATWNATLLDSFCNDLHQQLNDLKACVMQEPPLTQEDSLLAVRTYFHRITVYLRKKKHSLCAWEVIRAEVWRALSSSTNLLARLSEEKESGGGGSGGGGSGGGGSGGGGRTMHGDTPTLHEYMLDLQPETTDLYCYEQLNDSSEEEDEIDGPAGQAEPDRAHYNIVTFCCKCDSTLRLCVQSTHVDIRTLEDLLMGTLGIVCPICSQKPFEEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGKHV SEQ ID NO: 15: Amino acid sequence of the first chain mIFNα-Fc-hole in a heterodimer CDLPHTYNLGNKRALTVLEEMRRLPPLSCLKDRKDFGFPLEKVDNQQIQKAQAILVLRDLTQQILNLFTSKDLSATWNATLLDSFCNDLHQQLNDLKACVMQEPPLTQEDSLLAVRTYFHRITVYLRKKKHSLCAWEVIRAEVWRALSSSTNLLARLSEEKESGGGGSGGGGSGGGGSGGGGRTDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 16: Amino acid sequence of the second chain Pres1-Fc-knob in the heterodimer mIFNα-Pres1-Fc MGQNLSTSNPLGFFPDHQLDPAFRANTANPDWDFNPNKDTWPDANKVGAGAFGLGFTPPHGGLLGWSPQAQGILQTLPANPPPASTNRQTGRQPTPLSPPLRNTHPQAFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 17: Amino acid sequence of the second chain RBD(SARS-CoV-2)-Fc-knob in the heterodimer mIFNα-RBD(SARS-CoV-2)-Fc RVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 18: Amino acid sequence of the second chain HA-Fc-knob in the heterodimer mIFNα-HA-Fc DTICIGYHANNSTDTVDTVLEKNVTVTHSVNLLEDSHNGKLCRLKGIAPLQLGKCNIAGWLLGNPECDPLLPVRSWSYIVETPNSENGICYPGDFIDYEELREQLSSVSSFERFEIFPKESSWPNHNTNGVTAACSHEGKSSFYRNLLWLTEKEGSYPKLKNSYVNKKGKEVLVLWGIHHPPNSKEQQNLYQNENAYVSVVTSNYNRRFTPEIAERPKVRDQAGRMNYYWTLLKPGDTIIFEANGNLIAPMYAFALSRGFGSGIITSNASMHECNTKCQTPLGAINSSLPYQNIHPVTIGECPKYVRSAKLRMVTGLRNNPSIQSRFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 19: Amino acid sequence of the second chain E7-Fc-knob in the heterodimer mIFNα-E7(HPV)-Fc MHGDTPTLHEYMLDLQPETTDLYCYEQLNDSSEEEDEIDGPAGQAEPDRAHYNIVTFCCKCDSTLRLCVQSTHVDIRTLEDLLMGTLGIVCPICSQKPFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK 3. Vaccine IFNα-Pan-antigen-Fc sequence containing the Pan epitope of murine-derived IFN SEQ ID NO: 29: mIFNα-Pan-Pres1-Fc amino acid sequence in a homodimer CDLPHTYNLGNKRALTVLEEMRRLPPLSCLKDRKDFGFPLEKVDNQQIQKAQAILVLRDLTQQILNLFTSKDLSATWNATLLDSFCNDLHQQLNDLKACVMQEPPLTQEDSLLAVRTYFHRITVYLRKKKHSLCAWEVIRAEVWRALSSSTNLLARLSEEKEGGGGSGGGGSGGGGSRTAKFVAAWTLKAAAGSGSGSMGQNLSTSNPLGFFPDHQLDPAFRANTANPDWDFNPNKDTWPDANKVGAGAFGLGFTPPHGGLLGWSPQAQGILQTLPANPPPASTNRQTGRQPTPLSPPLRNTHPQAFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGK SEQ ID NO: 30: Amino acid sequence of mIFNα-Pan-RBD(SARS-CoV-2)-Fc in homodimer CDLPHTYNLGNKRALTVLEEMRRLPPLSCLKDRKDFGFPLEKVDNQQIQKAQAILVLRDLTQQILNLFTSKDLSATWNATLLDSFCNDLHQQLNDLKACVMQEPPLTQEDSLLAVRTYFHRITVYLRKKKHSLCAWEVIRAEVWRALSSSTNLLARLSEEKEGGGGSGGGGSGGGGSRTAKFVAAWTLKAAAGSGSGSRVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGK SEQ ID NO: 31: Amino acid sequence of mIFNα-Pan-HA-Fc in homodimer CDLPHTYNLGNKRALTVLEEMRRLPPLSCLKDRKDFGFPLEKVDNQQIQKAQAILVLRDLTQQILNLFTSKDLSATWNATLLDSFCNDLHQQLNDLKACVMQEPPLTQEDSLLAVRTYFHRITVYLRKKKHSLCAWEVIRAEVWRALSSSTNLLARLSEEKEGGGGSGGGGSGGGGSRTAKFVAAWTLKAAAGSGSGSDTICIGYHANNSTDTVDTVLEKNVTVTHSVNLLEDSHNGKLCRLKGIAPLQLGKCNIAGWLLGNPECDPLLPVRSWSYIVETPNSENGICYPGDFIDYEELREQLSSVSSFERFEIFPKESSWPNHNTNGVTAACSHEGKSSFYRNLLWLTEKEGSYPKLKNSYVNKKGKEVLVLWGIHHPPNSKEQQNLYQNENAYVSVVTSNYNRRFTPEIAERPKVRDQAGRMNYYWTLLKPGDTIIFEANGNLIAPMYAFALSRGFGSGIITSNASMHECNTKCQTPLGAINSSLPYQNIHPVTIGECPKYVRSAKLRMVTGLRNNPSIQSRFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGK SEQ ID NO: 32: Amino acid sequence of mIFNα-Pan-E7(HPV)-Fc in a homodimer CDLPHTYNLGNKRALTVLEEMRRLPPLSCLKDRKDFGFPLEKVDNQQIQKAQAILVLRDLTQQILNLFTSKDLSATWNATLLDSFCNDLHQQLNDLKACVMQEPPLTQEDSLLAVRTYFHRITVYLRKKKHSLCAWEVIRAEVWRALSSSTNLLARLSEEKEGGGGSGGGGSGGGGSRTAKFVAAWTLKAAAGSGSGSMHGDTPTLHEYMLDLQPETTDLYCYEQLNDSSEEEDEIDGPAGQAEPDRAHYNIVTFCCKCDSTLRLCVQSTHVDIRTLEDLLMGTLGIVCPICSQKPFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGK SEQ ID NO: 33: Amino acid sequence of the first chain mIFNα-Fc-hole in a heterodimer CDLPHTYNLGNKRALTVLEEMRRLPPLSCLKDRKDFGFPLEKVDNQQIQKAQAILVLRDLTQQILNLFTSKDLSATWNATLLDSFCNDLHQQLNDLKACVMQEPPLTQEDSLLAVRTYFHRITVYLRKKKHSLCAWEVIRAEVWRALSSSTNLLARLSEEKESGGGGSGGGGSGGGGSGGGGRTDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 34: Amino acid sequence of the second chain Pan-Pres1-Fc-knob in the heterodimeric mIFN-Pan-Pres1-Fc AKFVAAWTLKAAAGSGSGSMGQNLSTSNPLGFFPDHQLDPAFRANTANPDWDFNPNKDTWPDANKVGAGAFGLGFTPPHGGLLGWSPQAQGILQTLPANPPPASTNRQTGRQPTPLSPPLRNTHPQAFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 35: Amino acid sequence of the second chain Pan-RBD(SARS-CoV-2)-Fc-knob in the heterodimeric mIFNα-Pan-RBD(SARS-CoV-2)-Fc AKFVAAWTLKAAAGSGSGSRVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 36: Amino acid sequence of the second chain Pan-HA-Fc-knob in the heterodimeric mIFNα-Pan-HA-Fc AKFVAAWTLKAAAGSGSGSDTICIGYHANNSTDTVDTVLEKNVTVTHSVNLLEDSHNGKLCRLKGIAPLQLGKCNIAGWLLGNPECDPLLPVRSWSYIVETPNSENGICYPGDFIDYEELREQLSSVSSFERFEIFPKESSWPNHNTNGVTAACSHEGKSSFYRNLLWLTEKEGSYPKLKNSYVNKKGKEVLVLWGIHHPPNSKEQQNLYQNENAYVSVVTSNYNRRFTPEIAERPKVRDQAGRMNYYWTLLKPGDTIIFEANGNLIAPMYAFALSRGFGSGIITSNASMHECNTKCQTPLGAINSSLPYQNIHPVTIGECPKYVRSAKLRMVTGLRNNPSIQSRFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 37: Amino acid sequence of the second chain Pan-E7-Fc-knob in the heterodimeric mIFNα-Pan-E7(HPV)-Fc AKFVAAWTLKAAAGSGSGSMHGDTPTLHEYMLDLQPETTDLYCYEQLNDSSEEEDEIDGPAGQAEPDRAHYNIVTFCCKCDSTLRLCVQSTHVDIRTLEDLLMGTLGIVCPICSQKPFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK 4. Human IFN Vaccine hIFNα - Antigen - Fc Sequence: SEQ ID NO: 38: hIFNα - Pres1 - Fc Amino Acid Sequence in Homodimer CDLPQTHSLGSRRTLMLLAQMRRISLFSCLKDRHDFGFPQEEFGNQFQKAETIPVLHEMIQQIFNLFSTKDSSAAWDETLLDKFYTELYQQLNDLEACVIQGVGVTETPLMKEDSILAVRKYFQRITLYLKEKKYSPCAWEVVRAEIMRSFSLSTNLQESLRSKEGGGGSGGGGSGGGGSRTMGQNLSTSNPLGFFPDHQLDPAFRANTANPDWDFNPNKDTWPDANKVGAGAFGLGFTPPHGGLLGWSPQAQGILQTLPANPPPASTNRQTGRQPTPLSPPLRNTHPQAFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGK SEQ ID NO: 39: Amino acid sequence of hIFNα-RBD(SARS-CoV-2)-Fc in homodimer CDLPQTHSLGSRRTLMLLAQMRRISLFSCLKDRHDFGFPQEEFGNQFQKAETIPVLHEMIQQIFNLFSTKDSSAAWDETLLDKFYTELYQQLNDLEACVIQGVGVTETPLMKEDSILAVRKYFQRITLYLKEKKYSPCAWEVVRAEIMRSFSLSTNLQESLRSKEGGGGSGGGGSGGGGSRTRVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGK SEQ ID NO: 40: Amino acid sequence of hIFNα-HA-Fc in homodimer CDLPQTHSLGSRRTLMLLAQMRRISLFSCLKDRHDFGFPQEEFGNQFQKAETIPVLHEMIQQIFNLFSTKDSSAAWDETLLDKFYTELYQQLNDLEACVIQGVGVTETPLMKEDSILAVRKYFQRITLYLKEKKYSPCAWEVVRAEIMRSFSLSTNLQESLRSKEGGGGSGGGGSGGGGSRTDTICIGYHANNSTDTVDTVLEKNVTVTHSVNLLEDSHNGKLCRLKGIAPLQLGKCNIAGWLLGNPECDPLLPVRSWSYIVETPNSENGICYPGDFIDYEELREQLSSVSSFERFEIFPKESSWPNHNTNGVTAACSHEGKSSFYRNLLWLTEKEGSYPKLKNSYVNKKGKEVLVLWGIHHPPNSKEQQNLYQNENAYVSVVTSNYNRRFTPEIAERPKVRDQAGRMNYYWTLLKPGDTIIFEANGNLIAPMYAFALSRGFGSGIITSNASMHECNTKCQTPLGAINSSLPYQNIHPVTIGECPKYVRSAKLRMVTGLRNNPSIQSRFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGK SEQ ID NO: 41: Amino acid sequence of hIFNα-E7(HPV)-Fc in a homodimer CDLPQTHSLGSRRTLMLLAQMRRISLFSCLKDRHDFGFPQEEFGNQFQKAETIPVLHEMIQQIFNLFSTKDSSAAWDETLLDKFYTELYQQLNDLEACVIQGVGVTETPLMKEDSILAVRKYFQRITLYLKEKKYSPCAWEVVRAEIMRSFSLSTNLQESLRSKEGGGGSGGGGSGGGGSRTMHGDTPTLHEYMLDLQPETTDLYCYEQLNDSSEEEDEIDGPAGQAEPDRAHYNIVTFCCKCDSTLRLCVQSTHVDIRTLEDLLMGTLGIVCPICSQKPFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGK SEQ ID NO: 42: Amino acid sequence of the first chain hIFN-Fc-hole in a heterodimer CDLPQTHSLGSRRTLMLLAQMRRISLFSCLKDRHDFGFPQEEFGNQFQKAETIPVLHEMIQQIFNLFSTKDSSAAWDETLLDKFYTELYQQLNDLEACVIQGVGVTETPLMKEDSILAVRKYFQRITLYLKEKKYSPCAWEVVRAEIMRSFSLSTNLQESLRSKESGGGGSGGGGSGGGGSGGGGRTDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFKLVSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 43: Amino acid sequence of the second chain Pres1-Fc-knob in the heterodimer hIFNα-Pres1-Fc MGQNLSTSNPLGFFPDHQLDPAFRANTANPDWDFNPNKDTWPDANKVGAGAFGLGFTPPHGGLLGWSPQAQGILQTLPANPPPASTNRQTGRQPTPLSPPLRNTHPQAFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 44: Amino acid sequence of the second chain RBD(SARS-CoV-2)-Fc-knob in the heterodimer hIFNα-RBD(SARA-CoV-2)-Fc RVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 45: Amino acid sequence of the second chain HA-Fc-knob in the heterodimer hIFNα-HA-Fc DTICIGYHANNSTDTVDTVLEKNVTVTHSVNLLEDSHNGKLCRLKGIAPLQLGKCNIAGWLLGNPECDPLLPVRSWSYIVETPNSENGICYPGDFIDYEELREQLSSVSSFERFEIFPKESSWPNHNTNGVTAACSHEGKSSFYRNLLWLTEKEGSYPKLKNSYVNKKGKEVLVLWGIHHPPNSKEQQNLYQNENAYVSVVTSNYNRRFTPEIAERPKVRDQAGRMNYYWTLLKPGDTIIFEANGNLIAPMYAFALSRGFGSGIITSNASMHECNTKCQTPLGAINSSLPYQNIHPVTIGECPKYVRSAKLRMVTGLRNNPSIQSRFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 46: Amino acid sequence of the second chain E7(HPV)-Fc-knob in the heterodimer hIFNα-E7(HPV)-Fc MHGDTPTLHEYMLDLQPETTDLYCYEQLNDSSEEEDEIDGPAGQAEPDRAHYNIVTFCCKCDSTLRLCVQSTHVDIRTLEDLLMGTLGIVCPICSQKPFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK 5. Vaccine IFNα-Pan-antigen-Fc sequence containing the Pan epitope of human IFN SEQ ID NO: 47: Amino acid sequence of hIFNα-Pan-Pres1-Fc in a homodimer CDLPQTHSLGSRRTLMLLAQMRRISLFSCLKDRHDFGFPQEEFGNQFQKAETIPVLHEMIQQIFNLFSTKDSSAAWDETLLDKFYTELYQQLNDLEACVIQGVGVTETPLMKEDSILAVRKYFQRITLYLKEKKYSPCAWEVVRAEIMRSFSLSTNLQESLRSKEAKFVAAWTLKAAAGSGSGSMGQNLSTSNPLGFFPDHQLDPAFRANTANPDWDFNPNKDTWPDANKVGAGAFGLGFTPPHGGLLGWSPQAQGILQTLPANPPPASTNRQTGRQPTPLSPPLRNTHPQAFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGK SEQ ID NO: 48: Amino acid sequence of hIFNα-Pan-RBD(SARS-CoV-2)-Fc in homodimer CDLPQTHSLGSRRTLMLLAQMRRISLFSCLKDRHDFGFPQEEFGNQFQKAETIPVLHEMIQQIFNLFSTKDSSAAWDETLLDKFYTELYQQLNDLEACVIQGVGVTETPLMKEDSILAVRKYFQRITLYLKEKKYSPCAWEVVRAEIMRSFSLSTNLQESLRSKEAKFVAAWTLKAAAGSGSGSRVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGK SEQ ID NO: 49: Amino acid sequence of hIFNα-Pan-HA-Fc in homodimer CDLPQTHSLGSRRTLMLLAQMRRISLFSCLKDRHDFGFPQEEFGNQFQKAETIPVLHEMIQQIFNLFSTKDSSAAWDETLLDKFYTELYQQLNDLEACVIQGVGVTETPLMKEDSILAVRKYFQRITLYLKEKKYSPCAWEVVRAEIMRSFSLSTNLQESLRSKEAKFVAAWTLKAAAGSGSGSDTICIGYHANNSTDTVDTVLEKNVTVTHSVNLLEDSHNGKLCRLKGIAPLQLGKCNIAGWLLGNPECDPLLPVRSWSYIVETPNSENGICYPGDFIDYEELREQLSSVSSFERFEIFPKESSWPNHNTNGVTAACSHEGKSSFYRNLLWLTEKEGSYPKLKNSYVNKKGKEVLVLWGIHHPPNSKEQQNLYQNENAYVSVVTSNYNRRFTPEIAERPKVRDQAGRMNYYWTLLKPGDTIIFEANGNLIAPMYAFALSRGFGSGIITSNASMHECNTKCQTPLGAINSSLPYQNIHPVTIGECPKYVRSAKLRMVTGLRNNPSIQSRFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGK SEQ ID NO: 50: Amino acids of hIFNα-Pan-E7(HPV)-Fc in a homodimer CDLPQTHSLGSRRTLMLLAQMRRISLFSCLKDRHDFGFPQEEFGNQFQKAETIPVLHEMIQQIFNLFSTKDSSAAWDETLLDKFYTELYQQLNDLEACVIQGVGVTETPLMKEDSILAVRKYFQRITLYLKEKKYSPCAWEVVRAEIMRSFSLSTNLQESLRSKEAKFVAAWTLKAAAGSGSGSMHGDTPTLHEYMLDLQPETTDLYCYEQLNDSSEEEDEIDGPAGQAEPDRAHYNIVTFCCKCDSTLRLCVQSTHVDIRTLEDLLMGTLGIVCPICSQKPFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGK SEQ ID NO: 51: Amino acid sequence of the first chain hIFNα-Fc-hole in a heterodimer CDLPQTHSLGSRRTLMLLAQMRRISLFSCLKDRHDFGFPQEEFGNQFQKAETIPVLHEMIQQIFNLFSTKDSSAAWDETLLDKFYTELYQQLNDLEACVIQGVGVTETPLMKEDSILAVRKYFQRITLYLKEKKYSPCAWEVVRAEIMRSFSLSTNLQESLRSKESGGGGSGGGGSGGGGSGGGGRTDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 52: Amino acid sequence of the second chain Pan-Pres1-Fc-knob in the heterodimer hIFNα-Pan-Pres1-Fc AKFVAAWTLKAAAGSGSGSMGQNLSTSNPLGFFPDHQLDPAFRANTANPDWDFNPNKDTWPDANKVGAGAFGLGFTPPHGGLLGWSPQAQGILQTLPANPPPASTNRQTGRQPTPLSPPLRNTHPQAFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 53: Amino acid sequence of the second chain Pan-RBD(SARS-CoV-2)-Fc-knob in the heterodimer hIFNα-Pan-RBD(SARS-CoV-2)-Fc AKFVAAWTLKAAAGSGSGSRVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 54: Amino acid sequence of the second chain Pan-HA-Fc-knob in the heterodimer hIFNα-Pan-HA-Fc AKFVAAWTLKAAAGSGSGSDTICIGYHANNSTDTVDTVLEKNVTVTHSVNLLEDSHNGKLCRLKGIAPLQLGKCNIAGWLLGNPECDPLLPVRSWSYIVETPNSENGICYPGDFIDYEELREQLSSVSSFERFEIFPKESSWPNHNTNGVTAACSHEGKSSFYRNLLWLTEKEGSYPKLKNSYVNKKGKEVLVLWGIHHPPNSKEQQNLYQNENAYVSVVTSNYNRRFTPEIAERPKVRDQAGRMNYYWTLLKPGDTIIFEANGNLIAPMYAFALSRGFGSGIITSNASMHECNTKCQTPLGAINSSLPYQNIHPVTIGECPKYVRSAKLRMVTGLRNNPSIQSRFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 55: Amino acid sequence of the second chain Pan-HA-Fc-knob in the heterodimer hIFNα-Pan-E7(HPV)-Fc AKFVAAWTLKAAAGSGSGSMHGDTPTLHEYMLDLQPETTDLYCYEQLNDSSEEEDEIDGPAGQAEPDRAHYNIVTFCCKCDSTLRLCVQSTHVDIRTLEDLLMGTLGIVCPICSQKPFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK 6. Human mutant IFN vaccine hmIFNα-Pan-antigen-Fc sequence: SEQ ID NO: 56: hmIFNα-Pres1-Fc amino acid sequence in homodimer CDLPQTHSLGSRRTLMLLAQMRRISLFSCLKDRHDFGFPQEEFGNQFQKAETIPVLHEMIQQIFNLFSTKDSSAAWDETLLDKFYTELYQQLNDLEACVIQGVGVTETPLMKEDSILAVRKYFRRITLYLKEKKYSPCAWEVVRAEIMRSFSLSTNLQESLRSKEGGGGSGGGGSGGGGSRTMGQNLSTSNPLGFFPDHQLDPAFRANTANPDWDFNPNKDTWPDANKVGAGAFGLGFTPPHGGLLGWSPQAQGILQTLPANPPPASTNRQTGRQPTPLSPPLRNTHPQAFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGK SEQ ID NO: 57: Amino acid sequence of hmIFNα-RBD(SARS-CoV-2)-Fc in homodimer CDLPQTHSLGSRRTLMLLAQMRRISLFSCLKDRHDFGFPQEEFGNQFQKAETIPVLHEMIQQIFNLFSTKDSSAAWDETLLDKFYTELYQQLNDLEACVIQGVGVTETPLMKEDSILAVRKYFRRITLYLKEKKYSPCAWEVVRAEIMRSFSLSTNLQESLRSKEGGGGSGGGGSGGGGSRTRVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGK SEQ ID NO: 58: Amino acid sequence of hmIFNα-HA-Fc in homodimer CDLPQTHSLGSRRTLMLLAQMRRISLFSCLKDRHDFGFPQEEFGNQFQKAETIPVLHEMIQQIFNLFSTKDSSAAWDETLLDKFYTELYQQLNDLEACVIQGVGVTETPLMKEDSILAVRKYFRRITLYLKEKKYSPCAWEVVRAEIMRSFSLSTNLQESLRSKEGGGGSGGGGSGGGGSRTDTICIGYHANNSTDTVDTVLEKNVTVTHSVNLLEDSHNGKLCRLKGIAPLQLGKCNIAGWLLGNPECDPLLPVRSWSYIVETPNSENGICYPGDFIDYEELREQLSSVSSFERFEIFPKESSWPNHNTNGVTAACSHEGKSSFYRNLLWLTEKEGSYPKLKNSYVNKKGKEVLVLWGIHHPPNSKEQQNLYQNENAYVSVVTSNYNRRFTPEIAERPKVRDQAGRMNYYWTLLKPGDTIIFEANGNLIAPMYAFALSRGFGSGIITSNASMHECNTKCQTPLGAINSSLPYQNIHPVTIGECPKYVRSAKLRMVTGLRNNPSIQSRFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGK SEQ ID NO: 59: Amino acid sequence of hmIFNα-E7(HPV)-Fc in homodimer CDLPQTHSLGSRRTLMLLAQMRRISLFSCLKDRHDFGFPQEEFGNQFQKAETIPVLHEMIQQIFNLFSTKDSSAAWDETLLDKFYTELYQQLNDLEACVIQGVGVTETPLMKEDSILAVRKYFRRITLYLKEKKYSPCAWEVVRAEIMRSFSLSTNLQESLRSKEGGGGSGGGGSGGGGSRTMHGDTPTLHEYMLDLQPETTDLYCYEQLNDSSEEEDEIDGPAGQAEPDRAHYNIVTFCCKCDSTLRLCVQSTHVDIRTLEDLLMGTLGIVCPICSQKPFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGK SEQ ID NO: 60: Amino acid sequence of the first chain hmIFN-Fc-hole in a heterodimer CDLPQTHSLGSRRTLMLLAQMRRISLFSCLKDRHDFGFPQEEFGNQFQKAETIPVLHEMIQQIFNLFSTKDSSAAWDETLLDKFYTELYQQLNDLEACVIQGVGVTETPLMKEDSILAVRKYFRRITLYLKEKKYSPCAWEVVRAEIMRSFSLSTNLQESLRSKESGGGGSGGGGSGGGGSGGGGRTDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 61: Amino acid sequence of the second chain Pres1-Fc-knob in the heterodimer hmIFNα-Pres1-Fc MGQNLSTSNPLGFFPDHQLDPAFRANTANPDWDFNPNKDTWPDANKVGAGAFGLGFTPPHGGLLGWSPQAQGILQTLPANPPPASTNRQTGRQPTPLSPPLRNTHPQAFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 62: Amino acid sequence of the second chain RBD(SARS-CoV-2)-Fc-knob in the heterodimer hmIFNα-RBD(SARA-CoV-2)-Fc RVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 63: Amino acid sequence of the second chain HA-Fc-knob in the heterodimer hmIFNα-HA-Fc DTICIGYHANNSTDTVDTVLEKNVTVTHSVNLLEDSHNGKLCRLKGIAPLQLGKCNIAGWLLGNPECDPLLPVRSWSYIVETPNSENGICYPGDFIDYEELREQLSSVSSFERFEIFPKESSWPNHNTNGVTAACSHEGKSSFYRNLLWLTEKEGSYPKLKNSYVNKKGKEVLVLWGIHHPPNSKEQQNLYQNENAYVSVVTSNYNRRFTPEIAERPKVRDQAGRMNYYWTLLKPGDTIIFEANGNLIAPMYAFALSRGFGSGIITSNASMHECNTKCQTPLGAINSSLPYQNIHPVTIGECPKYVRSAKLRMVTGLRNNPSIQSRFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 64: Amino acid sequence of the second chain HA-Fc-knob in the heterodimer hmIFNα-E7(HPV)-Fc MHGDTPTLHEYMLDLQPETTDLYCYEQLNDSSEEEDEIDGPAGQAEPDRAHYNIVTFCCKCDSTLRLCVQSTHVDIRTLEDLLMGTLGIVCPICSQKPFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK 7. Vaccine containing the Pan epitope of human mutant IFN, hmIFNα-Pan epitope-antigen-Fc sequence SEQ ID NO: 65: Amino acid sequence of hmIFNα-Pan-Pres1-Fc in a homodimer CDLPQTHSLGSRRTLMLLAQMRRISLFSCLKDRHDFGFPQEEFGNQFQKAETIPVLHEMIQQIFNLFSTKDSSAAWDETLLDKFYTELYQQLNDLEACVIQGVGVTETPLMKEDSILAVRKYFRRITLYLKEKKYSPCAWEVVRAEIMRSFSLSTNLQESLRSKEAKFVAAWTLKAAAGSGSGSMGQNLSTSNPLGFFPDHQLDPAFRANTANPDWDFNPNKDTWPDANKVGAGAFGLGFTPPHGGLLGWSPQAQGILQTLPANPPPASTNRQTGRQPTPLSPPLRNTHPQAFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGK SEQ ID NO: 66: Amino acid sequence of hmIFNα-Pan-RBD(SARS-CoV-2)-Fc in homodimer CDLPQTHSLGSRRTLMLLAQMRRISLFSCLKDRHDFGFPQEEFGNQFQKAETIPVLHEMIQQIFNLFSTKDSSAAWDETLLDKFYTELYQQLNDLEACVIQGVGVTETPLMKEDSILAVRKYFRRITLYLKEKKYSPCAWEVVRAEIMRSFSLSTNLQESLRSKEAKFVAAWTLKAAAGSGSGSRVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGK SEQ ID NO: 67: Amino acid sequence of hmIFNα-Pan-HA-Fc in homodimer CDLPQTHSLGSRRTLMLLAQMRRISLFSCLKDRHDFGFPQEEFGNQFQKAETIPVLHEMIQQIFNLFSTKDSSAAWDETLLDKFYTELYQQLNDLEACVIQGVGVTETPLMKEDSILAVRKYFRRITLYLKEKKYSPCAWEVVRAEIMRSFSLSTNLQESLRSKEAKFVAAWTLKAAAGSGSGSDTICIGYHANNSTDTVDTVLEKNVTVTHSVNLLEDSHNGKLCRLKGIAPLQLGKCNIAGWLLGNPECDPLLPVRSWSYIVETPNSENGICYPGDFIDYEELREQLSSVSSFERFEIFPKESSWPNHNTNGVTAACSHEGKSSFYRNLLWLTEKEGSYPKLKNSYVNKKGKEVLVLWGIHHPPNSKEQQNLYQNENAYVSVVTSNYNRRFTPEIAERPKVRDQAGRMNYYWTLLKPGDTIIFEANGNLIAPMYAFALSRGFGSGIITSNASMHECNTKCQTPLGAINSSLPYQNIHPVTIGECPKYVRSAKLRMVTGLRNNPSIQSRFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGK SEQ ID NO: 68: Amino acid sequence of hmIFNα-Pan-E7(HPV)-Fc in homodimer CDLPQTHSLGSRRTLMLLAQMRRISLFSCLKDRHDFGFPQEEFGNQFQKAETIPVLHEMIQQIFNLFSTKDSSAAWDETLLDKFYTELYQQLNDLEACVIQGVGVTETPLMKEDSILAVRKYFRRITLYLKEKKYSPCAWEVVRAEIMRSFSLSTNLQESLRSKEAKFVAAWTLKAAAGSGSGSMHGDTPTLHEYMLDLQPETTDLYCYEQLNDSSEEEDEIDGPAGQAEPDRAHYNIVTFCCKCDSTLRLCVQSTHVDIRTLEDLLMGTLGIVCPICSQKPFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDQLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFLYSKLTVDKSRWQQGNVFSCSVLHEALHNHYTQKSLSLSPGK SEQ ID NO: 69: Amino acid sequence of the first chain hmIFNα4-Fc-hole in a heterodimer CDLPQTHSLGSRRTLMLLAQMRRISLFSCLKDRHDFGFPQEEFGNQFQKAETIPVLHEMIQQIFNLFSTKDSSAAWDETLLDKFYTELYQQLNDLEACVIQGVGVTETPLMKEDSILAVRKYFRRITLYLKEKKYSPCAWEVVRAEIMRSFSLSTNLQESLRSKESGGGGSGGGGSGGGGSGGGGRTDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 70: Amino acid sequence of the second chain Pan-Pres1-Fc-knob in the heterodimer hmIFNα-Pan-Pres1-Fc AKFVAAWTLKAAAGSGSGSMGQNLSTSNPLGFFPDHQLDPAFRANTANPDWDFNPNKDTWPDANKVGAGAFGLGFTPPHGGLLGWSPQAQGILQTLPANPPPASTNRQTGRQPTPLSPPLRNTHPQAFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 71: Amino acid sequence of the second chain Pan-RBD(SARS-CoV-2)-Fc-knob in the heterodimer hmIFNα-Pan-RBD(SARS-CoV-2)-Fc AKFVAAWTLKAAAGSGSGSRVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 72: Amino acid sequence of the second chain Pan-HA-Fc-knob in the heterodimer hmIFNα-Pan-HA-Fc AKFVAAWTLKAAAGSGSGSDTICIGYHANNSTDTVDTVLEKNVTVTHSVNLLEDSHNGKLCRLKGIAPLQLGKCNIAGWLLGNPECDPLLPVRSWSYIVETPNSENGICYPGDFIDYEELREQLSSVSSFERFEIFPKESSWPNHNTNGVTAACSHEGKSSFYRNLLWLTEKEGSYPKLKNSYVNKKGKEVLVLWGIHHPPNSKEQQNLYQNENAYVSVVTSNYNRRFTPEIAERPKVRDQAGRMNYYWTLLKPGDTIIFEANGNLIAPMYAFALSRGFGSGIITSNASMHECNTKCQTPLGAINSSLPYQNIHPVTIGECPKYVRSAKLRMVTGLRNNPSIQSRFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO: 73: Amino acid sequence of the second chain Pan-HA-Fc-knob in the heterodimer hmIFNα-Pan-E7(HPV)-Fc AKFVAAWTLKAAAGSGSGSMHGDTPTLHEYMLDLQPETTDLYCYEQLNDSSEEEDEIDGPAGQAEPDRAHYNIVTFCCKCDSTLRLCVQSTHVDIRTLEDLLMGTLGIVCPICSQKPFEDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK 8. Antibody sequence replacing Fc SEQ ID NO: 20: Amino acid sequence of ScFv(PD-L1) DIQMTQSPSSLSASVGDRVTITCRASQDVSTAVAWYQQKPGKAPKLLIYSASFLYSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQYLYHPATFGQGTKVEIKRGGGGSGGGGSGGGGSEVQLVESGGGLVQPGGSLRLSCAASGFTFSDSWIHWVRQAPGKGLEWVAWISPYGGSTYYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARRHWPGGFDYWGQGTLVTVSA SEQ ID NO: 74: Amino acid sequence of Anti-PD-L1 VH EVQLVESGGGLVQPGGSLRLSCAASGFTFSDSWIHWVRQAPGKGLEWVAWISPYGGSTYYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARRHWPGGFDYWGQGTLVTVSA SEQ ID NO: 75: Amino acid sequence of Anti-PD-L1 VL DIQMTQSPSSLSASVGDRVTITCRASQDVSTAVAWYQQKPGKAPKLLIYSASFLYSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQYLYHPATFGQGTKVEIKR 9. Other viral antigen sequences SEQ ID NO: 76: Amino acid sequence of the SARS-CoV-2 Spike protein VNLTTRTQLPPAYTNSFTRGVYYPDKVFRSSVLHSTQDLFLPFFSNVTWFHAIHVSGTNGTKRFDNPVLPFNDGVYFASTEKSNIIRGWIFGTTLDSKTQSLLIVNNATNVVIKVCEFQFCNDPFLGVYYHKNNKSWMESEFRVYSSANNCTFEYVSQPFLMDLEGKQGNFKNLREFVFKNIDGYFKIYSKHTPINLVRDLPQGFSALEPLVDLPIGINITRFQTLLALHRSYLTPGDSSSGWTAGAAAYYVGYLQPRTFLLKYNENGTITDAVDCALDPLSETKCTLKSFTVEKGIYQTSNFRVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFNFNGLTGTGVLTESNKKFLPFQQFGRDIADTTDAVRDPQTLEILDITPCSFGGVSVITPGTNTSNQVAVLYQDVNCTEVPVAIHADQLTPTWRVYSTGSNVFQTRAGCLIGAEHVNNSYECDIPIGAGICASYQTQTNSPRRARSVASQSIIAYTMSLGAENSVAYSNNSIAIPTNFTISVTTEILPVSMTKTSVDCTMYICGDSTECSNLLLQYGSFCTQLNRALTGIAVEQDKNTQEVFAQVKQIYKTPPIKDFGGFNFSQILPDPSKPSKRSFIEDLLFNKVTLADAGFIKQYGDCLGDIAARDLICAQKFNGLTVLPPLLTDEMIAQYTSALLAGTITSGWTFGAGAALQIPFAMQMAYRFNGIGVTQNVLYENQKLIANQFNSAIGKIQDSLSSTASALGKLQD VVNQNAQALNTLVKQLSSNFGAISSVLNDILSRLDKVEAEVQIDRLITGRLQSLQTYVTQQLIRAAEIRASANLAATKMSECVLGQSKRVDFCGKGYHLMSFPQSAPHGVVFLHVTYVPAQEKNFTTAPAICHDGKAHFPREGVFVSNGTHWFVTQRNFYEPQIITTDNTFVSGNCDVVIGIVNNTVYDPLQPELDSFKEELDKYFKNHTSPDVDLGDISGINASVVNIQKEIDRLNEVAKNLNESLIDLQELGKYEQYIKWPWYIWLGFIAGLIAIVMVTIMLCCMTSCCSCLKGCCSCGSCCKFDEDDSEPVLKGVKLHYT SEQ ID NO: 77: Amino acid sequence of the SARS-CoV-2 S1 protein VNLTTRTQLPPAYTNSFTRGVYYPDKVFRSSVLHSTQDLFLPFFSNVTWFHAIHVSGTNGTKRFDNPVLPFNDGVYFASTEKSNIIRGWIFGTTLDSKTQSLLIVNNATNVVIKVCEFQFCNDPFLGVYYHKNNKSWMESEFRVYSSANNCTFEYVSQPFLMDLEGKQGNFKNLREFVFKNIDGYFKIYSKHTPINLVRDLPQGFSALEPLVDLPIGINITRFQTLLALHRSYLTPGDSSSGWTAGAAAYYVGYLQPRTFLLKYNENGTITDAVDCALDPLSETKCTLKSFTVEKGIYQTSNFRVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFNFNGLTGTGVLTESNKKFLPFQQFGRDIADTTDAVRDPQTLEILDITPCSFGGVSVITPGTNTSNQVAVLYQDVNCTEVPVAIHADQLTPTWRVYSTGSNVFQTRAGCLIGAEHVNNSYECDIPIGAGICASYQTQTNSPRRAR SEQ ID NO: 78: Amino acid sequence of the RBD protein of the original SARS-CoV-2 strain RVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNF SEQ ID NO: 79: Amino acid sequence of the RBD protein of the SARS-CoV-2 UK variant (B.1.1.7, Alpha) RVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTYGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNF SEQ ID NO: 80: Amino acid sequence of the RBD protein of the SARS-CoV-2 South African variant (B.1.351, Beta) RVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGNIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVKGFNCYFPLQSYGFQPTYGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNF SEQ ID NO: 81: Amino acid sequence of the RBD protein of the SARS-CoV-2 Brazilian variant ((P.1) RVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGTIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVKGFNCYFPLQSYGFQPTYGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNF SEQ ID NO: 82: Amino acid sequence of the RBD protein of the SRAS-CoV-2 California variant (B.1.429) RVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYRYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNF SEQ ID NO: 83: Amino acid sequence of the RBD protein of the SARS-CoV-2 Indian B.1.617, B.1.617.1 (Kappa), and B.1.617.3 mutant strains RVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYRYRLFRKSNLKPFERDISTEIYQAGSTPCNGVQGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNF SEQ ID NO: 84: Amino acid sequence of the RBD protein of the SARS-CoV-2 Indian variant II B.1.617.2 (Delta) strain RVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYRYRLFRKSNLKPFERDISTEIYQAGSKPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNF 10. Other tumor antigen sequences: Amino acid sequences of murine-derived Her2 extracellular domains II, III, and IV according to the examples SEQ ID NO: 85: Mouse Her2 - extracellular domain 2: SRACPPCAPACKDNHCWGESPEDCQILTGTICTSGCARCKGRLPTDCCHEQCAAGCTGPKHSDCLACLHFNHSGICELHCPALVTYNTDTFESMHNPEGRYTFGASCVTTCPYNYLSTEVGSCTLVCPPNNQEVTAEDGTQRCEKCSKPC SEQ ID NO: 86: Mouse Her2 - extracellular domain 3: GCKKIFGSLAFLPESFDGDPSSGIAPLRPEQLQVFETLEEITGYLYISAWPDSLRDLSVFQNLRIIRGRILHDGAYSLTLQGLGIHSLGLRSLRELGSGLALIHRNAHLCFVHTVPWDQLF SEQ ID NO: 87: Mouse Her2 - extracellular domain 4: VCNSLCAHGHCWGPGPTQCVNCSHFLRGQECVEECRVWKGLPREYVSDKRCLPCHPECQPQNSSETCFGSEADQCAACAHYKDSSSCVARCPSGVKPDLSYMPIWKYPDEEGICQPCPINCTHSCVDLDERGCP Amino acid sequences of human Her2 extracellular domains II, III and IV according to the examples: SEQ ID NO: 88: Human Her2 - extracellular domain 2: SRACHPCSPMCKGSRCWGESSEDCQSLTRTVCAGGCARCKGPLPTDCCHEQCAAGCTGPKHSDCLACLHFNHSGICELHCPALVTYNTDTFESMPNPEGRYTFGASCVTACPYNYLSTDVGSCTLVCPLHNQEVTAEDGTQRCEKCSKPC SEQ ID NO: 89: Human Her2 - extracellular domain 3: GCKKIFGSLAFLPESFDGDPASNTAPLQPEQLQVFETLEEITGYLYISAWPDSLPDLSVFQNLQVIRGRILHNGAYSLTLQGLGISWLGLRSLRELGSGLALIHHNTHLCFVHTVPW SEQ ID NO: 90: Human Her2 - extracellular domain 4: CHQLCARGHCWGPGPTQCVNCSQFLRGQECVEECRVLQGLPREYVNARHCLPCHPECQPQNGSVTCFGPEADQCVACAHYKDPPFCVARCPSGVKPDLSYMPIWKFPDEEGACQPCPINCTHSCVDLDDKGCP 11. Herpes virus antigen sequence according to the example: SEQ ID NO: 91: VZV Envelope glycoprotein E (aa 31 - 538) SVLRYDDFHTDEDKLDTNSVYEPYYHSDHAESSWVNRGESSRKAYDHNSPYIWPRNDYDGFLENAHEHHGVYNQGRGIDSGERLMQPTQMSAQEDLGDDTGIHVIPTLNGDDRHKIVNVDQRQYGDVFKGDLNPKPQGQRLIEVSVEENHPFTLRAPIQRIYGVRYTETWSFLPSLTCTGDAAPAIQHICLKHTTCFQDVVVDVDCAENTKEDQLAEISYRFQGKKEADQPWIVVNTSTLFDELELDPPEIEPGVLKVLRTEKQYLGVYIWNMRGSDGTSTYATFLVTWKGDEKTRNPTPAVTPQPRGAEFHMWNYHSHVFSVGDTFSLAMHLQYKIHEAPFDLLLEWLYVPIDPTCQPMRLYSTCLYHPNAPQCLSHMNSGCTFTSPHLAQRVASTVYQNCEHADNYTAYCLGISHMEPSFGLILHDGGTTLKFVDTPESLSGLYVFVVYFNGHVEAVAYTVVSTVDHFVNAIEERGFPPTAGQPPATTKPKEITPVNPGTSPLLRY SEQ ID NO: 92: EBV Envelope glycoprotein GP350 (aa 1-425) MEAALLVCQYTIQSLIHLTGEDPGFFNVEIPEFPFYPTCNVCTADVNVTINFDVGGKKHQLDLDFGQLTPHTKAVYQPRGAFGGSENATNLFLLELLGAGELALTMRSKKLPINVTTGEEQQVSLESVDVYFQDVFGTMWCHHAEMQNPVYLIPETVPYIKWDNCNSTNITAVVRAQGLDVTLPLSLPTSAQDSNFSVKTQMLGNEIDIECIMEDGEISQVLPGDNKFNITCSGYESHVPSGGILTSTSPVVTPIPGTGYAYSLRLTPRPVSRFLGNNSILYVFYSGNGPKASGGDYCIQSNIVFSDEIPASQDMPTNTTDITYVGDNATYSVPMVTSEDANSPNVTVTAFWAWPNNTETDFKCKWTLTSGTPSGCENISGAFASNRTFDITVSGLGTAPKTLIITRTATNATTTTHKVIFSKAP SEQ ID NO: 93: HSV-2 Envelope glycoprotein gD (aa 26-339) KYALADPSLKMADPNRFRGKNLPVLDRLTDPPGVKRVYHIQPSLEDPFQPPSIPITVYYAVLERACRSVLLHAPSEAPQIVRGASDEARKHTYNLTIAWYRMGDNCAIPITVMEYTECPYNKSLGVCPIRTQPRWSYYDSFSAVSEDNLGFLMHAPAFETAGTYLRLVKINDWTEITQFILEHRARASCKYALPLRIPPAACLTSKAYQQGVTVDSIGMLPRFIPENQRTVALYSLKIAGWHGPKPPYTSTLLPPELSDTTNATQPELVPEDPEDSALLEDPAGTVSSQIPPNWHIPSIQDVAPHHAPAAPSNP

Brief Description of the Drawings

[0045]

Figure 1

Figure 2

Figure 3

Figure 4

Figure 5

Figure 6

Figure 7

Figure 8

Figure 9

Figure 10

Figure 11

Figure 12

Figure 13

Figure 14

Figure 15

Figure 16

Figure 17

Figure 18

Figure 19

Figure 20

Figure 21

Figure 22

Figure 23

Figure 24

Figure 25

Figure 26

Figure 27

Figure 28

Mode for Carrying Out the Invention

[0046] To make the objectives, technical solutions, and advantages of the present invention clearer, the present invention will be described in detail below with reference to the embodiments and the accompanying drawings. The described embodiments are only examples of the present invention and are not intended to limit the scope of the present invention. The embodiments are only a part of the present invention and not all embodiments of the present invention. The scope of the present invention is defined by the appended claims.

[0047] Example 1. Design of Vaccine Platform The vaccine platform of the interferon-target antigen-immunoglobulin Fc (or antibody) structural unit consists of three structural units. The first structural unit is the interferon part, the second structural unit is the immunoglobulin Fc region (or antibody), and the third structural unit is the target antigen. In actual construction, the three structural units can be arranged and combined in any form, and the target antigen can be linked to the Th cell helper epitope via the linker sequence 2. Its typical form was as follows.

[0048] Figure 1 was a schematic diagram of combining the above vaccine platform in the form of a homodimer in the order of interferon-linker fragment 1-target antigen-immunoglobulin Fc.

[0049] Figure 2 was a schematic diagram of combining the above vaccine platform in the form of a heterodimer in the order of interferon-linker fragment 1-IgG1-hole and target antigen-IgG1-knob respectively.

[0050] Figure 3 was a schematic diagram of combining the above vaccine platform in the form of a heterodimer in the order of interferon-linker fragment 1-IgG1-knob and target antigen-IgG1-hole.

[0051] Next, the target antigen was linked to the cell helper epitope by the linker fragment 2, and then combined with the other two vaccine platform components. Its typical form was as follows.

[0052] Figure 4 was a schematic diagram of combining the above vaccine platform in the form of a homodimer in the order of interferon-linker fragment 1-Th cell helper epitope-linker fragment 2-target antigen-immunoglobulin Fc.

[0053] FIG. 5 is a schematic diagram of combining the vaccine platform in the form of a heterodimer in the order of interferon-linked fragment 1-IgG1-hole and Th cell helper epitope-linked fragment 2-target antigen-IgG1-knob, respectively.

[0054] FIG. 6 is a schematic diagram of combining the vaccine platform in the form of a heterodimer in the order of interferon-linked fragment 1-IgG1-knob and Th cell helper epitope-linked fragment 2-target antigen-IgG1-hole, respectively.

[0055] Example 2. Construction, purification and production of the vaccine platform Taking the homodimers of hepatitis B virus Pres1 and coronavirus SARS-CoV-2 RBD protein as examples, the expression and production of the vaccine platform will be described.

[0056] 1. Construction of vectors, transfection of host cells and induced expression 1.1 Using PEE12.4 as the vector, a vaccine structural unit was constructed into the vector by molecular cloning to obtain a plasmid capable of expressing a fusion protein. Then, 293F cells were transiently transfected, the culture supernatant was collected, and finally the target protein was purified by a protein A affinity chromatography column. Construction of vectors (taking the example of containing HBV preS1) (1) PEE12.4-HindIII-signal peptide 1-interferon-BsiwI-Pres1-BstbI-hIgG1-EcoRI (2) PEE12.4-HindIII-signal peptide 1-interferon-BsiwI-RBD(SARS-CoV-2)-BstbI-hIgG1-EcoRI (3) PEE12.4-HindIII-signal peptide 1-interferon-Bsiwi-PADER-Pres1-hIgG1-EcoRI (4) PEE12.4-HindIII-Signal Peptide 1-Interferon-Bsiwi-PADER-RBD(SARS-CoV-2)-hIgG1-EcoRI The linker sequences between the respective fusion protein fragments were as follows. (1) The linker fragment 1 was between interferon and Pres1. (2) The linker fragment 1 was between interferon and RBD(SARS-CoV-2). (3) The linker sequence between interferon and PADER was linker segment 1, and the linker segment between PADER and Pres1 was linker segment 2. (4) The linker sequence between interferon and PADER was linker segment 1, and the linker segment between PADER and RBD(SARS-CoV-2) was linker segment 2.

[0057] 1.2, Rapid expression of target proteins by transient transfection: (1) Cell resuscitation: Freestyle 293F cells were cryopreserved in CD OptiCHOTM media (containing 10% DMSO) at a concentration of 3×10 7 cells / ml. After being taken out from liquid nitrogen, they were quickly dissolved in a 37°C water bath and added to a 15 ml centrifuge tube containing 10 ml of OptiCHOTM media, and centrifuged at 1000 rpm for 5 minutes. The supernatant was discarded, and the cell precipitate was suspended in 30 ml of OptiCHOTM media and cultured under the conditions of 37°C, 8% CO2, and 135 rpm. Four days later, the cells were expanded, and the concentration of the expanded culture was made not to exceed 3×10 6 cells / ml. (2) Two days before transfection, suspension-cultured 293F cells were prepared for transient transfection (200 ml) at a seeding density of 0.6 - 0.8×10 6 cells / ml. (3) Two days later, the cell suspension to be transfected was counted, and the expected cell density was 2.5 - 3.5×10 6cells / ml, and then the cell suspension was centrifuged at 1000 rpm for 5 minutes, and the supernatant was discarded. (4) The cells were resuspended in 50 ml of fresh Freestyle 293 media, centrifuged again at 1000 rpm for 5 minutes, and the supernatant was discarded. (5) The 293F cells were resuspended in 200 ml of Freestyle 293 media. (6) 600 μg of plasmid was diluted in 5 ml of Freestyle 293 media, and bacteria were removed using a 0.22 μM filter. (7) 1.8 mg of PEI was diluted in 5 ml of Freestyle 293 media, and bacteria were removed using a 0.22 μM filter. Immediately afterwards, 5 ml of plasmid and 5 ml of PEI were mixed and left standing at room temperature for 5 minutes. (8) The plasmid / PEI mixture was added to the cell suspension, placed in an incubator at 37 °C, 8% CO2, 85 rpm, and cultured while supplementing with 50 μg / L of the growth factor LONG TM R3IGF-1. (9) After 4 hours, 200 ml of EX-CELLTM 293 media supplemented with 2 mM Glutamine was added, the rotation speed was set to 135 rpm, and the culture was continued. (10) After 24 hours, 3.8 mM VPA, a cell growth inhibitor, was added. After 72 hours, 40 ml of medium D was added and the culture was continued for 6 - 8 days (cell viability less than 70%). Then the supernatant was collected and placed until used in the next purification step.

[0058] 1.3, Recovery, Purification and Electrophoresis Verification of the Fusion Protein

[0059] 2. Purification of the Target Protein Using Protein A: (1) Sample Preparation: The suspended cell culture was transferred to a 500 ml centrifuge bucket, centrifuged at 8000 rpm for 20 minutes, the precipitate was discarded, the supernatant was filtered through a 0.45 μM filter to remove impurities, and then NaN3 with a final concentration of 0.05% was added to prevent bacterial contamination during purification. Assembly of the chromatography column: Take an appropriate amount of Protein A Agarose (calculated by purifying 20 mg of human Fc fusion protein per 1 ml of Protein A), mix well, add it to the chromatography column, let it stand at room temperature for about 10 minutes, overlay Protein A with a 20% ethanol solution, then open the lower outlet and slowly let the ethanol solution flow out by gravity. (3) Wash and equilibrate the chromatography column with distilled water and Binding buffer (20 mM sodium phosphate + 0.15 M NaCl, pH 7.0) at 10 times the column volume. (4) Using a constant flow pump, load the sample at a flow rate of 10 times the column volume per hour, collect the flow-through, and repeat the sample loading twice. (5) Wash the column with Binding buffer at more than 10 times the column volume to remove impurities, and wash until no protein is detected in the effluent. (6) Elute using Elution Buffer (0.1 M Glycine, pH 2.7), collect into separate tubes one by one per 1 ml, and observe the elution peak using a protein indicator solution (Bio-Rad protein assay). Mix the collection tubes of the elution peak, add an appropriate amount of 1 M Tris, pH 9.0 for neutralization (adjust the pH to 6 - 8, which is at least 0.5 higher than the isoelectric point of the purified protein). (7) Using a Zeba desalting and concentration centrifugal column, replace the target protein solution with the target buffer (adjust the pH of the buffer while avoiding the isoelectric point of the protein). Using BSA as a standard, determine the protein concentration by SDS-PAGE electrophoresis and NanoDrop2000. (8) After elution, wash sequentially with 20 times the column volume of distilled water and 10 times the column volume of 20% ethanol, and finally immerse the ethanol solution in the gel medium and store at 4°C.

[0060] 3. The identification of the protein by SDS-PAGE electrophoresis is shown in Figure 7.

[0061] Example 3. Compared with the simple Pres1 antigen, FNα-Pres1-Fc and Pres1-Fc can induce a stronger immune response in mice.

[0062] Materials: C57BL / 6 male mice (5 - 8 weeks old) were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd. Horseradish peroxidase (HRP)-labeled goat anti-mouse IgG was purchased from Beijing ComWin Biotech Co., Ltd. The 96-well ELISA measurement board was purchased from Corning Costar. The ELISA chromogenic solution was purchased from eBioscience. The microplate reader SPECTRA max PLUS 384 used was purchased from Molecular, USA. The aluminum adjuvant used was purchased from SIGMA.

[0063] Methods: (1) Immunization of mice with Pres1 fusion protein: 80 pmol of IFN-Pres1-Fc or 80 pmol of Pres1-Fc and Pres1 protein were mixed with aluminum adjuvant and then used to immunize mice subcutaneously. Antibody detection was performed by collecting mouse serum by orbital bleeding at the designated time points. (2) The antibodies produced by IFNα-Pres1-Fc have a broad neutralizing effect against HBV viruses of different genotypes. Five-week-old male C57BL / 6 mice were infected via the tail vein with 1x10 11 μg of AAV-HBV 1.3 (the HBV genotypes are B and C). Six weeks later, mice with stable expression of HBV antigen were screened and tested. The screened mice (4 mice / group) were injected intravenously with 200 μl / mouse of serum from IFNα-Pres1-Fc-immunized mice. Twelve hours later, mouse serum was collected, and the changes in the Pres1 antigen of the mice before and after administration of the antiserum were detected by ELISA. (3) Detection of anti-Pres1 specific antibodies in serum by ELISA A Pres1 (2 μg / ml) coating solution was added to an ELISA plate (Corning 9018) at 50 μl / well and coated overnight at 4°C. The plate was washed once with 260 μl of PBS per well. Blocking was performed at 37°C for 2 hours with a 5% blocking solution (5% FBS). Serum samples (1:10, 1:100, 1:1000, 1:10000) were diluted with PBS and added to the blocked ELISA plate at 50 μl / well, followed by incubation at 37°C for 1 hour. Each well was washed 5 times with 260 μl of PBST each time, and 50 μl of an enzyme-conjugated secondary antibody (enzyme-conjugated anti-mouse IgG-HRP diluted 1:5000 with PBS) was added to each well and incubated at 37°C for 1 hour. Each well was washed 5 times with 260 μl of PBST each time, 100 μl / well of substrate TMB was added, and the mixture was incubated at room temperature in the dark. After waiting for the substrate to develop color, 50 μl of a termination solution (2N H2SO4) was added to each well to terminate the color development. The plate was read using a microplate reader, and the OD was 450 - 630.

[0064] Results: The immunogenicity of free Pres1 is weak. When IFNα and the Fc portion are added to Pres1 to form the IFNα-Pres1-Fc fusion protein, as shown in Figure 8(a), its immunogenicity is significantly improved. Also, as shown in Figure 8(b), the antibodies produced by IFNα-Pres1-Fc have a broad neutralizing effect against different HBV genotype viruses.

[0065] Example 4. IFNα-Pres1-Fc can be used as a hepatitis B preventive vaccine. Materials: Male C57BL / 6 mice (6 - 8 weeks old) were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd. The HBsAg detection kit was purchased from Shanghai Kehua Bio-Engineering Co., Ltd. The AAV-HBV 1.3 virus was purchased from Guangzhou PuriBio Technology Co., Ltd. Other experimental materials were the same as in Example 3.

[0066] Methods: (1) 80 pmol of different forms of Pres1 vaccine containing simple Pres1, Pes1-Fc, and IFNα-Pres1-Fc proteins were subcutaneously immunized into mice. Serum was collected on the 28th day after immunization, and 1x10 11 μg of AAV-HBV 1.3 virus was infected. Then, serum was collected weekly, and the detection of anti-Pres1 antibody, HBsAg, and Pres1 antigen in the serum was continuously performed for 4 weeks. At the 3rd week, the level of HBV-DNA in the mouse periphery was detected. (2) Detection of Pres1-specific antigen in serum by ELISA Coating of antigen: The Pres1 antibody XY007 (4 μg / ml) coating solution was added to an Elisa plate (Corning 9018) at 50 μl / well and coated overnight at 4°C. The plate was washed once with 260 μl of PBS per well. Blocking was performed with 5% blocking solution (5% FBS) at 37°C for 2 hours. Serum samples (1:10, 1:100) were diluted with PBS and added to the blocked Elisa plate at 50 μl / well. Two replicate wells were set for each dilution and incubated at 37°C for 1 hour. Each time, it was washed 5 times with 260 μl of PBST. 50 μl of enzyme conjugate (from Kehua HBsAg detection Kit) was added to each well and incubated at 37°C for 1 hour. Each time, it was washed 5 times with 260 μl of PBST. 100 μl / well of substrate TMB was added, incubated at room temperature in the dark, waited for the substrate to develop color, 50 μl of termination solution (2N H2SO4) was added to each well to terminate the color development, and the plate was read with an enzyme marker, and the result was OD450 - 630.

[0067] Results: As shown in Fig. 9(a), mice in the IFNα-Pres1-Fc immunized group could produce high levels of Pres1 antibodies before virus inoculation and maintained high levels throughout the virus infection. Immunization with the IFN-Pres1-Fc vaccine could significantly prevent HBV infection compared to the group without protein immunization. The anti-preS1 antibodies produced after immunization could rapidly and completely remove preS1 antigen in the serum (Fig. 9(b)). Moreover, most of the mice infected with the virus in the IFN-Pres1-Fc immunized group showed peripheral HBsAg negativity (Fig. 9(c, d)). The above experimental results, as shown in Fig. 9, indicate that IFN-Pres1-Fc as a vaccine can effectively prevent HBV infection.

[0068] Example 5. IFNα-Pres1-Fc was used as a therapeutic vaccine for chronic hepatitis B infection. Materials: C57BL / 6 male mice (4 weeks old) were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd. AAV-HBV 1.3 was purchased from Guangzhou Pieshine Biotechnology Co., Ltd. The HBsAg detection kit was purchased from Shanghai Kehua Biotechnology Co., Ltd. Other experimental materials were the same as in Example 4.

[0069] Methods: (1) Screening of HBV carrier mice: 1x10 11 μg of AAV-HBV 1.3 virus was injected into 4-week-old HBV C57BL / 6 mice via the tail vein. HBV antigen HBsAg was detected at 1 - 6 weeks, and mice with stable HBsAg expression were screened and used as HBV carrier mice. (2) The screened mice were subcutaneously injected with 80 pmol of different forms of Pres1 protein once every two weeks for a total of three immunizations. Mouse serum was collected 14 days after the first immunization and then weekly. The levels of anti-Pres1 antibodies, HBsAG, and Pres1 antigen in the mouse serum were detected by ELISA. The HBV-DNA content in the peripheral blood of the mice was detected after the last blood collection.

[0070] Results: Changes in preS1 antigen, Pres1 antibody in the serum, and HBsAg in the serum of Carrier mice immunized with the IFN-Pres1-Fc vaccine were detected. The results showed that after immunization with the IFNα-Pres1-Fc vaccine, the mice produced high levels of anti-Pres1 antibody (as shown in Figure 10(a)), and the preS1 antigen in the serum could be completely removed (as shown in Figure 10(b)). At the same time, HBsAg in the serum also decreased to some extent (as shown in Figure 10(c)), but there was no therapeutic effect in either the untreated control group or the simple Pres1 vaccine immunization group (as shown in Figure 10).

[0071] Example 6. T cell helper epitopes enhanced the antibody response to the IFNα-Pres1-Fc vaccine. The materials were the same as in Example 3.

[0072] Method: (1) Immunization of mice with the Pres1 fusion protein: Mice were subcutaneously immunized with 80 pmol of IFN-Pan-Pres1-Fc or 80 pmol of IFN-Pan-Pres1-Fc, Pres1-Fc, Pres1 protein containing the Pan epitope. Antibody detection was performed by collecting mouse serum by orbital bleeding at the designated time points. (2) Detection of anti-Pres1 specific antibodies in the serum by ELISA was the same as in Example 3.

[0073] Results: Compared with fusion protein vaccines such as IFN-preS1-Fc, IFN-Pan-preS1-Fc can significantly enhance the immunogenicity of antigen molecules and induce the production of a wide range of neutralizing antibodies. C57 / BL6 (n = 8 / group) mice were subcutaneously inoculated with hepatitis B Pres1, Pres1-Fc, IFNα-Pres1-Fc proteins without aluminum adjuvant, and the levels of Pres1 specific antibodies in the serum were detected by Elisa at the designated time.

[0074] Example 7. IFNα-Pan-Pres1-Fc was used as a preventive vaccine for chronic hepatitis B infection. Materials: C57BL / 6 male mice (4 weeks old) were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd. AAV-HBV 1.3 was purchased from Guangzhou Biomed Gene Technology Co., Ltd. The HBsAg detection kit was purchased from Shanghai Kehua Biotechnology Co., Ltd. Other experimental materials were the same as those in Example 4.

[0075] Methods: (1) Screening of HBV carrier mice: 1×10 11 μg of AAV-HBV 1.3 virus was injected into 4-week-old HBV C57BL / 6 mice via the tail vein. HBV antigen HBsAg was detected between 1 and 6 weeks, and mice with stable HBsAg expression were screened and used as HBV carrier mice. (2) The screened mice were subcutaneously injected with 80 pmol of different forms of Pres1 protein once every 2 weeks for a total of 3 immunizations. Mouse serum was collected 14 days after the first immunization and then weekly, and the levels of anti-Pres1 antibody, HBsAG, and Pres1 antigen in the mouse serum were detected by ELISA. After the last blood collection, the HBV-DNA content in the peripheral blood of the mice was detected.

[0076] Results: Changes in preS1 antigen, Pres1 antibody in serum, and HBsAg in the serum of carrier mice immunized with the IFN-Pan-Pres1-Fc vaccine were detected. The results showed that after immunization with the IFN-Pan-Pres1-Fc vaccine, the mice produced high levels of anti-Pres1 antibody (as shown in Figure 12(a)). And the preS1 antigen in the serum could be completely removed (as shown in Figure 12(b)), and the HBsAg in the serum also decreased to some extent (as shown in Figure 12(c)), but there was no therapeutic effect in either the untreated control group or the simple Pres1 vaccine immunization group. And the HBV DNA also decreased significantly in the IFNα-Pan-Pres1-Fc immunization group (as shown in Figure 12(d)).

[0077] Example 8. By combining IFNα-Pan-Pres1-Fc with a commercially available HBsAg vaccine, immune tolerance induced by HBsAg can be disrupted, and HBsAg-HBsAb seroconversion can be induced. Materials: C57BL / 6 male mice (4 weeks old) were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd. AAV-HBV 1.3 was purchased from Guangzhou Pisen Biotechnology Co., Ltd. The HBsAg detection kit was purchased from Shanghai Kehua Biotechnology Co., Ltd., and the Anti-HBsAg kit was purchased from Beijing Wantai Biological Pharmacy Co., Ltd. The commercially available HBsAg vaccine was purchased from Hanyin Vaccine (Dalian) Co., Ltd. Other experimental materials were the same as those in Example 7.

[0078] Methods: (1) Screening of HBV carrier mice: 1×10 11 μg of AAV-HBV 1.3 virus was injected into 4-week-old HBV C57BL / 6 mice via the tail vein. HBV antigen HBsAg was detected at 1 - 6 weeks, and mice with stable HBsAg expression were screened and used as HBV carrier mice. (2) The screened HBV carrier mice were immunized with 80 pmol of IFNα-pan-Pres1-Fc, and at the same time, 2 μg of the commercially available HBsAg vaccine was continuously immunized twice at 14-day intervals. Mouse serum was collected 14 days after the first immunization, and then weekly. Changes in anti-Pres1, Pres1, anti-HBsAg, and HBsAg in mouse serum were detected. And when the mouse serum was collected for the last time, the level of HBV-DNA in the serum was detected.

[0079] Results: As a treatment strategy for chronic hepatitis B, combining IFNα-Pan-Pres1-Fc with commercially available HBsAg can ultimately disrupt the immune tolerance caused by HBsAg. The immune response generated in HBV-resistant mice can completely remove the preS1 antigen in the serum (as shown in Figure 13(a)), and high concentrations of Pres1 antibodies are present in the serum (as shown in Figure 13(c)). Interestingly, the IFN-Pan-Pres1-Fc vaccine can effectively remove HBsAg in the serum while inducing a partial conversion of serological HBsAb (as shown in Figures 13(b) and 14(d)). This is considered an important clinical indicator for HBV cure. In addition, as a result of detecting the expression levels of HBV-related DNA in peripheral blood by fluorescence quantitative PCR (real-time PCR) method, compared with the control group, the immune method combining IFNα-Pan-Pres1-Fc and commercially available HBsAg can ultimately reduce the peripheral HBV DNA level (as shown in Figure 13(e)). Based on the above results, a vaccine strategy for the treatment of chronic hepatitis B with a vaccine combining IFNα-Pan-Pres1-Fc and commercially available HBsAg was invented.

[0080] Example 9. Compared with IFNα-RBD(SARS-CoV2)-Fc, the free SARS-Cov2 RBD protein can elicit a stronger antibody response. Materials: Balb / c male and female mice (6 - 8 weeks old) were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd., and the SARS-CoV-2 RBD protein used was purchased from Beijing Keyuan Zhongkai Biotechnology Co., Ltd. 293-hACE2 cells were provided by Professor Zhang Zheng (Shenzhen Third People's Hospital). The Luciferase Reporter detection kit was purchased from Promega. Other experimental materials were the same as those in Example 3.

[0081] Methods: (1) Immunization of mice with IFNα-RBD(SARS-Cov-2)-Fc fusion protein: 10 μg of IFNα-RBD-Fc, RBD-Fc or 10 μg of RBD protein was mixed with aluminum adjuvant and then mice were immunized subcutaneously. 28 days after immunization, mouse sera were collected by orbital bleeding for detection of novel coronavirus-specific antibodies. (2) Detection of SARS-cov2 RBD antibodies in sera. Antigen coating: RBD (1.5 μg / ml) coating solution was added to an Elisa plate (Corning 9018) at 100 μl / well and coated overnight at 4°C. The plate was washed once with 260 μl of PBS per well. Blocking was performed with 100 μl of 5% blocking solution (5% FBS) at 37°C for 2 hours. Serum samples (1:10, 1:100, 1:1000, 1:10000, 1:100000…) were diluted with PBS and added to the blocked Elisa plate at 100 μl / well, and incubated at 37°C for 1 hour. Each time, the plate was washed 5 times with 260 μl of PBST, 100 μl of enzyme-conjugated secondary antibody (enzyme-conjugated anti-mouse IgG-HRP diluted 1:5000 with PBS) was added to each well, and incubated at 37°C for 1 hour. Each time, the plate was washed with 260 μl of PBST, substrate TMB was added at 100 μl / well, and incubated at room temperature for 15 minutes in the dark to wait for the substrate to develop color. 50 μl of termination solution (2N H2SO4) was added to each well to terminate the color development, and the plate was read with a microplate reader at OD450-630. Method for calculating titer: Select the maximum dilution multiple that gave a positive result, and multiply the dilution multiple by the OD value corresponding to that dilution multiple / Cutoff value (0.1) (X) value as the antibody titer corresponding to that serum. (3) The in vitro neutralization experiment of SARS-CoV-2 protein pseudovirus: The antiserum was diluted 1:3 and added to a 96-well plate. 50 μl of pseudovirus particles containing luciferase spike protein were added to the wells. The virus-antibody mixture was left standing at 37°C for 1 hour, and 293-hACE2 cells were added to the 96-well plate at 10^4 / well. The 96-well plate was placed in a cell culture incubator at 37°C, and luciferase activity was detected after 48 hours.

[0082] Results: The immunogenicity of the free novel coronavirus is weak. When IFNα and the Fc portion are added to the novel coronavirus RBD polypeptide protein region to form the IFNα-RBD-Fc fusion protein, as shown in Figure 14, its immunogenicity is significantly improved. And, as shown in Figure 15, the antibodies produced by IFNα-RBD-Fc can block the infection of cells by the pseudovirus of the SARS-CoV-2 S protein in vitro.

[0083] Example 10. Detection of RBD-specific antibodies in antiserum produced by immunizing with IFNα-Pan-RBD (original strain)-Fc and IFNα-RBD (SARS-CoV-2 South African mutant strain)-Fc. Materials: Balb / c male and female mice (6 - 8 weeks old) were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd. The SARS-CoV-2 original strain RBD protein used was purchased from Beijing Key Laboratory of Biotechnology Co., Ltd. The SARS-CoV-2 South African mutant strain RBD protein used was purchased from Sino Biological, Inc. Other experimental materials were the same as those in Example 3.

[0084] Methods: (1) The construction and expression methods of IFNα-Pan-RBD (original strain)-Fc and IFNα-RBD (SARS-CoV-2 South African mutant strain)-Fc proteins were the same as those in Example 2. (2) Immunization of mice with IFNα-Pan-RBD (original strain)-Fc and IFNα-Pan-RBD (SARS-CoV-2 South African mutant strain)-Fc fusion proteins: 10 μg of IFNα-Pan-RBD (original strain)-Fc or IFNα-Pan-RBD (SARS-CoV-2 South African mutant strain)-Fc protein was mixed with an aluminum adjuvant and then used to immunize mice subcutaneously. Fourteen days after immunization, mouse sera were collected by orbital bleeding for detection of SARS-CoV-2-specific antibodies. (3) Analysis of the antibody response by ELISA was the same as in Example 9.

[0085] Results: The results of SDS-PAGE showed the exact band sizes of IFNα-Pan-RBD (SARS-CoV-2 original strain)-Fc, indicating the successful construction, expression, and purification of the mutant SARS-CoV-2 IFNα-RBD (SARS-CoV-2 original strain)-Fc vaccine protein (Figure 16a). The results of SDS-PAGE also showed the exact band sizes of IFNα-Pan-RBD (SARS-CoV-2 South African mutant strain)-Fc, indicating the successful construction, expression, and purification of the mutant SARS-CoV-2 IFNα-RBD (SARS-CoV-2 South African mutant strain)-Fc vaccine protein (Figure 16b). The results of ELISA showed that the antibodies induced in IFNα-Panan-RBD (original strain)-Fc-immunized mice, similar to those in IFNα-Pan-RBD (SARS-CoV-2 South African mutant strain)-Fc-immunized mice, were both able to bind to the RBD protein of the original strain of SARS-CoV-2, and there was no significant difference in the binding ability between the induced antibodies of both and the original strain RBD (Figure 16c). The results of ELISA for the South African mutant strain RBD showed that the antibodies induced in IFNα-Panan-RBD (original strain)-Fc-immunized mice, similar to those in IFNα-Pan-RBD (SARS-CoV-2 South African mutant strain)-Fc-immunized mice, were both able to bind to the South African mutant strain RBD, and their binding abilities were equivalent (Figure 16c).

[0086] Example 11. Materials: C57BL / 6 female mice (6 - 8 weeks old) were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd. The SARS-CoV-2 RBD protein used in ELISA was purchased from Beijing Key Laboratory of Biotechnology Co., Ltd. The Mouse IFNα-RBD-Fc, Mouse IFNα-Pan-RBD-Fc, Human IFNα-RBD-Fc, and Human IFNα-Pan-RBD-Fc proteins for immunization were all produced in our laboratory, and other experimental materials were the same as those in Example 3.

[0087] Methods: (1) The design of the fusion protein, plasmid construction, and protein purification methods were the same as those in Examples 1 and 2. (2) Immunization of mice with vaccine proteins 10 μg of Mouse IFNα-RBD-Fc, Mouse IFNα-Pan-RBD-Fc or 10 μg of Human IFNα-RBD-Fc, Human IFNα-Pan-RBD-Fc vaccine proteins were each mixed with 20 μg of aluminum adjuvant overnight, and then the mice were inoculated with the vaccine by intramuscular immunization. A booster immunization was performed 14 days after the first inoculation. Mouse sera were collected on the 7th, 14th, and 28th days after immunization, and the levels of RBD-specific antibodies in the mouse sera were detected by ELISA. (3) Detection antigen coating of SARS-cov2 RBD antibody in serum: 100 μl / well of RBD (1.5 μg / ml) coating solution was added to an Elisa plate (Corning 9018) and coated overnight at 4°C. The plate was washed once with 260 μl of PBS per well. It was blocked with 100 μl of 5% blocking solution (5% FBS) at 37°C for 2 hours. Serum samples (1:10, 1:100, 1:1000, 1:10000, 1:100000…) were diluted with PBS and added to the blocked Elisa plate at 100 μl / well and incubated at 37°C for 1 hour. Each time, it was washed 5 times with 260 μl of PBST, 100 μl of enzyme-labeled secondary antibody (enzyme-conjugated anti-mouse IgG-HRP diluted 1:5000 with PBS) was added to each well and incubated at 37°C for 1 hour. Each time, it was washed with 260 μl of PBST, 100 μl / well of substrate TMB was added, incubated at room temperature for 15 minutes avoiding light, and waited for the substrate to develop color. 50 μl of termination solution (2N H2SO4) was added to each well to terminate the color development, and the plate was read with an enzyme marker, and it was OD450-630. Method for calculating titer: Select the maximum dilution multiple that gave a positive result, and the (X) value obtained by multiplying the OD value corresponding to that dilution multiple by the dilution multiple / Cutoff value (0.1) corresponding to the serum was taken as the antibody titer corresponding to that serum.

[0088] Results: As shown in Figure 17, after expressing and purifying the protein, the results of SDS-PAGE showed that the size of the protein was as expected and a single band was displayed at the target position. As shown in Fig. 18, the addition of the Pan (Pan DR-binding epitope) CD4 T cell helper epitope can enhance the immunity of Mouse IFNα-RBD-Fc and Human IFNα-RBD-Fc. The experimental results show that regardless of whether it is the 7th day, 14th day, or 28th day after immunization with the vaccine protein, compared with Mouse IFNα-RBD-Fc, Mouse IFNα-Pan-RBD-Fc, and compared with Human IFNα-RBD-Fc, Human IFNα-Pan-RBD-Fc, the addition of the pan epitope results in higher production of RBD-specific antigens in both cases.

[0089] Example 12. Materials: C57BL / 6 female mice (6 - 8 weeks old) were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd., and the SARS-CoV-2 RBD protein used in ELISA was purchased from Beijing Key Laboratory Biotechnology Co., Ltd. The Human IFNα-RBD-Fc and Human IFNα-Pan-RBD-Fc proteins for immunization were both produced in our laboratory. Other experimental materials were the same as in Example 3.

[0090] Methods: (1) Immunization of mice with Human IFNα-RBD-Fc and Human IFNα-Pan-RBD-Fc proteins: 10 μg of Human IFNα-RBD-Fc or Human IFNα-Pan-RBD-Fc protein was mixed with aluminum adjuvant overnight and used as a vaccine sample containing aluminum adjuvant. In contrast, 10 μg of Human IFNα-RBD-Fc or Human IFNα-Pan-RBD-Fc protein was diluted with PBS and used as a vaccine sample without aluminum adjuvant. In the presence or absence of aluminum adjuvant, 10 μg of Human IFNα-RBD-Fc and Human IFNα-Pan-RBD-Fc proteins were inoculated into mice by intramuscular immunization, and booster immunization was performed 14 days after inoculation. Mouse sera were collected on the 7th, 14th, and 28th days after immunization, and the levels of RBD-specific antibodies in the mouse sera were detected by ELISA. (2) Detection of SARS-cov2 RBD antibodies in serum. Antigen coating: An RBD (1.5 ug / ml) coating solution was added to an Elisa plate (Corning 9018) at 100 ul / well and coated overnight at 4°C. The plate was washed once with 260 ul of PBS per well. Blocking was performed with 100 ul of 5% blocking solution (5% FBS) at 37°C for 2 hours. Serum samples (1:10, 1:100, 1:1000, 1:10000, 1:100000…) were diluted with PBS and added to the blocked Elisa plate at 100 ul / well, then incubated at 37°C for 1 hour. Each time, it was washed 5 times with 260 ul of PBST, 100 ul of an enzyme-conjugated secondary antibody (enzyme-conjugated anti-mouse IgG-HRP diluted 1:5000 with PBS) was added to each well, and incubated at 37°C for 1 hour. Each time, it was washed with 260 ul of PBST, substrate TMB was added at 100 ul / well, incubated at room temperature for 15 minutes avoiding light, and waited for the substrate to develop color. 50 ul of termination solution (2N H2SO4) was added to each well to terminate the color development, and the plate was read with an enzyme marker, and it was OD450-630. Method for calculating titer: Select the maximum dilution multiple that gave a positive result, and multiply the dilution multiple by the OD value corresponding to that dilution multiple / Cutoff value (0.1) (X) value as the antibody titer corresponding to that serum.

[0091] Results: As shown in Figure 19, the application of aluminum adjuvant can enhance the immunogenicity of Human IFNα-RBD-Fc and Human IFNα-Pan-RBD-Fc proteins. Without adjuvant, the Human IFNα-RBD-Fc and Human IFNα-Pan-RBD-Fc vaccines can produce a high-titer antibody response, but the Human IFNα-RBD-Fc and Human IFNα-Pan-RBD-Fc proteins assisted by aluminum adjuvant can further increase the level of RBD-specific antibody response on the 7th, 14th, and 28th days after vaccination compared with the non-adjuvant-assisted group.

[0092] Example 13. Materials: The experimental animals were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd. The animals used were 6 - 8 week - old C57BL / 6 mice; the animal certificate number was: No.110011200106828974; the RBD protein for immunization was purchased from Beijing Keyuan Zhongkai Biotechnology Co., Ltd., and the RBD - Fc, IFNα - RBD - Fc and IFN - pan - RBD - Fc proteins were produced in our laboratory; all adjuvants were purchased from SERVA, Germany; horseradish peroxidase (HRP) - labeled goat anti - mouse IgG was purchased from Beijing Kangwei Biotechnology Co., Ltd.; the 96 - well ELISA measurement board was purchased from Corning Costar; the ELISA chromogenic solution was purchased from eBioscience; the microplate reader SPECTRA max PLUS 384 used was purchased from Molecular, USA; the tissue homogenizer was purchased from Beijing Haonuosi Technology Co., Ltd.

[0093] Methods: 6 - 8 week - old mice were divided into 5 groups of 10 mice each. By intranasal immunization, 10 μg of IFNα - pan - RBD - Fc or an equimolar amount of RBD, RBD - Fc, and IFNα - RBD - Fc proteins were immunized respectively, and the nasal drop dosage was 10 μL / mouse. On day 0 and day 14, the mice were immunized using two immunization procedures. Mouse sera were collected on days 7, 14, 21, 28, 35, and 42 after immunization respectively, and the content of SARS - CoV - 2 RBD - specific antibodies in the sera of each group was detected by ELISA; sera were collected on day 28 for in vitro neutralization experiments of SARS - CoV - 2 pseudovirus.

[0094] Results: As shown in Figure 20, two intranasal immunizations with RBD and RBD-Fc proteins can induce a certain degree of antibody response. At the same time point after two intranasal immunizations with IFNα-pan-RBD-Fc, the levels of both serum IgG and IgA induced were significantly higher than those in the RBD, RBD-Fc, and IFN-RBD-Fc groups. The results of the pseudovirus neutralization experiment indicate that IFN-RBD-Fc can induce a higher level of neutralizing antibody production compared to the RBD and RBD-Fc immunized groups.

[0095] Example 14. Materials: The same as in Example 10.

[0096] Methods: Five 6- to 8-week-old mice were divided into four groups of five each. By intranasal immunization, each group was immunized with 10 μg of IFNα-pan-RBD-Fc or an equimolar amount of RBD, RBD-Fc, or IFNα-RBD-Fc protein, and the dosage was 10 μL / mouse. On days 0 and 14, the mice were immunized using a two-dose immunization procedure. On day 28 after immunization, the nasal mucosa supernatant and lung lavage fluid of the mice were collected. The content of SARS-CoV-2 RBD-specific antibodies in the serum of each group was detected by ELISA, and the SARS-CoV-2 pseudovirus in the serum and nasal mucosa supernatant was detected using a SARS-CoV-2 pseudovirus neutralization experiment. Obtaining nasal mucosa supernatant and lung lavage fluid from immunized experimental animal mice: The mice were anesthetized and sacrificed, and the nasal mucosa of the mice was collected and ground with a tissue homogenizer. The homogenized liquid was centrifuged at 13,000 rpm for 10 minutes at high speed, and the supernatant was used as the nasal mucosa supernatant (NMDS). In the lungs of the mice, approximately 0.8 mL of HBSS + 100 μM EDTA was aspirated with a 1 mL syringe, injected into the endotracheal tube, gently blown three times, and then the liquid was aspirated and collected in a centrifuge tube. This operation was repeated three times, and finally, approximately 2 mL of lung lavage fluid was obtained. The 500 g of mouse lung lavage fluid was centrifuged for 5 minutes, and the supernatant was used as the mouse lung lavage fluid (BALF). The precipitate was lymphocytes in the mouse lung and could be further analyzed.

[0097] Results: As shown in Figure 21, compared with the RBD and RBD-Fc proteins, the IFNα-pan-RBD-Fc protein can induce strong local IgG antibody responses and IgA mucosal immunity in the nasal mucosa by two intranasal immunizations. The intensity of the IFNα-pan-RBD-Fc protein response was stronger than that of the RBD and RBD-Fc groups. The results of the pseudovirus neutralization experiment indicate that the IFNα-pan-RBD-Fc protein immunization group can induce the production of higher titers of neutralizing antibodies in the nasal mucosa. As shown in Figure 22, two intranasal immunizations of C57BL / 6 mice with the IFNα-pan-RBD-Fc fusion protein also caused strong secretion of IgG and IgA antibodies in the local lung tissue. The results of the pseudovirus neutralization experiment show that IFNα-pan-RBD-Fc induced higher titers of neutralizing antibodies than RBD and RBD-Fc.

[0098] Example 15. Her2 belongs to the HER family of type I transmembrane growth factor receptors and consists of an extracellular ligand-binding domain, a transmembrane domain, and an intracellular tyrosine kinase domain. When a ligand binds to the extracellular domain, the HER proteins dimerize, transphosphorylate their intracellular domains, and the phosphorylated tyrosine residues bind to various intracellular signaling molecules, activating downstream signaling pathways and regulating gene transcription. Most of the genes regulated are related to cell proliferation, survival, differentiation, angiogenesis, invasion, and metastasis. The extracellular segment of the Her2 protein is relatively large, containing more than 600 amino acids, and can be divided into four structural domains, namely domain I, II, III, and IV. Currently approved Trastuzumab binds mainly to domain IV, Pertuzumab binds mainly to domain II, and the peptide vaccine E75 in clinical trials targets domain III. There are several important sites in different structural domains, indicating that they may mediate antitumor effects. To study a vaccine platform for tumor prevention and treatment, in this patent, the tumor antigen Her2 was targeted and selected, IFN-Her2-Fc and IFN-Pan-Her2-Fc were constructed, and a fusion protein vaccine was constructed, and its in vivo antitumor activity and vaccine immunological activity were analyzed.

[0099] Materials and methods: Materials: BALB / c female mice (6 - 8 weeks old) were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd., TUBO cells were derived from TCGA, and other materials were the same as in Example 3. Methods: (1) The design of the fusion protein, plasmid construction, and protein purification methods were the same as in Examples 1 and 2. First, expression plasmids were constructed for domains III and IV of the extracellular segment of mouse Her2 (designated as IFNα-3-Fc, IFNα-pan-3-Fc, IFNα-pan-4-Fc, and IFNα-4-Fc respectively), and the related proteins were expressed and purified in the human 293F cell line. The size and purity of the proteins were identified by SDS-PAGE and Coomassie Brilliant Blue staining. (2) Analysis of the direct antitumor activity of IFNα-3-Fc and IFNα-pan-3-Fc TUBO is a breast cancer cell line derived from BALB-NeuT mice and is used in the study of the growth and treatment of Her2-positive breast cancer. Using TUBO tumors, the antitumor activity of IFNα in the protein was detected. TUBO breast cancer model mice were constructed, and 5×105 TUBO cells were subcutaneously inoculated into BALB / C mice. When the tumor size reached 50 - 80 mm3, treatment was administered once a week for a total of 3 times. The dosage was 10 μg / mouse for IFNα-3-Fc, and other drugs were administered at equimolar amounts. CpG was used as an adjuvant. The tumor size was measured, and the tumor growth curve was plotted. (3) Analysis of the improvement of the immunogenicity of the Her2 vaccine by IFNα and Pan Six- to eight-week-old female BALB / C mice were subcutaneously inoculated once a week with the HER2 domain V fusion protein vaccines 4-Fc, IFNα-4-Fc, and IFNα-pan-4-Fc without adjuvant, for a total of 3 times. The immunization dosage was 10 μg / mouse for IFNα-4-Fc, and other proteins were inoculated at equimolar amounts. Venous blood was collected 14 days and 21 days after immunization, and the level of specific IgG antibodies against Her2 was detected by ELISA.

[0100] Results: (1) As shown in Figure 23, the size of the Her2 fusion protein was approximately as expected, and the purity also met the experimental requirements. IFNα-3-Fc (62.6 kDa), IFNα-pan-3-Fc (63.9 kDa), IFNα-pan-4-Fc (74.9 kDa), and IFNα-4-Fc (73.6 kDa). Under non-denaturing conditions, the protein was in a dimeric state, consistent with the auto-dimerization of the Fc fragment. (2) As shown in Figure 24, compared with the control group, intratumoral injection of Her2 fusion proteins IFNα-pan-3-Fc and IFNα-3-Fc significantly inhibited the growth of TUBO tumors, and the control effect was comparable to that of the IFNα-Fc group. The good IFNα activity in the protein vaccine indicates that there are no factors such as steric hindrance that affect IFNα activity, and it can be used to further explore its effectiveness and mechanism in anti-tumor immunity. (3) As shown in Figure 25, compared with the control group, 4-Fc, IFNα-4-Fc, and IFNα-pan-4-Fc could induce obvious Her2-specific IgG antibody responses 14 days and 21 days after Her2 fusion protein vaccine immunization. Compared with 4-Fc, the antibody titers induced by IFNα-4-Fc and IFNα-pan-4-Fc showed an upward trend. Also, on the 21st day after immunization, the antibody titer induced by IFNα-pan-4-Fc was significantly higher than that of the 4-Fc group. The addition of IFNα and pan contributed to the increase in the immunogenicity of 4-Fc and induced a stronger antigen-specific antibody response. Therefore, IFN-Pan-HER2-Fc and IFN-Pan-HER2-Fc were potentially effective tumor vaccines against Her2-positive tumors.

[0101] Example 16. Materials: BALB / c female mice (6 - 8 weeks old) were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd.; the HA1 (A / PR8) protein used in ELISA was purchased from Sino Biological, Inc.; the HA1 protein (A / PR 8) for immunization was purchased from Sino Biological, Inc.; IFNα-HA1-Fc was produced in our laboratory; the H1N1 (A / PR8) influenza virus used to infect mice was produced in our laboratory; other experimental materials were the same as those in Example 3.

[0102] Methods: (1) The design of IFNα-HA1-Fc protein, plasmid construction, and protein purification were the same as those in Examples 1 and 2. (2) Immunization of mice with HA1 and IFNα-HA1-Fc protein. 10 μg of IFNα-HA1-Fc or an equimolar amount of HA1 protein was mixed with 20 μg of aluminum adjuvant overnight and then inoculated into mice by intramuscular immunization. A booster immunization was performed 14 days after the first inoculation. Mouse sera were collected on the 28th day after immunization, and the levels of HA1-specific antibodies in the mouse sera were detected by ELISA. (3) Detection of HA1 antibodies in serum Coating of antigen: HA1 (2 μg / ml) coating solution was added to an ELISA plate (Corning 9018) at 100 μl / well and coated overnight at 4°C. The plate was washed once with 260 μl of PBS per well. Blocking was performed with 100 μl of 5% blocking solution (5% FBS) at 37°C for 2 hours. Serum samples (1:10, 1:100, 1:1000, 1:10000, 1:100000...) were diluted with PBS and added to the blocked ELISA plate at 100 μl / well, and incubated at 37°C for 1 hour. Each time, the plate was washed 5 times with 260 μl of PBST, 100 μl of enzyme-conjugated secondary antibody (enzyme-conjugated anti-mouse IgG-HRP diluted 1:5000 with PBS) was added to each well, and incubated at 37°C for 1 hour. Each time, the plate was washed with 260 μl of PBST, substrate TMB was added at 100 μl / well, and incubated at room temperature for 15 minutes in the dark to wait for the color development of the substrate. 50 μl of termination solution (2N H2SO4) was added to each well to terminate the color development, and the plate was read with a microplate reader at OD450-630. Calculation method of titer: Select the maximum dilution multiple that gave a positive result, and multiply the dilution multiple by the OD value corresponding to that dilution multiple / Cutoff value (0.1) (X) value as the antibody titer corresponding to that serum. (4) The mice were anesthetized 42 days after immunization, and 1000 PFU of A / PR8 influenza virus was infected into the mice by intranasal infection. The mice were observed every other day starting from the 3rd day after virus infection, and the changes in body weight were recorded.

[0103] Results: As shown in Fig. 26, after expressing and purifying the protein, the size and purity of the protein were detected by SDS-PAGE. As a result, a single band was shown at the position of the target band size. As shown in Fig. 27, compared with the HA1 protein, IFNα-HA1-Fc can induce higher-titer HA1-specific antibodies (Fig. 27a), indicating that the vaccine platform can enhance the immunogenicity of the HA1 protein. After the mice were challenged, the body weight changed significantly, but the body weight of the mice in the IFNα-HA1-Fc immunized group could recover rapidly compared with the PBS group and the HA1 protein immunized group. This indicates that the immunity induced by the IFNα-HA1-Fc vaccine has good protection against influenza infection (Fig. 27b).

[0104] Example 17 Materials and methods: The design of the IFNa-Pan-VZV-gE-Fc, IFNa-Pan-EBV-gp350-Fc, and IFNa-Pan-HSV-2-gD-Fc proteins, the construction of plasmids, and the purification of proteins were the same as in Examples 1 and 2.

[0105] Results: As shown in Fig. 28, after expressing and purifying the IFNa-Pan-VZV-gE-Fc, IFNa-Pan-EBV-gp350-Fc, and IFNa-Pan-HSV-2-gD-Fc fusion proteins, the measurement results of the size and purity of the proteins by SDS-PAGE indicate that the target bands are in the correct positions.

[0106] References: [1] WHO W. Global hepatitis report 2017 [J]. Geneva: World Health Organization, 2017, [2] RAZAVI-SHEARER D, GAMKRELIDZE I, NGUYEN M H, et al. Global prevalence, treatment, and prevention of hepatitis B virus infection in 2016: a modelling study [J]. The lancet Gastroenterology & hepatology, 2018, 3(6): 383-403. [3] SCHWEITZER A, HORN J, MIKOLAJCZYK R T, et al. Estimations of worldwide prevalence of chronic hepatitis B virus infection: a systematic review of data published between 1965 and 2013 [J]. The Lancet, 2015, 386(10003): 1546-55. [4] VOS T, ABAJOBIR A A, ABATE K H, et al. Global, regional, and national incidence, prevalence, and years lived with disability for 328 diseases and injuries for 195 countries, 1990-2016: a systematic analysis for the Global Burden of Disease Study 2016 [J]. The Lancet, 2017, 390(10100): 1211-59. [5] KRAMMER F, SMITH G J D, FOUCHIER R A M, et al. Influenza [J]. Nature reviews Disease primers, 2018, 4(1): 3. [6] HANNOUN C. The evolving history of influenza viruses and influenza vaccines [J]. Expert review of vaccines, 2013, 12(9): 1085-94. [7] HOUSER K, SUBBARAO K. Influenza vaccines: challenges and solutions [J]. Cell host & microbe, 2015, 17(3): 295-300. [8] HU B, GUO H, ZHOU P, et al. Characteristics of SARS-CoV-2 and COVID-19 [J]. Nature Reviews Microbiology, 2020, 1-14. [9] LURIE N, SAVILLE M, HATCHETT R, et al. Developing Covid-19 vaccines at pandemic speed [J]. New England Journal of Medicine, 2020, 382(21): 1969-73.

[10] LI Q, WU J, NIE J, et al. The impact of mutations in SARS-CoV-2 spike on viral infectivity and antigenicity [J]. Cell, 2020, 182(5): 1284-94. e9.

[11] KONTERMANN R E. Strategies for extended serum half-life of protein therapeutics [J]. Current opinion in biotechnology, 2011, 22(6): 868-76.

[12] MEKHAIEL D N, CZAJKOWSKY D M, ANDERSEN J T, et al. Polymeric human Fc-fusion proteins with modified effector functions [J]. Scientific reports, 2011, 1(1): 1-11.

[13] ROOPENIAN D C, AKILESH S. FcRn: the neonatal Fc receptor comes of age [J]. Nature reviews immunology, 2007, 7(9): 715-25.

[14] VILCEK J. Fifty years of interferon research: aiming at a moving target [J]. Immunity, 2006, 25(3): 343-8.

[15] BRACCI L, LA SORSA V, BELARDELLI F, et al. Type I interferons as vaccine adjuvants against infectious diseases and cancer [J]. Expert review of vaccines, 2008, 7(3): 373-81.

[16] HAHM B, TRIFILO M J, ZUNIGA E I, et al. Viruses evade the immune system through type I interferon-mediated STAT2-dependent, but STAT1-independent, signaling [J]. Immunity, 2005, 22(2): 247-57.

[17] ITO T, AMAKAWA R, INABA M, et al. Differential regulation of human blood dendritic cell subsets by IFNs [J]. The Journal of Immunology, 2001, 166(5): 2961-9.

[18] MONTOYA M, SCHIAVONI G, MATTEI F, et al. Type I interferons produced by dendritic cells promote their phenotypic and functional activation [J]. Blood, 2002, 99(9): 3263-71.

[19] LE BON A, ETCHART N, ROSSMANN C, et al. Cross-priming of CD8+ T cells stimulated by virus-induced type I interferon [J]. Nature immunology, 2003, 4(10): 1009-15.

[20] LE BON A, DURAND V, KAMPHUIS E, et al. Direct stimulation of T cells by type I IFN enhances the CD8+ T cell response during cross-priming [J]. The Journal of Immunology, 2006, 176(8): 4682-9.

[21] SPADARO F, LAPENTA C, DONATI S, et al. IFN-α enhances cross-presentation in human dendritic cells by modulating antigen survival, endocytic routing, and processing [J]. Blood, The Journal of the American Society of Hematology, 2012, 119(6): 1407-17.

[22] PARLATO S, SANTINI S M, LAPENTA C, et al. Expression of CCR-7, MIP-3β, and Th-1 chemokines in type I IFN-induced monocyte-derived dendritic cells: importance for the rapid acquisition of potent migratory and functional activities [J]. Blood, The Journal of the American Society of Hematology, 2001, 98(10): 3022-9.

[23] ROUZAUT A, GARASA S, TEIJEIRA A, et al. Dendritic cells adhere to and transmigrate across lymphatic endothelium in response to IFN‐α [J]. European journal of immunology, 2010, 40(11): 3054-63.

[24] LE BON A, SCHIAVONI G, D'AGOSTINO G, et al. Type I interferons potently enhance humoral immunity and can promote isotype switching by stimulating dendritic cells in vivo [J]. Immunity, 2001, 14(4): 461-70.

Claims

A vaccine comprising a fusion protein containing interferon, a target antigen, and an immunoglobulin Fc region, wherein: the interferon is a first structural unit and is selected from IFN-α and IFN-γ; the immunoglobulin Fc region is a second structural unit, and the target antigen is a third structural unit, the vaccine.

2. The following features: 1) The first structural unit is IFN-α; 2) The first structural unit is derived from human or murine; 3) The first structural unit is selected from mouse IFN-α4, human IFN-α2, and variants of human IFN-α2 that bind to human and mouse IFN receptors; 4) The first structural unit is selected from amino acid sequences SEQ ID NO: 1, SEQ ID NO: 21, SEQ ID NO: 22; 5) The second structural unit is selected from the Fc regions of IgG1, IgG2, IgG3, IgG4, and IgM; 6) The second structural unit is selected from Fc regions represented by amino acid sequences SEQ ID NO: 2, SEQ ID NO: 23, SEQ ID NO: 24; 7) The third structural unit is a tumor antigen or a pathogen antigen; 8) The third structural unit is a viral or bacterial antigen; 9) The vaccine is a target vaccine; and 10) The fusion protein contains one or more Th cell helper epitopes and / or linker fragments, The vaccine according to claim 1, having any of the above.

3. The following features: 1) The third structural unit is a viral antigen, and the virus is HBV, HPV, VZV, EBV, HSV-2, HIV, influenza virus, or coronavirus; 2) The third structural unit is a viral antigen, and the virus is SARS-COV, SARS-COV-2, or MERS-CoV; 3) The third structural unit is an HBV Pres1 antigen, an HBsAg antigen, or a peptide fragment; 4) The third structural unit is an ad subtype or ay subtype HBV Pres1 antigen; 5) The third structural unit is an ad subtype HBV Pres1 antigen represented by the amino acid sequence of SEQ ID NO: 6, an ay subtype HBV Pres1 antigen represented by the amino acid sequence of SEQ ID NO: 26; an adr subtype HBV HBsAg antigen represented by the amino acid sequence of SEQ ID NO: 7, an adw subtype HBV HBsAg antigen represented by the amino acid sequence of SEQ ID NO: 27, an ayw subtype HBV HBsAg antigen represented by the amino acid sequence of SEQ ID NO: 28; 6) The third structural unit is a SARS-COV2 RBD antigen; 7) The third structural unit is a SARS-COV2 RBD antigen represented by the amino acid sequence of SEQ ID NO: 8; 8) The third structural unit is an influenza virus antigen; 9) The third structural unit is an influenza virus HA antigen; 10) The third structural unit is an influenza virus HA antigen represented by the amino acid sequence of SEQ ID NO: 9; 11) The third structural unit is an HPV E7 antigen represented by the amino acid sequence of SEQ ID NO: 10; 12) The third structural unit is a gE antigen; 13) The third structural unit is a varicella-zoster virus (VZV) gE antigen represented by the amino acid sequence of SEQ ID NO: 91; 14) The third structural unit is EBV-gp350; 15) The third structural unit is an Epstein-Barr virus (EBV) gp350 protein represented by the amino acid sequence of SEQ ID NO: 92; 16) The third structural unit is the gD antigen; 17) The third structural unit is the glycoprotein D (gD) antigen of herpes simplex virus 2 (HSV-2) represented by the amino acid sequence of SEQ ID NO: 93; 18) The third structural unit is the EBV EBNA1 / LMP2, VZV-IE62, HSV-2 ICP0, or HIV gp120 antigen; 19) The third structural unit is a mutant virus antigen; 20) The third structural unit is a variant of SARS-CoV-2; 21) The third structural unit is a natural point mutation / deletion mutation / insertion mutation / truncation of a SARS-CoV-2 protein, an artificial point mutation / deletion mutation / insertion mutation / truncation, any combination of natural or artificial mutations, or a subtype generated after mutation; 22) The third structural unit is one or more variants of the S protein, N protein, M protein, or E protein; 23) The third structural unit is a variant of the full-length S protein of SEQ ID NO: 76, the S1 region of SEQ ID NO: 77, or the RBD region of SEQ ID NO: 78 of wild-type SARS-CoV-2; 24) The third structural unit contains at least one mutation selected from the group consisting of the NTD region 69-70 deletion, Y144 deletion, 242-244 deletion, L18F, D80A, D215, R246I mutations, the RBD region K417, E484, N501Y, L452R mutations, D614G, and H655Y mutations of the SARS-CoV-2 S protein; 25) The third structural unit contains mutations derived from the B.1.1.7 (501Y.1) variant strain in the UK, the B.1.351 (501Y.2) variant strain in South Africa, the P1 (501Y.3) variant strain in Brazil, or the B.1.429 variant strain in California; 26) The third structural unit contains a variant of a mutation represented by any one of SEQ ID NO: 79, SEQ ID NO: 80, SEQ ID NO: 81, or SEQ ID NO: 82; 27) The third structural unit comprises a variant of the sequence represented by any one of SEQ ID NO: 79, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 82; 28) The third structural unit enhances the responses of B cells and T cells and fuses a helper polypeptide epitope located at the N-terminus or C-terminus of the antigen epitope; 29) The third structural unit fuses a Pan HLA DR-binding epitope (PADER); 30) The third structural unit fuses the one represented by the amino acid sequence of SEQ ID NO: 3; 31) The fusion protein contains one or more linking fragments, and the linking fragment of each structural unit is a flexible polypeptide sequence; 32) The fusion protein contains one or more linking fragments represented by the amino acid sequences of SEQ ID NO: 4, SEQ ID NO: 25; 33) At the N-terminus of each polypeptide sequence consisting of the said structural unit, there is included a corresponding signal peptide capable of promoting protein secretion; and 34) At the N-terminus of each polypeptide sequence consisting of the said structural unit, there is included a corresponding signal peptide represented by the amino acid sequence of SEQ ID NO: 5, The vaccine according to claim 1 or 2, having any one of the above.

4. The following features: 1) The third structural unit is a tumor antigen containing a tumor cell highly expressed protein molecule; 2) The third structural unit is selected from human epidermal growth factor receptor 2 (HER2 / neu) and Epidermal growth factor receptor (EGFR); 3) The third structural unit is selected from the tumor cell highly expressed protein molecule Her2 and its functional regions and cleavage products; and 4) The third structural unit is selected from the antigens represented by SEQ ID NO: 85, 86, 87, 88, 89, 90, and has any one of the following, the vaccine according to claim 1 or 2.

5. The fusion protein is a homodimeric or heterodimeric fusion protein comprising a first polypeptide chain and a second polypeptide chain, the vaccine according to any one of claims 1 to 4.

6. The following features: 1) The fusion protein comprises one or more Th cell helper epitopes and / or linker fragments in the homodimeric or heterodimeric first polypeptide chain and / or the second polypeptide chain; 2) The first polypeptide chain and the second polypeptide chain are completely identical; 3) The first polypeptide chain and the second polypeptide chain form a polypeptide in the order from the N-terminus to the C-terminus, including IFN, a target antigen, and an immunoglobulin Fc region, or in any combination order of the three structural units to generate a homodimer; 4) The first polypeptide chain and the second polypeptide chain include IFN, a target antigen, and an immunoglobulin Fc region in the order from the N-terminus to the C-terminus; 5) The first polypeptide chain and the second polypeptide chain are not the same polypeptide chain; 6) The first polypeptide chain includes IFN and an immunoglobulin Fc region in the order from the N-terminus to the C-terminus, or includes an immunoglobulin Fc region and IFN in the order from the N-terminus to the C-terminus, the second polypeptide chain includes a target antigen and an immunoglobulin Fc region, where the target antigen is located at the N-terminus and the immunoglobulin Fc region is located at the C-terminus, or the immunoglobulin Fc region is located at the N-terminus and the target antigen is located at the C-terminus, or forms a polypeptide in any combination order of the three structural units to generate a heterodimer; 7) The IFN and the target antigen are each located at the N-terminus of the first polypeptide chain and the second polypeptide chain, respectively, and the immunoglobulin Fc region is located at the C-terminus of the first polypeptide chain and the second polypeptide chain. The vaccine according to claim 5, having any one of .

7. The following features: 1) The first polypeptide and the second polypeptide of the homodimer are selected from the amino acid sequences represented by SEQ ID NO: 11, 12, 13, 14, 29, 30, 31, 32, 38, 39, 40, 47, 48, 49, 50, 51, 56, 57, 59, 58, 65, 66, 67, 68; 2) The first polypeptide of the heterodimer is selected from the amino acid sequences represented by SEQ ID NO: 15, 33, 42, 51, 60, 69, and the second polypeptide is selected from the amino acid sequences represented by SEQ ID NO: 16, 17, 18, 19, 34, 35, 36, 37, 43, 44, 45, 46, 52, 53, 54, 55, 61, 62, 63, 64, 70, 71, 72, 73. The vaccine according to claim 5, having any one of .

8. A nucleic acid molecule encoding a fusion protein in the vaccine according to any one of claims 1 to 7.

9. An expression vector containing the nucleic acid molecule according to claim 8.

10. A host cell containing the nucleic acid molecule according to claim 8 or the expression vector according to claim 9.

11. The host cell according to claim 10, which is a eukaryotic cell.

12. Use of the fusion protein in the vaccine according to any one of claims 1 to 7 in the preparation of a composition.

13. The following features: 1) The composition is in a kit; 2) The composition is a pharmaceutical composition or an immunogenic composition; and 3) The fusion protein is expressed in a recombinant microorganism or cell line, The use according to claim 12, having any one of the above.

14. The composition is as follows: 1) Prevention or treatment of tumors or pathogens; 2) Prevention or treatment of viruses or bacteria, where the virus is selected from HBV, HPV, EBV, influenza virus, HIV, and coronavirus; 3) Prevention or treatment of SARS-CoV, SARS-CoV-2, or MERS-CoV; 4) The composition is used as a hepatitis B prevention or treatment vaccine, HBV prevention or treatment vaccine, influenza prevention or treatment vaccine, SARS-CoV2 prevention or treatment vaccine, HPV prevention or treatment vaccine, HPV-related tumor prevention or treatment vaccine, EBV prevention or treatment vaccine, EBV-related tumor prevention or treatment vaccine, or HIV prevention or treatment vaccine. The use according to claim 12, used for any one of the above.

15. The following characteristics: 1) The vaccine is inoculated by an immune route such as intramuscular, intravenous, transdermal, subcutaneous, or nasal; 2) The vaccine contains an adjuvant; and 3) The vaccine contains an adjuvant, and the adjuvant is selected from aluminum adjuvant (Alum), Toll-like receptor 4 activator ligand MPLA, Toll-like receptor 9 ligand, oligodeoxynucleotide (CpG-ODN), M59, and Freund's adjuvant. The use according to any one of claims 12 to 14, having any one of the above.

16. The following characteristics: 1) The vaccine is used in combination with another prevention or treatment therapy; 2) The vaccine is a HBV treatment vaccine, and the HBV treatment vaccine is used in combination with another prophylactic or therapeutic HBV therapy; 3) The vaccine is a HBV treatment vaccine, and the HBV treatment vaccine is used in combination with a hepatitis B virus envelope protein HBsAg vaccine and is used for the treatment of chronic hepatitis B virus infection; 4) The vaccine is a HBV treatment vaccine, and the HBV treatment vaccine is used in combination with a nucleoside or nucleotide analog and is used for the treatment of chronic hepatitis B virus infection; 5) The vaccine is an influenza, SARS-COV2, HPV, EBV, HIV prophylactic or therapeutic vaccine; 6) The vaccine is used for combined use with antiviral drugs and other therapies; 7) The vaccine is an HPV or EBV-related tumor prophylactic or therapeutic vaccine and is used for combined use with antiviral and tumor drugs and therapies; 8) The vaccine is combined with other virus or pathogen vaccines or tumor vaccines to form a multivalent mixed vaccine; 9) The vaccine is a SARS-COV-2 vaccine and is combined with other vaccines to form a multivalent mixed vaccine; 10) The vaccine is a SARS-COV-2 vaccine and is combined with an influenza vaccine to form a multivalent mixed vaccine; 11) The vaccine and an adenovirus vaccine or mRNA vaccine or inactivated vaccine or DNA vaccine of the same virus, pathogen or tumor are immunized by sequential or simultaneous immunization procedures; and 12) The vaccine is a SARS-COV-2 fusion protein vaccine, and the vaccine and an adenovirus vaccine or mRNA vaccine or inactivated vaccine or DNA vaccine of SARS-COV-2 are used for immunization in a sequential or simultaneous immunization procedure. The order of immunization is as follows: 1) First, immunize with the SARS-COV-2 fusion protein vaccine, and then immunize with an adenovirus vaccine or mRNA vaccine or inactivated vaccine or DNA vaccine of SARS-COV-2; 2) First, immunize with an adenovirus vaccine or mRNA vaccine or inactivated vaccine or DNA vaccine of SARS-COV-2, and then immunize with the SARS-COV-2 fusion protein vaccine; 3) Simultaneously immunize with the SARS-COV-2 fusion protein vaccine and an adenovirus vaccine or mRNA vaccine or inactivated vaccine or DNA vaccine of SARS-COV-2. The use according to any one of claims 12 to 14, having any one of the above.

Images