Fusion proteins comprising varicella zoster virus proteins and uses thereof

Abstract

The present application discloses a fusion protein comprising a varicella zoster virus protein and a ferritin, a nucleic acid encoding the fusion protein, a pharmaceutical composition comprising the fusion protein, a method of preparing the fusion protein, and a use of the fusion protein.

Description

Technical Field

[0001] The present application generally relates to the fields of genetic engineering, biology and medicine. Specifically, the present application relates to a fusion protein comprising varicella zoster virus protein and ferritin and its use. Background Art

[0002] Varicella-zoster virus (VZV) is a widespread virus that is prevalent worldwide. The first infection with VZV causes chickenpox in infants and young children. Subsequently, VZV remains latent in human spinal ganglion cells and may relapse when the immune system is weakened and cause the symptoms of herpes zoster. Herpes zoster can be complicated by chronic pain (postherpetic neuralgia (PHN)) and other serious neurological and ocular diseases (e.g., meningoencephalitis, myelitis, cranial nerve palsies, vasculopathy, keratitis, and retinopathy), as well as a variety of visceral and gastrointestinal diseases (e.g., ulcers, hepatitis, and pancreatitis). 1 .

[0003] Innate and adaptive immunity are critical to limiting VZV-mediated diseases. After acute VZV infection or varicella vaccination, the immune response formed can maintain a long-term immune response to VZV and play a role in preventing varicella. 2,3 For the prevention of herpes zoster caused by VZV reactivation, VZV-specific cellular immunity may be more critical. 4,5 In elderly subjects or immunocompromised patients, the risk of herpes zoster virus infection increases as VZV-specific T cell immunity decreases. 6,7 .

[0004] Currently, two varicella zoster virus vaccines are available on the market, including the live attenuated vaccine Zostavax, which was launched in 2006, and the recombinant subunit vaccine SHINGRIX, which was launched in 2017. The SHINGRIX vaccine contains a single gE glycoprotein and a special AS01B adjuvant system, and studies have shown that it has higher safety and better protection than Zostavax. 8,9 The adjuvant component significantly enhanced the immunogenicity of gE, especially cellular immunity.

[0005] Therefore, the preparation of new vaccines against herpes zoster virus has important research and application value. SUMMARY OF THE INVENTION

[0007] In a first aspect, the present application provides a fusion protein comprising a varicella-zoster virus protein and ferritin.

[0008] In some embodiments of the first aspect, the varicella-zoster virus protein is a varicella-zoster virus glycoprotein or a varicella-zoster virus interstitial protein.

[0009] In some embodiments of the first aspect, the ferritin is selected from the group consisting of classical ferritin, heme-containing bacterial ferritin, and DNA-binding ferritin of starved cells.

[0010] In some embodiments of the first aspect, the varicella-zoster virus protein comprises or consists of a sequence selected from:

[0011] (a) the amino acid sequence shown in SEQ ID NO: 1; and

[0012] (b) A variant obtained by deleting 1 to 50 amino acids from the amino acid sequence shown in SEQ ID NO: 1 at the C-terminus.

[0013] In some embodiments of the first aspect, the ferritin comprises or consists of a sequence selected from:

[0014] (c) the amino acid sequence shown in SEQ ID NO: 2; and

[0015] (d) A variant obtained by subjecting the amino acid sequence shown in SEQ ID NO: 2 to one or more substitutions, deletions or additions.

[0016] In some embodiments of the first aspect, the ferritin is linked to the C-terminus of the varicella-zoster virus protein.

[0017] In some embodiments of the first aspect, the fusion protein further comprises a purification tag; optionally, the purification tag is connected to the N-terminus or C-terminus of the varicella-zoster virus protein.

[0018] In some embodiments of the first aspect, the fusion protein comprises or consists of a sequence selected from the group consisting of:

[0019] (e) an amino acid sequence as shown in any one of SEQ ID NOs: 4-11; and

[0020] (f) A variant obtained by subjecting the amino acid sequence shown in any one of SEQ ID NOs: 4-11 to one or more substitutions, deletions or additions.

[0021] In a second aspect, the present application provides a nucleic acid encoding the fusion protein described in the first aspect.

[0022] In a third aspect, the present application provides a pharmaceutical composition comprising the fusion protein described in the first aspect, and a pharmaceutically acceptable excipient, diluent or carrier.

[0023] In a fourth aspect, the present application provides a method for preparing the fusion protein described in the first aspect, comprising: transferring the nucleic acid described in the second aspect into a host cell, culturing the host cell; and harvesting the fusion protein.

[0024] In a fifth aspect, the present application provides use of the fusion protein described in the first aspect or the pharmaceutical composition described in the third aspect in the preparation of a medicament for preventing varicella-zoster virus infection or a disease caused by the infection. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 A schematic diagram of the structure of a fusion protein comprising varicella-zoster virus protein and ferritin prepared in the present application is shown.

[0027] Figure 2 The results of double sandwich ELISA for detecting the expression of gE-His, His-ferritin, His-gE-ferritin and gE-His-ferritin in cell supernatant are shown.

[0028] Figure 3 The results of double sandwich ELISA detection of the expression of gE-His, gE-ferritin-A, gE-ferritin-B, gE-ferritin-C, gE-ferritin-D, gE-ferritin-E and gE-ferritin-F in the cell supernatant are shown.

[0029] Figure 4 The SEC results of the fusion proteins are shown, where AC represents His-ferritin, gE-ferritin-D, and gE-ferritin-E, respectively.

[0030] Figure 5 The DLS results of the fusion proteins are shown, where AC represents His-ferritin, gE-ferritin-D, and gE-ferritin-E, respectively.

[0031] Figure 6 The results of ELISA for detecting the serum titer of each fusion protein are shown, wherein A represents the results of gE-His coating for detecting the serum titer of each fusion protein, and B represents the results of His-ferritin coating for detecting the serum titer of each fusion protein.

[0032] Figure 7 The results of T cell immune response to gE-ferritin fusion protein detected by ELISPOT are shown.

[0033] Sequence Description

[0034] SEQ ID NO: 1 shows the amino acid sequence of the extracellular region of glycoprotein E (gE) of varicella-zoster virus of the Oka strain.

[0035] SEQ ID NO: 2 shows the amino acid sequence of Helicobacter pylori non-heme iron protein.

[0036] SEQ ID NO: 3 shows the amino acid sequence of the His tag.

[0037] SEQ ID NO: 4 shows the amino acid sequence of the fusion protein His-gE-ferritin.

[0038] SEQ ID NO: 5 shows the amino acid sequence of the fusion protein gE-His-ferritin.

[0039] SEQ ID NO: 6 shows the amino acid sequence of the fusion protein gE-ferritin-A.

[0040] SEQ ID NO: 7 shows the amino acid sequence of the fusion protein gE-ferritin-B.

[0041] SEQ ID NO: 8 shows the amino acid sequence of the fusion protein gE-ferritin-C.

[0042] SEQ ID NO:9 shows the amino acid sequence of the fusion protein gE-ferritin-D.

[0043] SEQ ID NO: 10 shows the amino acid sequence of the fusion protein gE-Ferritin-E.

[0044] SEQ ID NO: 11 shows the amino acid sequence of the fusion protein gE-ferritin-F.

[0045] SEQ ID NO: 12 shows the amino acid sequence of the extracellular region of glycoprotein E (gE) of the Oka strain varicella-zoster virus after 17 amino acids are truncated from the C-terminus.

[0046] SEQ ID NO: 13 shows the amino acid sequence of the extracellular region of glycoprotein E (gE) of the Oka strain varicella-zoster virus after truncating 31 amino acids at the C-terminus.

[0047] SEQ ID NO: 14 shows the amino acid sequence of the extracellular region of glycoprotein E (gE) of the Oka strain varicella-zoster virus after 41 amino acids are truncated from the C-terminus.

[0048] SEQ ID NO: 15 shows the amino acid sequence of linker 1.

[0049] SEQ ID NO: 16 shows the amino acid sequence of linker 2.

[0050] SEQ ID NO: 17 shows the amino acid sequence of linker 4. DETAILED DESCRIPTION OF THE INVENTION

[0052] The inventors of the present application have obtained a new fusion protein comprising varicella zoster virus protein and ferritin through genetic engineering technology. In various aspects of the present application, a new fusion protein comprising varicella zoster virus protein and ferritin, a nucleic acid encoding the fusion protein, a pharmaceutical composition comprising the fusion protein, a method for preparing the fusion protein, and the medical and biological applications of the fusion protein are provided. According to the amino acid sequence of the fusion protein provided in the present application, the fusion protein can be constructed as a drug for preventing varicella zoster virus infection or diseases caused by infection (e.g., chickenpox, herpes zoster, or chronic pain concurrent with chickenpox or herpes zoster, neurological and eye diseases, or visceral and gastrointestinal diseases).

[0053] The practice of the present application employs, unless otherwise indicated, conventional techniques of molecular biology, microbiology, cell biology, biochemistry, and immunology within the skill of the art.

[0054] Unless otherwise specified, the terms used in this application have the meanings commonly understood by those skilled in the art.

[0055] definition

[0056] As used herein, the term "varicella-zoster virus protein" refers to a protein encoded by the varicella-zoster virus (VZV) genome. The VZV genome is stable and has a low mutation rate. It has 69 open reading frames (ORFs), 7 of which encode glycoproteins. In some embodiments of the present application, the varicella-zoster virus protein is a varicella-zoster virus glycoprotein or a varicella-zoster virus interstitial protein. In some embodiments, the varicella-zoster virus protein is a gE, gI, gB, gH, gL or gC glycoprotein. The envelope glycoprotein gE encoded by ORF68 in the VZV genome has the highest abundance and the strongest immunogenicity, contains important T cell and B cell epitopes, and stimulates the production of high-abundance neutralizing antibodies in the body during natural infection, and gE can also stimulate cellular immunity. In some specific embodiments of the present application, the varicella-zoster virus protein is a gE glycoprotein.

[0057] As used herein, the term "ferritin" is a widely-existing iron storage protein. Ferritin superfamily members are divided into three subclasses: classical ferritins (Ftn), heme-containing bacterioferritins (Bfr) and DNA-binding proteins from starved cells (Dps). Ftn exists in plants, animals and bacteria, while Bfr and Dps exist only in prokaryotes. Ftn and Bfr are composed of 24 subunits, which self-assemble into hollow symmetrical octahedral nanoparticles with an inner diameter and an outer diameter of 8 nm and 12 nm, respectively. The particle shell is hydrophilic, and a hydrophobic symmetry axis is formed inside, which has good stability. Although ferritins from different sources vary greatly in primary structure (the amino acid sequence similarity sometimes reaches 14%), and some bacterial ferritins even contain heme, they essentially have the same system architecture, very similar tertiary structures, and functional subdomains are also very conserved in sequence between different species. The size and structure of naturally self-assembled ferritin nanoparticles are similar to those of pathogens, making them an attractive antigen display platform. Self-assembly into nanoparticles is a common feature of all ferritins, so in principle all ferritins can be used in nanoparticle vaccines. Since the vaccine prepared in this application is used in humans, considering that ferritin immunization with sequences similar to human ferritin may cause autoimmune reactions, ferritin from species distantly related to humans, such as bacterial ferritin, is preferred.

[0058] As used herein, the term "fusion protein", also known as chimeric protein, refers to a protein produced by connecting two or more genes that originally encode proteins independently.

[0059] As used herein, the term "nanoparticle" refers to an artificially manufactured microparticle with a size not exceeding 100 nanometers. In some embodiments of the present application, the nanoparticle is a nanoparticle self-assembled from a fusion protein comprising varicella zoster virus protein and ferritin.

[0060] As used herein, the term "immunogenicity" refers to the ability to induce an immune response, i.e., the ability of an antigen to stimulate specific immune cells, activate, proliferate, and differentiate immune cells, and ultimately produce immune effector substances such as antibodies and sensitized lymphocytes. The immunogenicity of an antigen is first determined by its own chemical properties, but the immunogenicity of the same antigen to different species of animals or different individuals of the same species can vary greatly, so the immunogenicity of an antigen is determined by its chemical properties and host factors.

[0061] As used herein, the term "immunogen" refers to a substance that has immunogenic properties.

[0062] In this article, a variant "does not substantially change" the activity or function of the parent means that, in terms of the desired activity or function, the variant is identical to the parent at a qualitative level and is at least similar in a quantitative sense, such as by ±5%, ±10%, ±15%, ±20%, ±25% or ±30%. For example, the amino acid sequence shown in SEQ ID NO: 1 is deleted at one or more positions without substantially changing the activity of the amino acid sequence shown in SEQ ID NO: 1, which means that the ability of the variant to induce an immune response is ±5%, ±10%, ±15%, ±20%, ±25% or ±30% different from the amino acid sequence shown in SEQ ID NO: 1.

[0063] As used herein, the amino acid residues are abbreviated as follows: alanine is Ala or A; arginine is Arg is R; asparagine is Asn or N; aspartic acid is Asp or D; cysteine ​​is Cys or C; glutamic acid is Glu or E; glutamine is Gln or Q; glycine is Gly or G; histidine is His or H; isoleucine is Ile or I; leucine is Leu or L; lysine is Lys or K; methionine is Met or M; phenylalanine is Phe or F; proline is Pro or P; serine is Ser or S; threonine is Thr or T; tryptophan is Trp or W; tyrosine is Tyr or Y; and valine is Val or V.

[0064] It is well known to those skilled in the art that certain substitutions can result in retention of activity. For example, amino acids can be described as, for example, polar, non-polar, acidic, basic, aromatic, or neutral. Polar amino acids are amino acids that can interact with water through hydrogen bonds at biological or near-neutral pH. The polarity of an amino acid is an indication of the degree of hydrogen bonding at biological or near-neutral pH. Examples of polar amino acids include serine, proline, threonine, cysteine, asparagine, glutamine, lysine, histidine, arginine, aspartic acid, tyrosine, and glutamic acid. Examples of non-polar amino acids include glycine, alanine, valine, leucine, isoleucine, methionine, phenylalanine, and tryptophan. Acidic amino acids have a net negative charge at neutral pH. Examples of acidic amino acids include aspartic acid and glutamic acid. Basic amino acids have a net positive charge at neutral pH. Examples of basic amino acids include arginine, lysine, and histidine.

[0065] In a first aspect, the present application provides a fusion protein comprising a varicella-zoster virus protein and ferritin.

[0066] In some embodiments of the first aspect, the fusion protein is in the form of nanoparticles. In some embodiments, the varicella zoster virus protein and the ferritin can self-assemble to form nanoparticles after fusion.

[0067] In some embodiments of the first aspect, the varicella-zoster virus protein is a varicella-zoster virus glycoprotein, such as gE, gI, gB, gH, gL, or gC glycoprotein.

[0068] In some specific embodiments of the first aspect, the varicella-zoster virus protein is gE glycoprotein, such as the extracellular region of gE glycoprotein.

[0069] In some embodiments of the first aspect, the varicella-zoster virus protein is a varicella-zoster virus metastatic protein.

[0070] In some embodiments of the first aspect, the ferritin is selected from the group consisting of classical ferritin, heme-containing bacterial ferritin, and DNA-binding ferritin of starved cells.

[0071] In some embodiments of the first aspect, the ferritin is bacterial ferritin, such as Helicobacter pylori ferritin.

[0072] In some specific embodiments of the first aspect, the ferritin is Helicobacter pylori non-heme ferritin.

[0073] In some embodiments of the first aspect, the varicella-zoster virus protein comprises or consists of a sequence selected from:

[0074] (a) the amino acid sequence shown in SEQ ID NO: 1; and

[0075] (b) a variant obtained by deleting 1-50 amino acids from the amino acid sequence shown in SEQ ID NO: 1 at the C-terminus (e.g., the membrane-proximal region), for example, by deleting 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49 or 50 amino acids, or an integer range or integer between any two of the above values.

[0076] In some embodiments of the first aspect, the varicella-zoster virus protein comprises or consists of a sequence selected from the following: a variant obtained by deleting 1-5 amino acids at the C-terminus (e.g., the membrane-proximal region) of the amino acid sequence shown in SEQ ID NO:1, such as 1, 2, 3, 4, 5, or an integer range or integer between any two of the above values.

[0077] In some embodiments of the first aspect, the varicella-zoster virus protein comprises or consists of a sequence selected from the following: a variant obtained by deleting 6-10 amino acids at the C-terminus (e.g., the membrane-proximal region) of the amino acid sequence shown in SEQ ID NO:1, for example, 6, 7, 8, 9, 10, or an integer range or integer between any two of the above values.

[0078] In some embodiments of the first aspect, the varicella-zoster virus protein comprises or consists of a sequence selected from the following: a variant obtained by deleting 11-15 amino acids at the C-terminus (e.g., the membrane-proximal region) of the amino acid sequence shown in SEQ ID NO:1, such as 11, 12, 13, 14, 15, or an integer range or integer between any two of the above values.

[0079] In some embodiments of the first aspect, the varicella-zoster virus protein comprises or consists of a sequence selected from the following: a variant obtained by deleting 16-20 amino acids at the C-terminus (e.g., the membrane-proximal region) of the amino acid sequence shown in SEQ ID NO:1, such as 16, 17, 18, 19, 20, or an integer range or integer between any two of the above values.

[0080] In some embodiments of the first aspect, the varicella-zoster virus protein comprises or consists of a sequence selected from the following: a variant obtained by deleting 21-25 amino acids at the C-terminus (e.g., the membrane-proximal region) of the amino acid sequence shown in SEQ ID NO:1, such as 21, 22, 23, 24, 25, or an integer range or integer between any two of the above values.

[0081] In some embodiments of the first aspect, the varicella-zoster virus protein comprises or consists of a sequence selected from the following: a variant obtained by deleting 26-30 amino acids at the C-terminus (e.g., the membrane-proximal region) of the amino acid sequence shown in SEQ ID NO:1, such as 26, 27, 28, 29, 30, or an integer range or integer between any two of the above values.

[0082] In some embodiments of the first aspect, the varicella-zoster virus protein comprises or consists of a sequence selected from the following: a variant obtained by deleting 31-35 amino acids at the C-terminus (e.g., the membrane-proximal region) of the amino acid sequence shown in SEQ ID NO:1, such as 31, 32, 33, 34, 35, or an integer range or integer between any two of the above values.

[0083] In some embodiments of the first aspect, the varicella-zoster virus protein comprises or consists of a sequence selected from the following: a variant obtained by deleting 36-40 amino acids at the C-terminus (e.g., the membrane-proximal region) of the amino acid sequence shown in SEQ ID NO:1, such as 36, 37, 38, 39, 40, or an integer range or integer between any two of the above values.

[0084] In some embodiments of the first aspect, the varicella-zoster virus protein comprises or consists of a sequence selected from the following: a variant obtained by deleting 41-45 amino acids at the C-terminus (e.g., the membrane-proximal region) of the amino acid sequence shown in SEQ ID NO:1, such as 41, 42, 43, 44, 45, or an integer range or integer between any two of the above values.

[0085] In some embodiments of the first aspect, the varicella-zoster virus protein comprises or consists of a sequence selected from the following: a variant obtained by deleting 46-50 amino acids at the C-terminus (e.g., the membrane-proximal region) of the amino acid sequence shown in SEQ ID NO:1, such as 46, 47, 48, 49, 50, or an integer range or integer between any two of the above values.

[0086] In some embodiments of the first aspect, the varicella-zoster virus protein comprises or consists of a sequence selected from the following: a variant obtained by deleting 17, 31 or 41 amino acids at the C-terminus (e.g., the membrane-proximal region) of the amino acid sequence shown in SEQ ID NO:1.

[0087] In some embodiments of the first aspect, 17 amino acids are deleted at the C-terminus (eg, the membrane-proximal region) of the amino acid sequence shown in SEQ ID NO:1 to obtain the amino acid sequence shown in SEQ ID NO:12.

[0088] In some embodiments of the first aspect, 31 amino acids are deleted at the C-terminus (eg, the membrane-proximal region) of the amino acid sequence shown in SEQ ID NO:1 to obtain the amino acid sequence shown in SEQ ID NO:13.

[0089] In some embodiments of the first aspect, the amino acid sequence shown in SEQ ID NO: 14 is obtained by deleting 41 amino acids at the C-terminus (eg, the membrane-proximal region) of the amino acid sequence shown in SEQ ID NO: 1.

[0090] In some embodiments of the first aspect, the variant obtained by deleting a certain number of amino acids at the C-terminus of the amino acid sequence shown in SEQ ID NO: 1 does not substantially change the immunogenicity of the varicella-zoster virus protein.

[0091] In some embodiments of the first aspect, the varicella-zoster virus protein comprises an amino acid sequence as shown in any one of SEQ ID NOs: 1 and 12-14, or consists of an amino acid sequence as shown in any one of SEQ ID NOs: 1 and 12-14.

[0092] In some embodiments of the first aspect, the amino acid sequence of the varicella-zoster virus protein has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or higher homology with the amino acid sequence shown in any one of SEQ ID NOs: 1 and 12-14, or a homology of a value or range between any two of the above values.

[0093] In some embodiments of the first aspect, a fusion protein is prepared by truncating the C-terminal membrane-proximal amino acid sequence of the extracellular region of the varicella-zoster virus gE glycoprotein shown in SEQ ID NO: 1 by a certain number of amino acids, for example, 17 or 31 amino acids (such as the amino acid sequence shown in SEQ ID NO: 12 or 13), so that the His tag is exposed, which is beneficial to the purification of the fusion protein.

[0094] In some embodiments of the first aspect, the ferritin comprises or consists of a sequence selected from:

[0095] (a) the amino acid sequence shown in SEQ ID NO: 2; and

[0096] (b) A variant obtained by subjecting the amino acid sequence shown in SEQ ID NO: 2 to one or more substitutions, deletions or additions.

[0097] In some embodiments of the first aspect, the variant obtained by one or more substitutions, deletions or additions of the amino acid sequence shown in SEQ ID NO:2 does not substantially change the function of the ferritin, such as the function of enabling the fusion protein to self-assemble into nanoparticles after being fused with varicella-zoster virus protein.

[0098] In some embodiments of the first aspect, the ferritin comprises or consists of the amino acid sequence shown in SEQ ID NO:2.

[0099] In some embodiments of the first aspect, the amino acid sequence of the ferritin has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or higher homology with the amino acid sequence shown in SEQ ID NO:2, or a homology of a value or range between any two of the above values.

[0100] In some embodiments of the first aspect, the amino acid sequence of the ferritin protein differs from the amino acid sequence of SEQ ID NO: 2 by about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 28, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50 or more amino acid substitutions, deletions and / or additions while still maintaining similar functions of the ferritin protein.

[0101] In some embodiments of the first aspect, the C-terminal or N-terminal region of the amino acid sequence shown in SEQ ID NO:2 can also be truncated by about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 28, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50 or more amino acids while still maintaining similar functions of the ferritin.

[0102] In some embodiments of the first aspect, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 28, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50 or more amino acids can be added to the C-terminal or N-terminal region of the amino acid sequence shown in SEQ ID NO: 2, and the resulting amino acid sequence still maintains similar function of the ferritin.

[0103] In some embodiments of the first aspect, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 28, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50 or more amino acids may be added or deleted in the region other than the C-terminus or N-terminus of the amino acid sequence shown in SEQ ID NO: 2, as long as the altered amino acid sequence substantially maintains the similar function of the ferritin.

[0104] In some embodiments of the first aspect, the ferritin is linked to the C-terminus of the varicella-zoster virus protein.

[0105] In some embodiments of the first aspect, the ferritin is connected to the C-terminus of the varicella zoster virus protein via a linker. In some embodiments, the linker is a GS-type linker, such as (GGGGS)n, where n is an integer ≥ 1; GGSGGSGS; GS. In some embodiments, the linker is EKAAKAEEAA. In some embodiments, the linker is any combination of the above-mentioned GS-type linkers or EKAAKAEEAA.

[0106] In some embodiments of the first aspect, the amino acid sequence of linker 1 is (GGGGS)3 (SEQ ID NO: 15).

[0107] In some embodiments of the first aspect, the amino acid sequence of linker 2 is GGSGGSGS (SEQ ID NO: 16).

[0108] In some embodiments of the first aspect, the amino acid sequence of linker 3 is GS.

[0109] In some embodiments of the first aspect, the amino acid sequence of linker 4 is: EKAAKAEEAA (SEQ ID NO: 17).

[0110] In some embodiments of the first aspect, the fusion protein further comprises a purification tag.

[0111] In some embodiments of the first aspect, the purification tag is a His tag, a GST tag, an MBP tag, an HA tag, a c-Myc tag, or a Flag tag.

[0112] In some specific embodiments of the first aspect, the purification tag is a His tag.

[0113] In some embodiments of the first aspect, the purification tag is linked to the N-terminus of the varicella-zoster virus protein.

[0114] In some embodiments of the first aspect, the purification tag is linked to the C-terminus of the varicella-zoster virus protein.

[0115] In some embodiments of the first aspect, the structural form of the fusion protein is any one of the following: purification tag-linker-varicella zoster virus protein-linker-ferritin, varicella zoster virus protein-linker-purification tag-linker-ferritin, varicella zoster virus protein-linker-ferritin, and varicella zoster virus protein-purification tag-linker-ferritin.

[0116] In some embodiments of the first aspect, the fusion protein comprises or consists of a sequence selected from the group consisting of:

[0117] (e) an amino acid sequence as shown in any one of SEQ ID NOs: 4-11; and

[0118] (f) A variant obtained by subjecting the amino acid sequence shown in any one of SEQ ID NOs: 4-11 to one or more substitutions, deletions or additions.

[0119] In some embodiments of the first aspect, the variant obtained by one or more substitutions, deletions or additions of the amino acid sequence shown in any one of SEQ ID NOs: 4-11 does not substantially change the function of the fusion protein, such as immunogenicity.

[0120] In some embodiments of the first aspect, the fusion protein comprises an amino acid sequence as shown in any one of SEQ ID NOs: 4-11, or consists of an amino acid sequence as shown in any one of SEQ ID NOs: 4-11.

[0121] In some embodiments of the first aspect, the amino acid sequence of the fusion protein has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or higher homology with the amino acid sequence shown in any one of SEQ ID NOs: 4-11, or a homology of a value or range between any two of the above values.

[0122] In some embodiments of the first aspect, the amino acid sequence of the fusion protein differs from the amino acid sequence shown in any one of SEQ ID NOs:4-11 by about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 28, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50 or more amino acid substitutions, deletions and / or additions, while still maintaining similar functions of the fusion protein.

[0123] In some embodiments of the first aspect, the C-terminal or N-terminal region of the amino acid sequence shown in any one of SEQ ID NOs: 4-11 can also be truncated by about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 28, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50 or more amino acids while still maintaining the function of the similar fusion protein.

[0124] In some embodiments of the first aspect, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 28, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50 or more amino acids can be added to the C-terminal or N-terminal region of the amino acid sequence shown in any one of SEQ ID NOs: 4-11, and the resulting amino acid sequence still maintains the function of the similar fusion protein.

[0125] In some embodiments of the first aspect, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 28, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50 or more amino acids can be added or deleted in the region other than the C-terminus or N-terminus of the amino acid sequence shown in any one of SEQ ID NOs: 4-11, as long as the altered amino acid sequence substantially maintains the function of the similar fusion protein.

[0126] In a second aspect, the present application provides a nucleic acid encoding the fusion protein described in the first aspect.

[0127] In some embodiments of the second aspect, the nucleic acid molecule may include a DNA molecule and an RNA molecule. The nucleic acid molecule may be single-stranded or double-stranded, and may be a cDNA.

[0128] In some embodiments of the second aspect, the nucleic acid molecule is operably linked to a regulatory nucleotide sequence that can be recognized by a host cell transformed with the vector.

[0129] In a third aspect, the present application provides a pharmaceutical composition comprising the fusion protein described in the first aspect, and a pharmaceutically acceptable excipient, diluent or carrier.

[0130] In some embodiments of the third aspect, the pharmaceutical composition is used to prevent varicella-zoster virus infection or a disease caused by the infection.

[0131] In some embodiments of the third aspect, the disease is varicella, herpes zoster, or chronic pain complicated by varicella or herpes zoster (e.g., postherpetic neuralgia (PHN)), neurological and ocular diseases (e.g., meningoencephalitis, myelitis, cranial nerve palsies, vasculopathy, keratitis and retinopathy), or visceral and gastrointestinal diseases (ulcers, hepatitis and pancreatitis).

[0132] In some embodiments of the third aspect, the pharmaceutical composition of the present application can be delivered by any physiologically acceptable administration method, including but not limited to oral administration, intravascular injection, etc.

[0133] In some embodiments of the third aspect, pharmaceutical compositions for prevention or detection purposes can be prepared for storage in the form of lyophilized preparations or aqueous solutions by mixing reagents having the desired purity with pharmaceutically acceptable carriers, excipients, etc. as appropriate.

[0134] In a fourth aspect, the present application provides a method for preparing the fusion protein described in the first aspect, comprising: transferring the nucleic acid described in the second aspect into a host cell, culturing the host cell; and harvesting the fusion protein.

[0135] In some embodiments of the fourth aspect, the method further comprises purifying the fusion protein. In some embodiments, the fusion protein is purified by a purification tag (eg, a His tag) contained in the fusion protein.

[0136] In some embodiments described in the fourth aspect, the fusion protein of the present application can be screened and prepared by any suitable method known to those skilled in the art.

[0137] In some embodiments described in the fourth aspect, a suitable host cell refers to a host cell suitable for expression of an expression vector or a target nucleic acid. A suitable culture medium refers to a culture medium suitable for growth of host cells or induction of expression thereof.

[0138] In some embodiments of the fourth aspect, various conventional culture media can be selected according to the host cell used. Cultivate under conditions suitable for host cell growth. Preferably, the host cell of through engineering approaches can be cultivated in a conventional nutrient medium modified to be suitable for activating a promoter, to screen transformants or to increase the polynucleotides of the present application. After transforming a suitable host cell and growing the host cell to a suitable cell density, induce the promoter selected with a suitable method (such as temperature conversion or chemical induction), and the cell is cultivated for a period of time to allow it to produce a target fusion protein or its fragment.

[0139] In some embodiments of the fourth aspect, the fusion protein produced by the host cell can be coated in the cell or expressed on the cell membrane or secreted outside the cell. If necessary, the fusion protein can be separated and purified by various separation methods using its physical, chemical and other characteristics. For example, the expressed fusion protein or its fragment can be recovered and purified from the recombinant cell culture by the following methods well known in the art: conventional renaturation treatment, protein precipitation agent treatment (salting out method), centrifugation, osmotic bacteria breaking, ultrasonic treatment, ultracentrifugation, molecular sieve chromatography (gel filtration), adsorption chromatography, ion exchange chromatography, high performance liquid chromatography (HPLC) and other various liquid chromatography techniques and the combination of these methods.

[0140] In some embodiments of the fourth aspect, the fusion protein of the present application can also be synthesized by chemosynthesis. The chemical synthesis method of fusion protein is also well known to those skilled in the art, for example, the fusion protein of the present application and variants thereof (Merrifield, J.Am.Chem.Soc.85:2149-2154 (1963)) can be produced by the directed peptide synthesis using solid phase technology. Protein synthesis can be performed manually or by automation. For example, automated synthesis can be realized by the 431A peptide synthesizer (Perkin Elmer) of Applied Biosystems. Alternatively, different sizes can be synthesized by chemical means respectively and combined to prepare required molecules using chemical methods.

[0141] In a fifth aspect, the present application provides use of the fusion protein described in the first aspect or the pharmaceutical composition described in the third aspect in the preparation of a medicament for preventing varicella-zoster virus infection or a disease caused by the infection.

[0142] In some embodiments of the fifth aspect, the medicament is a vaccine, such as a multivalent vaccine.

[0143] In some embodiments of the fifth aspect, the disease is varicella, herpes zoster, or chronic pain complicated by varicella or herpes zoster (e.g., postherpetic neuralgia (PHN)), neurological and ocular diseases (e.g., meningoencephalitis, myelitis, cranial nerve palsies, vasculopathy, keratitis and retinopathy), or visceral and gastrointestinal diseases (ulcers, hepatitis and pancreatitis).

[0144] In a sixth aspect, the present application provides a method for generating an immune response against varicella-zoster virus in a subject, comprising administering the fusion protein described in the first aspect or the pharmaceutical composition described in the third aspect to the subject.

[0145] In some embodiments of the sixth aspect, the subject is a mammal.

[0146] In some embodiments of the sixth aspect, the subject is a human.

[0147] In a seventh aspect, the present application provides a method for preventing varicella-zoster virus infection or a disease caused by the infection, comprising administering the fusion protein described in the first aspect or the pharmaceutical composition described in the third aspect to a subject in need.

[0148] In some embodiments of the seventh aspect, the subject is a mammal.

[0149] In some embodiments of the seventh aspect, the subject is a human.

[0150] In some embodiments of the seventh aspect, the disease is varicella, herpes zoster, or chronic pain complicated by varicella or herpes zoster (e.g., postherpetic neuralgia (PHN)), neurological and ocular diseases (e.g., meningoencephalitis, myelitis, cranial nerve palsies, vasculopathy, keratitis and retinopathy), or visceral and gastrointestinal diseases (ulcers, hepatitis and pancreatitis).

[0151] In other aspects, the present application also provides a vector comprising the nucleic acid described in the second aspect, a host cell comprising the nucleic acid described in the second aspect or the vector, and a method for producing the fusion protein. In some embodiments, the nucleic acid described in the second aspect is operably linked to a regulatory nucleotide sequence, and the regulatory nucleotide sequence can be recognized by a host cell transformed with the vector.

[0152] In some embodiments of the present application, using the fusion protein described in the first aspect or the pharmaceutical composition described in the third aspect as an immunogen can effectively activate the immune system in the body.

[0153] In some embodiments of the present application, the fusion protein described in the first aspect or the pharmaceutical composition described in the third aspect is used as an immunogen to effectively produce antibodies in vivo.

[0154] In some embodiments of the present application, the use of the fusion protein described in the first aspect or the pharmaceutical composition described in the third aspect as an immunogen can effectively stimulate a T cell immune response, thereby preventing varicella-zoster virus infection or diseases caused by infection, such as chickenpox, herpes zoster, or chronic pain complicated by chickenpox or herpes zoster (such as postherpetic neuralgia (PHN)), neurological and eye diseases (such as meningoencephalitis, myelitis, cranial nerve palsy, vasculopathy, keratitis and retinopathy), or visceral and gastrointestinal diseases (ulcers, hepatitis and pancreatitis).

[0155] It should be understood that the above detailed description is only for the purpose of enabling those skilled in the art to more clearly understand the content of the present application, and is not intended to be limiting in any aspect. Those skilled in the art can make various modifications and changes to the embodiments. Example

[0156] The following examples are for illustration purposes only and are not intended to limit the scope of this application.

[0157] Example 1: Preparation and identification of fusion protein

[0158] In the process of preparing and identifying gE-ferritin nanoparticles, a variety of different recombinant proteins are needed, including the extracellular region of glycoprotein E of Oka strain varicella-zoster virus (gE, SEQ ID NO: 1) and Helicobacter pylori non-heme ferritin (ferritin, SEQ ID NO: 2). Adding His tags (His, SEQ ID NO: 3) to the C-terminus or N-terminus of these recombinant proteins helps in the purification and functional identification of the recombinant proteins. With reference to the amino acid sequence in the Uniprot database, codon-optimized recombinant protein genes were designed and synthesized. Using conventional molecular biological techniques, gE-His, His-ferritin, and two gE-ferritin genes in which His tags were fused to the N-terminus (His-gE-ferritin, SEQ ID NO: 4) and the C-terminus (gE-His-ferritin, SEQ ID NO: 5) of the gE protein were cloned into suitable eukaryotic expression vectors (such as pcDNA3.1 of Invitrogen, etc.), and then the prepared recombinant protein expression plasmids were transfected into HEK293 cells (such as HEK293F of Invitrogen) using liposomes (such as 293fectin of Invitrogen, etc.) or other cationic transfection reagents (such as PEI, etc.), cultured under serum-free suspension culture conditions for 3-4 days, and then the culture supernatant was harvested by centrifugation or the like.

[0159] The culture supernatant harvested by centrifugation was analyzed and identified by ELISA. 10Prepare gE monoclonal antibodies with two different epitopes, 301-1G5 and 303-1C9, and coat the prepared 303-1C9-mIgG2a monoclonal antibody on a 96-well ELISA plate at 5 μg / mL, 100 μL / well, and coat overnight at 4°C. After blocking with blocking solution PBS-0.1% Tween 20-3% milk (PBST-3% milk) at 37°C for 1 hour, add equal volumes of gE-His, His-ferritin, His-gE-ferritin, and gE-His-ferritin culture supernatant, respectively, and bind at 37°C for 1 hour. Wash the ELISA plate with PBS-0.1% Tween 20 (PBST) buffer, then add the prepared 301-1G5-IgG1 protein, and bind at 37°C for 1 hour. Add HRP mouse anti-human IgG (Bioss, bsm-0297M-HRP) and bind at 37°C for 1 hour. Wash the ELISA plate with PBS-0.1% Tween 20 (PBST) buffer, add OPD substrate colorimetric solution, and terminate the color development with 1M H2SO4 after 5-10 minutes. Measure the optical density value with a dual wavelength of 492nm / 630nm using an ELISA reader. Figure 2 As shown, the results showed that both His-gE-ferritin and gE-His-ferritin were expressed, among which the expression level of gE-His-ferritin was significantly higher by about 10 times.

[0160] Example 2: Structural optimization of fusion protein

[0161] In order to obtain gE-ferritin-based fusion protein nanoparticles, a series of structural optimizations were performed on the above-mentioned His-gE-ferritin and gE-His-ferritin, and gE-ferritin-A (SEQ ID NO: 6), gE-ferritin-B (SEQ ID NO: 7), gE-ferritin-C (SEQ ID NO: 8), gE-ferritin-D (SEQ ID NO: 9), gE-ferritin-E (SEQ ID NO: 10) and gE-ferritin-F (SEQ ID NO: 11) expression plasmids were prepared, respectively, and after parallel transfection of HEK293F cells, expression and purification studies were performed.

[0162] Referring to Example 1, the above culture supernatant was subjected to ELISA analysis, and the results were as follows: Figure 3 As shown: All fusion proteins can be expressed normally and recognize two antibody epitopes of gE protein at the same time. Similar to the results in Example 1, the expression level of GE-ferritin-C fused to the N-terminus of gE protein with His tag is 2-3 times lower than that of other proteins.

[0163] Subsequently, the fusion protein in the culture supernatant was purified in one step using a metal chelate affinity chromatography column (such as GE's HisTrap FF, etc.). Then, the recombinant protein storage buffer was replaced with PBS (pH 7.0) or other suitable buffer using a desalting column (such as GE's Hitrap desaulting, etc.). The experimental results showed that the target protein could only be purified from the supernatants of gE-ferritin-D and gE-ferritin-E cells using a nickel column.

[0164] Example 3: Identification of fusion proteins

[0165] The His-ferritin, gE-ferritin-D and gE-ferritin-E protein samples purified above were centrifuged at 13000g for 5 minutes and then used for SEC analysis. The detection was performed using Thermo U3000, the chromatographic column was Xbridge BEH450A SEC 3.5μm 7.8×300mm, and the mobile phase was 50mM PB, 300mM NaCl, pH7.0. The column temperature was 25°C, the injection volume was 0.02mL, the flow rate was 0.8mL / min, and the absorbance was detected at 214nm. The experimental results are shown in Figure 4 As shown, the peak time of His-ferritin is 12.185 minutes, and the protein purity is 92.552%; the peak time of gE-ferritin-D is 8.395 minutes, and the protein purity is 92.438%; the peak time of gE-ferritin-E is 8.605 minutes, and the protein purity is 99.125%; all three proteins form uniform protein aggregates.

[0166] DLS data were detected using a Malvern Zetasizer μV, with a detection temperature of 25°C and a detection volume of 60uL. A ZEN0118 detection cell was used, and the automatic interval and cycle number were measured 3 times. The data analysis mode was Multiple narrow modes (high resolution), and the data processing display interval was 10 to 300nm. The experimental results are shown in Figure 5 As shown: the average particle size of His-ferritin is 20.33 nm; the average particle size of gE-ferritin-D is 55.56 nm; and the average particle size of gE-ferritin-E is 42.87 nm.

[0167] Example 4: Study on immunogenicity of gE-ferritin-D nanoparticles

[0168] 1. Mouse Immunization

[0169] 40 BALB / c male mice aged 6-8 weeks were randomly divided into 4 groups (Table 1), with 10 mice in each group, and each mouse was numbered independently. Equimolar gE protein was used for immunization, and the adjuvant was Poly I:C, and the immunization method was intramuscular injection. Booster immunization was performed once every 28 days, and blood was collected on the 28th and 56th days after the primary immunization. The mice were killed on D59 and spleen cells were collected, and spleen lymphocytes were separated and frozen for later use.

[0170] Table 1. Mouse immunization schedule

[0171]

[0172] 2. Serum titer detection

[0173] The prepared His-ferritin and gE-His were coated on 96-well ELISA plates, 1 μg / mL, 100 μL / well, and coated overnight at 4°C. After blocking with blocking solution PBS-0.1% Tween 20-3% milk at 37°C for 1 hour, the immunized mouse serum was added, and the serum was diluted 3 times with PBST buffer starting at 1:500, and combined at 37°C for 1 hour. The ELISA plate was washed with PBST buffer, and HRP-labeled goat anti-mouse polyclonal antibody (Zhongshan Jinqiao / ZB-5305 / 133006) was added, and combined at 37°C for 1 hour. The ELISA plate was washed with PBST buffer, and OPD substrate colorimetric solution was added. After 5-10 minutes, 1M H2SO4 was used to stop the color development, and the optical density value was measured by a microplate reader at a dual wavelength of 492nm / 630nm. The ELISA result data processing scheme adopts: (immunized mouse serum signal value - blank mouse serum signal value) / blank mouse serum signal value, and the minimum serum dilution ratio value with a value ≥ 2 is used as the data for plotting. Figure 6 As shown, the serum titer of gE-ferritin-D is about 2 times higher than that of gE-His immunization, and can effectively stimulate the body to produce antibodies.

[0174] 3. Serum neutralization activity detection

[0175] Neutralization activity was determined by detecting the reduction of plaque formation in MRC5 cells infected with serum after mixing with VZV. MRC-5 cells were inoculated into a 12-well cell culture plate at 1.5E+5 cells / well and cultured for 48 hours to form a monolayer of cells. When 90% confluent, the culture supernatant was removed and virus maintenance solution (MEM medium containing 2% serum) was added, 400 μL per well. VZV (Oka) was diluted to 1000 pfu / mL with virus maintenance solution, that is, 50 pfu of VZV per 100 μL after mixing with an equal volume of antibody; the serum of groups G1-G4 shown in Table 1 was diluted and filtered with virus diluent, and 100 μL was mixed with an equal volume of VZV. The dilution multiples of the serum after mixing were 200 times, 400 times, 800 times, and 1600 times, respectively. The mixed sample of VZV and serum was neutralized for 1 hour at 25°C. Infect MRC5 cells in 12-well plates, add 100 μL of neutralization sample to each well, adsorb at 37°C for 1 hour, add 25 μL of complement, and continue incubation at 37°C for 30 minutes; discard the supernatant, wash twice with PBS, and add 3 mL of maintenance solution to each well. Maintain culture at 37°C for 5-7 days, remove the supernatant, wash once with PBS, add 1 mL of Coomassie Brilliant Blue staining solution to each well, act for 10 minutes, wash once with PBS, place in a bright place to count plaques, and calculate the serum dilution multiple (PRNT50) for 50% reduction in plaques using the Reed and Muench method. The experimental results are shown in Table 2. Compared with gE-His immunization, the serum neutralization activity of gE-ferritin-D is about 2 times higher.

[0176] Table 2. Serum dilutions required to reduce plaques by 50% (PRNT50)

[0177] Experimental Group G1 G2 G3 G4 PRNT50 (dilution factor) 400 941.12 NA NA

[0178] Note: NA means not available.

[0179] 4. T cell immune response activity detection

[0180] The activity of gE-ferritin nanoparticles in inducing T cell immune response was evaluated using a mouse IFN-γ pre-coated ELISPOT kit. The mouse spleen lymphocytes that had been separated and frozen were resuscitated, and after being treated with the super nuclease Benzonase, the cells were resuspended in serum-containing RPMI1640 medium to adjust the cell density to 2.0E+7 cells / mL. The pre-coated strips were treated with 250μL of RPMI1640 medium, and the mouse spleen lymphocytes of G1-G4 shown in Table 1 of the experimental group were added to the pre-coated strips at 100μL, 2.0E+6 cells / well, respectively, and VZV was mixed with the experimental group cells at 50μL / well, and the final concentration of VZV was 900pfu / well. Incubate at 37℃ for 24h, where the experimental wells with PMA added were used as positive controls (PC), and the experimental wells with culture medium added were used as negative controls (NC). After incubation, wash with washing solution six times, incubate with detection antibody at room temperature for 1 hour, add enzyme-linked avidin and incubate at room temperature for 1 hour, AEC color development, terminate the reaction according to the appearance of spots in each experimental well, and read the number of reactive spots with ELISPOT spot counter. The results are shown as follows Figure 7 As shown, gE-ferritin-D significantly increased the T cell immune response compared with gE-His immunization, thereby effectively preventing varicella-zoster virus infection or diseases caused by infection.

[0181] The exemplary embodiments of the various inventions of the present application are described above; however, without departing from the essence and scope of the present application, those skilled in the art will be able to modify or improve the exemplary embodiments described in the present application, and the resulting variations or equivalents also fall within the scope of the present application.

[0182] Sequence information

[0183] SEQ ID NO:1

[0184] SVLRYDDFHIDEDKLDTNSVYEPYYHSDHAESSWVNRGESSRKAYDHNSPYIWPRNDYDGFLENAHEHHGVYNQGRGIDSGERLMQPTQMSAQEDLGDDTGIHVIPTLNGDDRHKIVNVDQRQYGDVFKGDLNPKPQGQRLIEVSVEENHPFTLRAPIQRIYGVRYTETWSFLPSLTCTGDAAPAIQHICLKHTTCFQDVVVDVDCAENTKEDQLAEISYRFQGKKEADQPWIVVNTSTLFDELELDPPEIEPGVLKVLRTEKQYLGVYIWNMRGSDGTSTYATFLVTWKGDEKTRNPTPAVTPQPRGAEFHMWNYHSHVFSVGDTFSLAMHLQYKIHEAPFDLLLEWLYVPIDPTCQPMRLYSTCLYHPNAPQCLSHMNSGCTFTSPHLAQRVASTVYQNCEHADNYTAYCLGISHMEPSFGLILHDGGTTLKFVDTPESLSGLYVFVVYFNGHVEAVAYTVVSTVDHFVNAIEERGFPPTAGQPPATTKPKEITPVNPGTSPLIRYAAWTGGLA

[0185] SEQ ID NO:2

[0186] DIIKLLNEQVNKEMQSSNLYMSMSSWCYTHSLDGAGLFLFDHAAEEYEHAKKLIIFLNENNVPVQLTSISAPEHKFEGLTQIFQKAYEHEQHISESINNIVDHAIKSKDHATFNFLQWYVAEQHEEEVLFKDILDKIELIGNENHGLYLADQYVKGIAKSRKS

[0187] SEQ ID NO:3

[0188] HHHHHH

[0189] SEQ ID NO:4

[0190] HHHHHHHHGGGGSSVLRYDDFHIDEDKLDTNSVYEPYYHSDHAESSWVNRGESSRKAYDHNSPYIWPRNDYDGFLENAHEHHGVYNQGRGIDSGERLMQPTQMSAQEDLGDDTGIHVIPTLNGDDRHKIVNVDQRQYGDVFKGDLNPKPQGQRLIEVSVEENHPFTLRAPIQRIYGVRYTETWSFLPSLTCTGDAAPAIQHICLKHTTCFQDVVVDVDCAENTKEDQLAEISYRFQGKKEADQPWIVVNTSTLFDELELDPPEIEPGVLKVLRTEKQYLGVYIWNMRGSDGTSTYATFLVTWKGDEKTRNPTPAVTPQPRGAEFHMWNYHSHVFSVGDTFSLAMHLQYKIHEAPFDLLLEWLYVPIDPTCQPMRLYSTCLYHPNAPQCLSHMNSGCTFTSPHLAQRVASTVYQNCEHADNYTAYCLGISHMEPSFGLILHDGGTTLKFVDTPESLSGLYVFVVYFNGHVEAVAYTVVSTVDHFVNAIEERGFPPTAGQPPATTKPKEITPVNPGTSPLIRYAAWTGGLAGGSGGSGSEKAAKAEEAADIIKLLNEQVNKEMQSSNLYMSMSSWCYTHSLDGAGLFLFDHAAEEYEHAKKLIIFLNENNVPVQLTSISAPEHKFEGLTQIFQKAYEHEQHISESINNIVDHAIKSKDHATFNFLQWYVAEQHEEEVLFKDILDKIELIGNENHGLYLADQYVKGIAKSRKS

[0191] SEQ ID NO:5

[0192] SVLRYDDFHIDEDKLDTNSVYEPYYHSDHAESSWVNRGESSRKAYDHNSPYIWPRNDYDGFLENAHEHHGVYNQGRGIDSGERLMQPTQMSAQEDLGDDTGIHVIPTLNGDDRHKIVNVDQRQYGDVFKGDLNPKPQGQRLIEVSVEENHPFTLRAPIQRIYGVRYTETWSFLPSLTCTGDAAPAIQHICLKHTTCFQDVVVDVDCAENTKEDQLAEISYRFQGKKEADQPWIVVNTSTLFDELELDPPEIEPGVLKVLRTEKQYLGVYIWNMRGSDGTSTYATFLVTWKGDEKTRNPTPAVTPQPRGAEFHMWNYHSHVFSVGDTFSLAMHLQYKIHEAPFDLLLEWLYVPIDPTCQPMRLYSTCLYHPNAPQCLSHMNSGCTFTSPHLAQRVASTVYQNCEHADNYTAYCLGISHMEPSFGLILHDGGTTLKFVDTPESLSGLYVFVVYFNGHVEAVAYTVVSTVDHFVNAIEERGFPPTAGQPPATTKPKEITPVNPGTSPLIRYAAWTGGLAGGGGSGGGGSGGGGSHHHHHHHHGGSGGSGSEKAAKAEEAADIIKLLNEQVNKEMQSSNLYMSMSSWCYTHSLDGAGLFLFDHAAEEYEHAKKLIIFLNENNVPVQLTSISAPEHKFEGLTQIFQKAYEHEQHISESINNIVDHAIKSKDHATFNFLQWYVAEQHEEEVLFKDILDKIELIGNENHGLYLADQYVKGIAKSRKS

[0193] SEQ ID NO:6

[0194] SVLRYDDFHIDEDKLDTNSVYEPYYHSDHAESSWVNRGESSRKAYDHNSPYIWPRNDYDGFLENAHEHHGVYNQGRGIDSGERLMQPTQMSAQEDLGDDTGIHVIPTLNGDDRHKIVNVDQRQYGDVFKGDLNPKPQGQRLIEVSVEENHPFTLRAPIQRIYGVRYTETWSFLPSLTCTGDAAPAIQHICLKHTTCFQDVVVDVDCAENTKEDQLAEISYRFQGKKEADQPWIVVNTSTLFDELELDPPEIEPGVLKVLRTEKQYLGVYIWNMRGSDGTSTYATFLVTWKGDEKTRNPTPAVTPQPRGAEFHMWNYHSHVFSVGDTFSLAMHLQYKIHEAPFDLLLEWLYVPIDPTCQPMRLYSTCLYHPNAPQCLSHMNSGCTFTSPHLAQRVASTVYQNCEHADNYTAYCLGISHMEPSFGLILHDGGTTLKFVDTPESLSGLYVFVVYFNGHVEAVAYTVVSTVDHFVNAIEERGFPPTAGQPPATTKPKEITPVNPGTSPLIRYAAWTGGLAGGGGSGGGGSGGGGSDIIKLLNEQVNKEMQSSNLYMSMSSWCYTHSLDGAGLFLFDHAAEEYEHAKKLIIFLNENNVPVQLTSISAPEHKFEGLTQIFQKAYEHEQHISESINNIVDHAIKSKDHATFNFLQWYVAEQHEEEVLFKDILDKIELIGNENHGLYLADQYVKGIAKSRKS

[0195] SEQ ID NO:7

[0196] SVLRYDDFHIDEDKLDTNSVYEPYYHSDHAESSWVNRGESSRKAYDHNSPYIWPRNDYDGFLENAHEHHGVYNQGRGIDSGERLMQPTQMSAQEDLGDDTGIHVIPTLNGDDRHKIVNVDQRQYGDVFKGDLNPKPQGQRLIEVSVEENHPFTLRAPIQRIYGVRYTETWSFLPSLTCTGDAAPAIQHICLKHTTCFQDVVVDVDCAENTKEDQLAEISYRFQGKKEADQPWIVVNTSTLFDELELDPPEIEPGVLKVLRTEKQYLGVYIWNMRGSDGTSTYATFLVTWKGDEKTRNPTPAVTPQPRGAEFHMWNYHSHVFSVGDTFSLAMHLQYKIHEAPFDLLLEWLYVPIDPTCQPMRLYSTCLYHPNAPQCLSHMNSGCTFTSPHLAQRVASTVYQNCEHADNYTAYCLGISHMEPSFGLILHDGGTTLKFVDTPESLSGLYVFVVYFNGHVEAVAYTVVSTVDHFVNAIEERGFPPTAGQPPATTKPKEITPVNPGTSPLIRYAAWTGGLAGGGGSGGGGSGGGGSEKAAKAEEAADIIKLLNEQVNKEMQSSNLYMSMSSWCYTHSLDGAGLFLFDHAAEEYEHAKKLIIFLNENNVPVQLTSISAPEHKFEGLTQIFQKAYEHEQHISESINNIVDHAIKSKDHATFNFLQWYVAEQHEEEVLFKDILDKIELIGNENHGLYLADQYVKGIAKSRKS

[0197] SEQ ID NO:8

[0198] HHHHHHHHGGGGSGGGGSGGGGSSVLRYDDFHIDEDKLDTNSVYEPYYHSDHAESSWVNRGESSRKAYDHNSPYIWPRNDYDGFLENAHEHHGVYNQGRGIDSGERLMQPTQMSAQEDLGDDTGIHVIPTLNGDDRHKIVNVDQRQYGDVFKGDLNPKPQGQRLIEVSVEENHPFTLRAPIQRIYGVRYTETWSFLPSLTCTGDAAPAIQHICLKHTTCFQDVVVDVDCAENTKEDQLAEISYRFQGKKEADQPWIVVNTSTLFDELELDPPEIEPGVLKVLRTEKQYLGVYIWNMRGSDGTSTYATFLVTWKGDEKTRNPTPAVTPQPRGAEFHMWNYHSHVFSVGDTFSLAMHLQYKIHEAPFDLLLEWLYVPIDPTCQPMRLYSTCLYHPNAPQCLSHMNSGCTFTSPHLAQRVASTVYQNCEHADNYTAYCLGISHMEPSFGLILHDGGTTLKFVDTPESLSGLYVFVVYFNGHVEAVAYTVVSTVDHFVNAIEERGFPPTAGQPPATTKPKEITPVNPGTSPLIRYAAWTGGLAGGSGGSGSEKAAKAEEAADIIKLLNEQVNKEMQSSNLYMSMSSWCYTHSLDGAGLFLFDHAAEEYEHAKKLIIFLNENNVPVQLTSISAPEHKFEGLTQIFQKAYEHEQHISESINNIVDHAIKSKDHATFNFLQWYVAEQHEEEVLFKDILDKIELIGNENHGLYLADQYVKGIAKSRKS

[0199] SEQ ID NO:9

[0200] SVLRYDDFHIDEDKLDTNSVYEPYYHSDHAESSWVNRGESSRKAYDHNSPYIWPRNDYDGFLENAHEHHGVYNQGRGIDSGERLMQPTQMSAQEDLGDDTGIHVIPTLNGDDRHKIVNVDQRQYGDVFKGDLNPKPQGQRLIEVSVEENHPFTLRAPIQRIYGVRYTETWSFLPSLTCTGDAAPAIQHICLKHTTCFQDVVVDVDCAENTKEDQLAEISYRFQGKKEADQPWIVVNTSTLFDELELDPPEIEPGVLKVLRTEKQYLGVYIWNMRGSDGTSTYATFLVTWKGDEKTRNPTPAVTPQPRGAEFHMWNYHSHVFSVGDTFSLAMHLQYKIHEAPFDLLLEWLYVPIDPTCQPMRLYSTCLYHPNAPQCLSHMNSGCTFTSPHLAQRVASTVYQNCEHADNYTAYCLGISHMEPSFGLILHDGGTTLKFVDTPESLSGLYVFVVYFNGHVEAVAYTVVSTVDHFVNAIEERGFPPTAGQPPATTKPKEITPVNHHHHHHHHGSEKAAKAEEAADIIKLLNEQVNKEMQSSNLYMSMSSWCYTHSLDGAGLFLFDHAAEEYEHAKKLIIFLNENNVPVQLTSISAPEHKFEGLTQIFQKAYEHEQHISESINNIVDHAIKSKDHATFNFLQWYVAEQHEEEVLFKDILDKIELIGNENHGLYLADQYVKGIAKSRKS。

[0201] SEQ ID NO:10

[0202] SVLRYDDFHIDEDKLDTNSVYEPYYHSDHAESSWVNRGESSRKAYDHNSPYIWPRNDYDGFLENAHEHHGVYNQGRGIDSGERLMQPTQMSAQEDLGDDTGIHVIPTLNGDDRHKIVNVDQRQYGDVFKGDLNPKPQGQRLIEVSVEENHPFTLRAPIQRIYGVRYTETWSFLPSLTCTGDAAPAIQHICLKHTTCFQDVVVDVDCAENTKEDQLAEISYRFQGKKEADQPWIVVNTSTLFDELELDPPEIEPGVLKVLRTEKQYLGVYIWNMRGSDGTSTYATFLVTWKGDEKTRNPTPAVTPQPRGAEFHMWNYHSHVFSVGDTFSLAMHLQYKIHEAPFDLLLEWLYVPIDPTCQPMRLYSTCLYHPNAPQCLSHMNSGCTFTSPHLAQRVASTVYQNCEHADNYTAYCLGISHMEPSFGLILHDGGTTLKFVDTPESLSGLYVFVVYFNGHVEAVAYTVVSTVDHFVNAIEERGFPPTAGQHHHHHHHHGSEKAAKAEEAADIIKLLNEQVNKEMQSSNLYMSMSSWCYTHSLDGAGLFLFDHAAEEYEHAKKLIIFLNENNVPVQLTSISAPEHKFEGLTQIFQKAYEHEQHISESINNIVDHAIKSKDHATFNFLQWYVAEQHEEEVLFKDILDKIELIGNENHGLYLADQYVKGIAKSRKS

[0203] SEQ ID NO:11

[0204] SVLRYDDFHIDEDKLDTNSVYEPYYHSDHAESSWVNRGESSRKAYDHNSPYIWPRNDYDGFLENAHEHHGVYNQGRGIDSGERLMQPTQMSAQEDLGDDTGIHVIPTLNGDDRHKIVNVDQRQYGDVFKGDLNPKPQGQRLIEVSVEENHPFTLRAPIQRIYGVRYTETWSFLPSLTCTGDAAPAIQHICLKHTTCFQDVVVDVDCAENTKEDQLAEISYRFQGKKEADQPWIVVNTSTLFDELELDPPEIEPGVLKVLRTEKQYLGVYIWNMRGSDGTSTYATFLVTWKGDEKTRNPTPAVTPQPRGAEFHMWNYHSHVFSVGDTFSLAMHLQYKIHEAPFDLLLEWLYVPIDPTCQPMRLYSTCLYHPNAPQCLSHMNSGCTFTSPHLAQRVASTVYQNCEHADNYTAYCLGISHMEPSFGLILHDGGTTLKFVDTPESLSGLYVFVVYFNGHVEAVAYTVVSTVDHFVNAIEHHHHHHHHGSEKAAKAEEAADIIKLLNEQVNKEMQSSNLYMSMSSWCYTHSLDGAGLFLFDHAAEEYEHAKKLIIFLNENNVPVQLTSISAPEHKFEGLTQIFQKAYEHEQHISESINNIVDHAIKSKDHATFNFLQWYVAEQHEEEVLFKDILDKIELIGNENHGLYLADQYVKGIAKSRKS

[0205] SEQ ID NO:12

[0206] SVLRYDDFHIDEDKLDTNSVYEPYYHSDHAESSWVNRGESSRKAYDHNSPYIWPRNDYDGFLENAHEHHGVYNQGRGIDSGERLMQPTQMSAQEDLGDDTGIHVIPTLNGDDRHKIVNVDQRQYGDVFKGDLNPKPQGQRLIEVSVEENHPFTLRAPIQRIYGVRYTETWSFLPSLTCTGDAAPAIQHICLKHTTCFQDVVVDVDCAENTKEDQLAEISYRFQGKKEADQPWIVVNTSTLFDELELDPPEIEPGVLKVLRTEKQYLGVYIWNMRGSDGTSTYATFLVTWKGDEKTRNPTPAVTPQPRGAEFHMWNYHSHVFSVGDTFSLAMHLQYKIHEAPFDLLLEWLYVPIDPTCQPMRLYSTCLYHPNAPQCLSHMNSGCTFTSPHLAQRVASTVYQNCEHADNYTAYCLGISHMEPSFGLILHDGGTTLKFVDTPESLSGLYVFVVYFNGHVEAVAYTVVSTVDHFVNAIEERGFPPTAGQPPATTKPKEITPVN

[0207] SEQ ID NO:13

[0208] SVLRYDDFHIDEDKLDTNSVYEPYYHSDHAESSWVNRGESSRKAYDHNSPYIWPRNDYDGFLENAHEHHGVYNQGRGIDSGERLMQPTQMSAQEDLGDDTGIHVIPTLNGDDRHKIVNVDQRQYGDVFKGDLNPKPQGQRLIEVSVEENHPFTLRAPIQRIYGVRYTETWSFLPSLTCTGDAAPAIQHICLKHTTCFQDVVVDVDCAENTKEDQLAEISYRFQGKKEADQPWIVVNTSTLFDELELDPPEIEPGVLKVLRTEKQYLGVYIWNMRGSDGTSTYATFLVTWKGDEKTRNPTPAVTPQPRGAEFHMWNYHSHVFSVGDTFSLAMHLQYKIHEAPFDLLLEWLYVPIDPTCQPMRLYSTCLYHPNAPQCLSHMNSGCTFTSPHLAQRVASTVYQNCEHADNYTAYCLGISHMEPSFGLILHDGGTTLKFVDTPESLSGLYVFVVYFNGHVEAVAYTVVSTVDHFVNAIEERGFPPTAGQ

[0209] SEQ ID NO:14

[0210] SVLRYDDFHIDEDKLDTNSVYEPYYHSDHAESSWVNRGESSRKAYDHNSPYIWPRNDYDGFLENAHEHHGVYNQGRGIDSGERLMQPTQMSAQEDLGDDTGIHVIPTLNGDDRHKIVNVDQRQYGDVFKGDLNPKPQGQRLIEVSVEENHPFTLRAPIQRIYGVRYTETWSFLPSLTCTGDAAPAIQHICLKHTTCFQDVVVDVDCAENTKEDQLAEISYRFQGKKEADQPWIVVNTSTLFDELELDPPEIEPGVLKVLRTEKQYLGVYIWNMRGSDGTSTYATFLVTWKGDEKTRNPTPAVTPQPRGAEFHMWNYHSHVFSVGDTFSLAMHLQYKIHEAPFDLLLEWLYVPIDPTCQPMRLYSTCLYHPNAPQCLSHMNSGCTFTSPHLAQRVASTVYQNCEHADNYTAYCLGISHMEPSFGLILHDGGTTLKFVDTPESLSGLYVFVVYFNGHVEAVAYTVVSTVDHFVNAIE

[0211] SEQ ID NO:15

[0212] GGGGSGGGGSGGGGS

[0213] SEQ ID NO:16

[0214] GGSGGSGS

[0215] SEQ ID NO:17

[0216] EKAAKAEEAA

[0217] References

[0218] 1.Gershon AA,Breuer J,Cohen JI,Cohrs RJ,Gershon MD,Gilden D,Grose C,Hambleton S,Kennedy PG,Oxman MN,Seward JF,Yamanishi K.Varicella zoster virusinfection.Nat Rev Dis Primers.2015Jul 2;1:15016.

[0219] 2.Takahashi M,Otsuka T,Okuno Y,Asano Y,Yazaki T.Live vaccine used toprevent the spread of varicella in children in hospital.Lancet.1974Nov 30;2(7892):1288-90.

[0220] 3.Zaia JA,Levin MJ,Preblud SR,Leszczynski J,Wright GG,Ellis RJ,CurtisAC,Valerio MA,LeGore J.Evaluation of varicella-zoster immune globulin:protection of immunosuppressed children after household exposure tovaricella.J Infect Dis.1983Apr;147(4):737-43.

[0221] 4.Arvin AM,Koropchak CM.Immunoglobulins M and G to varicella-zostervirus measured by solid-phase radioimmunoassay:antibody responses tovaricella and herpes zoster infections.J Clin Microbiol.1980Sep;12(3):367-74.

[0222] 5.Webster A,Grint P,Brenner MK,Prentice HG,Griffiths PD.Titration ofIgG antibodies against varicella zoster virus before bone marrowtransplantation is not predictive of future zoster.J Med Virol.1989Feb;27(2):117-9.

[0223] 6.Levin MJ,Oxman MN,Zhang JH,Johnson GR,Stanley H,Hayward AR,Caulfield MJ,Irwin MR,Smith JG,Clair J,Chan IS,Williams H,Harbecke R,MarcheseR,Straus SE,Gershon A,Weinberg A;Veterans Affairs Cooperative Studies ProgramShingles Prevention Study Investigators.Varicella-zoster virus-specificimmune responses in elderly recipients of a herpes zoster vaccine.J InfectDis.2008Mar 15;197(6):825-35.

[0224] 7.Weinberg A,Zhang JH,Oxman MN,Johnson GR,Hayward AR,Caulfield MJ,Irwin MR,Clair J,Smith JG,Stanley H,Marchese RD,Harbecke R,Williams HM,ChanIS,Arbeit RD,Gershon AA, F,Morrison VA,Kauffman CA,Straus SE,SchmaderKE,Davis LE,Levin MJ;USDepartment of Veterans Affairs(VA)Cooperative StudiesProgram Shingles Prevention Study Investigators.Varicella-zoster virus-specific immune responses to herpes zoster in elderly participants in a trialof a clinically effective zoster vaccine.J Infect Dis.2009Oct 1;200(7):1068-77.

[0225] 8.Heineman TC,Cunningham A,Levin M.Understanding the immunology ofShingrix,a recombinant glycoprotein E adjuvanted herpes zoster vaccine.CurrOpin Immunol.2019Aug;59:42-48.

[0226] 9.James SF,Chahine EB,Sucher AJ,Hanna C.Shingrix:The New AdjuvantedRecombinant Herpes Zoster Vaccine.Ann Pharmacother.2018Jul;52(7):673-680.

[0227] 10.Sullivan NL,Reuter-Monslow MA,Sei J,Durr E,Davis CW,Chang C,McCausland M,Wieland A,Krah D,Rouphael N,Mehta AK,Mulligan MJ,Pulendran B,Ahmed R,Vora KA.Breadth and Functionality of Varicella-Zoster VirusGlycoprotein-Specific Antibodies Identified after Zostavax Vaccination inHumans.J Virol.2018Jun 29;92(14):e00269-18.

Claims

1. A fusion protein comprising a varicella-zoster virus protein and ferritin; preferably, the fusion protein is in the form of nanoparticles.

2. The fusion protein of claim 1, wherein The varicella zoster virus protein is a varicella zoster virus glycoprotein or a varicella zoster virus interstitial protein; preferably, the varicella zoster virus protein is a gE, gI, gB, gH, gL or gC glycoprotein; more preferably, the varicella zoster virus protein is a gE glycoprotein; and / or The ferritin is selected from the group consisting of classical ferritin, heme-containing bacterial ferritin and DNA-binding ferritin of starved cells; preferably, the ferritin is bacterial ferritin, such as Helicobacter pylori ferritin.

3. The fusion protein according to claim 1 or 2, wherein The varicella zoster virus protein comprises or consists of a sequence selected from the following: (a) the amino acid sequence shown in SEQ ID NO: 1; and (b) a variant obtained by deleting 1-50 amino acids from the amino acid sequence shown in SEQ ID NO: 1 at the C-terminus; preferably, a variant obtained by deleting 1-5, 6-10, 11-15, 16-20, 21-25, 26-30, 31-35, 36-40, 41-45, or 46-50 amino acids from the amino acid sequence shown in SEQ ID NO: 1 at the C-terminus; more preferably, a variant obtained by deleting 17, 31 or 41 amino acids from the amino acid sequence shown in SEQ ID NO: 1 at the C-terminus; most preferably, the amino acid sequence of the variant is as shown in any one of SEQ ID NO: 12-14; and / or The ferritin protein comprises or consists of a sequence selected from the group consisting of: (c) the amino acid sequence shown in SEQ ID NO: 2; and (d) A variant obtained by subjecting the amino acid sequence shown in SEQ ID NO: 2 to one or more substitutions, deletions or additions.

4. The fusion protein according to any one of claims 1 to 3, wherein The ferritin is linked to the C-terminus of the varicella zoster virus protein; Preferably, the ferritin is connected to the C-terminus of the varicella zoster virus protein via a linker; More preferably, the linker is a GS-type linker (eg, (GGGGS)n, wherein n is an integer ≥ 1; GGSGGSGS or GS); EKAAKAEEAA; or any combination thereof.

5. The fusion protein according to any one of claims 1 to 4, further comprising a purification tag, preferably a His tag, a GST tag, an MBP tag, an HA tag, a c-Myc tag or a Flag tag, more preferably a His tag; Optionally, the purification tag is linked to the N-terminus or C-terminus of the varicella-zoster virus protein.

6. The fusion protein according to any one of claims 1 to 5, comprising or consisting of a sequence selected from the following: (e) an amino acid sequence as shown in any one of SEQ ID NOs: 4-11; and (f) A variant obtained by subjecting the amino acid sequence shown in any one of SEQ ID NOs: 4-11 to one or more substitutions, deletions or additions.

7. A nucleic acid encoding the fusion protein of any one of claims 1 to 6.

8. A pharmaceutical composition comprising the fusion protein according to any one of claims 1 to 6, and a pharmaceutically acceptable excipient, diluent or carrier.

9. A method for preparing the fusion protein according to any one of claims 1 to 6, comprising: Transforming the nucleic acid of claim 7 into a host cell, and culturing the host cell; and harvesting the fusion protein; Preferably, the method further comprises purifying the fusion protein.

10. Use of the fusion protein according to any one of claims 1 to 6 or the pharmaceutical composition according to claim 8 in the preparation of a medicament for preventing varicella zoster virus infection or a disease caused by the infection; Preferably, the drug is a vaccine; and / or The disease is chickenpox, herpes zoster, or chronic pain complicated by chickenpox or herpes zoster, neurological and eye diseases, or visceral and gastrointestinal diseases.

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