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CN-122005773-A - African swine fever mRNA vaccine composition capable of synergistically activating and enhancing humoral immunity and cellular immunity and application thereof

CN122005773ACN 122005773 ACN122005773 ACN 122005773ACN-122005773-A

Abstract

The invention discloses an African swine fever mRNA vaccine composition capable of synergistically activating and enhancing humoral immunity and cellular immunity and application thereof, and belongs to the technical field of biological medicines. Specifically, the invention takes a plurality of key antigen proteins of African swine fever virus as targets, and forms single-chain mRNA through the cascade connection of connectors, so that multiple immunogens can be translated from the same transcript and multiple antibodies can be induced, and a conservative recombinant T cell immunogen capable of activating cellular immune response is introduced, so that the cellular immune response can be enhanced, the ADE risk can be reduced, the cross protection capability can be improved, and in addition, a molecular adjuvant IL-12 is added into the mRNA vaccine to enhance the immune response of the African swine fever vaccine, so that stronger immune protection efficacy can be induced.

Inventors

  • XU JIANQING
  • ZHANG XIAOYAN
  • Bai Shimeng

Assignees

  • 复旦大学附属中山医院

Dates

Publication Date
20260512
Application Date
20260210

Claims (10)

  1. 1. A nucleic acid composition for inducing an immune response against African Swine Fever Virus (ASFV), characterized in that the nucleic acid composition comprises a combination of 1) and 2) or a combination of 1) and 2) and 3): 1) At least one mRNA encoding an ASFV immunogen 2) At least one mRNA encoding a recombinant T cell immunogen; 3) At least one mRNA encoding an immunomodulatory molecule; wherein the mRNA is encapsulated in or complexed with a delivery system to effect in vivo delivery.
  2. 2. The nucleic acid composition of claim 1, wherein the mRNA encoding an ASFV immunogen encodes any one of the following immunogens: 1) The amino acid sequence of the fusion protein (CD 2v-p54-p 30) formed by connecting CD2v/pEP402R, p/pE 183L and p30/CP204L through a connector is shown as SEQ ID NO. 15; 2) The amino acid sequence of the fusion protein P49-B602L formed by connecting P49/B438L and B602L through a linker is shown as SEQ ID NO. 16; 3) The amino acid sequence of the fusion protein P49-Penton formed by connecting P49/B438L and Penton/H240R through a linker is shown as SEQ ID NO. 17; 4) The amino acid sequence of the fusion protein P72-Penton formed by connecting P72/B646L and Penton/H240R through a linker is shown as SEQ ID NO. 18; 5) The amino acid sequence of the fusion protein P72-B602L formed by connecting P72/B646L and B602L through a linker is shown in SEQ ID NO. 19; And/or mRNA encoding recombinant T cell immunogen encodes fusion protein p17-T formed by connecting recombinant conservative T cell antigen and p17/D117L through a connector, and the amino acid sequence of the fusion protein p17-T is shown as SEQ ID NO. 20.
  3. 3. The nucleic acid composition of claim 2, wherein the mRNA encoding an ASFV immunogen encodes any one of the following codon optimized immunogens: i) Encoding the candidate immunogen 1 with optimized codons, wherein the nucleic acid sequence after codon optimization is shown as SEQ ID NO. 21; ii) encodes a codon optimised candidate immunogen 2, the codon optimised nucleic acid sequence being shown in SEQ ID NO. 22; iii) Encoding the candidate immunogen 3 with optimized codons, wherein the nucleic acid sequence after codon optimization is shown as SEQ ID NO. 23; iv) encoding a codon optimised candidate immunogen 4, the codon optimised nucleic acid sequence being shown in SEQ ID NO. 24; v) encoding a codon optimised candidate immunogen 5, the codon optimised nucleic acid sequence being as shown in SEQ ID NO. 25; And/or, the mRNA encoding the recombinant T cell immunogen encodes the p17-T with optimized codon, and the nucleic acid sequence after codon optimization is shown as SEQ ID NO. 26.
  4. 4. The nucleic acid composition of any one of claims 1-3, wherein the mRNA encoding the ASFV immunogen is a single-stranded mRNA and the open reading frame thereof encodes a multi-antigen fusion protein.
  5. 5. The nucleic acid composition of any one of claims 1-4, wherein the immunoregulatory molecule is selected from one or more of an adjuvant and/or a cytokine; preferably, the adjuvant is selected from one or more of aluminium adjuvants, cholera toxin and subunits thereof, oligodeoxynucleotides, manganese ion adjuvants, colloidal manganese adjuvants, freund's adjuvant, MF59, QS-21, poly I: C and other TLR ligands; preferably, the cytokine is selected from one or more of GM-CSF, IL-2, IL-3, IL-7, IL-11, IL-12, IL-18, IL-21 and IFN- α, more preferably, the immunoregulatory molecule is IL-12.
  6. 6. The nucleic acid composition of claim 5, wherein the IL-12 is mouse-derived IL-12 and/or pig-derived IL-12; preferably, the mRNA encoding IL-12 comprises a codon optimized coding sequence of: 1) The coding sequence of mouse IL-12 is shown as SEQ ID NO 27, and/or 2) The coding sequence of the pig IL-12 is shown as SEQ ID NO. 28.
  7. 7. The nucleic acid composition of any one of claims 1-6, wherein the mRNA encoding an ASFV immunogen and the mRNA encoding an immunomodulatory molecule are administered at an equal mass and/or an equal molar mass; and/or, the nucleic acid composition is administered in a manner that satisfies any of the following: a) The mRNA in the combination is respectively wrapped in a delivery system and then is singly administered, or B) The mRNA in the combination is respectively wrapped in a delivery system and then mixed for administration, or C) mRNA in the combination is combined after being respectively prepared, or D) The mRNAs in the combination are co-packaged in the same delivery system and then administered.
  8. 8. The nucleic acid composition of any one of claims 1-7, wherein the delivery system is a carrier system capable of delivering mRNA into animal cells, the carrier system selected from the group consisting of liposomes, lipid nanoparticles, polymer nanoparticles, cationic polymer complexes, inorganic nanocarriers, emulsions, or combinations thereof; and/or, the mRNA is an unmodified mRNA and/or an mRNA containing a nucleotide modification.
  9. 9. An immunogenic peptide derived from African Swine Fever Virus (ASFV), characterized in that the immunogenic peptide is selected from the group consisting of any one or more of the following 1), 2), 3), 4), 5) and 6): 1) A fusion protein CD2v-p54-p30 formed by connecting CD2v/pEP402R, p/pE 183L and p30/CP204L through a connector or a variant thereof, wherein the amino acid sequence of the CD2v-p54-p30 is shown as SEQ ID NO. 15, the variant is a sequence with at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQ ID NO. 15, and the function of the fusion protein shown as SEQ ID NO. 15 is reserved; 2) A fusion protein P49-B602L formed by connecting P49/B438L and B602L through a connector, wherein the amino acid sequence of the P49-B602L is shown as SEQ ID NO. 16, the variant is a sequence with at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQ ID NO. 16, and the function of the fusion protein shown as SEQ ID NO. 16 is reserved; 3) A fusion protein P49-Penton formed by connecting P49/B438L and Penton/H240R through a connector, wherein the amino acid sequence of the P49-Penton is shown as SEQ ID NO. 17, the variant is a sequence with at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQ ID NO. 17, and the function of the fusion protein shown as SEQ ID NO. 17 is reserved; 4) Fusion protein P72-Penton formed by connecting P72/B646L and Penton/H240R through a connector, wherein the amino acid sequence of P72-Penton is shown as SEQ ID NO. 18, and the variant is a sequence with at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQ ID NO. 18, and the function of the fusion protein shown as SEQ ID NO. 18 is reserved; 5) A fusion protein P72-B602L formed by connecting P72/B646L and B602L through a connector, wherein the amino acid sequence of the P72-B602L is shown as SEQ ID NO. 19, the variant is a sequence with at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQ ID NO. 19, and the function of the fusion protein shown as SEQ ID NO. 19 is reserved; 6) The amino acid sequence of the p17-T is shown as SEQ ID NO. 20, and the variant is a sequence with at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQ ID NO. 20, and retains the function of the fusion protein shown as SEQ ID NO. 20.
  10. 10. Use of the nucleic acid composition of any one of claims 1-8 or the immunogenic peptide of claim 9 in the manufacture of a medicament for the prevention and/or treatment of an ASFV infection, wherein the medicament is administered alone or in combination with an anti-ASFV antibody and/or an antiviral medicament; Preferably, the medicament is administered in combination with an anti-ASFV antibody and/or an antiviral medicament.

Description

African swine fever mRNA vaccine composition capable of synergistically activating and enhancing humoral immunity and cellular immunity and application thereof Technical Field The invention belongs to the technical field of biological medicines, and relates to an African swine fever mRNA vaccine composition capable of synergistically activating and enhancing humoral immunity and cellular immunity and application thereof. Background African Swine Fever Virus (ASFV) is a large linear double-stranded DNA virus, the genome is about 170-194 kb, the encoding of more than 150 Open Reading Frames (ORFs), the structure is complex in level, and severe hemorrhagic fever can be caused in domestic pigs and wild pigs, and the acute illness death rate can be close to 100%. ASFV can be transmitted through various ways such as toxic pigs and secretions thereof, pollution devices/feeds, pork products and the like, is not easy to inactivate in polluted meat products, and is easy to cause long-distance diffusion and regional continuous popularity. The broad spectrum and rapid renewal of vaccines are continually challenged by the large number of ASFV genotypes, significant epidemic strain lineage differences, and the possibility of large-scale genomic variations such as deletions, recombinations, etc. The current vaccine development route for ASFV mainly comprises attenuated live vaccine, inactivated vaccine, subunit vaccine, viral vector vaccine, DNA vaccine, mRNA vaccine and the like. Although the above routes are researched and tried, the problems of unstable immune protection, difficulty in considering safety and effectiveness, limited cross protection on heterologous strains, insufficient industrial consistency and the like are still faced, and the comprehensive requirements of 'safety, effectiveness, large-scale and rapid iteration' on long-term prevention and control of serious animal epidemic diseases are still difficult to meet. A limitation of live attenuated vaccines is that live attenuated vaccines typically rely on deletion of specific virulence related genes to reduce pathogenicity. Due to the complex immune escape mechanism of ASFV, single or limited gene deletion does not necessarily ensure sufficient safety, and multiple gene deletion may lead to reduced immunogenicity and insufficient or unstable protective power. In addition, attenuated strains may still present genetic stability risks during serial passage or in vivo replication, and exhibit virulence reversion or residual pathogenicity, thereby bringing biosafety and regulatory assessment pressure. Attenuated live vaccines have certain dependence on strain background, and the attenuation effect and protection effect among different genotypes/isolates may be obviously different, so that the universal application of the attenuated live vaccines in epidemic strains in different areas is further limited. The limitation of inactivated vaccines and subunit vaccines is that it is often difficult for an inactivated vaccine to induce a sufficient protective immune response, especially in terms of cellular immunity, and to provide stable protection against challenge. Subunit vaccines have safety advantages, but the immunogen screening and combination are highly dependent on the accurate recognition of protective antigens/epitopes, and single or few structural protein antigens are difficult to realize stable broad-spectrum protection under the ASFV diversity background, and the cross protection capability between different strains is limited. The limitation of viral vector vaccines and DNA vaccines is that viral vector vaccines can induce both humoral and cellular immunity and are considered to be one of the important directions of ASFV vaccines. The earlier filed patent of the team (African swine fever virus immunogen combination and application thereof) discloses an African swine fever virus immunogen composition and application thereof, wherein the vaccine can be a viral vector vaccine (such as recombinant vaccinia virus rTV vaccine) or a nucleic acid vaccine and the like, and the scheme emphasizes that humoral immunity and T cell immune response are simultaneously activated from the basis of an immunological mechanism, so that the African swine fever virus immunogen composition has important technical significance and application value. However, although the existing viral vector vaccine and DNA vaccine can induce effective immune response, the existing viral vector vaccine and DNA vaccine still have the following defects that firstly, ASFV protection has obvious strain/region specificity, the vector vaccine or DNA vaccine has relatively long iteration cycle in terms of antigen matching construction, verification and amplification, and is difficult to form a rapid response loop for new epidemic strains, secondly, multi-antigen loading can promote immune coverage, but the increase of exogenous gene quantity possibly brings higher requirements on construction effici