Nipah virus immunogens and their use
Inventors
Graham, Barney • Loomis, Rebecca • Stewart-Jones, Guillaume • Mascola, John • McLellan, Jason
Assignees
Dartmouth College • US Department of Health and Human Services
Publication Number
US-11890339-B2
Publication Date
2024-02-06
Expiration Date
2039-08-05
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Abstract
Embodiments of immunogens comprising a recombinant Nipah virus (NiV) F ectodomain trimer stabilized in a prefusion conformation are provided. Also provided are embodiments of immunogens comprising chimeric proteins comprising the recombinant NiV F ectodomain trimer and one or more G ectodomains, a multimer of NiV G ectodomains, and protein nanoparticles comprising the recombinant NiV F ectodomain trimer or an NiV G ectodomain. Also disclosed are nucleic acids encoding the immunogens and methods of their production. Methods for inducing an immune response in a subject by administering a disclosed immunogen to the subject are also provided. In some embodiments, the immune response treats or inhibits NiV infection in a subject.
Core Innovation
The invention provides immunogens comprising a recombinant Nipah virus (NiV) F ectodomain trimer stabilized in a prefusion conformation by one or more amino acid substitutions in protomers of the trimer. These substitutions include cysteine substitutions forming non-natural intra-protomer disulfide bonds, proline, phenylalanine, glycine substitutions, and deletions linked by glycine-serine linkers, resulting in improved stability and immunogenicity. The invention also includes chimeric proteins of the recombinant NiV F ectodomain trimer linked to one or more G ectodomains, multimers of NiV G ectodomains, protein nanoparticles containing the recombinant NiV F ectodomain trimer or NiV G ectodomain, nucleic acids encoding these immunogens, and methods for their production and use in inducing an immune response to treat or inhibit NiV infection.
NiV infection causes severe disease in humans, ranging from asymptomatic to fatal encephalitis, and also affects domestic animals like pigs. The NiV viral envelope includes membrane proteins F and G, which mediate fusion and attachment to host cells, respectively. The F protein naturally exists in a metastable prefusion conformation which triggers membrane fusion upon conformational change. However, no vaccine is available despite the illness burden caused by NiV. The invention addresses the need for effective immunogens eliciting a protective immune response by providing prefusion-stabilized NiV F ectodomain trimers with improved stability and immunogenicity compared to unstabilized forms.
The disclosed recombinant NiV F ectodomain trimers and their chimeras with G ectodomains show superior immune responses in animal models, with surprisingly good combinations of stability, homogeneity, yield, and immunogenicity. The invention provides methods to design such stabilization via specific amino acid substitutions or deletions, trimerization domains (such as GCN4 or T4 fibritin), and fusion to heterologous proteins. It also provides nucleic acids and vectors encoding these immunogens and methods to induce an immune response in a subject to prevent or treat NiV infection.
Claims Coverage
The patent presents one independent claim encompassing an immunogen comprising a recombinant NiV F ectodomain trimer stabilized in a prefusion conformation by specific amino acid substitutions. The claim covers main inventive features related to stabilization mutations, structural compositions, conjugations, nucleic acid molecules encoding the immunogen, methods of production, immunogenic compositions, and methods of inducing immune responses.
Prefusion stabilization of NiV F ectodomain trimer
The immunogen comprises a recombinant NiV F ectodomain trimer stabilized in a prefusion conformation by one or more amino acid substitutions, including cysteine substitutions that form intra-protomer disulfide bonds at positions 104 and 114, or 114 and 426; a proline substitution at position 191; a phenylalanine substitution at position 172; a glycine substitution at position 70; and deletion of positions 102-113 with positions 101 and 114 linked by a glycine-serine linker, based on the reference NiV F sequence SEQ ID NO: 52.
Specific exemplary stabilizing mutation combinations
Stabilization is particularly obtained with cysteine substitutions at positions 104 and 114, combined with the proline substitution at 191 and phenylalanine substitution at 172, as well as various other combinations involving these cysteine substitutions with proline, phenylalanine, glycine substitutions, or deletions linked by glycine-serine linker.
Recombinant protomers composition and sequence identity
Protomers comprise an F2 protein encompassing NiV F positions 25-109 and an F1 ectodomain including positions 110-488, comprising amino acid sequences at least 90% identical or consisting of residues 21-486 or 21-477 from a list of specific SEQ ID NOs, reflecting their stabilized forms.
Trimerization domain fusion and composition
The NiV F ectodomain trimer protomers are fused C-terminally to a trimerization domain, such as a GCN4 trimerization domain or a T4 fibritin trimerization domain, including a specific GCN4 sequence (residues 485-519 of SEQ ID NO: 1), with the protomers comprising amino acid sequences of residues 21-519 or 21-510 of specific SEQ ID NOs.
Conjugation to heterologous proteins including NiV G ectodomains
The recombinant NiV F ectodomain trimer protomers can be fused to heterologous proteins, including NiV G ectodomains, optionally linked to at least three NiV G ectodomains fused at the N-terminus of the protomers or C-terminus of trimerization domains, with specific sequences of the F-G chimeras exemplified by SEQ ID NOs.
Nucleic acids, vectors and methods of production
Nucleic acid molecules encoding the disclosed immunogens are claimed, including those operably linked to promoters and contained in vectors such as RNA vectors, and methods of producing the immunogen by expressing the nucleic acid molecules in host cells followed by purification.
Immunogenic compositions and methods of immune response induction
Immunogenic compositions comprising the immunogen and pharmaceutically acceptable carriers are claimed, together with methods of administering an effective amount of such compositions to a subject to elicit immune responses preventing or inhibiting NiV infection.
The claims cover the stabilized recombinant NiV F ectodomain trimer immunogen defined by precise amino acid modifications conferring prefusion stability, fusion to trimerization domains and heterologous proteins such as NiV G, nucleic acid and vector compositions encoding the immunogen, methods for its production, and compositions for administration to induce protective immune responses against NiV infection.
Stated Advantages
The disclosed prefusion-stabilized NiV F ectodomain trimers produce a superior immune response in animal models compared to corresponding trimers not stabilized in the prefusion conformation.
The immunogens including chimeras of NiV F ectodomain trimers and NiV G ectodomains provide a surprisingly good combination of stability, homogeneity, yield, and immunogenicity.
The immunogens elicit neutralizing immune responses effective to treat or inhibit NiV infection.
The recombinant NiV F ectodomain trimers and multimeric G constructs are stable in solution and maintain their prefusion conformation upon storage and incubation at specified temperatures, being suitable for use in vaccines.
Documented Applications
The disclosed immunogens are used to induce an immune response in a subject to prevent, inhibit, or treat Nipah virus infection.
Immunogens are used in immunogenic compositions for vaccination of humans and domestic animals such as pigs.
The immunogens are administered by various routes including parenteral injection, mucosal, or respiratory tract delivery, and can be used in prime-boost vaccine regimens with mRNA or protein-based platforms.
Recombinant NiV F ectodomain trimers and chimeric NbV F-G multimers are applicable in vaccine formulations to elicit neutralizing antibodies against NiV.
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