Glycan-masked engineered outer domains of HIV-1 gp120 and their use
Inventors
Chen, Xuejun • Boyington, Jeffrey • Holdsworth, Hongying Duan • Cheng, Cheng • Mascola, John R.
Assignees
US Department of Health and Human Services
Publication Number
US-12053520-B2
Publication Date
2024-08-06
Expiration Date
2038-03-26
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Abstract
Embodiments of immunogens based on the outer domain of HIV-1 gp120 and methods of their use and production are disclosed. Nucleic acid molecules encoding the immunogens are also provided. In several embodiments, the immunogens can be used to prime an immune response to gp120 in a subject, for example, to treat or prevent an HIV-1 infection in the subject.
Core Innovation
Embodiments of immunogens based on the engineered outer domain (eOD) of HIV-1 gp120 are disclosed, which include specific amino acid substitutions to introduce N-linked glycan sequons that mask non-CD4 binding site (non-CD4bs) epitopes. These engineered immunogens specifically bind VRC01-class broadly neutralizing antibodies (bnAbs) and their inferred germline revertants, aiming to induce an immune response focused on the CD4 binding site (CD4bs) of gp120.
The invention addresses the challenge that most broadly neutralizing antibodies to HIV-1 undergo extensive somatic mutation and their germline precursors bind poorly or not at all to viral antigens. Prior germline-targeting immunogens, such as eOD-GT8, elicit significant off-target immune responses that reduce their effectiveness in priming neutralizing antibodies. Therefore, there is a need for immunogens that selectively activate rare bnAb precursors while reducing off-target immune activation.
The immunogens disclosed include eOD variants of HIV-1 gp120 with engineered N-linked glycosylation sites at specific residues to mask non-CD4bs surfaces, thereby substantially focusing the immune response toward the CD4bs. These modifications result in improved antigenicity by reducing binding to non-CD4bs antibodies and maintaining strong binding to VRC01-class antibodies and their precursors. The immunogens can be linked to self-assembling protein nanoparticles, such as lumazine synthase 60mers, enhancing immune presentation.
Claims Coverage
The claims include one independent claim directed to an isolated immunogen comprising an engineered gp120 outer domain (eOD) with specific amino acid modifications, including glycan sequons, and additional dependent claims addressing preferred features, linkage to nanoparticles and carriers, and methods of inducing immune responses.
Engineered gp120 outer domain with specific glycan sequons
An engineered gp120 outer domain comprising an amino acid sequence based on SEQ ID NO: 1, further modified by amino acid substitutions and insertions to introduce N-linked glycan sequons at one or more of residues 8, 14, 56, 70, 71, 74, 102, and 153. A glycine insertion is introduced between residues 10 and 11 if the sequon at residue 8 is introduced, and an E70G substitution is introduced if the sequon at residue 71 is introduced. Optionally, a GG substitution at positions 70 and 71 is included if sequons at these positions are not introduced.
Retention of native glycan sequons and critical residues
The engineered gp120 outer domain retains N-linked glycan sequons at residues corresponding to 18, 65, 92, 98, 106, 113, 129, 146, 165, and 170 of SEQ ID NO: 1. Residues corresponding to 25-45, 79-85, and 126-145 of SEQ ID NO: 1 are identical to SEQ ID NO: 1, and remaining residues are at least 90% identical to SEQ ID NO: 1.
Specific binding to VRC01-class antibodies and immune focus
The engineered gp120 outer domain specifically binds to VRC01-class broadly neutralizing antibodies and their inferred germline revertants, and induces an immune response targeting the CD4 binding site, including activation of memory B cells where at least 50% bind to the VRC01 binding site.
Linkage to self-assembling protein nanoparticles
The engineered gp120 outer domain is linked, directly or via a peptide linker, to a subunit of a self-assembling protein nanoparticle, preferably a lumazine synthase subunit, which forms multimers such as 60mers.
Carrier protein linkage and peptide linker usage
The engineered gp120 outer domain can be linked to a carrier protein, optionally via a peptide linker such as a glycine-serine peptide linker with the amino acid sequence of SEQ ID NO: 50 (GGSGGSGGSGGSGGG).
Multimeric presentation in protein nanoparticles or virus-like particles
The immunogen may comprise multimers of the engineered gp120 outer domain assembled into protein nanoparticles or virus-like particles, including lumazine synthase 60mer nanoparticles.
Administration methods and immune response induction
Methods of generating an immune response to the CD4 binding site of gp120 include administering the immunogen, protein nanoparticle, virus-like particle, nucleic acid encoding the immunogen, or immunogenic composition to a subject to prime or boost the immune response, leading to production of IGHV1-2*02 antibodies that bind to the engineered gp120 outer domain and inhibit or treat HIV-1 infection.
The claims cover engineered HIV-1 gp120 outer domain immunogens with specific glycan-masking modifications to reduce off-target immune responses while focusing the response on the CD4 binding site, including their presentation on protein nanoparticles, linkage to carriers, and use in methods to induce CD4bs-targeted immune responses to prevent or treat HIV-1 infection.
Stated Advantages
The immunogens provide a substantially more focused immune response to the VRC01-class precursor antibodies compared to prior eOD immunogens.
The glycan-masking reduces production of off-target immune responses while maintaining strong binding to CD4bs-specific broadly neutralizing antibodies and their germline revertants.
Selected glycan masked eOD-GT8 60mer mutants elicit an increased frequency of CD4bs-specific B cells among antigen-specific B cells, improving the potential to induce broadly neutralizing antibodies.
Documented Applications
The immunogens can be used to prime an immune response to gp120 in a subject, for example, to treat or prevent an HIV-1 infection.
The immunogens can be used to induce an immune response that activates memory B cells targeting the CD4 binding site of gp120.
The disclosed immunogens can be used in coordinate prime-boost vaccination protocols, where the prime includes the engineered eOD immunogen and the boost includes HIV-1 Env protein or variants thereof.
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