Structure-guided design of native-like HIV Env Single Chain Trimers for enhanced stability, immunogenicity, and versatile vaccine delivery

Xuduo Li; Ipsita Krishnamurthy; Nitesh Mishra; Nicole E. James; Rohan Roy Chowdhury; Sean Callaghan; Anjali Somanathan; Gabriel Avillion; Khaled Amereh; Tharunika Sekar; Prabhgun Oberoi; Yuxin Zhu; Jonathan L. Torres; Wen-Hsin Lee; Joel D. Allen; Bo Liang; Gabriella Giese; Anh L. Vo; Sharaf Andrabi; Ishita Jain; Asanti Mergiya; Tazio Capozzola; Wanting He; Fabio Anzanello; Rumi Habib; Frédéric Bibollet‐Ruche; Rebecca Nedellec; Sara Dutton Sackett; Gabriel Ozorowski; Dennis R. Burton

doi:10.1101/2025.10.25.684215

ImmunologybioRxivOct 25, 2025

Preprint

Structure-guided design of native-like HIV Env Single Chain Trimers for enhanced stability, immunogenicity, and versatile vaccine delivery

Xuduo Li iD, Ipsita Krishnamurthy iD, Nitesh Mishra iD, Nicole E. James, Rohan Roy Chowdhury iD, Sean Callaghan iD, Anjali Somanathan, Gabriel Avillion iD, Khaled Amereh iD, Tharunika Sekar, Prabhgun Oberoi, Yuxin Zhu iD, Jonathan L. Torres iD, Wen-Hsin Lee iD, Joel D. Allen iD, Bo Liang iD, Gabriella Giese, Anh L. Vo iD, Sharaf Andrabi, Ishita Jain, Asanti Mergiya, Tazio Capozzola iD, Wanting He iD, Fabio Anzanello iD, Rumi Habib iD, , Raiees Andrabi iD

Abstract

Stabilizing the HIV envelope (Env) trimer in its native prefusion conformation is key to eliciting broadly neutralizing antibodies (bnAbs). We present a generalizable single-chain trimer-(SCT) design platform that enables the production of stable, native-like Env trimers across diverse HIV strains. Using a structure-guided approach, we developed CAP256.SU-SCT9, based on a CAP256.SU Env shown to induce V2-apex bnAbs in humans and rhesus macaques. CAP256.SU-SCT9 trimer exhibited native-like structure, bnAb-specific antigenicity, authentic glycosylation, and compatibility with nucleic acid and nanoparticle delivery. We extended this design strategy to a panel of HIV Envs representing global diversity, and immunogens known to initiate bnAb responses in humans. Structural analyses confirmed that SCTs consistently maintained prefusion-closed conformations, favorable glycan profiles, strong binding to trimer-specific bnAbs, and low reactivity with non-neutralizing antibodies. These findings establish SCT as a versatile platform for HIV trimer immunogen design, with potential to support next-generation vaccines aimed at eliciting protective bnAb responses.

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