Engineering well-expressed, V2-immunofocusing HIV-1 envelope glycoprotein membrane trimers for use in heterologous prime-boost vaccine regimens.

Autor:	Crooks ET; San Diego Biomedical Research Institute, San Diego, California, United States of America., Almanza F; San Diego Biomedical Research Institute, San Diego, California, United States of America., D'Addabbo A; School of Biological Sciences, University of Southampton, Southampton, United Kingdom., Duggan E; Scintillon Institute, San Diego, California, United States of America.; Cellarcus BioSciences, La Jolla, California, United States of America., Zhang J; San Diego Biomedical Research Institute, San Diego, California, United States of America., Wagh K; Theoretical Biology & Biophysics, Los Alamos National Laboratory, Los Alamos, New Mexico, United States of America., Mou H; Department of Immunology and Microbial Science, The Scripps Research Institute, Jupiter, Florida, United States of America., Allen JD; School of Biological Sciences, University of Southampton, Southampton, United Kingdom., Thomas A; San Diego Biomedical Research Institute, San Diego, California, United States of America., Osawa K; San Diego Biomedical Research Institute, San Diego, California, United States of America., Korber BT; Theoretical Biology & Biophysics, Los Alamos National Laboratory, Los Alamos, New Mexico, United States of America., Tsybovsky Y; Frederick National Laboratory for Cancer Research, Leidos Biomedical Research, Inc., Frederick, Maryland, United States of America., Cale E; Vaccine Research Center, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, United States of America., Nolan J; Scintillon Institute, San Diego, California, United States of America.; Cellarcus BioSciences, La Jolla, California, United States of America., Crispin M; School of Biological Sciences, University of Southampton, Southampton, United Kingdom., Verkoczy LK; San Diego Biomedical Research Institute, San Diego, California, United States of America., Binley JM; San Diego Biomedical Research Institute, San Diego, California, United States of America.
Jazyk:	angličtina
Zdroj:	PLoS pathogens [PLoS Pathog] 2021 Oct 22; Vol. 17 (10), pp. e1009807. Date of Electronic Publication: 2021 Oct 22 (Print Publication: 2021).
DOI:	10.1371/journal.ppat.1009807
Abstrakt:	HIV-1 vaccine immunofocusing strategies may be able to induce broadly-reactive neutralizing antibodies (NAbs). Here, we engineered a panel of diverse, membrane-resident native HIV-1 trimers vulnerable to two broad targets-the V2 apex and fusion peptide (FP). Selection criteria included i) high expression and ii) infectious function, so that trimer neutralization sensitivity can be profiled in pseudovirus (PV) assays. Initially, we boosted the expression of 17 candidate trimers by truncating gp41 and introducing a gp120-gp41 SOS disulfide to prevent gp120 shedding. "Repairs" were made to fill glycan holes and eliminate other strain-specific aberrations. A new neutralization assay allowed PV infection when our standard assay was insufficient. Trimers with exposed V3 loops, a target of non-NAbs, were discarded. To try to increase V2-sensitivity, we removed clashing glycans and modified the C-strand. Notably, a D167N mutation improved V2-sensitivity in several cases. Glycopeptide analysis of JR-FL trimers revealed near complete sequon occupation and that filling the N197 glycan hole was well-tolerated. In contrast, sequon optimization and inserting/removing glycans at other positions frequently had global "ripple" effects on glycan maturation and sequon occupation throughout the gp120 outer domain and gp41. V2 MAb CH01 selectively bound to trimers with small high mannose glycans near the base of the V1 loop, thereby avoiding clashes. Knocking in a rare N49 glycan was found to perturb gp41 glycans, increasing FP NAb sensitivity-and sometimes improving expression. Finally, a biophysical analysis of VLPs revealed that i) ~25% of particles bear Env spikes, ii) spontaneous particle budding is high and only increases 4-fold upon Gag transfection, and iii) Env+ particles express ~30-40 spikes. Taken together, we identified 7 diverse trimers with a range of sensitivities to two targets to allow rigorous testing of immunofocusing vaccine concepts. Competing Interests: The authors have declared that no competing interests exist.
Databáze:	MEDLINE
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