Mapping the neutralizing specificity of human anti-HIV serum by deep mutational scanning

Understanding the specificities of human serum antibodies that broadly neutralize HIV can inform prevention and treatment strategies. Here we describe a deep mutational scanning system that can measure the effects of combinations of mutations to HIV envelope (Env) on neutralization by antibodies and polyclonal serum. We first show that this system can accurately map how all functionally tolerated mutations to Env affect neutralization by monoclonal antibodies. We then comprehensively map Env mutations that affect neutralization by a set of human polyclonal sera known to target the CD4-binding site that neutralize diverse strains of HIV. The neutralizing activities of these sera target different epitopes, with most sera having specificities reminiscent of individual characterized monoclonal antibodies, but one sera targeting two epitopes within the CD4 binding site. Mapping the specificity of the neutralizing activity in polyclonal human serum will aid in assessing anti-HIV immune responses to inform prevention strategies. ### Competing Interest Statement JDB is on the scientific advisory boards of Apriori Bio, Invivyd, Aerium Therapeutics, and the Vaccine Company. JDB, KHDC, BD, ASD, and CER receive royalty payments as inventors on Fred Hutch licensed patents related to viral deep mutational scanning. ASD is currently an employee of Apriori Bio, though his contributions to this manuscript were performed when he was an employee of the Fred Hutch before he started work at Apriori Bio. A patent application encompassing aspects of this work has been filed by the University of Cologne and lists P.S. and F.K. as inventors. P.S., and F.K. received payments from the University of Cologne for licensed antibodies.