P-B1 Characterization of broadly neutralizing nanobodies from dromedaries immunized with soluble trimeric subtype C SOSIP proteins

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Nanobodies or VHH are the smallest naturally occurring antibody fragments derived from heavy chain only antibodies from Camelidae. They are characterized by high stability, high affinity, and high specificity for their target antigens, as well as having extended CDR3 loops.These properties make nanobodies suitable tools for therapeutic and preventive applications. We immunized dromedaries with gp140 SOSIPs from HIV-1 subtype C, generated VHH phage libraries from these animals, and selected Env-specific VHHs on the autologous SOSIPs. After screening 1300 phage clones, we tested 80 Env-specific soluble VHHs for neutralizing activity against the autologous and a heterologous subtype B pseudovirus in the Tzm-bl assay. Eight neutralizing VHH were further tested against a panel of 21 HIV-1 pseudoviruses, which includes the 12 pseudoviruses from the extended global panel of HIV-1 reference strains for standardized assessment of vaccine-elicited neutralizing antibodies (deCamp et al, 2014). All 8 VHHs showed neutralization of Tier 2 pseudoviruses from at least 2 and up to 6 different subtypes including circulating recombinant forms (CRF). Although the neutralization breadth differed for the individual VHHs, some VHHs showed complementary neutralization patterns covering 19 of 21 pseudoviruses in our panel including the epidemiologically most relevant subtypes C and A. Preliminary epitope mapping data by competitive ELISAs with known bnAbs, as well as negative stain EM structures with trimeric SOSIPs, identified the CD4 binding site as the major target. The broadly neutralizing nanobodies identified here are promising candidates for further development as prophylactic/therapeutic treatments of HIV-1 infection.

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