PROJECT SUMMARY A major roadblock to rational HIV-1 vaccine design is the lack of a suitable primate model in which broadly neutralizing antibodies (bNAbs) can be commonly induced and the molecular, biological and immunological mechanisms responsible for eliciting such responses studied in a reproducible and iterative fashion. Recently, we demonstrated that primary HIV-1 Envs, when expressed by simian-human immunodeficiency viruses (SHIVs) in rhesus macaques (RMs), elicited patterns of Env-antibody coevolution strikingly similar to humans infected by homologous virus strains, leading to neutralization breadth. These similarities in Env-antibody coevolution in humans and rhesus included conserved immunogenetic, structural and chemical solutions to epitope recognition and precise Env amino acid substitutions, insertions and deletions leading to virus persistence. The structure of one rhesus bNAb, capable of neutralizing 49% of a 208-strain panel, revealed a V2-apex mode of recognition like that of human bNAbs PGT145 and PCT64-35M. Another rhesus antibody bound the CD4-binding site of HIV-1 Env by CD4 mimicry mirroring human bNAbs 8ANC131, CH235 and VRC01. Based on these observations supporting the relevance of the rhesus model to bNAb induction in humans, we propose here a novel “reverse vaccinology” strategy in SHIV infected RMs as a “molecular guide” to inform and accelerate HIV-1 vaccine design in humans. Specific aims are: (i) To isolate bNAb mAbs targeting CD4bs, fusion peptide, V3 glycan and V2 apex epitopes from a subset of 150 SHIV infected RMs and to characterize their breadth, potency, immunogenetics, target epitopes and structural solutions to epitope recognition. (ii) To characterize molecular patterns of Env-Ab coevolution from rhesus germline B cell unmutated common ancestors (UCAs) to mature bNAbs and to identify key Env intermediates, or “immunotypes,” that are responsible for driving bNAb lineage affinity maturation to breadth. (iii) To design, construct and characterize novel SOSIP Env trimers that mimic key Env “immunotypes” and demonstrate that they bind preferentially to bNAb UCAs and intermediate stage Abs. (iv) To conduct a proof- of-concept preclinical vaccine trial in 24 RMs to test the hypothesis that reverse-engineered, B lineage-designed SOSIP Env trimers can prime, boost and affinity mature bNAb responses in RMs to an extent that is superior to conventional SOSIP Env immunogens and comparable to SOSIP-SHIV or SHIV-only immunizations.