Circulating (tier 2) HIV-1 variants are highly resistant to antibody-mediated neutralization, making broadly effective vaccine design a major challenge. Encouragingly, work in our previous HIVRAD program demonstrated that Ab responses capable of cross-neutralizing multiple heterologous tier 2 viruses were elicited by targeted N-glycan deletion priming and heterologous glycan restorative Env trimer-liposome boosting. The isolation of two monoclonal antibodies with broadly neutralizing activity from these studies and high-resolution structures in complex with native-like trimers revealed that one mAb targeted the conserved CD4 binding site (CD4bs) and the other targeted the gp41:gp120 interface region, substantiated this result. In the current application, we will build on these efforts by evaluating responses elicited by well-ordered, novel, trimer-based immunogens inoculated into Indian origin rhesus macaques. In addition to the use pf protein-based trimers, we will test trimer platforms based on administration of mRNA lipid nanoparticles as described in Project 1. In Project 2, we will characterize B cell responses elicited by vaccine regimens evaluated in Indian origin rhesus macaques in Core B by rapid, high-throughput monoclonal antibody (mAb) isolation to define the targeted epitopes and guide both the choice of boosting immunogens and, if needed, trimer redesign by eliminating or masking unwanted non-neutralizing immunodominant responses for subsequent immunization studies. We will interact with the investigators in Core C, who also will generate information about immunodominant Ab responses through their negative stain electron microscopy (nsEM)-based method for evaluation of vaccine-induced serum responses. Using the Env-specific mAbs, we will further determine how Env-specific antibody lineages evolve over time using Next Generation Sequencing (NGS) and IgDiscover, a computational tool optimized for use in rhesus macaques. We will generate individualized databases of macaque germline VDJ alleles for precise gene assignments, which is necessary for correct conclusions about Ab affinity maturation and SHM levels. We will further use the IgDiscover results and the new haplotype module to investigate if certain alleles, or combinations of alleles, predispose to elicitation of neutralizing Ab responses in either adult of juvenile macaques. We will investigate the level of expansion of neutralizing and non-neutralizing Ab lineages over time and ask if vaccine-induced neutralizing Ab lineages persist in long-lived immune compartments such as memory B cells and plasma cells, which are critical for the long-term protective effects of vaccines.
