The long term goal is to elicit broadly neutralizing antibodies, but to forward this objective by defining the Env-specific, vaccine-elicited B cell responses in primates primarily directed at the CD4 binding site. Using this information, through iterative design, we will improve responses directed at this and other conserved neutralizing determinants. Project 1, entitled Immunogen Design and Selection, will generate the HIV envelope glycoprotein-based candidate vaccine immunogens to be analyzed in the HIVRAD. The immunogens will be derived by structure-based design, as well as by selection, and responses to them will be analyzed in the other program components. Project 2, B cell Analysis and Ig Genetics, will examine the diversity of the memory B cell repertoire against selected regions of Env elicited by soluble trimers at both short and longer intervals, by newly designed trimeric imijnunogens, as well as long-lived plasma cells in the bone marrow to examine the impact of regimen on V gene usage and somatic hypermutation. Project 3, Memory B Cell Isolation and Antibody Characterization, will isolate B cells using Env-specific FACS and will characterize the isolated Env-specific antibodies for their epitope specificity to aid immunogen redesign and will also analyze the Env-specific memory IgM compartment following Env-trimer inoculation to look for defects in transition to the IgG compartment. Project 4, Bioinformatics and Structure, will use bioinformatics analysis of deep sequencing of B cell transcripts, obtained from the NHP immunogenicity experiments and to characterize antigen-specific antibody populations that arise in response to immunization. The second focus will be to use x-ray crystallography to determine atomic-level structures of high-interest antibodies in complex with gp120. These efforts will be supported by the Administrative Core A, which will institute a management plan to ensure clear communications within the HIVRAD and the NHP Core B which will house the animals, perform immunogenicity experiments with immunogens from Project 1 and provide samples for the analysis of B cell responses for Projects 2, 3 and 4. RELEVANCE: The overall goal of this HIVRAD, and this Overview, is to elicit broadly neutralizing antibodies to the human pathogen HIV-1. The successful elicitation of such antibodies would be a large step forward toward the goal of generating a broadly effective HIV-1 vaccine and would have a substantial impact on improving human public health.