Modified Project Summary/Abstract Section The development of a prophylactic vaccine for HIV would benefit from the generation of immunogens and vaccine delivery modalities that can induce a combination of highly functional CD8+ T cell responses towards mutation constrained epitopes and broadly neutralizing antibody responses to the HIV envelope trimer. Thus, in this proposal, we plan to leverage several components towards this goal: an approach known as structure-based network analysis that was used to identify a set of protective epitopes preferentially targeted by CD8+ T cell responses in spontaneous HIV controllers, 2) an envelope trimer that is capable of eliciting heterologous neutralizing antibody responses and 3) and a heterologous vaccine delivery platform of a replication-incompetent chimpanzee adenoviral (ChAd) vector and lipid-nanoparticle encapsulated messenger RNA (LNP-mRNA) that can elicit both robust CD8+ T cell and humoral immunity. Having applied structure-based network analysis to SIV protein structures, we have already identified a set of mutation-constrained (‘highly networked’) SIV epitopes restricted by an array of Mamu class I alleles. In addition, we have also produced compelling data demonstrating the ability of ChAd vector and LNP-mRNA to induce CD8+ T cell responses to highly networked SIV epitopes and strong trimer-specific antibody responses in rhesus macaques. What remains to be determined is whether functional CD8+ T cell responses towards highly networked SIV epitopes have a protective benefit against heterologous challenge, either individually or in tandem with antibodies. Thus, in Aim 1 of this proposal, we will assess the protective efficacy of heterologous ChAd and LNP-mRNA vaccines encoding highly networked SIV epitopes in a group of 20 rhesus macaques. These immunized macaques will be divided equally between those receiving mock immunization or highly networked SIV epitope immunization (10 per group). Following immunization, vaccine-induced CD8+ T cell responses directed against highly networked SIV epitopes will be assessed, and vaccinated animals will then be subjected to repeated low-dose SHIV.BG505 challenge. These SHIV-challenged macaques will have viral load, CD4 count, and CD8+ T cell phenotype assessments every 2 weeks during the challenge period and for 20 weeks following the challenge period as well. Go/No-Go criteria regarding T cell immunogenicity and the effects of vaccination on SHIV viral loads will be established and evaluated at the end of year 2. In Aim 2, we will assess the immunogenicity and efficacy of a combined ChAd highly networked SIV epitope immunogen and LNP-mRNA HIV Env trimer immunogen. These immunized macaques received either mock immunization, HIV Env trimer + mock immunization or both HIV Env Trimer and highly networked SIV epitope immunogens (10 per group). These studies will help determine whether vaccine-induced highly networked CD8+ T cell responses can provide a protective...