Project Summary Anti-retroviral therapy (ART) has dramatically altered the HIV pandemic landscape by rendering the disease manageable - but still incurable. Significant barriers are associated with the global implementation of ART that limit its utility for sustainable prevention of HIV. Moreover, the intersection of aging-related conditions and the consequences of long-term ART will have a substantial impact on the healthcare system as well as on HIV patients. These challenges underscore the need to pursue strategies that will provide both sustainable HIV prevention and a functional HIV cure. The results of the RV144 trial indicate that vaccination may prevent HIV transmission in humans and that durability of anti-envelope (Env) HIV antibodies may be the key to this protection. Efforts to improve upon the RV144 trial have demonstrated that booster immunizations increase serum anti-Env antibody titers only transiently. This “anti-Env antibody persistence problem” impedes our efforts to develop an effective HIV vaccine. This proposal is designed to improve our functional understanding of the immunological mechanism involved in producing durable HIV Env antibodies in both the systemic circulation and the rectal and vaginal mucosa. To accomplish this, we will be using a DNA-prime/Protein-boost vaccine regimen in rhesus macaques. Aim 1 of this research project is focused on establishing how Th1 versus Th1/Th17 priming impacts innate inflammatory response, Env TFH frequencies, phenotype, transcriptional profile, and B cell helper capacity, as well as systemic and mucosal anti-Env antibody titers. Aim 2 will determine whether combining the state-of-the-art soluble Env mimic, an Env SOSIP trimer protein immunogen with a highly potent and robust adjuvant (to induce Th1/Th17 response, results in increased production of a high-quality, long- lasting anti-Env antibodies relative to Th1 vaccine regimen. We will further investigate whether vaccine-mediated induction of the polyfunctional TFH response improves protection when faced with a mucosal simian HIV challenge. These preclinical studies will establish the mechanistic and experimental foundations to identify how TFH helper cell profiles impact anti-Env antibody longevity and vaccine efficacy and will lay the necessary, evidence-based foundation for the design of a successful HIV vaccine.