PROGRAM SUMMARY In preliminary data we show the results of two studies demonstrating that a sustained state of ART-free virologic remission, a ?functional cure?, is possible in non-human primates. First, the Desrosiers laboratory has shown that long-term expression of two broadly neutralizing antibodies (bNAbs) can suppress an untreated SHIV-AD8 infection in three rhesus macaques, in one case, to undetectable (<15 copies/ml) for over two years. Second, the Farzan laboratory has shown that long-term expression of the antibody-like entry inhibitor eCD4-Ig can also efficiently suppress viral replication in five of six SHIV-AD8-infected rhesus macaques for more than a year after cessation of anti-retroviral therapy (ART). We propose to build on these initial results by making these functional cures safer, more consistent, and more robust. In the process, we will establish a useful platform for evaluating latency-reversing agents (LRAs), a so-called ?kick?, by providing an environment in which a potent ?kill? is always present, and develop a way to halt transgene expression, thereby enabling time-to-rebound measurements. We will determine how best to limit anti-drug antibodies (ADA) that emerge with AAV-expressed bNAbs. We will determine how to best use ART or the long-lasting integrase inhibitor cabotegravir to establish AAV-mediated functional cures. Finally, we will determine whether a sustained and potent kill can by itself change the decay rate of latently infected cells. To do so, we have assembled a team with deep experience in HIV and SIV studies, years of experience working together, and a long-term commitment to understanding and improving AAV-based functional cures. These effort are accomplished with four projects and three cores. Project 1 will establish robust functional cures in SHIV-AD8- and SIVmac239-infected macaques using AAV-expressed eCD4-Ig and evaluate the impact of sustained eCD4-Ig on the viral reservoir. Project 2 will develop and test multiple approaches for eliminating anti-drug antibodies that frequently emerge with AAV-expressed bNAbs. Project 3 will assess the impact of triple therapy and long-acting cabotegravir on the establishment of eCD4-Ig- mediated functional cures, and determine how cabotegravir might best be combined with eCD4-Ig. Project 4 will develop a permanent off- and on-switches for AAV transgenes, facilitating time-of-rebound studies after sustained bNAb expression and increasing the safety of AAV-mediated functional cures. These projects are organized around a uniform experimental pipeline of assays supporting a series of non-human primate studies, established and implemented by Core B. Core A will manage regulatory and logistical aspects of the Program, and Core C will provide produce high-quality AAV particles for these studies, and improve its capacity for doing so. Collectively these efforts will develop and improve a viable approach to functional cures in humans, and provide tools and insight useful for complete eradication of the virus.