Abstract In the era of combined antiretroviral therapy (cART), mortality and morbidity associated with HIV-1 infection has been reduced. Nevertheless, a wide range of AIDS-related conditions as well as serious non-AIDS events (SNAEs) continue to afflict people living with HIV (PLWH). Improved long-term strategies are needed, ranging from easier administration to better suppression to functional cure. Barriers to functional cure include a limited ability to deliver activators and other bioactive molecules into tissue reservoirs of HIV, particularly the possible reservoirs in the central nervous system (CNS). Representing a novel mode of delivery, extracellular vesicles (EVs) are double membrane-bound particles, released by all known cell types, that engage in intercellular communication by shuttling components of the parent cell (such as proteins and RNAs) to target cells. EVs contribute to disease pathogenesis and are actively investigated, especially in cancers, as biomarkers, actors in disease processes, and potential therapeutics. Importantly, EVs have been shown to cross biological barriers, even the blood-brain barrier, and can be easily delivered to the brain. EVs thus provide an exceptional opportunity to deliver components of HIV control or reactivation/cure to tissue reservoirs, with potential for cell-specificity. To this end, we have assembled a multidisciplinary team with two major, unique assets. The first is a novel small EV-transcriptional activator (sEVTA) tool that has already passed in vitro and in vivo tests. In this system, retroviral transactivator proteins are specifically packaged into EVs, which can be further functionalized with tracers and surface peptides for cell targeting. The second is our well established SIV/macaque model, which has been used successfully to study retroviral latency, rebound, and retrovirus-associated CNS disease. We will conduct careful nonhuman primate dosing and distribution studies followed by optimized intravenous and intranasal delivery of sEVTAs. Effects of sEVTAs on viral rebound will be assessed with innovative tools in circulating and peripheral tissue reservoirs (Aim 1), followed by reservoirs in the CNS (Aim 2). In Aim 3, we will investigate the potential toxicity of these approaches, with particular focus on the central nervous system, and explore more cell-targeted approaches. The goal of these studies is to use sEVTAs to reactivate latent retroviruses in the CNS and the periphery. However, the project will also provide much-needed information about EV delivery, distribution, and efficacy in primates that can be harnessed in development of a variety of therapies for HIV infection and disease.