Project 1: Dissecting Persistent Virus Reservoirs in Tissues

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PROJECT SUMMARY/ABSTRACT Without a complete knowledge of the formation of the HIV reservoir, its persistence, and potential rebound, it will be difficult to develop therapies to achieve a cure for HIV. A major limitation that the HIV field needs to overcome in order to advance our understanding of viral rebound is the inability to systemically study the reservoir in tissues. With the support of a P01 focused on viral rebound of the early reservoir, we have recently optimized a new workflow that we named PET-CT guided necropsy. This workflow is based on the use of a 64Cu-labeled anti-SIV env FAB2 (64Cu-7D3FAB2) to guide the sampling and/or necropsy process to identify and collect pieces of tissues containing foci of 64Cu signal revealing sites of SIV replication. The presence of virus infected cells in these “hot” tissues is now validated by 4 markers (Gag and Env expression and CD4 and MHC-I downregulation). Unexpectedly, we only detect SIVmac239 infection of myeloid cells during the first 7-10 days after ATI. No SIV infected T cells are observed although CD4 T cells are abundant in these foci. Finally, our 64Cu-7D3FAB2 probe led us to tissues where we can reproducibly visualize virion producing cells, with a myeloid cell morphology, by transmission electron microscopy. This multiscaling imaging, leveraging PET/CT guided necropsy, allows us to peak into the elusive tissue reservoir. In this Project 1 of the Program, we propose to leverage our advancements with PET-CT-guided sampling of tissues as well as additional innovative sampling techniques in non-human primate (NHP) models to dissect the dynamics of the persistent SIV reservoir. Specifically, we have planned 3 different type of NHP studies that, with the help of ad-hoc interventions, will give us an unprecedented resolution into the dynamics of the reservoir and its rebound. We have planned interventions that aim at “tickling” and disturbing the reservoir during cART, which will allow PET-CT-signal increase and collection of tissue samples that will be analyzed with different techniques including multiscale imaging and distributed to the other 2 projects. We will “mulch” the reservoir by favoring its seeding into mucosal tissues with DSS treatment before infection and then again an in-depth investigation of the tissue reservoir at the time of cART initiation and after several months of suppressive cART. Finally, we will deplete or stimulate myeloid cells, which our studies have revealed as main contributor to viral rebound after ATI and study the impact of these interventions on the reservoir dynamics and timing of rebound viremia. In summary, our project will use different strategies in NHP studies to clarify the nature and source of the SIV reservoir with particular focus at the mucosal tissue and myeloid cells.