Latently infected, quiescent CD4 T cells represent a central obstacle to eradication of HIV-1. Major hurdles are the low frequencies of latently infected cells in peripheral blood and the lack of known phenotypic markers that can distinguish them from uninfected ones. These impediments have prompted the development of in vitro cell models of latency. Such models allow manipulation of cellular and viral characteristics to gain a mechanistic understanding of how latency is established and regulated. However, no single experimental system of HIV latency is perceived to completely recapitulate the biology of the latent viral reservoir in vivo. This is mainly because (a) the mechanisms for establishment of latency by HIV are multiple;and (b) the types of cells harboring latent infection are also multiple. A wealth of knowledge has been gained regarding how specific stimuli (e.g. activation/differentiation, homeostatic proliferation, cytokines) and viral factors (e.g. integration site, Tat-driven expression) influence the dynamics of latent reservoirs in experimental systems. Project 2.2 will perform """"""""secondary level"""""""" characterization of novel small molecule compounds and combinations of compounds that have been identified to induce HIV transcription through primary screening by Dr. Hazuda's group in Project 2.1.
In Aims 1 and 2, we propose to use a panel of well-characterized primary cell systems and one Jurkat-based cell line clone to validate candidate drug compounds.
A third Aim will be the development of additional primary cell models to enable the study of HIV latency in other relevant cell types, not previously explored ex vivo (e.g. transitional memory T cells and macrophages). Treatments that are active in at least one of the primary culture systems, without showing overt toxicity or induction of cellular activation/proliferation, will be moved to subsequent phases of characterization and development within the Delaney Collaborative program. This will include detailed mechanistic studies (Objective 1), testing for activity in humanized mice and non-human primates (Objective 3) and in patient cells ex vivo (Objective 4). Our ultimate goal is to identify novel drugs and drug combinations that will have a high probability for success in activating HIV latent viruses when applied to treatment strategies in patients.
Within the Delaney Collaboratory program, the overall goal of Project 2.2 is the application of in vitro primary cell culture systems to identify new small molecule drugs and drug combinations that will activate latent HIV in biological systems, prior to studies conducted in animals and humans. A second aspect of these studies is to initiate identification of the mechanisms of action of such drugs to enable further studies that can focus on the identification of novel cellular targets and pathways, which will broaden of the scope of available therapeutic possibilities in the near future.
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