In vivo HSC gene therapy using a multi-modular HDAd vector for HIV cure

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Hematopoietic stem cell (HSC) transplantation can provide durable HIV elimination as exemplified in the “Berlin” patient, the “London” patient, and recently, in a third (“New York”) patient. This gives a strong rationale for HSC gene therapy of HIV/AIDS. Current clinical HSC gene therapy protocols (e.g. for hemoglobinopathies) involve high-dose chemotherapy to make space in the bone marrow, and the transplantation of HSCs after ex vivo gene transfer. Because of the risk, cost, and technical complexity, it is unlikely that ex vivo protocols will be widely applicable, specifically in developing countries where the greatest demand for HIV/AIDS therapy lies. We have developed an in vivo HSC transduction approach that requires only intravenous injections and could be provided as an outpatient treatment. In this approach, HSCs are mobilized from the bone marrow into the peripheral blood stream and transduced with intravenously injected in vivo gene transfer vectors (helper-dependent adenovirus vectors) that target receptors present on primitive HSCs. HSCs transduced in the periphery return to the bone marrow, persist there long-term, and contribute to all blood cell lineages. The central goal of this application is to further develop our in vivo approach toward HIV prophylaxis and therapy with persistent eradication of HIV in target/reservoir cells. The Specific aims are. 1. Optimize HSC mobilization regimens, HSC homing, and HDAd vectors/expression systems to achieve i) efficient bone marrow homing of mobilized HSCs, ii) efficient trafficking of transduced HSC progeny cells, specifically to the brain, a main HIV reservoir tissue that is difficult to target by therapeutics, and iii) increase the level and safety of transgene expression. 2. Prevent HIV/SIV escape mutants and eliminate virus from reservoirs by a multi-modular in vivo HSC gene therapy approach. Modules will exert anti-HIV activity based on different mechanisms (e.g. opsonization of virus in blood by eCD4-Ig, protection of target cells by co-receptor knockout through in vivo genome editing, and killing of infected cells independently of MHC-I presentation by a CD4 chimeric antigen receptor (CD4-CAR) expressed on immune effector cells. 3. Demonstrate in NHPs that the optimized in vivo HSC gene therapy approach will allow for i) complete protection against SIV challenges (absence of escape mutants) and ii) SIV elimination in infected animals (including the brain). Model systems to test the safety and antiviral efficacy of the approaches will include primary HSCs/HSC- derived cells, transgenic and humanized mouse models (with and without SIV infection), as well as NHPs (in prophylaxis and therapy setting). Our efforts will address important biological obstacles in HIV therapy in the context of a technically simple, cost-efficient, and portable approach.