In vivo hematopoietic stem cell engineering for HIV cure

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) with CCR5Δ32 hematopoietic stem cells (HSCs) can replace the hematopoietic system with HIV-resistant blood cells and has been the only curative therapy for HIV to date.1-3 While an important proof-of-concept, allo-HSCT for HIV cure is hindered by the scarcity of CCR5Δ32 donors and a toxicity profile that restricts its application to a subset of people living with HIV (PLWH) with hematological malignancies. Gene therapy presents a highly promising approach that with significantly enhanced safety and accessibility by modifying a patient’s own (autologous) HSCs to confer HIV-resistance. HSC gene therapy approaches continue to evolve to improve CCR5-editing efficiency, eliminate the need for laborious and costly ex vivo gene editing procedures, and avoid toxic chemotherapy conditioning regimens. Our proposal addresses these challenges by direct modification of HSCs in vivo to inactivate CCR5 for HIV- resistance and enrichment of edited cells using novel CD45 epitope editing methods to protect them from CD45- specific immunotherapy. We anticipate our in vivo gene editing strategy will surpass previous approaches in editing efficiency, safety, and accessibility, making it suitable for widespread application including among healthy PLWH and low- and middle-income (LMIC) countries where HIV is most prevalent. We will harness our in vivo gene therapy to enhance HIV protection and elimination by sustained delivery of promising anti-HIV effectors, including the synergistic combination of eCD4-Ig and 10-1074, as well as the Brec1 recombinase that will excise proviruses. In our well-established NHP model of suppressed HIV infection, we have a breadth of data demonstrating the feasibility of our approach including efficient CCR5 gene editing in HSCs, robust depletion of CD45+ cells using anti-CD45 antibodies, HSC modification to express eCD4-Ig and bNAbs, and in vivo delivery of the Brec1 recombinase to excise proviruses. Our in vivo gene therapy approach will use our latest generation helper dependent adenoviral (HDAd) vector, which we have demonstrated to safely and efficiently modify HSCs in vivo. Overall, our goal is to apply next-generation in vivo CCR5 editing and selection in combination with anti-HIV effector delivery to achieve a safe and accessible HIV cure.