In vitro and in vivo base editing of CCR5 in hematopoietic stem cells confers HIV-1 resistance.

Here, we present an HIV gene therapy strategy by in vivo precision gene editing in hematopoietic stem cells (HSCs). We successfully generated a panel of helper-dependent adenoviral vectors expressing all-in-one base editors (HDAd-BEs) targeting the CCR5 gene. In an HIV-permissive cell line, transduction with the HDAd-BE vector led to near-complete target site editing, CCR5 knockout, and inhibition of HIV infection. In HSC-enriched human CD34+ cells from mobilized donors or cord blood, we measured efficient precision base editing after HDAd-BE transduction and selection. Following in vitro T cell differentiation and HIV infection, we demonstrated that HIV genome titers were significantly lower with precision CCR5 base editing. Importantly, in a humanized mouse model, in vivo transduction with the HDAd-BE vector followed by selection resulted in ∼50% base editing at the CCR5 target site in bone marrow mononuclear cells, which conferred ∼12-fold lower HIV plasma titers than control animals after HIV challenge. No significant off-target editing and adverse effects associated with the treatment were observed. By targeting HSCs to precisely engineer HIV-resistant cells in vivo, our strategy represents an efficient, durable, and affordable approach to achieve a functional cure for HIV infection.