Human T cell generation is restored in CD3δ severe combined immunodeficiency through adenine base editing

CD3δ SCID is a devastating inborn error of immunity caused by mutations in CD3D, encoding the invariant CD3δ chain of the CD3/TCR complex necessary for normal thymopoiesis. We demonstrate an adenine base editing (ABE) strategy to restore CD3δ in autologous hematopoietic stem and progenitor cells (HSPCs). Delivery of mRNA encoding a laboratory-evolved ABE and guide RNA into a CD3δ SCID patient’s HSPCs resulted in a 71.2% ± 7.85% (n = 3) correction of the pathogenic mutation. Edited HSPCs differentiated in artificial thymic organoids produced mature T cells exhibiting diverse TCR repertoires and TCR-dependent functions. Edited human HSPCs transplanted into immunodeficient mice showed 88% reversion of the CD3D defect in human CD34+ cells isolated from mouse bone marrow after 16 weeks, indicating correction of long-term repopulating HSCs. These findings demonstrate the preclinical efficacy of ABE in HSPCs for the treatment of CD3δ SCID, providing a foundation for the development of a one-time treatment for CD3δ SCID patients. [Display omitted] •BE targets CD3δ SCID-causing mutations•BE of CD3δ SCID stem cells restores T lymphopoiesis and TCR diversity•Development of CD3δ SCID stem cells is blocked at the double positive precursor stage•BE is superior to homology-directed repair in correction of CD3δ SCID The genetic mutation underlying a severe immune system deficiency can be corrected by adenine base editing of human patient stem cells, restoring the ability of these cells to develop into functional T cells in model systems, and paving the path for future treatment of CD3δ SCID in patients.