Base Editing Hematopoietic Stem/Progenitor Cell Gene Therapy for Treatment of X-Linked Severe Combined Immunodeficiency

Abstract

X-linked Severe Combined Immunodeficiency (XSCID) due to IL2RG loss-of-function mutations typically cause a severe combined cellular and humoral immunodeficiency that is diagnosed and transplanted in infancy. Hypomorphic IL2RG variants with milder phenotypes are generally diagnosed later in life. Early stem cell transplants (allogeneic or autologous gene therapy) performed without conditioning often results in partial immune reconstitution with limited donor engraftment in T cell lineage alone. Defective host B, natural killer (NK), and other cell lineages remain and can cause progressive multi-organ disease. Ex vivo gene therapy using lentivector-transduced autologous hematopoietic stem/progenitor cells (HSPCs) has provided clinical benefit for many previously transplanted older XSCID patients (NCT01306019) and transplant-naïve infants (NCT01512888, NCT03311503). Lentivector transduction functionally corrects XSCID stem cells by semi-random insertion of IL2RG cDNA under the regulation of an engineered promoter. A conversion to gene-corrected immune cells in their respective compartment can take years, particularly in older adult patients. The exact mechanism for this slow progression, whether this is attributable to a relatively weak promoter in the transgenes, is unclear. CRISPR/Cas9 base editing provides the potential for precise gene mutation correction that is devoid of random virus integrations and, critically, restores physiological gene regulation by endogenous promoter. We developed a highly efficient base editing strategy using adenine base editor and respective guide RNAs to repair respective IL2RG mutations (IL2RG p.289X, IL2RG p.Gln235X, IL2RG p.Q144X, and IL2RG p.R226H). Preclinical efficacy and safety data supported a Phase I/II clinical trial to treat XSCID patients with base-edited HSPCs (IND 31037; NCT 06851767). Three patients have been treated to date with base-edited (BE) HSPCs (25 million to 50 million cells per kg/weight). BE HSPC products were well tolerated, with early myeloid recovery within 3 weeks after cell infusion. Gene correction frequencies in the study cell products averaged >80%. Follow-up data at 6 months in the first patient showed robust levels of gene correction in myeloid (70%), B (94%), NK (100%), and, surprisingly, in his T cell compartment (65%) as well, with a corresponding decline in donor T cell chimerism (99% at baseline, 47% at 6 months). Monitoring of the other treated patients is ongoing. We conclude from our preliminary data that gene therapy using BE HSPCs appear very promising for achieving an earlier and more robust multi-lineage immune reconstitution in adult patients using BE HSPCs.