Development of CRISPR_SCD001, an autologous hematopoietic stem cell gene therapy for sickle cell disease

We developed CRISPR_SCD001, an autologous cellular therapy in which the pathogenic HBB sickle allele is corrected by CRISPR-Cas9-mediated homology-directed repair, using a single-stranded oligodeoxyribonucleotide as DNA donor template. The drug product (DP) from plerixafor-mobilized peripheral blood CD34⁺ cells from sickle cell disease patients and healthy donors were used for (1) pharmacodynamic assessments to interrogate therapeutic potential, (2) evaluation of genotoxicity, and (3) in vivo characterization by a non-GLP toxicology study in NBSGW mice. Large-scale DP lots yielded 89% ± 5% CD34⁺ purity and had a viability of 84% ± 6%. Mean gene correction at HBB was 22% ± 4% of alleles, with 51% ± 6% of allelic disruption. Edited erythroid cultures produced 39% ± 5% adult hemoglobin and 40% ± 6% fetal hemoglobin, reducing sickle hemoglobin from 89% to 22% ± 6%. In NOD,B6.SCID Il2rg ^-/- Kit^W41/W41 (NBSGW) mice, CRISPR_SCD001 engrafted robustly (59% ± 25%), maintained editing frequencies comparable to injected cells (∼20% gene correction), and showed polyclonal distribution of the edited cells; no noteworthy treatment-related toxicity or pathologic evidence was found by a blinded histopathology assessment. Our protocol generates a clinical-grade, cryopreserved CD34⁺ cell product that corrects the sickle mutation, restores anti-sickling hemoglobins, and meets pre-clinical safety criteria, establishing a reproducible manufacturing process suitable for clinical translation. These data support initiation of a first-in-human phase 1 trial of CRISPR_SCD001 for sickle cell disease patients presenting with severe pathology.