Preclinical Evaluation of a Novel Lentiviral Vector Driving Lineage-Specific BCL11A Knockdown for Sickle Cell Gene Therapy

Abstract In this work we provide preclinical data to support initiation of a first-in-human trial for sickle cell disease using an approach which relies on reversal of the developmental fetal-to-adult hemoglobin switch. Erythroid specific knock down of BCL11A via a lentiviral encoded shRNAmiR leads to reactivation of the gamma-globin gene while simultaneously reducing expression of the pathogenic adult sickle β-globin. We generated a refined lentiviral vector BCH-BB694, that was developed to overcome poor vector titers observed in the manufacturing scale-up of the original research-grade LVV. Healthy or sickle cell donor CD34+ cells transduced with GMP grade BCH-BB694 LVV achieved high vector copy numbers (VCN) >5 and gene marking of >80% resulting in a 3-5 fold induction of HbF compared with mock transduced cells without affecting growth, differentiation and engraftment of gene modified cells in vitro or in vivo. In vitro immortalization assays, which are designed to measure vector-mediated genotoxicity, showed no increased immortalization compared with mock-transduced cells. Together these data demonstrate that BCH-BB694 LVV is (i) non-toxic and efficacious in preclinical studies, and (ii) can be generated at clinically relevant scale in a GMP setting at high titer to support clinical testing for the treatment of SCD.