Gene Therapy for Lad-I Immunodeficiency: Preclinical Evaluation of HSC Transduction Under Optimized GMP-Conditions

The efficacy of lentiviral gene therapy for the treatment of primary immunodeficiencies (PIDs) has been extensively demonstrated in diseases such as X1-SCID, ADA-SCID, X-CGD and WAS. Leukocyte Adhesion Deficiency type I (LAD-I) has also emerged as a PID that is potentially correctable by gene therapy. To this aim during the recent years we have completed all the pre-clinical work required for the initiation of a gene therapy trial for patients with severe LAD-I. Mutations in the ITGB2 gene (encoding for the β2 subunit of integrins, also known as CD18) impair leukocyte extravasation to inflamed areas. LAD-I patients thus suffer from recurrent and life-threatening bacterial and fungal infections. We have previously demonstrated that the ectopic expression of CD18 driven by a myeloid chimeric promoter in LAD-I leukocytes restores their ability to migrate to inflamed sites. Because the efficacy of hematopoietic gene therapy relies on the stable engraftment of gene-corrected HSCs in treated patients, we have implemented the transduction conditions of human HSCs with this therapeutic lentiviral vector in healthy donor CD34+ cells, either from cord blood or mobilized peripheral blood sources. CD34+ cells were transduced at relatively low multiplicities of infection (MOIs) of 50 i.u./cell in the presence of transduction enhancers. Using an optimized transduction protocol, high transduction efficacies were obtained with minimal loss in hematopoietic progenitors. On average, transduction efficiencies of 70% and mean vector copy numbers of 2-5 copies/cell were identified in hematopoietic progenitor cells. Similar results were obtained when large-scale GMP conditions were used. To assess the transduction efficacy of human HSCs, transduced CD34+ cells were transplanted into immunodeficient NSG mice. In these studies, stable repopulation levels of human cells harboring 1-3 copies of the therapeutic vector per cell were observed up to three months post transplantation. In some cases, secondary transplants were also performed, confirming long-term and stable engraftment of transduced cells. These results have allowed us to obtain the approval for the initiation of the first lentiviral-mediated gene therapy clinical trial for the treatment of LAD-I patients. Almarza:Rocket Pharmaceuticals: Equity Ownership, Patents & Royalties, Research Funding. Rio:Rocket Pharmaceuticals: Equity Ownership, Patents & Royalties, Research Funding. Law:Rocket Pharmaceuticals: Employment, Equi