Cell and gene therapy in the murine model of adenosine deaminase deficiency

UnrestrictedI have investigated three different therapeutic approaches for the treatment of adenosine deaminase (ADA)-deficient severe combined immunodeficiency (SCID) using a murine model of ADA deficiency. The first two approaches were performed in neonates (d1-3) prior to enzyme replacement therapy (ERT) and investigated the use of hematopoietic stem cell transplantation (HSCT) and in vivo gene therapy (GT). The last approach were performed in young adults, after 8 weeks of ERT, and investigated the role of cytoreductive conditioning and ERT in ex vivo retroviral mediated gene therapy. ADA-deficient neonates who received whole marrow HSCT, without cytoreductive conditioning, had immune reconstitution, but had very low donor chimerism (5-10%), and chimerism was multi-lineage. With increasing doses of cytoreductive of total body irradiation, donor chimerism increased significantly (30-50%), however, chimerism remained multi-lineage. These results are in contrast to the lymphoid specific donor chimerism observed in another murine model of SCID (X-link) that has a T lymphoid intrinsic defect. When ADA-deficient neonates received a single intravenous dose (1x108 transducing units (TU)/ml) of a lentiviral vector expressing human ADA, they had long term survival and good immune reconstitution. Proviral marking was mostly in the liver and lung, and secondary transplants showed no evidence of hematopoietic stem cell (HSC) transduction, suggesting the mice were corrected by systemic detoxification in trans. In an effort to delimit the role of cytoreductive conditioning and concurrent ERT during ex vivo HSC gene therapy, mice were conditioned with either 200 or 900 cGy total body irradiation (TBI) and ERT was either continued or discontinued. Conditioning dose was very important for engraftment of transduced HSC, with 100-1000 fold more engraftment at 900 cGy compared to 200 cGy. ERT did not decrease the amount engraftment; as neither did 1 month of ERT post GT.; These studies have been useful in understanding the important aspects required for a good clinical outcome and lasting benefit, in current and future cell and gene therapy for ADA-SCID