RETROVIRAL TRANSFER OF THE GLUCOCEREBROSIDASE GENE INTO CD34(-DISEASE- IN-VIVO DETECTION OF TRANSDUCED CELLS WITHOUT MYELOABLATION() CELLSFROM PATIENTS WITH GAUCHER)

Retroviral gene transfer of the glucocerebrosidase gene to hematopoiet ic progenitor and stem cells has shown promising results in animal mod els and corrected the enzyme deficiency in cells from Gaucher patients in vitro, Therefore, a clinical protocol was initiated to explore the safety and feasibility of retroviral transduction of peripheral blood (PB) or bone marrow (BM) CD34(+) cells with the G1Gc vector, This vec tor uses the viral LTR promoter to express the human glucocerebrosidas e cDNA, Three adult patients have been entered with follow-up of 6-15 months. Target cells were G-CSF-mobilized and CD34-enriched PB cells o r CD34-enriched steady state BM cells, and were transduced ex vivo for 72 hr, Patient 1 had PB cells transduced in the presence of autologou s stromal marrow cells. Patient 2 had PB cells transduced in the prese nce of autologous stroma, IL-3, IL-6, and SCF, Patient 3 had BM cells transduced in the presence of autologous stroma, IL-3, IL-6, and SCF, At the end of transduction, the cells were collected and infused immed iately without any preparative treatment of the patients. The transduc tion efficiency of the CD34(+) cells at the end of transduction was ap proximately 1, 10, and I for patients 1, 2, and 3, respectively, as es timated by semiquantitative PCR on bulk samples and PCR analysis of in dividual hematopoietic colonies, Gene marking in vivo was demonstrated in patients 2 and 3. Patient 2 had vector-positive PB granulocytes an d mononuclear bone marrow cells at I month postinfusion and positive P B mononuclear cells at 2 and 3 months postinfusion, Patient 3 had a po sitive BM sample at 1 month postinfusion but was negative thereafter. These results indicate that gene-marked cells can engraft and persist for at least 3 months postinfusion, even without myeloablation, Howeve r, the level of corrected cells (<0.02%) is too low to result in any c linical benefit, and glucocerebrosidase enzyme activity did not increa se in any patient following infusion of transduced cells, Modification s of vector systems and transduction conditions, along with partial my eloablation to allow higher levels of engraftment, may be necessary to achieve beneficial levels of correction in patients with Gaucher dise ase.