RETROVIRAL TRANSFER OF A BACTERIAL ALKYLTRANSFERASE GENE (ADA) INTO HUMAN BONE-MARROW CELLS PROTECTS AGAINST O-6-BENZYLGUANINE PLUS 1,3-BIS(2-CHLOROETHYL)-1-NITROSOUREA CYTOTOXICITY

The antitumor activity of 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) is limited by the O-6-alkylguanine-DNA alkyltransferase (ATase) in tum or cells and by delayed myelosuppression, Inactivation of neoplastic A Tase by O-6-benzylguanine (BG) improves the therapeutic index for BCNU . We have demonstrated previously that BG + BCNU-induced myelosuppress ion in mice is reduced by expression of the BG-resistant ATase adn in murine bone marrow. We have now generated an amphotropic retrovirus co ntaining the ada gene and tested the effectiveness of ada expression i n preventing BG + BCNU cytotoxicity in human hematopoietic progenitor cells. A retroviral producer clone with a biological titer of 6.5 x 10 (4) colony-forming units/ml and 4.4 pmol ATase/mg protein was used for transduction of bone marrow. Cocultivation of these ada producer cell s with progenitor cells from six normal individuals resulted in 1.9-3. 9-fold protection against BG + BCNU-induced cytotoxicity in committed progenitor cell assays, Furthermore, this cytoprotective effect was as sociated with a high transduction efficiency (40%) and a 2-fold increa se of ATase activity in the surviving committed progenitor cell coloni es. These data provide a basis for testing the clinical effectiveness of retroviral ada gene transfer into hematopoietic cells to increase t he therapeutic index of BG + BCNU.