RETROVIRAL TRANSDUCTION OF A MUTANT METHYLGUANINE DNA METHYLTRANSFERASE GENE INTO HUMAN CD34 CELLS CONFERS RESISTANCE TO O-6-BENZYLGUANINE PLUS 1,3-BIS(2-CHLOROETHYL)-1-NITROSOUREA

Human CD34 cells express low levels of the DNA repair protein O-6-alky lguanine-DNA alkyltransferase (AGT) and are sensitive to 1,3-bis(2-chl oroethyl)-1-nitrosourea (BCNU). Gene transfer of the AGT gene, methylg uanine DNA methyltransferase (MGMT), results in only modest BCNU resis tance, Recently, an AGT inhibitor, O-6-benzylguanine (BG), entered cli nical trials, In preclinical studies, BG potentiated the cytotoxic eff ect of BCNU in tumors but increased toxicity to normal CD34 cells, We transferred a mutant MGMT containing a glycine-to-alanine mutation at position 156, resulting in marked resistance to BG, into Chinese hamst er cells; the K562 cell line and human CD34 cells used the retroviral backbone MFG. In each instance, cells expressed increased AGT and mere much more resistant to the combination of BG and BCNU than the parent al cells or cells transduced with wild-type MGMT. Furthermore, the tra nsduction efficiency in human CD34 cells was in excess of 70%, and the proportion of CD34 transduced cells resistant to the combination was >30%. Thus, retroviral-mediated transduction of a mutant MGMT into CD3 4 cells appears to be an effective may to induce selective resistance to a drug combination designed to overcome a significant resistance me chanism to nitrosoureas in tumors.