Characterization of a chlorambucil-resistant human ovarian carcinoma cell line overexpressing glutathione S-transferase mu.

Ovarian carcinoma cells 10-fold resistant to the alkylating agent chlorambu cil (CBL) were isolated after repeated exposure of the parent cells to grad ually escalating concentrations of the drug. The resistant variant, A2780(1 00), was highly cross-resistant (9-fold) to melphalan and showed lower-leve l resistance to other cross-linking agents. The resistant A2780(100) cells had almost 5-fold higher glutathione S-transferase (GST) activity than the parental A2780 cells with 1-chloro-2,4-dinitrobenzene (CDNB) as substrate. The pi-class GST(s) was the major isoform(s) in both cell lines. However, t he resistant A2780(100) cells had at least 11-fold higher GST mu as compare d with the parental cells, in which this isoform was barely detectable. A s ignificant induction of GST mu was observed in A2780 cells, but not in the resistant cells, 18 hr after a single exposure to 100 mu M CBL. The inducti on of GST mu by CBL was both time and concentration-dependent. Assays of th e conjugation of CBL with GSH showed that the human CL-class GST had 3.6- a nd 5.2-fold higher catalytic efficiency relative to the pi- and alpha-class GSTs, respectively. This difference was reflected in the relatively higher (about 6-fold) efficiency of CBL conjugation in A2780(100) cells as compar ed with the parental cells. These results have demonstrated for the first t ime a near-linear correlation between CBL resistance and overexpression of mu-class GSTs and suggest that this overexpression maybe responsible, at le ast in part, for the acquired resistance of ovarian carcinoma cells to CBL, and possibly the other bifunctional alkylating agents. Consistent with thi s hypothesis, we found evidence for decreased formation of DNA lesions in A 2780(100) compared with the drug sensitive A2780 cells after exposure to CB L. (C) 1999 Elsevier Science Inc.