Increased DT-diaphorase expression and cross-resistance to mitomycin C in a series of cisplatin-resistant human ovarian cancer cell lines.

In a series of ovarian carcinoma cell lines selected in vitro for resistance to cisplatin by continuous exposure to increasing drug concentrations, the level of resistance is proportional to the expression of gamma-glutamylcysteine synthetase (gamma-GCS). To determine if other detoxicating genes are coordinately expressed, we measured the activity of DT-diaphorase and cytochrome P450 reductase. The specific activity of DT-diaphorase, but not that of cytochrome P450 reductase, increased with increasing resistance to cisplatin. Steady-state mRNA levels for DT-diaphorase correlated with enzyme activity and hence with cisplatin resistance. Since the activity of DT-diaphorase has been associated with sensitivity to quinones, we studied the cytotoxicity of mitomycin C under oxic conditions. Unexpectedly, resistance to mitomycin C increased proportionally with that to cisplatin (r = 0.997). Pretreatment with buthionine sulfoximine, which inhibits glutathione (GSH) synthesis, failed to sensitize either the sensitive or the resistant lines to mitomycin C. Thus, the basis for collateral resistance to mitomycin C in the cisplatin-resistant lines under oxic conditions is unrelated to overproduction of GSH. Under hypoxia, the toxicity of mitomycin C to the most sensitive (A2780) cell line was unchanged. However, the most resistant (C200) line was 2-fold more resistant to mitomycin C under hypoxic conditions. The coordinate overexpression of DT-diaphorase and gamma-GCS in the resistant cell lines is thus associated with hypoxic cell resistance, and supports the involvement of shared mechanisms of gene regulation in the observed resistant phenotype.