Metallothionein-mediated cisplatin resistance in human ovarian carcinoma cells

We have determined the ability of two human ovarian carcinoma cells to over-express metallothioneins (MTs) and the subsequent effect this elevation has on DDP cytotoxicity. Cells of 2008 and COLO 316 human ovarian carcinomas that were resistant to CdCl2 were obtained by stepwise selection and chronic culture in CdCl2 and ZnCl2. The 2008/MT cells were 3.2-fold resistant to CdCl2 and 4.1-fold resistant to DDP; they had 23-fold elevated MTs. The COLO/MT cells were 1.2-fold resistant to CdCl2 and 3.3-fold resistant to DDP, and they had 9-fold elevated MTs. Glutathione (GSH) was also elevated in the Cd-resistant sublines. However, four times more intracellular thiols were contributed by the MTs than by the GSH. 2008 and 2008/MT cells were examined in more detail to elucidate the mechanism of DDP resistance. Depletion of GSH with D,L-buthionine-S,R-sulfoximine (BSO) had no effect on the sensitivity of these cells to either CdCl2 or DDP. Uptake of [195m Pt]DDP in 2008 and 2008/MT cells was identical. Fractionation of the cytosol from [195mPt]DDP-exposed cells on Sephadex G-75 revealed that 17% of the total cellular Pt in 2008/MT cells was associated with the MT fraction, as against 4% in the parent 2008 cells. This increase corresponded to a concomitant loss of Pt from the particulate fraction. Fractionation of 2008 cells selected with DDP (2008/DDP cells) indicated that elevated MTs did not contribute to the DDP resistance of these cells. Only 2% of the total cellular Pt was in the MT fraction in 2008/DDP cells. These results showed that elevation of MTs may be one mechanism of DDP resistance in ovarian carcinoma; however, in vitro selection with DDP does not trigger this mechanism.