IN-VIVO ETOPOSIDE-RESISTANT C6 GLIOMA CELL-LINE - SIGNIFICANCE OF ALTERED DNA TOPOISOMERASE-II ACTIVITY IN MULTIDRUG-RESISTANCE

We have established an in vivo etoposide-resistant glioma cell line (C 6/VP) from C6 rat glioma cells by stepwise exposure to increasing dose s of etoposide. The C6/VP cells were 10 times more resistant to etopos ide than the parental C6 cells. In addition C6/VP cells demonstrated c ross-resistance to vincristine and vinblastine, but not to ADM or ni-A MSA. Interestingly, the cells had collateral sensitivity to ACNU, cisD DP and Ara-C. The C6/VP cells did not express the MDR gene or p-glycop rotein, while they showed 16 times less topoisomerase II catalytic act ivity compared to the C6 cells. Although there was no significant diff erence between C6 and C6/VP cells in amounts of topoisomerase LI hi nu clear extracts, the C6/VP cells had 2.9 times higher amounts of the en zyme than C6 cells in nuclear scaffold prepared from a relatively low- salt buffer (0.5 M NaCl). Northern blot analysis demonstrated that mRN As of topoisomerase II alpha isoforms were expressed both in C6 and C6 /VP cells, and that the amounts of topoisomerase II alpha in C6/VP cel ls were 14 times greater than in C6 cells. The total uptake of etoposi de in tumor tissues derived from C6/VP cells was 3 times less than tho se derived from parental C6 cells. These results indicate that the C6/ VP acquired a multi-drug resistance phenotype by a reduction of the ca talytic activity of topoisomerase II and/or diminished accumulation of drugs. This phenotype did not involve the p-glycoprotein. Alterations of topoisomerase II in the C6/VP cells also were accompanied by an in creased amount of the topoisomerase II alpha isoform, most of which wa s localized in the nuclear scaffold (matrix). This suggests that alter ed binding of topoisomerase II to topologically organized DNAs in the nuclear scaffold may be the molecular basis of this multi-drug resista nce phenotype.