Effects of the combination of camptothecin and doxorubicin or etoposide on rat glioma cells and camptothecin-resistant variants.

From the rat C6 glioma cell line in culture, we selected camptothecin-resistant variants by growth in the presence of increasing amounts of this drug (C6(CPT10), C6(CPT50)and C6(CPT100), growing respectively with 10, 50 and 100 ng ml(-1)camptothecin). The degree of resistance to camptothecin ranged between 15-fold (C6(CPT10)) and 30-fold (C6(CPT50)and C6(CPT100)). The C6(CPT10)cell line presented a collateral sensitivity to etoposide (3.6-fold), while the C6(CPT50)and C6(CPT100)cell lines were cross-resistant to etoposide (1.8-fold) The resistant lines were characterised by a two-fold reduced content and catalytic activity of topoisomerase I, and C6(CPT50)and C6(CPT100)presented a significant increase in topoisomerase IIalpha content and catalytic activity and a marked overexpression of P-glycoprotein. We explored the cytotoxicity of combinations of a topoisomerase I inhibitor (camptothecin) and a topoisomerase II inhibitor (doxorubicin or etoposide) at several molar ratios, allowing the evaluation of their synergistic or antagonistic effects on cell survival using the median effect principle. The simultaneous combination of camptothecin and doxorubicin or etoposide was additive or antagonistic in C6 cells, slightly synergistic in the C6(CPT10)line and never more than additive in the C6(CPT50)and C6(CPT100)cell lines. The sequential combination of doxorubicin and camptothecin gave additivity in the order camptothecin --> doxorubicin and antagonism in the order doxorubicin --> camptothecin. Clinical protocols combining a topoisomerase I and a topoisomerase II inhibitor should be considered with caution because antagonistic effects have been observed with combinations of camptothecin and doxorubicin.

Combination of irinotecan (CPT11) and 5-fluorouracil with an analysis of cellular determinants of drug activity.

We evaluated the combination SN38 (7-ethyl-10-hydroxycamptothecin) -5fluorouracil (5FU) +/- folinic acid (FA) on six human colon cancer cell lines expressing spontaneous sensitivity to both drugs. Tumoral parameters potentially related to drug sensitivity were investigated: topoisomerase I (topo I) cleavable complexes formed with SN38, thymidylate synthase (TS) activity, folylpolyglutamate synthetase activity and dihydropyrimidine dehydrogenase activity. Drugs (SN38 and/or 5FU +/- FA) were applied for 72 hr, either sequentially or together. The concentration ratio between SN38 and 5FU was 100. Cytotoxicity (MTT [3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide] test), DNA flow cytometry and isobologram analysis (Chou and Talatay) were performed. Based on 5FU IC50 values and isobologram analyses, the most cytotoxic schedule was SN38 followed by 5FU - FA, with high synergistic effects. Flow cytometry indicated that SN38 induced a more or less marked S-G2 block in all cell lines. Sensitivity to SN38, 5FU +/- FA, or combinations were not linked to the potential above-cited tumoral parameters. Interestingly, an inverse correlation was demonstrated between TS activity and topo I cleavable complexes (r2 = 0.78, P = 0.019). These data emphasize the critical importance of the irinotecan-5FU schedule and strongly support this association for the treatment of potentially 5FU-sensitive tumors.