Mechanisms of synergism between cisplatin and gemcitabine in ovarian and non-small-cell lung cancer cell lines

2',2'-Difluorodeoxycytidine (gemcitabine, dFdC) and cis-diammine-dichloropl atinum (cisplatin, CDDP) are active agents against ovarian cancer and non-s mall-cell lung cancer (NSCLC). CDDP acts by formation of platinum (Pt)-DNA adducts; dFdC by dFdCTP incorporation into DNA, subsequently leading to inh ibition of exonuclease and DNA repair. Previously, synergism between both c ompounds was found in several human and murine cancer cell lines when cells were treated with these drugs in a constant ratio. In the present study we used different combinations of both drugs (one drug at its IC25 and the ot her in a concentration range) in the human ovarian cancer cell line A2780, its CDDP-resistant variant ADDP, its dFdC-resistant variant AG6000 and two NSCLC cell lines, H322 (human) and Lewis lung (LL) (murine). Cells were exp osed for 4, 24 and 72 h with a total culture time of 96 h, and possible syn ergism was evaluated by median drug effect analysis by calculating a combin ation index (CI; CI < 1 indicates synergism). With CDDP at its IC25, the av erage Cls calculated at the IC50, IC75 IC90 and IC95 after 4, 24 and 72 h o f exposure were < 1 for all cell lines, indicating synergism, except for th e CI after 4 h exposure in the LL cell line which showed an additive effect . With dFdC at its IC25, the Cls for the combination with CDDP after 24 h w ere < 1 in all cell lines, except for the Cls after 4 h exposure in the LL and H322 cell lines which showed an additive effect. At 72 h exposure all C ls were < 1. CDDP did not significantly affect dFdCTP accumulation in ail c ell lines. CDDP increased dFdC incorporation into both DNA and RNA of the A 2780 cell lines 33- and 79-fold (P < 0.01) respectively, and tended to incr ease the dFdC incorporation into RNA in all cell lines. In the AG6000 and L L cell lines, CDDP and dFdC induced > 25% more DNA strand breaks (DSB) than each drug alone; however, in the other cell lines no effect, or even a dec rease in DSB, was observed. dFdC increased the cellular Pt accumulation aft er 24 h incubation only in the ADDP cell line. However, dFdC did enhance th e Pt-DNA adduct formation in the A2780, AG6000, ADDP and LL cell lines (1.6 -, 1.4-, 2.9- and 1.6-fold respectively). This increase in Pt-DNA adduct fo rmation seems to be related to the incorporation of dFdC into DNA (r = 0.91 ). No increase in DNA platination was found in the H322 cell line, dFdC onl y increased Pt-DNA adduct retention in the A2780 and LL cell lines, but dec reased the Pt-DNA adduct retention in the AG6000 cell line. In conclusion, the synergism between dFdC and CDDP appears to be mainly due to an increase in Pt-DNA adduct formation possibly related to changes in DNA due to dFdC incorporation into DNA.