Cross-resistance in the 2 ',2 '-difluorodeoxycytidine (gemcitabine)-resistant human ovarian cancer cell line AG6000 to standard and investigational drugs

Gemcitabine (2'-2'-difluorodeoxycytidine; dFdC) is a deoxycytidine analogue which is effective against solid tumours, iucluding lung cancer and ovaria n cancer. dFdC requires phosphorylation by deoxycytidine kinase (dCK) for a ctivation. In the human ovarian cancer cell line A2780 and its 30 000-fold dFdC-resistant variant AG6000 (P < 0.001), we investigated the cross-resist ance profile to several drugs. AG6000, which has a complete dCK deficiency, was approximately 1000-10 000-fold resistant to other deoxynucleoside anal ogues such as 1-beta-D-arabinofuranosyl cytosine, 2-chloro-deoxyadenosine, aza-deoxycytidine and 2',2'-difluorodeoxyguanosine (dFdG) (P < 0.001). dFdG can be activated by dCK and deoxyguanosine kinase (dGK); but the latter en zyme was not altered in AG6000 cells. Thus dFdG resistance was only due to dCK deficiency. AG6000 was 1.6- and 46.7-fold resistant to 5-fluorouracil ( 5-FU) and ZD1694, respectively (the latter was significant, P < 0.01), whic h may be due to the 1.7-fold higher thymidylate synthase (TS) activity, but AG6000 cells were also 2.7-fold resistant to the lipophilic TS inhibitor A G337 (P < 0.05). Remarkably, AG6000 cells were 2.5-fold more sensitive to m ethotrexate (MTX) (P < 0.01) than A2780 cells, but 1.6-fold more resistant to trimetrexate (TMQ) (P < 0.10). However, no differences in reduced folate carrier activity, folylpolyglutamate synthetase (FPGS) activity and polygl utamation of MTX were found between the cell lines. AG6000 cells were appro ximately 2 to 7.5-fold more resistant to doxorubicin (DOX), daunorubicin (D AU), epirubicin and vincristine (VCR) (the latter was significant; P < 0.02 ) and approximately 4-fold more resistant to the microtubule inhibitors pac litaxel and docetaxel (P < 0.001). Fluorescent activated cell sorter (FACS) analysis revealed no P-glycoprotein (Pgp) or multidrug resistance-associat ed protein (MRP) expression, but less fluorescence of intercalated DAU in A G6000 cells. An approximately 2-fold resistance to the topoisomerase I and II inhibitors etoposide, CPT-I i and SN38 uas found in AG6000 cells. Topois omerase land II alpha RNA expression was decreased in AG6000 cells. AG6000 was 2.4, 2.4, 2.3 and 3.7-fold more resistant to EO9 (P < 0.02), mitomycin- C (MMC) (p < 0.05), cisplatin (CDDP) (P < 0.10) and maphosphamide (MAPH), r espectively. DT-diaphorase (DTD), which activates EO9, was 2.2-fold lower i n AG6000 cells. CDDP resistance might be related to a reduced retention of DNA adducts in AG6000. However, glutathione levels were equal in A2780 and AG6000 cells. A 24 h exposure to DOX, VCR and paclitaxel at equimolar and e quitoxic concentrations, resulted in more double-strand breaks (1.5- to 2-f old) in A2780 than in AG6000 cells. MAPH at 1120 nM and 17 nM of EO9 did no t cause DNA damage in either cell line. In conclusion, AG6000 is a cell lin e highly cross-resistant to a wide variety of drugs. This cross-resistance might be related to altered enzyme activities and;or increased DNA repair. (C) 2000 Elsevier Science Ltd. All rights reserved.