Gemcitabine (2'-2'-difluorodeoxycytidine; dFdC) is a
deoxycytidine analogue which is effective against solid tumours, including
lung cancer and
ovarian cancer. dFdC requires phosphorylation by
deoxycytidine kinase (dCK) for activation. In the human
ovarian cancer cell line A2780 and its 30,000-fold dFdC-resistant variant AG6000 (P<0.001), we investigated the cross-resistance profile to several drugs. AG6000, which has a complete dCK deficiency, was approximately 1000-10,000-fold resistant to other deoxynucleoside analogues 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
enzyme 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), which 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
AG337 (P<0.05). Remarkably, AG6000 cells were 2.5-fold more sensitive to
methotrexate (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 polyglutamation of MTX were found between the cell lines. AG6000 cells were approximately 2 to 7.5-fold more resistant to
doxorubicin (DOX),
daunorubicin (DAU),
epirubicin and
vincristine (VCR) (the latter was significant; P<0.02) and approximately 4-fold more resistant to the microtubule inhibitors
paclitaxel and
docetaxel (P<0.001). Fluorescent activated cell sorter (FACS) analysis revealed no
P-glycoprotein (Pgp) or
multidrug resistance-associated protein (MRP) expression, but less fluorescence of intercalated DAU in AG6000 cells. An approximately 2-fold resistance to the
topoisomerase I and II inhibitors
etoposide,
CPT-11 and SN38 was found in AG6000 cells.
Topoisomerase I and IIalpha
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), respectively.
DT-diaphorase (DTD), which activates
EO9, was 2.2-fold lower in 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 equitoxic concentrations, resulted in more double-strand breaks (1.5- to 2-fold) in A2780 than in AG6000 cells. MAPH at 1120 nM and 17 nM of
EO9 did not cause DNA damage in either cell line. In conclusion, AG6000 is a cell line highly cross-resistant to a wide variety of drugs. This cross-resistance might be related to altered
enzyme activities and/or increased DNA repair.