Schedule dependence of sensitivity to 2',2'-difluorodeoxycytidine (Gemcitabine) in relation to accumulation and retention of its triphosphate in solid tumour cell lines and solid tumours.
2',2'-Difluorodeoxycytidine (Gemcitabine, dFdC) is a relatively new deoxycytidine antimetabolite, with established activity against ovarian cancer and non-small-cell lung cancer. dFdC is assumed to exert its antitumour effect mainly by incorporation of the triphosphate dFdCTP into DNA. We determined the sensitivity to dFdC of six cell lines derived from solid tumours; two ovarian carcinoma (A2780 and OVCAR-3), two colon carcinoma (WiDr and C26-10) and two squamous cell carcinoma cell lines (UM-SCC-14C and UM-SCC-22B). In vitro sensitivity to dFdC was strongly time dependent. Under all conditions A2780 was the most sensitive cell line with an IC50 (the concentration of dFdC causing 50% growth inhibition) of 31 and 0.6 nM at 1 and 48 hr exposure, respectively. WiDr and C26-10 cells were relatively insensitive, with IC50s of 468 and 1133 nM, respectively, at 1 hr exposure, but of 11 and 6 nM at 48 hr exposure. Accumulation of the triphosphate dFdCTP was also time dependent. After 4 hr exposure to 10 microM dFdC, A2780, WiDr and C26-10 cells accumulated 223, 136 and 267 pmol/10(6) cells, respectively; after 24 hr exposure they accumulated 1045, 619 and 617 pmol/10(6) cells, respectively. A2780 cells retained the high dFdCTP concentration longer than 24 hr. For comparison purposes we also studied dFdCTP kinetics in the corresponding solid tumours, showing the same sensitivity pattern as the cell lines. In general, sensitivity to dFdC in vitro related with dFdCTP accumulation and retention, but in vivo this relation was less clear. Unexpectedly, remarkable in vitro and in vivo changes were observed in the ribonucleotide pools. The most predominant in vitro cell line dependent changes were a decrease in CTP concentrations, accompanied by an increase in UTP and GTP concentrations. In vivo CTP, UTP and GTP pools increased in all tumours. In conclusion, in this study we demonstrate that dFdCTP is accumulated and retained in solid tumours and cell lines. dFdCTP is not only important as a DNA precursor, but also appears to interfere with normal ribonucleotide metabolism.
Ruiz van Haperen
V W
VW
Department of Oncology, Free University Hospital, Amsterdam, The Netherlands.
Veerman
G
G
Boven
E
E
Noordhuis
P
P
Vermorken
J B
JB
Peters
G J
GJ
D000818:Animals; D000964:Antimetabolites, Antineoplastic / Q000378:metabolism*; D002455:Cell Division / Q000187:drug effects; D003110:Colonic Neoplasms / Q000378:metabolism*; D003841:Deoxycytidine / Q000008:administration & dosage / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D004354:Drug Screening Assays, Antitumor; D005260:Female; D006801:Humans; D051379:Mice; D010051:Ovarian Neoplasms / Q000378:metabolism*; D010710:Phosphates / Q000378:metabolism*; D012265:Ribonucleotides / Q000378:metabolism; D014407:Tumor Cells, Cultured / Q000378:metabolism; D000093542:Gemcitabine
D000964:Antimetabolites, Antineoplastic; D010710:Phosphates; D012265:Ribonucleotides; D003841:Deoxycytidine; D000093542:Gemcitabine