2',2'-Difluoro-deoxycytidine (gemcitabine) incorporation into RNA and DNA of tumour cell lines.
Gemcitabine (dFdC) is a new cytidine analogue which is active mainly by the incorporation of its triphosphate (dFdCTP) into DNA, leading to cell death. We determined incorporation of dFdC into nucleic acids of two solid tumour cell lines: the murine colon carcinoma cell line Colon 26-10, the human ovarian carcinoma cell line A2780, and the human leukemic cell line CCRF-CEM. dFdC was not only incorporated into DNA, but also into RNA. The extent of incorporation into DNA was highest in A2780 cells and lowest in CCRF-CEM cells (2-4-fold difference). The same pattern was observed for incorporation into RNA, but with a 10-20-fold difference. In A2780, incorporation into DNA was about twice that of the incorporation into RNA, in CEM cells 10-20-fold that of RNA. Incorporation into RNA was verified using two methods for separation of RNA and DNA, acid precipitation and CsCl-gradient centrifugation. Incorporation into DNA was time and concentration dependent, but incorporation into RNA seemed to be only concentration dependent. We also determined the effect of dFdC on DNA and RNA synthesis by measurement of thymidine and uridine incorporation, respectively, using similar conditions as for the incorporation studies. In all three cell lines DNA synthesis was inhibited almost completely, even at 0.1 microM dFdC and at 4-hr exposure. RNA synthesis inhibition did not exceed 50% in both solid tumour cell lines, even at 1 microM dFdC exposure for 24 hr. A clear concentration effect was only observed in the CCRF-CEM cell line and only after 24 hr exposure. At a 1 microM dFdC exposure for 24 hr, RNA synthesis was completely inhibited in these cells. Incorporation of dFdC into RNA and inhibition of RNA synthesis represent an unrecognized but possibly important mechanism of action of this drug.
Ruiz van Haperen
V W
VW
Department of Oncology, Free University Hospital, Amsterdam, The Netherlands.
Veerman
G
G
Vermorken
J B
JB
Peters
G J
GJ
D000818:Animals; D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology*; D004273:DNA, Neoplasm / Q000096:biosynthesis / Q000187:drug effects*; D003841:Deoxycytidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D006801:Humans; D051379:Mice; D012334:RNA, Neoplasm / Q000096:biosynthesis / Q000187:drug effects*; D014407:Tumor Cells, Cultured; D000093542:Gemcitabine
D000964:Antimetabolites, Antineoplastic; D004273:DNA, Neoplasm; D012334:RNA, Neoplasm; D003841:Deoxycytidine; D000093542:Gemcitabine