Ruiz van Haperen V W, Veerman G, Vermorken J B, Peters G J
Department of Oncology, Free University Hospital, Amsterdam, The Netherlands.
Biochem Pharmacol. 1993 Aug 17;46(4):762-6. doi: 10.1016/0006-2952(93)90566-f.
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.
吉西他滨（dFdC）是一种新型胞苷类似物，其主要通过将其三磷酸形式（dFdCTP）掺入DNA而发挥作用，从而导致细胞死亡。我们测定了dFdC在两种实体瘤细胞系核酸中的掺入情况：小鼠结肠癌细胞系Colon 26 - 10、人卵巢癌细胞系A2780以及人白血病细胞系CCRF - CEM。dFdC不仅掺入DNA，还掺入RNA。掺入DNA的程度在A2780细胞中最高，在CCRF - CEM细胞中最低（相差2 - 4倍）。掺入RNA也观察到相同模式，但相差10 - 20倍。在A2780细胞中，掺入DNA的量约为掺入RNA量的两倍，在CEM细胞中则为RNA的10 - 20倍。通过两种分离RNA和DNA的方法（酸沉淀和CsCl梯度离心）验证了其掺入RNA的情况。掺入DNA具有时间和浓度依赖性，但掺入RNA似乎仅具有浓度依赖性。我们还通过分别测量胸苷和尿苷的掺入情况，在与掺入研究相似的条件下，测定了dFdC对DNA和RNA合成的影响。在所有三种细胞系中，即使在0.1微摩尔/升dFdC浓度及4小时暴露时间下，DNA合成几乎完全被抑制。在两种实体瘤细胞系中，即使在1微摩尔/升dFdC暴露24小时的情况下，RNA合成抑制也未超过50%。仅在CCRF - CEM细胞系中且仅在24小时暴露后才观察到明显的浓度效应。在1微摩尔/升dFdC暴露24小时时，这些细胞中的RNA合成被完全抑制。dFdC掺入RNA以及对RNA合成的抑制代表了该药物一种未被认识但可能重要的作用机制。
