Shewach D S, Hahn T M, Chang E, Hertel L W, Lawrence T S
Department of Pharmacology, University of Michigan Medical Center, Ann Arbor 48109.
Cancer Res. 1994 Jun 15;54(12):3218-23.
Difluorodeoxycytidine (dFdCyd) is a new antimetabolite with clinical activity in patients with solid tumors but not leukemias. We have studied the metabolism, cytotoxicity, and radiosensitizing properties of dFdCyd in HT-29 human colon carcinoma cells. The results demonstrated that dFdCyd rapidly accumulated as the 5'-triphosphate dFdCTP in HT-29 cells, which was eliminated slowly in the absence of dFdCyd with a half-life of > 12 h. Accumulation of dFdCTP was associated with rapid depletion of cellular dATP pools. Exposure to the concentration that reduces cell survival by 50% of 30 nM dFdCyd decreased dATP levels by > 80% within 4 h. dGTP pools were depleted at higher concentrations of dFdCyd, whereas smaller decreases were effected in dTTP and dCTP pools. These results contrast with previous reports in leukemic cells which demonstrated that dFdCyd exposure depleted the endogenous dCTP pool to a greater extent than the dTTP, dATP, or dGTP pools. Based on these data, we suggest that the profound depletion by dFdCyd of dATP and dGTP pools in HT-29 compared to leukemic cells accounts for the superiority of this agent in solid tumors versus leukemias. Additional studies demonstrated that dFdCyd was a potent radiosensitizer in HT-29 cells. Maximal radiosensitization was observed when cells were irradiated immediately following dFdCyd exposure instead of prior to or in the middle of drug treatment. Radiation sensitization was dose and time dependent, with a noncytotoxic exposure to 10 nM dFdCyd for 24 h or 30 nM dFdCyd for 16 h producing a radiation enhancement ratio of approximately 2. Under these conditions, only the cellular dATP pool was depleted. When cells were exposed to higher concentrations of dFdCyd for 4 h, equivalent radiosensitization with a radiation enhancement ratio of 1.4 was obtained using 0.1, 1.0, or 10 microM dFdCyd. Despite a 15-fold increase in dFdCTP and depletion of dGTP and dCTP pools to < 25% of the control value with 10 microM compared to 0.1 microM dFdCyd, no increase in radiosensitization was observed. These results suggest that dATP depletion is an important factor in the radiosensitizing property of this promising new antitumor compound.
二氟脱氧胞苷（dFdCyd）是一种新型抗代谢物，对实体瘤患者具有临床活性，但对白血病患者无活性。我们研究了dFdCyd在HT - 29人结肠癌细胞中的代谢、细胞毒性和放射增敏特性。结果表明，dFdCyd在HT - 29细胞中迅速积累为5'-三磷酸dFdCTP，在没有dFdCyd的情况下，其消除缓慢，半衰期> 12小时。dFdCTP的积累与细胞内dATP池的快速消耗有关。暴露于使细胞存活率降低50%的30 nM dFdCyd浓度下，4小时内dATP水平降低> 80%。在较高浓度的dFdCyd下，dGTP池被耗尽，而dTTP和dCTP池的降低较小。这些结果与先前关于白血病细胞的报道形成对比，先前报道表明，暴露于dFdCyd会使内源性dCTP池比dTTP、dATP或dGTP池消耗得更多。基于这些数据，我们认为与白血病细胞相比，dFdCyd使HT - 29细胞中的dATP和dGTP池深度消耗，这解释了该药物在实体瘤中相对于白血病的优势。进一步的研究表明，dFdCyd在HT - 29细胞中是一种有效的放射增敏剂。当细胞在暴露于dFdCyd后立即照射，而不是在药物治疗之前或中间照射时，观察到最大放射增敏效果。放射增敏作用具有剂量和时间依赖性，非细胞毒性暴露于10 nM dFdCyd 24小时或30 nM dFdCyd 16小时产生的放射增强比约为2。在这些条件下，仅细胞内dATP池被耗尽。当细胞暴露于更高浓度的dFdCyd 4小时时，使用0.1、1.0或10 microM dFdCyd可获得等效的放射增敏效果，放射增强比为1.4。尽管与0.1 microM dFdCyd相比，10 microM dFdCyd使dFdCTP增加了15倍，dGTP和dCTP池减少至对照值的< 25%，但未观察到放射增敏作用增强。这些结果表明，dATP消耗是这种有前景的新型抗肿瘤化合物放射增敏特性的一个重要因素。
