Enhanced radiosensitization with gemcitabine in mismatch repair-deficient HCT116 cells.
Enhanced radiosensitization with gemcitabine in mismatch repair-deficient HCT116 cells.
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发表时间：
2003-10
期刊：
影响因子：
11.2
通讯作者：
B. Robinson;M. Im;M. Ljungman;F. Praz;D. Shewach
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作者：
B. Robinson;M. Im;M. Ljungman;F. Praz;D. Shewach
吉西他滨[2 '，2'-二氟-2 '-脱氧胞苷（dFdCyd）]是体外和体内实体瘤细胞中的有效电离辐射增敏剂。之前，我们已经证明了（Shewach等人，癌症研究所，五十四：3218-3223，1994）dATP的消耗（由dFdCyd二磷酸介导的核糖核苷酸还原酶抑制引起）与放射增敏之间的强相关性。此外，我们和其他人（Latz等人，国际辐射杂志Oncol.生物物理学，41：875-882，1998; Ostruszka和Shewach，Cancer Res.，60：6080-6088，2000）已经表明，在照射之前S期细胞的积累对于用dFdCyd进行放射增敏也是重要的。这使我们假设，由于dATP池不平衡而导致的不正确核苷酸的掺入对于dFdCyd的放射增敏作用很重要，因此，错配修复（MMR）缺陷的细胞将表现出更大的放射增敏作用。我们通过评估HCT 116结肠癌细胞系（MMR能力不同）对dFdCyd放射增敏的能力来验证这一假设。MMR熟练细胞系（HCT 116 + ch 3）比MMR缺陷细胞系（HCT 116、HCT 116 + ch 2和HCT 116 p53（-/-））对单独的dFdCyd更敏感。有趣的是，在dFdCyd IC（96）的浓度下，MMR熟练细胞不能被放射敏化，用放射增强细胞杀伤。与此相反，MMR缺陷细胞在dFdCyd浓度或dATP降低≥ 80%时在药物添加后4 h内放射增敏，并且这种低dATP水平再维持12-20 h。尽管dFdCyd的IC（50）不能维持MMR-熟练细胞中dATP水平降低>80%，但IC（90）确实实现了该dATP消耗水平;然而，它不能使MMR-熟练细胞放射增敏。用HCT 116细胞获得了类似的结果，其中通过用含有hMLH 1 cDNA的载体转染来校正MMR缺陷。此外，p53基因的缺失并没有增加放射增强率。这些结果表明，MMR缺陷促进dFdCyd的放射增敏作用。我们认为，dATP耗竭产生错误的复制在MMR缺陷的细胞，如果离开未修复，增强电离辐射的细胞死亡。
Gemcitabine [2',2'-difluoro-2'-deoxycytidine (dFdCyd)] is a potent ionizing radiation sensitizer in solid tumor cells in vitro and in vivo. Previously, we have demonstrated (Shewach et al., Cancer Res., 54: 3218-3223, 1994) a strong correlation between depletion of dATP (caused by dFdCyd diphosphate-mediated inhibition of ribonucleotide reductase) and radiosensitization. In addition, we and others (Latz et al., Int. J. Radiat. Oncol. Biol. Phys., 41: 875-882, 1998; Ostruszka and Shewach, Cancer Res., 60: 6080-6088, 2000) have shown that the accumulation of cells in S phase prior to irradiation is also important for radiosensitization with dFdCyd. This led us to hypothesize that the incorporation of incorrect nucleotides because of the dATP pool imbalance was important for radiosensitization with dFdCyd, and, therefore, cells deficient in mismatch repair (MMR) would exhibit greater radiosensitization. We tested this hypothesis by evaluating the ability of HCT116 colon carcinoma cell lines, which differ in MMR proficiency, to be radiosensitized by dFdCyd. The MMR-proficient cell line (HCT116 + ch3) was more sensitive to dFdCyd alone than were the MMR-deficient cell lines (HCT116, HCT116 + ch2, and HCT116 p53(-/-)). Interestingly, the MMR-proficient cells could not be radiosensitized at concentrations of dFdCyd IC(96)) enhanced cell killing with radiation. In contrast, the MMR-deficient cells were radiosensitized at concentrations of dFdCyd or=80% decrease in dATP within 4 h after drug addition, and this low dATP level was maintained for another 12-20 h. Although the IC(50) of dFdCyd was unable to sustain a >80% decrease in the dATP level in the MMR-proficient cells, the IC(90) did achieve this level of dATP depletion; however, it was unable to radiosensitize the MMR-proficient cells. Similar results were obtained with HCT116 cells, in which the MMR deficiency was corrected by transfection with a vector containing the hMLH1 cDNA. In addition, the deletion of p53 did not increase radiation enhancement ratios. These results demonstrate that MMR deficiency promotes radiosensitization with dFdCyd. We suggest that dATP depletion produces errors of replication in MMR-deficient cells, which, if left unrepaired, enhances cell death by ionizing radiation.