Mechanisms of acquired resistance to modulation of 5-fluorouracil by leucovorin in HCT-8 human ileocecal carcinoma cells.
Mechanisms of acquired resistance to modulation of 5-fluorouracil by leucovorin in HCT-8 human ileocecal carcinoma cells.
复制标题
HCT-8 人回盲部癌细胞对亚叶酸调节 5-氟尿嘧啶的获得性耐药机制。
DOI：
10.1016/s0006-2952(96)00900-8
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发表时间：
1997
影响因子：
5.8
通讯作者：
Rustum,YM
中科院分区：
文献类型：
--
作者：
Lu,K;McGuire,JJ;Slocum,HK;Rustum,YM
将HCT-8人回盲部癌细胞重复（10倍）暴露于5-氟尿嘧啶（5-FU）2或72小时，均在20 μM甲酰四氢叶酸（LV）持续存在下孵育，产生了两种稳定的抗调节亚系FL 2 h和FL 72 h。尽管LV在亲本HCT-8细胞中增强了5-FU的2倍生长抑制，但在FL 2 h或FL 72 h亚系中并未增强5-FU的作用。5-氟脱氧尿苷（5-FdUrd）的LV调节也减少（FL 72 h）或消除（FL 2 h）。在FL 2 h和FL 72 h亚系中，细胞提取物中胸苷酸合成酶（TS）蛋白水平和TS活性、原位TS活性、LV的细胞摄取和代谢速率以及5-FU掺入细胞总RNA的水平与亲本HCT-8细胞中的水平相似。LV对5-FU或5-FdUrd处理HCT-8细胞2 h后24 h的TS活性抑制作用明显增强（P < 0.01），而对FL 2 h和FL 72 h细胞则无此作用（P > 0.1）。LV对5-FU调节的抗性与LV不能增加细胞内TS-FdUMP-亚甲基四氢叶酸三元复合物的形成有关，并且在存在不同浓度的5，10-亚甲基四氢蝶酰五谷氨酸时，这些复合物解离更快（T1 2> 1.5 - 3倍）。因此，三元复合物的稳定性降低似乎是对氟嘧啶细胞毒性的LV调节的获得性抗性的机制，可能是由于这两种调节抗性HCT-8亚系中的TS突变。
Repeated (10x) exposure of HCT-8 human ileocecal carcinoma cells to 5-fluorouracil (5-FU) for 2 or 72 hr, both incubations in the continuous presence of 20 μM leucovorin (LV), yielded two stable modulation-resistant sublines, FL2h and FL72h. Although LV potentiated growth inhibition by 5-FU 2-fold in parental HCT-8 cells, it did not potentiate the effect of 5-FU in the FL2h or FL72h sublines. LV modulation of 5-fluorodeoxyuridine (5-FdUrd) was also reduced (FL72h) or eliminated (FL2h). In the FL2h and FL72h sublines, the level of thymidylate synthase (TS) protein and TS activity in cell extracts, TS activity in situ, the rate of cellular uptake and metabolism of LV, and the level of 5-FU incorporation into total cellular RNA were similar to those in parental HCT-8 cells. However, LV significantly (P < 0.01) potentiated the inhibition of TS activity insitu in HCT-8 cells at 24 hr after a 2-hr treatment with either 5-FU or 5-FdUrd, but had no such activity in the FL2h and FL72h sublines (P > 0.1). Resistance to modulation of 5-FU by LV was associated with the inability of LV to increase the formation of intracellular TS-FdUMP-methylenetetrahydrofolate ternary complexes, and these complexes dissociated more rapidly (T1 2> 1.5 - to 3-fold faster) in the presence of different concentrations of 5,10-methylenetetrahydropteroylpentaglutamate. Thus, decreased stability of ternary complexes appears to be the mechanism of acquired resistance to the LV modulation of fluoropyrimidine cytotoxicity, possibly due to mutation(s) of TS in these two modulation-resistant HCT-8 sublines.