Takara Kohji, Kitada Noriaki, Yoshikawa Eri, Yamamoto Kazuhiro, Horibe Sayo, Sakaeda Toshiyuki, Nishiguchi Kohshi, Ohnishi Noriaki, Yokoyama Teruyoshi
Department of Clinical Pharmacy, Division of Clinical Pharmaceutical Sciences, Kyoto Pharmaceutical University, 5 Nakauchi-cho, Misasagi, Yamashina-ku, Kyoto 607-8414, Japan.
Cancer Lett. 2009 Jun 8;278(1):88-96. doi: 10.1016/j.canlet.2008.12.033. Epub 2009 Feb 6.
It is important to clarify the molecular characteristics of tumor cells showing multidrug resistance (MDR) and to identify the novel targets or biomarkers for chemotherapy. The aim of this study is to establish resistant HeLa sublines through exposure to SN-38, an active metabolite of irinotecan hydrochloride, and to investigate their molecular changes. HeLa cells were exposed to SN-38 at 1, 10, or 100 nM, and resistant clones were isolated and named HeLa/SN1, HeLa/SN10, and HeLa/SN100, respectively. Their cellular changes were examined based on growth inhibition assays, the function of ABCG2/BCRP, and a RT-PCR analysis of MDR-related protein. The sublines showed a decrease in sensitivity to not only SN-38 but also other chemotherapeutic agents as compared with HeLa cells. mRNA and protein levels of ABCG2/BCRP were increased, and the transport activity of ABCG2/BCRP was enhanced, in the resistant cells. In addition, the expression levels of ABCC1/MRP1, ABCC3/MRP3, and ABCC5/MRP5 were higher than in HeLa cells. The mRNA levels of GGT1 encoding a gamma-glutamyl transferase, but not GCS encoding a gamma-glutamyl cysteine synthetase, were also higher. Other factors examined, i.e., topoisomerase, SLCO1B1, and apoptosis-regulating factors, were comparable among the cells. The overexpression of ABCG2/BCRP was involved in the mechanism of resistance in SN-38-tolerant cells, and ABCC1/MRP1, ABCC3/MRP3, ABCC5/MRP5, and GGT1 may also have participated.
阐明显示多药耐药性（MDR）的肿瘤细胞的分子特征，并确定化疗的新靶点或生物标志物非常重要。本研究的目的是通过暴露于盐酸伊立替康的活性代谢物SN-38来建立耐药的HeLa亚系，并研究它们的分子变化。将HeLa细胞暴露于1、10或100 nM的SN-38中，分离出耐药克隆并分别命名为HeLa/SN1、HeLa/SN10和HeLa/SN100。基于生长抑制试验、ABCG2/BCRP的功能以及MDR相关蛋白的RT-PCR分析来检测它们的细胞变化。与HeLa细胞相比，这些亚系不仅对SN-38，而且对其他化疗药物的敏感性均降低。耐药细胞中ABCG2/BCRP的mRNA和蛋白水平升高，并且ABCG2/BCRP的转运活性增强。此外，ABCC1/MRP1、ABCC3/MRP3和ABCC5/MRP5的表达水平高于HeLa细胞。编码γ-谷氨酰转移酶的GGT1的mRNA水平也较高，但编码γ-谷氨酰半胱氨酸合成酶的GCS的mRNA水平则不然。所检测的其他因素，即拓扑异构酶、SLCO1B1和凋亡调节因子，在细胞之间相当。ABCG2/BCRP的过表达参与了对SN-38耐受细胞的耐药机制，并且ABCC1/MRP1、ABCC3/MRP3、ABCC5/MRP5和GGT1也可能参与其中。
