Interdependence of Gemcitabine Treatment, Transporter Expression, and Resistance in Human Pancreatic Carcinoma Cells
Interdependence of Gemcitabine Treatment, Transporter Expression, and Resistance in Human Pancreatic Carcinoma Cells
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DOI：
10.1593/neo.10576
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发表时间：
2010-09-01
期刊：
影响因子：
4.8
通讯作者：
Loehr, Johannes Matthias
中科院分区：
文献类型：
--
作者：
Hagmann, Wolfgang;Jesnowski, Ralf;Loehr, Johannes Matthias
吉西他滨是胰腺癌治疗的一线化疗药物。我们以前的实验性化疗研究表明，5-氟尿嘧啶（5-FU）治疗人胰腺癌细胞改变了细胞转运蛋白的表达谱和多药耐药蛋白5（MRP 5; ABCC 5）的表达的调制影响这些肿瘤细胞的耐药性。在这里，我们研究了急性和慢性吉西他滨治疗胰腺癌细胞相关的摄取和输出转运蛋白的表达的影响，通过逆转录聚合酶链反应（RT-PCR），定量RT-PCR和免疫印迹分析。通过使用MRP 5过表达和MRP 5沉默细胞的细胞毒性试验，研究了MRP 5在细胞吉西他滨敏感性中的特定作用。暴露于吉西他滨（12 nM，3天）并没有改变MRP 1，MRP 3，MRP 5和平衡型核苷转运蛋白1（ENT 1）的信使RNA（mRNA）表达，而高剂量的药物（20 μ M，1小时）在大多数研究的细胞系中引起这些转运蛋白的上调。在获得性吉西他滨耐药（高达160 nM吉西他滨）的细胞中，吉西他滨转运蛋白MRP 5和ENT 1的mRNA或蛋白表达在几种细胞系中上调。5-FU和吉西他滨联合治疗导致MRP 5和ENT 1表达增加5- 40倍。使用MRP 5过表达（HEK和PANC-1）或MRP 5沉默（PANC 1/shMRP 5）细胞的细胞毒性试验表明，MRP 5有助于吉西他滨耐药性。因此，我们的新数据，不仅对药物诱导的转运蛋白表达的改变相关的吉西他滨的摄取和输出，但也对吉西他滨的敏感性和MRP 5表达之间的联系可能会导致未来的化疗方案使用吉西他滨在胰腺癌患者的改进策略。
Gemcitabine is widely used as first-line chemotherapeutic drug in the treatment of pancreatic cancer. Our previous experimental chemotherapy studies have shown that treatment of human pancreatic carcinoma cells with 5-fluorouracil (5-FU) alters the cellular transporter expression profile and that modulation of the expression of multidrug resistance protein 5 (MRP5; ABCC5) influences the chemoresistance of these tumor cells. Here, we studied the influence of acute and chronic gemcitabine treatment on the expression of relevant uptake and export transporters in pancreatic carcinoma cells by reverse transcription-polymerase chain reaction (RT-PCR), quantitative RT-PCR, and immunoblot analyses. The specific role of MRP5 in cellular gemcitabine sensitivity was studied by cytotoxicity assays using MRP5-overexpressing and MRP5-silenced cells. Exposure to gemcitabine (12 nM for 3 days) did not alter the messenger RNA (mRNA) expression of MRP1, MRP3, MRP5, and equilibrative nucleoside transporter 1 (ENT1), whereas high dosages of the drug (20 mu M for 1 hour) elicited up-regulation of these transporters in most cell lines studied. In cells with acquired gemcitabine resistance (up to 160 nM gemcitabine), the mRNA or protein expression of the gemcitabine transporters MRP5 and ENT1 was upregulated in several cell lines. Combined treatment with 5-FU and gemcitabine caused a 5- to 40-fold increase in MRP5 and ENT1 expressions. Cytotoxicity assays using either MRP5-overexpressing (HEK and PANC-1) or MRP5-silenced (PANC1/shMRP5) cells indicated that MRP5 contributes to gemcitabine resistance. Thus, our novel data not only on drug-induced alterations of transporter expression relevant for gemcitabine uptake and export but also on the link between gemcitabine sensitivity and MRP5 expression may lead to improved strategies of future chemotherapy regimens using gemcitabine in pancreatic carcinoma patients.