Department of Gastroenterology, Zhejiang Provincial People's Hospital, Hangzhou, Zhejiang, China.
People's Hospital of Hangzhou Medical College, Hangzhou, Zhejiang, China.
J Pharm Pharmacol. 2019 Feb;71(2):196-205. doi: 10.1111/jphp.12941. Epub 2019 Jan 2.
Chemotheraputic drug resistance is a critical factor associated with the poor survival in advanced/metastatic pancreatic cancer (PC) patients.
Human pancreatic cell lines Capan-1 and BXPC-3 were cultured with different concentrations of erlotinib (0, 10, 50, and 100 μm) for 48 h. The relative cell viability and apoptosis was detected using MTT assays and flow cytometry apoptosis analysis, respectively. Transfection of pcDNA-EphA2, si-EphA2 and miR-124 mimic/inhibitor was used to modulate the intracellular level of EphA2 and miR-124. The interaction between miR-124 and the 3'UTR of EphA2 was explored using dual luciferase reporter assay.
Compared with BXPC-3 cells, Capan-1 cells showed resistance to differential concentration treatment of erlotinib. The expression of EphA-2 was significantly increased and the expression of miR-124 was significantly decreased in Capan-1 cells. Overexpressing EphA2 induced resistance of BXPC-3 cells to erlotinib treatment. And EphA2 was identified as a novel target gene for miR-124. MiR-124 overexpression was able to sensitize the response of Capan-1 cells to erlotinib through inhibiting EphA2. Furthermore, both miR-124 overexpression and EphA2 inhibition sensitized Capan-1 cells to erlotinib in xenograft model.
Our study demonstrated that EphA2 rescued by miR-124 downregulation conferred the erlotinib resistance of PC cell Capan-1 with K-RAS mutation.
化疗药物耐药性是与晚期/转移性胰腺癌细胞（PC）患者生存不良相关的关键因素。
用不同浓度的厄洛替尼（0、10、50 和 100μm）培养人胰腺细胞系 Capan-1 和 BXPC-3 48 小时。分别用 MTT 法和流式细胞术凋亡分析检测相对细胞活力和细胞凋亡。转染 pcDNA-EphA2、si-EphA2 和 miR-124 模拟物/抑制剂，以调节 EphA2 和 miR-124 的细胞内水平。用双荧光素酶报告基因检测法探讨 miR-124 与 EphA2 的 3'UTR 之间的相互作用。
与 BXPC-3 细胞相比，Capan-1 细胞对厄洛替尼的不同浓度处理表现出耐药性。EphA-2 的表达在 Capan-1 细胞中显著增加，而 miR-124 的表达显著降低。过表达 EphA2 诱导 BXPC-3 细胞对厄洛替尼治疗产生耐药性。EphA2 被鉴定为 miR-124 的新靶基因。miR-124 过表达通过抑制 EphA2 使 Capan-1 细胞对厄洛替尼的反应敏感化。此外，miR-124 过表达和 EphA2 抑制均使携带 K-RAS 突变的 Capan-1 细胞对厄洛替尼敏感化。
本研究表明，miR-124 下调拯救的 EphA2 赋予了具有 K-RAS 突变的 PC 细胞 Capan-1 对厄洛替尼的耐药性。
