Department of Hepatobiliary Surgery, The First Hospital of Shangjiazhuang City, Shijiazhuang City, Hebei Province 050011, China.
Exp Biol Med (Maywood). 2013 Sep;238(9):1024-32. doi: 10.1177/1535370213497321. Epub 2013 Aug 7.
Multidrug resistance (MDR) has become a major impediment to a successful treatment for liver cancer patients, and one of the common reasons for MDR is the activation of ABCB1 gene, leading to the over-expression of P-glycoprotein (P-gp), which conferred cancer cells be resistant to a broad range of anticancer drugs. MicroRNAs (miRNAs) are a class of short, non-coding RNA moleculars that can regulate gene expression at the post-transcriptional level. In the current study, the aim is to explore whether miRNA participates in the regulation of MDR mediated by ABCB1. We found that the expression of ABCB1 was correlated with the doxorubicin IC50 dose in eight hepatocellular carcinoma (HCC) cell lines: Hep3B, HCC3, LM-6, SMMC7721, Huh-7, SK-Hep-1, HepG2 and BEL-7402. Using the bioinformatics, we discovered that there were several miRNAs that can bind to the 3'UTR of ABCB1 gene. Among these candidate miRNAs, miR-223 was chosen for further study. Then, EGFP reporter assay, real-time PCR and Western blot were performed to verify that miR-223 targeted ABCB1 3'UTR directly, and miR-223 downregulated ABCB1 at both mRNA and protein levels. Finally, we found that the over-expression of miR-223 increased the HCC cell sensitivity to anticancer drugs, and the inhibition of miR-223 had the opposite effect. Importantly, the over-expression or silencing of ABCB1 can rescue the cell response to the anticancer drugs mediated by miR-223 over-expression or inhibition, respectively. In conclusion, our findings indicated that miR-223 played an important role in the regulation of MDR mediated by ABCB1, and it suggests that miR-223 may be considered as a therapeutic biomarker for HCC patients who had MDR problems induced by high expression of ABCB1.
多药耐药（MDR）已成为肝癌患者成功治疗的主要障碍，MDR 的一个常见原因是 ABCB1 基因的激活，导致 P-糖蛋白（P-gp）的过度表达，使癌细胞对广泛的抗癌药物产生耐药性。微小 RNA（miRNA）是一类短的非编码 RNA 分子，可以在转录后水平调节基因表达。在本研究中，目的是探讨 miRNA 是否参与 ABCB1 介导的 MDR 调节。我们发现，在八种肝癌（HCC）细胞系：Hep3B、HCC3、LM-6、SMMC7721、Huh-7、SK-Hep-1、HepG2 和 BEL-7402 中，ABCB1 的表达与阿霉素 IC50 剂量相关。通过生物信息学，我们发现有几个 miRNA 可以与 ABCB1 基因的 3'UTR 结合。在这些候选 miRNA 中，选择 miR-223 进行进一步研究。然后，进行 EGFP 报告基因检测、实时 PCR 和 Western blot 实验，验证 miR-223 可以直接靶向 ABCB1 3'UTR，miR-223 可以下调 ABCB1 的 mRNA 和蛋白水平。最后，我们发现 miR-223 的过表达增加了 HCC 细胞对抗癌药物的敏感性，而 miR-223 的抑制作用则相反。重要的是，ABCB1 的过表达或沉默可以分别挽救 miR-223 过表达或抑制介导的抗癌药物对细胞反应。总之，我们的研究结果表明，miR-223 在 ABCB1 介导的 MDR 调节中发挥重要作用，并表明 miR-223 可能被认为是由 ABCB1 高表达引起的 MDR 问题的 HCC 患者的治疗性生物标志物。
