MRP3 as a novel resistance factor for sorafenib in hepatocellular carcinoma.

Tetsu Tomonari,Tetsuji Takayama,Shota Fujimoto,Hiroshi Miyamoto,Tatsuya Taniguchi,Koichi Okamoto,Takahiro Tanaka,Naoki Muguruma,Hironori Tanaka,Tetsuo Kimura,Shunsaku Takeishi

doi:10.18632/oncotarget.6889

Tetsu Tomonari, Tetsuji Takayama + Show 9 more

Open Access

https://doi.org/10.18632/oncotarget.6889

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Journal: Oncotarget	Publication Date: Jan 12, 2016
Citations: 84	License type: cc-by

Affiliation: Tokushima University

Abstract

The mechanism of resistance of hepatocellular carcinoma (HCC) to sorafenib is unknown and no useful predictive biomarker for sorafenib treatment has been reported. Accordingly, we established sorafenib-resistant HCC cells and investigated the underlying mechanism of resistance to sorafenib. Sorafenib-resistant cell lines were established from the HCC cell line PLC/PRF5 by cultivation under continuous exposure to increasing concentration of sorafenib. The IC50 values of the 2 resistant clones PLC/PRF5-R1 and PLC-PRF5-R2 were 9.2±0.47 μM (1.8-fold) and 25±5.1 μM (4.6-fold) respectively, which were significantly higher than that of parental PLC/PRF5 cells (5.4±0.17 μM) (p < 0.01 respectively), as determined by MTT assay. Western blot analysis of signal transduction-related proteins showed no significant differences in expression of AKT/pAKT, mTOR/pmTOR, or ERK/pERK between the 2 resistant clones versus parent cells, suggesting no activation of an alternative signal transduction pathway. Likewise, when expression of membrane transporter proteins was determined, there were no significant differences in expression levels of BSEP, MDR1, MRP2, BCRP, MRP4 and OCT1 between resistant clones and parent cells. However, the expression levels of MRP3 in the 2 resistant clones were significantly higher than that of parent cells. When MRP3 gene was knocked down by siRNA in PLC-PRF5-R2 cells, the sensitivity of the cells to sorafenib was restored. In the analysis of gene mutation, there was no mutation in the activation segment of Raf1 kinase in the resistant clones. Our data clearly demonstrate that the efflux transporter MRP3 plays an important role in resistance to sorafenib in HCC cells.

Highlights

Hepatocellular carcinoma (HCC) is the thirdleading cause of cancer-related death worldwide [1]
It became evident that the AKT/mTOR signaling pathway was not activated in our sorafenib-resistant clones
We established 2 sorafenib-resistant clones from the HCC cell line PLC/PRF5, and showed that themembrane transporter MRP3 was overexpressed in these clones

Summary

Introduction

Hepatocellular carcinoma (HCC) is the thirdleading cause of cancer-related death worldwide [1]. Most patients with HCC are diagnosed at an advanced stage, when curative therapies, such as surgical resection and percutaneous ablation, are of limited utility. Since a majority of HCC present intrinsic resistance to many cytotoxic anti-cancer agents, interventional treatment such as transarterial embolization had been applied for advanced HCC [2,3]. It has been reported that sorafenib, a multitargeted tyrosine kinase inhibitor that blocks Raf kinase, platelet-derived growth factor receptor (PDGFR) and vascular endothelial growth factor receptors (VEGFR), significantly improved overall survival in patients with advanced HCC in large-scaled multicenter phase III trials [4,5]. Systemic treatment with sorafenib is currently recommended for advanced stage HCC in the treatment algorithm of HCC worldwide [6]. The response rate for sorafenib in HCC is very low (i.e., 2 - 3%) [4,5]

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