Basic Study
Copyright ©The Author(s) 2015. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Sep 14, 2015; 21(34): 9945-9956
Published online Sep 14, 2015. doi: 10.3748/wjg.v21.i34.9945
β-elemene enhances the radiosensitivity of gastric cancer cells by inhibiting Pak1 activation
Jun-Song Liu, Xiang-Ming Che, Shuai Chang, Guang-Lin Qiu, Shi-Cai He, Lin Fan, Wei Zhao, Zheng-Liang Zhang, Shu-Feng Wang
Jun-Song Liu, Xiang-Ming Che, Shuai Chang, Guang-Lin Qiu, Shi-Cai He, Lin Fan, Wei Zhao, Zheng-Liang Zhang, Shu-Feng Wang, Department of General Surgery, First Affiliated Hospital of Xi’an Jiaotong University School of Medicine, Xi’an 710061, Shaanxi Province, China
Author contributions: Liu JS and Che XM contributed equally to this work and should be regarded as co-first authors; Liu JS and Che XM performed the majority of experiments and wrote the manuscript; Chang S, Qiu GL, He SC and Zhang ZL performed some of the experiments and helped edit the figures; Fan L and Zhao W provided analytical tools and analyzed the data; Wang SF designed the study and approved the final version of the manuscript.
Supported by National Nature Science Foundation of China, No. 81172357.
Conflict-of-interest statement: We have no conflicts of interest to declare.
Data sharing statement: We agree the data sharing. Technical appendix, statistical code and dataset are available from the corresponding author at dawn@mail.xjtu.edu.cn.
Open-Access: This article is an open-access article which was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/
Correspondence to: Shu-Feng Wang, MD, Department of General Surgery, First Affiliated Hospital of Xi’an Jiaotong University School of Medicine, Yanta West Road 277, Xi’an 710061, Shaanxi Province, China. dawn@mail.xjtu.edu.cn
Telephone: +86-29-82653900 Fax: +86-29-82653905
Received: March 5, 2015
Peer-review started: March 7, 2015
First decision: April 24, 2015
Revised: May 12, 2015
Accepted: July 18, 2015
Article in press: July 18, 2015
Published online: September 14, 2015
Abstract

AIM: To explore the potential of β-elemene as a radiosensitizer for gastric cancer cells and the underlying mechanisms.

METHODS: SGC7901, MKN45, MKN28, N87, and AGS human gastric cancer cell lines were used to screen for radioresistant gastric cancer cell lines. A 3-(4,5-dimeth-ylthiazol-2-yl)-2,5-diphenyltetrazolium (MTT) assay was used to determine the effects of β-elemene and IPA-3 on cell viability in MKN45 and SGC7901 gastric cancer cell lines. A clonogenic survival assay and annexin V-FITC/PI apoptosis detection assay were used to evaluate cellular radiosensitivity and radiation-induced cell death, respectively. A proteomic method, isobaric tags for relative and absolute quantitation (iTRAQ), was employed to screen the proteins regulated by β-elemene pretreatment prior to ionizing radiation (IR) in SGC7901 gastric cancer cell line. IPA-3 was used as a specific small molecule inhibitor of p21-activated protein kinase 1 (Pak1) to target Pak1 signaling. Protein levels of PAK1IP1 (p21-activated protein kinase-interacting protein 1), total Pak1 (t-Pak1), phospho-Pak1 (T423), phospho-ERK1/2 (Thr202/Tyr204), and cleaved caspase-3 (17 kDa) were assessed by western blotting.

RESULTS: MKN45 and SGC7901 gastric cancer cell lines were relatively more resistant to IR. β-elemene pretreatment decreased clonogenic survival following IR in MKN45 and SGC7901 gastric cancer cell lines. Additionally, β-elemene pretreatment prior to IR increased radiation-induced cell death compared with IR alone in MKN45 (10.4% ± 0.9% vs 34.8% ± 2.8%, P < 0.05) and SGC7901 (11.6% ± 0.9% vs 46.7% ± 5.2%, P < 0.05) human gastric cancer cell lines, respectively, consistent with the level of cleaved caspase-3 (17 kDa). Through iTRAQ analysis and western blot validation, we found that β-elemene upregulated PAK1IP1 and downregulated phospho-Pak1 (T423) and phospho-ERK1/2 in SGC7901 gastric cancer cells. IR increased the level of phospho-Pak1 (T423). Pretreatment with β-elemene decreased radiation-induced Pak1 and ERK1/2 phosphorylation. Inhibition of Pak1 using IPA-3 decreased clonogenic survival following IR. In addition, IPA-3 increased radiation-induced cell death in MKN45 (13.4% ± 0.3% vs 26.6% ± 1.0%, P < 0.05) and SGC7901 (16.0% ± 0.6% vs 37.3% ± 1.7%, P < 0.05) gastric cancer cell lines, respectively, consistent with the level of cleaved caspase-3 (17 kDa). Western blotting showed that IPA-3 decreased radiation-induced Pak1 and ERK1/2 phosphorylation.

CONCLUSION: This is the first demonstration that β-elemene enhances radiosensitivity of gastric cancer cells, and that the mechanism involves inhibition of Pak1 signaling.

Keywords: β-elemene, Radiosensitivity, Gastric cancer cells, Clonogenic survival, Apoptosis, p21-activated protein kinase 1

Core tip: In this study, we explore the potential of β-elemene, a novel anti-cancer drug isolated from the Chinese traditional herb Curcuma wenyujin, as a radiosensitizer for gastric cancer cells, and the underlying mechanisms. β-elemene pretreatment decreased clonogenic survival and increased apoptosis in response to ionizing radiation in SGC7901 and MKN45 gastric cancer cell lines. This is the first demonstration that β-elemene enhances radiosensitivity of gastric cancer cells. The underlying mechanism involves inhibition of p21-activated protein kinase 1 (Pak1) signaling. We also provide direct evidence for targeting Pak1 signaling as a means to sensitize cancer cells to ionizing radiation.