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ALEX1, a novel tumor suppressor gene, inhibits gastric cancer metastasis via the PAR-1/Rho GTPase signaling pathway

  • Original Article—Alimentary Tract
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Abstract

Background

The ALEX is a novel member of the armadillo family and ALEX1 was reported to be reduced or even lost in multiple solid tumors. However, its expression profile and oncogenic role in gastric cancer (GC) remains largely unknown.

Methods

ALEX1 expression was detected in 161 GC samples by immunohistochemistry staining. NCI-N87 cells transfected by ALEX1 lentivirus vectors and MKN28 cells transfected by ALEX1 shRNA were used for biological function investigation. Western blot was applied to explore the molecular mechanism and pull-down assays were applied to measure the activity of Rho GTPases. In vivo tumorigenicity, peritoneal and lung metastasis experiments were performed by tumor cell engraftment into nude mice. Bisulfite genomic sequencing and methylation-specific PCR were applied to check the methylation status of the ALEX1 gene.

Results

The expression rate of ALEX1 was significantly reduced in gastric tumor samples compared to non-tumor samples (43.5 vs. 90.2%), and its expression was closely related to the tumor differentiation, TNM staging, and lymph nodes metastasis. ALEX1 overexpression in NCI-N87 cells significantly inhibited cell proliferation, migration, and invasion in vitro, and disrupted the structure of the cytoskeleton. ALEX1 overexpression attenuated xenografts growth, peritoneal, and lung metastasis in nude mice. Mechanistically, the overexpression of ALEX1 inhibits thrombin-induced metastasis and Rho GTPases activation. Bisulfite genomic sequencing and methylation-specific PCR revealed that the promoter of ALEX1 is highly methylated in GC cells and tissues.

Conclusions

ALEX1 expression is reduced in GC and is involved in diverse cellular functions. ALEX1 inhibits metastasis through the PAR-1/Rho GTPase signaling pathway.

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Abbreviations

GC:

Gastric cancer

ALEX:

Arm protein lost in epithelial cancers, on chromosome X

PAR-1:

Protease-activated receptor 1

Rho GTPase:

Rho family of GTPase

EMT:

Epithelial-to-mesenchymal transition

CCK-8:

Cell Counting Kit-8

MSP:

Methylation-specific PCR

IHC:

Immunohistochemistry

qRT-PCR:

Quantitative reverse transcription-polymerase chain reaction

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Acknowledgements

We thank Dr. Wen-tao Dai for his help in statistical analysis. This study was supported by grants from National Natural Science foundation of China (No. 91529302, No. 81572798), and The National Key Technology R&D Program of China (No. 2014BAI09B03).

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Authors and Affiliations

  1. Shanghai Key Laboratory of Gastric Neoplasms, Department of Surgery, Shanghai Institute of Digestive Surgery, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200025, People’s Republic of China

    Li Pang, Jian-fang Li, Liping Su, Mingde Zang, Zhiyuan Fan, Beiqin Yu, Xiongyan Wu, Chen Li, Min Yan, Zheng-gang Zhu & Bingya Liu

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  1. Li Pang
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Correspondence to Bingya Liu.

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Pang, L., Li, Jf., Su, L. et al. ALEX1, a novel tumor suppressor gene, inhibits gastric cancer metastasis via the PAR-1/Rho GTPase signaling pathway. J Gastroenterol 53, 71–83 (2018). https://doi.org/10.1007/s00535-017-1329-y

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  • DOI: https://doi.org/10.1007/s00535-017-1329-y

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