한빛사논문
Hye Ri Ahn1,2, Geum Ok Baek1, Moon Gyeong Yoon1, Ju A Son1,2, Jung Hwan Yoon3, Jae Youn Cheong1, Hyo Jung Cho1, Ho Chul Kang2,4, Jung Woo Eun1* and Soon Sun Kim1*
1Department of Gastroenterology, Ajou University School of Medicine, Worldcup-ro 164, Yeongtong-Gu, Suwon 16499, Republic of Korea. 2Department of Biomedical Sciences, Ajou University Graduate School of Medicine, Worldcup-ro 164, Yeongtong-Gu, Suwon 16499, Republic of Korea. 3Department of Pathology, College of Medicine, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul 06591, Republic of Korea. 4Department of Physiology, Ajou University Graduate School of Medicine, Worldcup-ro 164, Yeongtong-Gu, Suwon 16499, Republic of Korea.
*Corresponding author.
Abstract
Background
Hepatocellular carcinoma (HCC) is one of the most common and lethal cancers worldwide. Wiskott–Aldrich syndrome protein family member 2 (WASF2) is an integral member of the actin cytoskeleton pathway, which plays a crucial role in cell motility. In this study, we aimed to explore the role of WASF2 in HCC carcinogenesis and its regulatory mechanism.
Methods
WASF2 expression in HCC was analyzed using six public RNA-seq datasets and 66 paired tissues from patients with HCC. The role of WASF2 in normal hepatocyte cell phenotypes was evaluated using a WASF2 overexpression vector in vitro; it was evaluated in HCC cell phenotypes using small interfering RNA (siRNA) in vitro and in vivo. Epigenetic regulatory mechanism of WASF2 was assessed in the Cancer Genome Atlas liver hepatocellular carcinoma project (TCGA_LIHC) dataset and also validated in 38 paired HCC tissues. Site mutagenesis, bisulfite sequencing polymerase chain reaction (BSP), methylation-specific polymerase chain reaction (MSP), and quantitative MSP (qMSP) were used for evaluating WASF2 methylation status.
Results
WASF2 is overexpressed in HCC and is clinically correlated with its prognosis. WASF2 overexpression promoted normal hepatocyte proliferation. WASF2 inactivation decreased the viability, growth, proliferation, migration, and invasion of Huh-7 and SNU475 HCC cells by inducing G2/M phase arrest. This induced cell death and inhibited epithelial–mesenchymal transition, hindering actin polymerization. In addition, WASF2 knockdown using siWASF2 in a xenograft mouse model and a lung metastasis model exerted tumor suppressive effect. There was a negative correlation between WASF2 methylation status and mRNA expression. The methylation pattern of CpG site 2 (− 726 bp), located in the WASF2 promoter, plays an important role in the regulation of WASF2 expression. Furthermore, the cg242579 CpG island in the WASF2 5′ promoter region was hypomethylated in HCC compared to that in the matched non-tumor samples. Patients with high WASF2 methylation and low WASF2 expression displayed the highest overall survival.
Conclusions
WASF2 is overexpressed and hypomethylated in HCC and correlates with patient prognosis. WASF2 inactivation exerts anti-tumorigenic effects on HCC cells in vitro and in vivo, suggesting that WASF2 could be a potential therapeutic target for HCC.
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