Genome-wide methylation analysis and epigenetic unmasking identify tumor suppressor genes in hepatocellular carcinoma.
Genome-wide methylation analysis and epigenetic unmasking identify tumor suppressor genes in hepatocellular carcinoma.
复制标题
DOI：
10.1053/j.gastro.2013.08.055
复制
发表时间：
2013-12
期刊：
影响因子：
29.4
通讯作者：
Powers S
中科院分区：
文献类型：
--
作者：
Revill K;Wang T;Lachenmayer A;Kojima K;Harrington A;Li J;Hoshida Y;Llovet JM;Powers S
肿瘤抑制基因的表观遗传沉默有助于肝细胞癌（HCC）的发病。为了鉴定HCC中被DNA甲基化沉默的临床相关肿瘤抑制基因，我们将来自人类原发性HCC样本的DNA甲基化数据与表观遗传揭开后基因表达上调的数据相结合。我们使用Illumina human headchip 27K阵列对71例人类HCC样本进行了全基因组甲基化分析；数据与4种肝癌细胞系暴露于逆转DNA甲基化试剂（表观遗传揭膜）后基因再表达的微阵列分析结果相结合。基于原发性HCC的DNA甲基化和表观遗传揭开后细胞系中基因的重新表达，我们确定了13个候选肿瘤抑制基因。随后的验证使我们专注于两种候选基因的功能特征，鞘磷脂二酯酶3 （SMPD3）和神经丝重多肽（NEFH），我们发现它们在HCC中起肿瘤抑制基因的作用。通过诱导构建物稳定转染肝癌细胞系，过量表达SMPD3和NEFH分别使细胞增殖降低50%和20% （SMPD3, P = 0.003， NEFH, P = 0.003）。相反，用小发夹RNA敲低这些基因的表达可促进细胞在体外的侵袭和迁移（SMPD3, P = 0.0001， NEFH, P = 0.022），并增加其皮下注射或原位移植到小鼠体内后形成肿瘤的能力，证实了它们在HCC中作为肿瘤抑制基因的作用。在一个独立的患者队列中，低水平的SMPD3与根治性手术后HCC早期复发相关（P = 0.001；风险比= 3.22；多因素分析95%置信区间：1.6-6.5）。综合基因组分析发现SMPD3和NEFH是HCC中的肿瘤抑制基因。我们提供的证据表明SMPD3是一种有效的肿瘤抑制基因，可以影响肿瘤的侵袭性；SMPD3水平降低是HCC早期复发的独立预后因素。
Epigenetic silencing of tumor suppressor genes contributes to the pathogenesis of hepatocellular carcinoma (HCC). To identify clinically relevant tumor suppressor genes silenced by DNA methylation in HCC, we integrated DNA methylation data from human primary HCC samples with data on up-regulation of gene expression after epigenetic unmasking. We performed genome-wide methylation analysis of 71 human HCC samples using the Illumina Human Beadchip 27K array; data were combined with those from microarray analysis of gene re-expression in 4 liver cancer cell lines after their exposure to reagents that reverse DNA methylation (epigenetic unmasking). Based on DNA methylation in primary HCC and gene re-expression in cell lines after epigenetic unmasking, we identified 13 candidate tumor suppressor genes. Subsequent validation led us to focus on functionally characterizing 2 candidates, sphingomyelin phosphodiesterase 3 (SMPD3) and neurofilament, heavy polypeptide (NEFH), which we found to behave as tumor suppressor genes in HCC. Overexpression of SMPD3 and NEFH by stable transfection of inducible constructs into an HCC cell line reduced cell proliferation by 50% and 20%, respectively (SMPD3, P = .003 and NEFH, P = .003). Conversely, knocking down expression of these genes with small hairpin RNA promoted cell invasion and migration in vitro (SMPD3, P = .0001 and NEFH, P = .022), and increased their ability to form tumors after subcutaneous injection or orthotopic transplantation into mice, confirming their role as tumor suppressor genes in HCC. Low levels of SMPD3 were associated with early recurrence of HCC after curative surgery in an independent patient cohort (P = .001; hazard ratio = 3.22; 95% confidence interval: 1.6–6.5 in multivariate analysis). Integrative genomic analysis identified SMPD3 and NEFH as tumor suppressor genes in HCC. We provide evidence that SMPD3 is a potent tumor suppressor gene that could affect tumor aggressiveness; a reduced level of SMPD3 is an independent prognostic factor for early recurrence of HCC.
登录
查看更多内容
影响因子：
4.2
作者：
Hoshida Y;Toffanin S;Lachenmayer A;Villanueva A;Minguez B;Llovet JM
通讯作者：
Llovet JM
影响因子：
3.7
作者：
Kim MS;Chang X;LeBron C;Nagpal JK;Lee J;Huang Y;Yamashita K;Trink B;Ratovitski EA;Sidransky D
通讯作者：
Sidransky D
影响因子：
8
作者：
Leiblich, A.;Cross, S. S.;Rehman, I.
通讯作者：
Rehman, I.
影响因子：
158.5
作者：
Llovet, Josep M.;Ricci, Sergio;Bruix, Jordi
通讯作者：
Bruix, Jordi
影响因子：
3.7
作者：
Kim, June-Hyung;Kim, Ei-Lyoung;Yoon, Gyesoon
通讯作者：
Yoon, Gyesoon