Background & Aims

Development of hepatocellular carcinoma (HCC) is associated with alterations in the transforming growth factor-beta (TGF-β) signaling pathway, which regulates liver inflammation and can have tumor suppressor or promoter activities. Little is known about the roles of specific members of this pathway at specific of HCC development. We took an integrated approach to identify and validate the effects of changes in this pathway in HCC and identify therapeutic targets.

Methods

We performed transcriptome analyses for a total of 488 HCCs that include data from The Cancer Genome Atlas. We also screened 301 HCCs reported in the Catalogue of Somatic Mutations in Cancer and 202 from Cancer Genome Atlas for mutations in genome sequences. We expressed mutant forms of spectrin beta, non-erythrocytic 1 (SPTBN1) in HepG2, SNU398, and SNU475 cells and measured phosphorylation, nuclear translocation, and transcriptional activity of SMAD family member 3 (SMAD3).

Results

We found somatic mutations in at least 1 gene whose product is a member of TGF-β signaling pathway in 38% of HCC samples. SPTBN1 was mutated in the largest proportion of samples (12 of 202, 6%). Unsupervised clustering of transcriptome data identified a group of HCCs with activation of the TGF-β signaling pathway (increased transcription of genes in the pathway) and a group of HCCs with inactivation of TGF-β signaling (reduced expression of genes in this pathway). Patients with tumors with inactivation of TGF-β signaling had shorter survival times than patients with tumors with activation of TGF-β signaling (P = .0129). Patterns of TGF-β signaling correlated with activation of the DNA damage response and sirtuin signaling pathways. HepG2, SNU398, and SNU475 cells that expressed the D1089Y mutant or with knockdown of SPTBN1 had increased sensitivity to DNA crosslinking agents and reduced survival compared with cells that expressed normal SPTBN1 (controls).

Conclusions

In genome and transcriptome analyses of HCC samples, we found mutations in genes in the TGF-β signaling pathway in almost 40% of samples. These correlated with changes in expression of genes in the pathways; up-regulation of genes in this pathway would contribute to inflammation and fibrosis, whereas down-regulation would indicate loss of TGF-β tumor suppressor activity. Our findings indicate that therapeutic agents for HCCs can be effective, based on genetic features of the TGF-β pathway; agents that block TGF-β should be used only in patients with specific types of HCCs.

中文翻译：

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肝细胞癌基因组和转录组分析鉴定转化生长因子-β 通路中的突变和基因表达变化

 背景与目标

肝细胞癌（HCC）的发展与转化生长因子-β（TGF-β）信号通路的改变有关，该通路调节肝脏炎症并具有肿瘤抑制或启动子活性。关于该途径的特定成员在 HCC 发展的特定过程中的作用知之甚少。我们采用综合方法来识别和验证该通路变化对 HCC 的影响并确定治疗靶点。

 方法

我们对总共 488 个 HCC 进行了转录组分析，其中包括来自癌症基因组图谱的数据。我们还筛查了癌症体细胞突变目录中报告的 301 个 HCC 和癌症基因组图谱中报告的 202 个 HCC 的基因组序列突变。我们在 HepG2、SNU398 和 SNU475 细胞中表达了血影蛋白 β、非红细胞 1 (SPTBN1) 的突变形式，并测量了 SMAD 家族成员 3 (SMAD3) 的磷酸化、核易位和转录活性。

 结果

我们在 38% 的 HCC 样本中发现至少 1 个基因发生体细胞突变，其产物是 TGF-β 信号通路的成员。 SPTBN1在最大比例的样本中发生突变（202 个样本中的 12 个，占 6%）。无监督转录组数据聚类确定了一组 TGF-β 信号通路激活（该通路中基因转录增加）的 HCC 和一组 TGF-β 信号通路失活（该通路中基因表达减少）的 HCC。 TGF-β 信号失活的肿瘤患者的生存时间比 TGF-β 信号激活的肿瘤患者的生存时间短 ( P = .0129)。 TGF-β 信号传导模式与 DNA 损伤反应和 Sirtuin 信号传导途径的激活相关。与表达正常 SPTBN1 的细胞（对照）相比，表达 D1089Y 突变体或敲低 SPTBN1 的 HepG2、SNU398 和 SNU475 细胞对 DNA 交联剂的敏感性增加，存活率降低。

 结论

在 HCC 样本的基因组和转录组分析中，我们发现近 40% 的样本中 TGF-β 信号通路基因发生突变。这些与通路中基因表达的变化相关；该通路中基因的上调将导致炎症和纤维化，而下调则表明 TGF-β 肿瘤抑制活性丧失。我们的研究结果表明，基于 TGF-β 途径的遗传特征，HCC 的治疗药物可能是有效的；阻断 TGF-β 的药物只能用于患有特定类型 HCC 的患者。