Genomic and Epigenomic Features of Primary and Recurrent Hepatocellular Carcinomas
Genomic and Epigenomic Features of Primary and Recurrent Hepatocellular Carcinomas
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DOI：
10.1053/j.gastro.2019.09.005
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发表时间：
2019-12-01
期刊：
影响因子：
29.4
通讯作者：
Wong, Nathalie
中科院分区：
文献类型：
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作者：
Ding, Xiaofan;He, Mian;Wong, Nathalie
背景与目的：原发性和复发性肝细胞癌（HCC）内和之间的肿瘤异质性和不同的克隆谱系对患者管理产生了挑战。我们研究了肝脏肿瘤内、肝脏病变间以及原发性和复发性肿瘤间的遗传和表观遗传变异。方法：肿瘤和匹配的非肿瘤肝脏标本收集自113例在香港2家医院接受原发性或复发性HCC部分肝切除术的患者。我们对从多个肿瘤区域收集的356例HCC标本进行了全基因组、全外显子组或靶向捕获测序分析，并匹配了初始和复发肿瘤。我们对95例标本进行了平行的DNA甲基化分析。分析了含有肝硬化或纤维化区域的非肿瘤组织的基因组和表观基因组。我们开发了内源性表达TP53（R249S）突变形式或过表达STAT3（D170 Y，K348 E和Y640 F）或JAK 1（S703 I和L910 P）突变形式的肝癌细胞系，并测试了药物降低活性的能力。通过免疫印迹和染色质免疫沉淀与定量聚合酶链反应分析细胞。研究结果：我们使用单一样本收集方法确定了单个肿瘤的单克隆起源，该方法捕获了在肿瘤所有区域检测到的超过90%的突变。系统发育和表观遗传分析揭示了HCC的基因组和表观基因组特征之间的相互作用和相互依赖。甲基化分析揭示了在肝硬化肝组织中的场效应，使其易于发生肿瘤。遗传特征的比较显示，52%的复发性HCC来自初始肿瘤的克隆谱系。如果复发性肝癌的克隆起源允许构建与肿瘤复发相关的遗传改变的时间图。JAK信号转导激活STAT是HCC进展的特征，通过与药物敏感性反应相关的突变。增加TP53功能的突变和17p染色体缺失的组合可能为肝癌细胞提供复制优势。染色质免疫沉淀分析TP53与R249S取代揭示了其与编码染色质调节因子（MLL1和MLL2）的基因的相互作用。我们验证了MLL1和MLL2作为TP53（R249S）的直接靶点，并在癌症基因组图谱数据集中确认了它们的关联。MLL复合物拮抗剂MI-2 - 2（蛋白质相互作用抑制剂）和OICR-9492（活性抑制剂）在纳摩尔浓度下特异性抑制表达TP53（R249S）的HCC细胞的增殖。结论：我们对HCC样本中肿瘤内和肿瘤间的遗传异质性进行了系统评价，并确定了与肿瘤进展和复发相关的遗传和表观遗传变化。我们鉴定了在HCC细胞中由突变型TP53上调的染色质调节剂和减少这些细胞增殖的抑制剂。肝硬化或肝纤维化组织中的DNA甲基化模式可用于识别HCC发展的风险。
BACKGROUND & AIMS: Intratumor heterogeneity and divergent clonal lineages within and among primary and recurrent hepatocellular carcinomas (HCCs) produce challenges to patient management. We investigated genetic and epigenetic variations within liver tumors, among hepatic lesions, and between primary and relapsing tumors. METHODS: Tumor and matched nontumor liver specimens were collected from 113 patients who underwent partial hepatectomy for primary or recurrent HCC at 2 hospitals in Hong Kong. We performed whole-genome, whole-exome, or targeted capture sequencing analyses of 356 HCC specimens collected from multiple tumor regions and matched initial and recurrent tumors. We performed parallel DNA methylation profiling analyses of 95 specimens. Genomes and epigenomes of nontumor tissues that contained areas of cirrhosis or fibrosis were analyzed. We developed liver cancer cell lines that endogenously expressed a mutant form of TP53 (R249S) or overexpressed mutant forms of STAT3 (D170Y, K348E, and Y640F) or JAK1 (S703I and L910P) and tested the abilities of pharmacologic agents to reduce activity. Cells were analyzed by immunoblotting and chromatin immunoprecipitation with quantitative polymerase chain reaction. RESULTS: We determined the monoclonal origins of individual tumors using a single sample collection approach that captured more than 90% of mutations that are detected in all regions of tumors. Phylogenetic and phyloepigenetic analyses revealed interactions and codependence between the genomic and epigenomic features of HCCs. Methylation analysis revealed a field effect in cirrhotic liver tissues that predisposes them to tumor development. Comparisons of genetic features revealed that 52% of recurrent HCCs derive from the clonal lineage of the initial tumor. The clonal origin if recurrent HCCs allowed construction of a temporal map of genetic alterations that associated with tumor recurrence. Activation of JAK signaling to STAT was a characteristic of HCC progression via mutations that associate with response to drug sensitivity. The combination of a mutation that increases the function of TP53 and the 17p chromosome deletion might provide liver cancer cells with a replicative advantage. Chromatin immunoprecipitation analysis of TP53 with the R249S substitution revealed its interaction with genes that encode chromatin regulators (MLL1 and MLL2). We validated MLL1 and MLL2 as direct targets of TP53(R249S) and affirmed their association in the Cancer Genome Atlas dataset. The MLL-complex antagonists MI-2-2 (inhibitor of protein interaction) and OICR-9492 (inhibitor of activity) specifically inhibited proliferation of HCC cells that express TP53(R249S) at nanomolar concentrations. CONCLUSIONS: We performed a systematic evaluation of intra- and intertumor genetic heterogeneity in HCC samples and identified genetic and epigenetic changes that associate with tumor progression and recurrence. We identified chromatin regulators that are upregulated by mutant TP53 in HCC cells and inhibitors that reduce proliferation of these cells. DNA methylation patterns in cirrhotic or fibrotic liver tissues might be used to identify those at risk of HCC development.