肝内胆管癌(ICC)是一种致命的癌症,尤其是在其扩散时,生存率较差。其罕见的胆管型导管内乳头状肿瘤(IPNB)的组织病理学特征是癌细胞起源于狭窄的胆管空间。这些细胞最终侵入并渗入附近的肝脏组织,使其成为研究局部入侵机制的良好模型,这是转移的最早步骤。为了发现ICC局部入侵的潜在抑制基因,我们分析了来自同一患者的11对有局部侵袭的大解剖IPNB组织(LI-IPNB)和无局部侵袭的IPNB组织的体细胞突变谱,并进行了克隆进化分析.我们在E3泛素连接酶中鉴定了一种蛋白质截短变体(PTV),RNF213(c.6967C>T;p.Gln2323X;chr17:78,319,102[hg19],外显子29),作为LI-IPNB样本中最常见的PTV事件(4/11患者)。敲除HuCCT1和YSCCC细胞中的RNF213显示增加的迁移和侵袭,减少血管生成拟态,但保持了正常的增殖。然后在HuCCT1,YSCCC,和KKU-100细胞。常见差异表达基因的基因本体论(GO)富集分析显示,细胞因子和氧化还原酶-氧化金属离子活性显着改变,正如西方印迹所证实的。基因集富集分析(GSEA)确定了最富集的途径是氧化磷酸化,脂肪酸代谢,活性氧,脂肪生成,和血管生成。总之,RNF213的功能丧失是LI-IPNB组织中常见的遗传改变。RNF213敲低导致ICC细胞的迁移和侵袭增加,可能通过与炎症相关的途径的故障,和能量代谢。
Intrahepatic cholangiocarcinoma (ICC) is a lethal cancer with poor survival especially when it spreads. The histopathology of its rare intraductal papillary neoplasm of the bile duct type (IPNB) characteristically shows cancer cells originating within the confined bile duct space. These cells eventually invade and infiltrate the nearby liver tissues, making it a good model to study the mechanism of local invasion, which is the earliest step of metastasis. To discover potential suppressor genes of local invasion in ICC, we analyzed the somatic mutation profiles and performed clonal evolution analyses of the 11 pairs of macrodissected locally invasive IPNB tissues (LI-IPNB) and IPNB tissues without local invasion from the same patients. We identified a protein-truncating variant in an E3 ubiquitin ligase, RNF213 (c.6967C>T; p.Gln2323X; chr17: 78,319,102 [hg19], exon 29), as the most common protein-truncating variant event in LI-IPNB samples (4/11 patients). Knockdown of RNF213 in HuCCT1 and YSCCC cells showed increased migration and invasion, and reduced vasculogenic mimicry but maintained normal proliferation. Transcriptomic analysis of the RNF213-knockdown vs control cells was then performed in the HuCCT1, YSCCC, and KKU-100 cells. Gene ontology enrichment analysis of the common differentially expressed genes revealed significantly altered cytokine and oxidoreductase-oxidizing metal ion activities, as confirmed by Western blotting. Gene Set Enrichment Analysis identified the most enriched pathways being oxidative phosphorylation, fatty acid metabolism, reactive oxygen species, adipogenesis, and angiogenesis. In sum, loss-of-function mutation of RNF213 is a common genetic alteration in LI-IPNB tissues. RNF213 knockdown leads to increased migration and invasion of ICC cells, potentially through malfunctions of the pathways related to inflammation and energy metabolisms.