基于MAP3K1突变的肝内胆管癌分子分型及其机制研究

Several recent reports have described rapidly increasing in the incidence and mortality of ICC with geographic variation. The lack of effective chemotherapies and molecular targeted therapies caused by poor molecular classification and pathobiological understanding of ICC should be responsible for this dismal situation. Hence, a better understanding of molecular classification of ICC is urgently needed. Previously, we have performed whole-exome sequencing of ICC tumors and matched normal tissues to detect somatic alterations and found that MAP3K1 had recurrent somatic mutations（2/7）and might be a drive gene in tumorigenesis and development of ICC. In the preliminary test, we sequenced all the exons of MAP3K1 in a cohort of 124 ICC patients and firstly identified somatic mutations in 50% ICC patients, more than 31.5% of TP53. In parallel, high expression of MAP3K1 promoted invasion and metastasis of ICC. Some of MAP3K1 mutations had a close relationship with ICC prognosis. Therefore, we propose a hypothesis: mutation of MAP3K1 can be used as the molecular classification basis and prognostic label of ICC. We are aimed to clarify the hypothesis through construction of mutant strains and wild strain of MAP3K1. Further, we compare the differences in cell function and molecular phenotype and potential molecular mechanisms between mutant strains and wild strain through the aid of molecular and cell biology techniques. Last, we find out the specific mutation used as the molecular classification basis、personalized therapeutic targets and prognostic label of ICC. Starting from MAP3K1 mutation, this study lay a foundation for the development of personalized therapy of ICC and provid a new mentality to improve the survival rates of the ICC.

肝内胆管癌(ICC)的发病率和死亡率逐年上升,主要原因是其对现有化疗药物的高度抵抗和缺乏分子分型而导致有效的靶向治疗药物的缺失。因此，探索ICC的分子分型成为迫在眉睫的问题。我们前期发现MAP3K1在ICC中存在频发突变(2/7)，可能是ICC发生发展的驱动基因。申请人预实验大样本验证首次发现MAP3K1在ICC中突变率高达50%，超过TP53的31.5%； MAP3K1在癌内高表达促进ICC侵袭转移；且突变与ICC临床特征和预后关系密切。为此，我们提出假说：MAP3K1突变可以作为ICC分子分型及预后判断标签。为验证这一假说，我们构建MAP3K1突变株和野生株，运用分子和细胞生物学技术，比较两者在细胞功能和分子表型上差别及潜在分子机制，明确可作为ICC分子分型及预后判断标签的具体突变位点。本研究从MAP3K1突变出发，为实现ICC个性化治疗奠定基础，也为提高ICC的生存率提供新的思路。

肝内胆管癌（ICC）发病率居肝脏原发性肿瘤的第二位，仅次于肝细胞癌。由于缺乏特异性临床表现，早期诊断困难，疗效差，近30年来ICC的5年总生存率一直<5％。在全球范围内，ICC的发病率和死亡率正在逐年增高，主要原因是其对现有化疗药物的高度抵抗和缺乏分子分型而导致有效的靶向治疗药物的缺失。MAP3K1是MAPK信号通路的重要组成成分，既往研究发现其拷贝数改变和体细胞突变在乳腺癌、肺癌、卵巢癌，前列腺癌、子宫癌、结肠癌等多种肿瘤基因组数据中占重要部分,但是它在ICC中的表达特点、功能状况、基因突变情况以及临床相关性和对预后的影响仍缺乏系统全面的分析研究。本项目通过对上述内容的研究发现：① MAP3K1在ICC癌组织内高表达，且与ICC患者的淋巴结转移及更差的TNM分期显著相关，具有促进ICC细胞克隆形成和增殖侵袭的能力；高表达MAP3K1的患者较低表达者预后更差（P=0.001）及复发率更多（P=0.026）。② MAP3K1在ICC癌组织中存在高频突变（50%），其部分突变株可以增强ICC细胞生长、侵袭迁移等的能力，以S637I位点突变最为显著。③具有S637I点突变表型的ICC患者更易表现出高CA199和淋巴结转移的临床特性。这些结果为后续以MAP3K1突变为基础，对ICC进行分子分型及预后判断提供了可能性，也给以MAP3K1为靶点的靶向治疗提供了新的方向。