PRCC靶向激活JNK促进肝癌干细胞自我更新加速肿瘤进展的作用和机制研究

Hepatocellular carcinoma (HCC) is characterized with high malignant and poor prognosis. The specific changes of genome, proteome, metabolome and epigenetics in HCC cells lead to cell excessive proliferation and malignant transformation. In previous study, we paid close attention to PRCC by screening TCGA database of HCC. Based on our preliminary research, we found PRCC was highly expressed in hepatocellular carcinoma tissue and the serum of HCC patients. The overexpression of PRCC promoted HCC cell proliferation, invasion and chemoradiotherapy resistance, up-regulated the protein level of OCT4, CD133, CD44, EpCAM and p-JNK, along with the Hep3B mammosphere forming efficiency increased. The detail mechanisms are only partially understood. In consideration of so many stem cell markers being altered, we made a hypothesis that PRCC was associated with liver cancer stem cell self-renewal and transformation. This project will study the clinical significance of high level PRCC in hepatoma tissue and serum of HCC patients through statistical analysis; Observe the influence of PRCC on the proliferation, invasion, chemoradiotherapy resistance and tumor growth in vivo and in vitro through cytology and animal model research. This research will reveal up- and down-stream regulatory mechanism of PRCC, especially its function through JNK signaling pathway; Uncover the definite signaling pathways and molecules it acts on, and illuminate the mechanism of self-renewal and transformation of liver cancer stem cells. This study will explore the potency of PRCC as a liver cancer diagnosis and prognosis predictive factor, and provide scientific basis for its clinical application as an effective therapeutic target for HCC patients with radiation and chemotherapy resistance.

肝癌恶性度高、预后差，其基因组、蛋白质组、代谢组和表观遗传的特异变化，引发癌细胞失控性增殖生长。我们前期利用TCGA数据库筛选出在肝癌组织和血液中高表达的PRCC基因进行研究，发现PRCC能促进肝癌细胞的增殖、侵袭、对抗失巢凋亡及诱发放化疗耐受，提高癌细胞成球率，上调OCT4、CD133、CD44、EpCAM的表达，提高JNK磷酸化水平，但具体机制不明。由于上述分子均为干细胞标志物，因此我们认为PRCC可能与肿瘤干细胞的更新活化有关。本课题将通过临床样本分析明确PRCC在肝癌组织及血液中高水平存在的作用和意义，通过细胞学和动物模型研究PRCC对癌细胞增殖转移、放化疗耐受、体内成瘤及肿瘤生长的影响，研究PRCC在肝癌中表达的上下游信号通路的调控，特别是JNK通路，从而阐明PRCC对肝癌干细胞转化及自我更新的作用及机制。本研究为PRCC作为肝癌诊断预后标志物以及放化疗耐受患者治疗靶点提供依据。

富含脯氨酸的有丝分裂检验点控制因子PRCC是参与pre-mRNA剪切过程的蛋白元件。本研究通过临床样本分析明确PRCC在肝癌组织及血液中高水平存在的作用和意义，通过细胞学和动物模型研究PRCC对癌细胞增殖转移、放化疗耐受的影响，研究PRCC在肝癌中表达所影响的信号通路，特别是JNK通路，从而阐明PRCC作为肝癌诊断预后标志物以及放化疗耐受患者治疗靶点的重要意义。得到如下重要结果：（1）通过组织蛋白及组织芯片分析，发现PRCC在肝癌患者的癌组织中高表达且与患者不良预后相关；（2）采用ELISA检测发现PRCC在血清中可检测到，且它作为肝癌诊断指标的敏感性及特异性都很高，可作为肝癌血清检测的候选指标；（3）PRCC高表达对肝癌细胞的增殖无显著影响，但可抑制肝癌细胞克隆形成及成球能力；（4）PRCC高表达在体外可抑制肝癌细胞的迁移及侵袭能力；（5）在体内，采用裸鼠尾静脉细胞注射模型也验证了PRCC高表达可抑制肝癌细胞的转移及成瘤性；（6）采用UV诱导DNA损伤及顺铂处理，发现PRCC高表达的肝癌细胞对UV及顺铂处理不敏感，处理后的细胞周期检测结果显示PRCC高表达组的G2/M期细胞比例减少，S期细胞比例增加；（7）通过对TCGA共表达数据分析，发现PRCC与DNA损伤修复过程的关键因子表达呈负相关，Western blot 进一步证实PRCC是通过抑制JNK/ATM/ATR/ATF2轴降低肝癌细胞的DNA损伤敏感性并增强其异质性。这些结果清晰地阐释了PRCC在肝癌发生发展中所扮演的角色。为肝癌诊断、治疗指导、预后预测及靶向治疗提供了新线索及新思路。