靶向肿瘤微环境SYK抑制肝癌复发转移的机制研究

The tumor microenvironment (TME) underlying liver fibrosis plays a key role in the recurrence and metastasis of hepatocellular carcinoma (HCC). ..In our previous study, we found that up-regulated SYK expression levels were associated with HBV or HCV infection in human liver biopsy. SYK promoted the activation of hepatic stellate cells (HSCs), and the progression of liver fibrosis and HCC formation in vivo. SYK inhibitor GS-9973 significantly reduced HSC and M2 macrophage activation in liver. Importantly, SYK antagonist treatment significantly reduced HCC invasion and metastasis in the orthtopic xenograft with liver fibrosis mouse compared to orthotopic xenograft normal mouse. We hypothesized that targeting SYK of tumor microenvironment as an effective therapy to attenuate tumor recurrence and metastasis of HCC. .. We plan to test SYK inhibitor in orthtopic xenograft mouse with liver fibrosis to investigate the effects of SYK inhibition on HCC recurrence and metastasis. We will isolate the following primary liver cell types including hepatocytes, HSCs, Kuffer and liver sinusoidalendothelial cells from normal or liver fibrosis mouse. We plan to perform RNA-sequencing, protein mass spectrometric analysis and other technologies to explore the mechanism by which SYK-mediation in tumor development via tumor microenvironment. In addition, we plan to monitor the anti-recurrence/metastasis effects of SYK-targeting therapy in the patient-derived HCC xenograft mouse model. .. The study in this grant proposal will help to elucidate the mechanism of SYK-mediated tumor microenvironment and the anti-recurrence/metastasis effects of SYK-targeted therapy. Our findings may achieve significant progress in clinical and translational settings for HCC treatment.

肝纤维化背景下的肿瘤微环境（TME）在肝癌复发转移中扮演重要角色。前期研究揭示：HBV或HCV等相关肝纤维化中肝星状细胞（HSC）SYK表达上调；SYK促进HSC激活，加速肝纤维化进程及肝癌形成；靶向SYK可显著抑制HSC激活，减少M2型巨噬细胞产生。最近对TME中SYK研究取得新进展：靶向SYK可抑制小鼠肝纤维化基础上肝原位移植瘤的侵袭转移，但对正常小鼠肝原位移植瘤效果不明显。据此推测：靶向SYK通过TME抑制肝癌复发转移。本项目采用肝纤维化基础上的肝原位移植瘤模型，研究靶向SYK对肿瘤复发转移的影响；利用肝脏原代细胞分离、RNA测序、蛋白质谱等技术，探讨SYK通过TME影响肝癌复发转移的调控机制；选用人源肝癌组织异种移植模型进行抗肿瘤治疗，验证SYK靶向治疗的临床价值。本研究旨在探讨SYK靶向治疗对肝癌复发转移的疗效，并阐明其对TME的调控机制，具有重要的临床转化应用价值。

慢性肝脏疾病诱导的炎症微环境是肝细胞癌（Hepatocellular carcinoma，以下简称HCC或肝癌）发生的主要诱因，但炎症微环境对肝癌复发转移的影响尚不明确。脾酪氨酸激酶（Spleen tyrosine kinase, SYK）是非受体酪氨酸激酶Src家族的一员，在多种自身炎症免疫相关性疾病中发挥重要作用。我们前期研究发现肝脏SYK促进了炎症诱导的肝癌发生。本项目拟进一步探讨SYK介导的肝脏炎症微环境对肝癌进展的影响及潜在机制，并在此基础上探索靶向SYK对肝癌进展的影响及其治疗价值。..研究发现：在伴有炎症及纤维化的小鼠肝癌原位移植瘤中，肝癌进展与炎症水平呈正相关（r=0.626, P=0.001），而与纤维化严重程度的相关性不明显（r=0.294, P=0.163）。在上述模型中，发现在癌旁炎症微环境中SYK表达显著升高。对158例肝癌患者分析，发现非肿瘤肝组织SYK高表达患者的肝脏炎症相关病理特征明显、生存期显著缩短。构建肝细胞特异性SYK敲除转基因小鼠，发现肝细胞SYK敲除可显著抑制炎症基础上的小鼠肝癌进展。体外实验提示：SYK可通过增强衔接蛋白ASC寡聚化促进肝细胞NLRP3 (NLR Family Pyrin Domain Containing 3)炎症小体激活，诱发肝细胞焦亡及促瘤肝脏炎症微环境形成。在多种伴有肝脏炎症微环境的小鼠肝癌原位移植瘤模型中，SYK抑制剂GS-9973单药或联合肝癌一线靶向药物（Sorafenib或Regorafenib），均可显著抑制肿瘤进展，并延长小鼠生存期。..结论：在慢性肝损伤因素作用下，肝细胞SYK促进衔接蛋白ASC寡聚化和NLRP3炎症小体激活，引起肝细胞焦亡并诱导促瘤肝脏炎症微环境形成。靶向SYK可通过改善肝脏炎症微环境抑制肝癌进展，并可提高临床肝癌一线靶向药物的疗效。