VASP在缺氧肿瘤微环境中调控肝细胞癌侵袭转移的机制研究

Postoperative recurrence and metastasis of hepatocellular carcinoma (HCC) are the leading cause for the patients with poor prognosis. Hypoxia-related tumor microenvironment is the main reason for the invasion and metastasis of HCC. Our prior clinical study demonstrated that tumors with high VASP expression levels indeed associated with poor overall survival and recurrence-free survival of HCC patients (HEPATOLOGY, 2015). However, the molecular mechanisms are poorly investigated. Our previous studies found that hypoxia induced VASP overexpression in HCC cells, while two hypoxia inducible factor (HIF)-binded hypoxia response elements were found in the promoter of VASP. Otherwise, VASP overexpression induced EMT and subsequently promoted migration and invasion of HCC cells. Based on our previous studies, clinical and biological experiments will be performed to verify that VASP overexpression is a independent prognostic factor for predicting poor prognosis of HCC patients. Hypoxia-related tumor microenvironment directly up-regulates VASP expression and indirectly increases VASP expression via regulating TGFβ/SMAD pathway and miR-20b. VASP promotes EMT by regulating actin cytoskeleton dynamics and accordingly facilitates HCC cell migration and invasion. Meanwhile, the anti-HCC effect of the tumor-selective VASP shRNA-expressing adenovirus will be investigated in a nude mice model. Altogether, our project is aimed to provide the basis for the development of new HCC targeted therapy drugs.

肝细胞癌（HCC）术后复发和转移是患者预后差的主要原因，缺氧肿瘤微环境是导致HCC侵袭转移的关键因素。课题组前期临床研究证实VASP过表达促进HCC术后复发和转移（HEPATOLOGY, 2015），但分子机制不明。我们前期实验发现缺氧诱导HCC细胞过表达VASP，VASP启动子区域存在两个HIF结合的缺氧反应元件，VASP过表达促进HCC细胞发生EMT及迁移和侵袭。本项目是既往工作的深入，拟通过临床标本和分子生物学实验证实：VASP过表达是HCC患者预后不良的独立预测因素；缺氧肿瘤微环境中HIF直接和通过调控TGFβ/SMAD通路及miR-20b间接上调VASP表达，VASP通过调控肌动蛋白细胞骨架动力学促进HCC细胞发生EMT，从而促进细胞迁移和侵袭。同时，本项目试图研究荷载VASP shRNA的肿瘤特异性增殖腺病毒在动物体内的抗HCC作用，为临床上开发新的HCC靶向治疗药物提供依据。

肝细胞癌是我国最为常见的恶性肿瘤之一，约占原发性肝癌的90%。其恶性程度高，侵袭转移能力强，易复发等因素导致肝癌患者预后较差。尽管肝癌的早期筛查逐步开展，诊断手段、手术技术以及综合治疗策略不断发展，患者的临床预后依然不佳。缺氧肿瘤微环境是多种恶性肿瘤的共同特征，是导致肝癌侵袭转移的关键因素。课题组前期临床研究证实VASP过表达促进肝癌术后复发和转移，但分子机制不明。本实验的目的在于探讨VASP在肝癌中的表达、临床意义、生物学功能及潜在分子机制，探究缺氧微环境对肝癌中VASP表达的调控机制，为寻找肝癌新的生物学标志物和潜在治疗靶点奠定基础。本项目结果证实：1. VASP在肝癌中存在显著高表达；VASP高表达与肝癌患者恶性病理特征及不良临床预后相关，VASP可作为肝癌诊断及预后评估的生物学标志物。2. VASP可结合CRKL SH3N结构域，激活细胞内的AKT和ERK通路，进而增强肝癌细胞在体外及体内的侵袭转移能力。缺氧微环境可导致肝癌中VASP的表达增加，缺氧可直接通过HIF-1α结合到VASP启动子区域促进转录；降低细胞内miR-204的水平；激活TGF-β/Smad通路，导致VASP表达水平增加。本项目旨在阐明VASP在肝癌中的分子机制，为临床开发新的肝癌靶向治疗药物和预后标志物奠定理论基础。