肿瘤血管靶向性凝血蛋白对原位肝癌动物模型的抗肿瘤活性与机理

Transcatheter arterial chemoembolization is the most effective method for the non operative treatment of liver cancer, but its long-term curative effect is poor, and there are defects such as incomplete embolization, lack of targeting, poor controllability, and large toxic and side effects. In order to overcome the above shortcomings, this study aims to develop a new therapeutic strategy for the liver cancer vascular embolization with the tumor vascular targeted blood coagulation proteins. we will observe the activity of vascular thrombosis and molecular targeting characteristics of the tumor vascular targeted blood coagulation proteins in orthotopic liver transplantation model, study the molecular mechanism of the their induced coagulation reactions and the main affecting factors, establish a new treatment model for the liver cancer vascular embolization, clarify the therapeutic mechanism, play theoretical basis and new ideas for the novel hepatocellular carcinoma vascular embolization agent development and application.

肝动脉化疗性栓塞是目前肝癌非手术治疗最有效的方法，但其远期疗效差，并存在栓塞不全、缺乏靶向性、可控性差、毒副作用大等缺陷。为克服以上不足，本课题拟利用自主研发的肿瘤血管靶向性凝血蛋白，发展一种新型肝癌血管栓塞的治疗策略。用原位移植性肝癌模型，比较研究不同肿瘤血管靶向性凝血蛋白的血管栓塞活性、分子靶向特性、诱发凝血反应的分子机制及其主要影响因素，探讨靶向性凝血蛋白治疗肝癌的模式，阐明其治疗机理，为新型肝癌血管栓塞剂研制与应用提供理论基础与新思路。

肝动脉化疗性栓塞是目前肝癌非手术治疗的有效方法，传统栓塞剂存在栓塞不全、缺乏靶向性、可控性差、毒副作用大等缺点。为克服以上问题，本研究制备了三种肿瘤血管靶向性凝血蛋白（tTF-EG3287、tTF-EG3287/CMC/Fe3O4和GNR@VSNP-tTF-EG3287），并对它们的理化特性进行鉴定。共聚焦荧光显微镜和流式细胞仪分析显示三种凝血蛋白均能靶向结合Neuroplin-1（NRP1）分子，并且具有较高的亲和力；凝血因子X活化试验显示它们均保持了Xa的活化能力。凝血反应影响因素实验显示tTF-EG3287促凝血活性受钙离子浓度、磷脂 浓度和温度变化的影响；改变这些因素可以调控凝血反应和血栓程度。荧光成像仪分析Cy5.5-tTF-EG3287在原位肝癌小鼠模型主要器官中的生物学分布，结果显示tTF-EG3287能够富集于肝脏肿瘤组织部位。利用HepG2皮下移植瘤模型探讨tTF-EG3287的使用剂量和方案，结果显示tTF-EG3287的抗肿瘤效果与药物的剂量、给药次数和疗程密切相关。静脉注射tTF-EG3287（ 2.5-5mg/kg，qdX4）对原位肝癌模型有显著的治疗效果。tTF-EG3287诱发肿瘤血管血栓栓塞能力与其凝血活性、靶向能力和在靶点的富集效率等有关。具有磁靶向的tTF-EG3287@OCMC/Fe3O4和具有光热效应的GNR@VSNP-tTF-EG3287纳米颗粒可以提高tTF-EG3287在肿瘤组织的靶向富集、诱发更强的血管血栓栓塞、产生更强的抗肿瘤作用。组织病理学分析未见tTF-EG3287融合蛋白有明显的毒性作用。结论:tTF-EG3287是一种新的肝癌血管靶向血栓栓塞剂，具有良好的靶向性和肿瘤血管栓塞作用，其抗肿瘤作用主要与其诱发肿瘤血管栓塞、肿瘤组织缺血、细胞变性坏死有关