TRAIL-CTP融合长效蛋白的表达筛选及生物活性分析

TRAIL能诱导多种肿瘤细胞凋亡，具有较好的有效性和安全性，是理想的抗肿瘤分子靶向生物靶点。但TRAIL易被肾小球清除，体内半衰期短，延长TRAIL生物半衰期能增强其治疗效能。CTP融合蛋白是近年兴起的一种新型蛋白长效化技术，CTP为内源性富含糖基化序列，该片段不增加蛋白质抗原性、不影响受体亲和力及蛋白生物活性，已成功用于长效蛋白开发。本项目拟构建TRAIL与CTP不同拷贝基因连接的融合蛋白真核表达载体，以期改善TRAIL药代动力学特性，实现TRAIL蛋白的长效表达，延长TRAIL在体内的半衰期，增强TRAIL的抗肿瘤疗效。本项目通过对真核表达载体加压筛选得到高表达细胞株，蛋白质纯化得到高纯度蛋白，在荷瘤小鼠模型和体外肿瘤细胞株中观察TRAIL-CTP融合蛋白抗肿瘤生物活性，通过药代动力学实验检测该融合蛋白半衰期，探索一种实现TRAIL蛋白长效化的可靠手段。

TRAIL could induce apoptosis in many kinds of cancer cells with preferable efficacy and safety which makes it an ideal target of cancer treatment. However, TRAIL is prone to be eradicated by glomerulus and the half-life period is short. So the elongation of its biological half-life could increase the treatment efficacy. CTP fusion protein is an emerging technology that prolongs the existence of proteins. CTP is an endogenous rich in glycosylation sequence. It didn't increase antigenicity or influence the receptor affinity and protein bioactivity. It has been successfully used in the development of long-term protein. This study tries to construct fusion protein eukaryotic expression vector of TRAIL and CTP in order to improve the pharmacokinetic characteristics and antitumor effects of TRAIL. First, we screen for high expression cell lines in eukaryotic expression vectors, then the purification of protein, and last, to observe the antitumor efficacy of TRAIL-CTP fusion protein. We intend to find a reliable method to make TRAIL long acting.