唾液酸酶NEU1调节纤连蛋白和整合素α5β1的相互作用抑制膀胱癌发展的机制研究

Abnormal high expression of sialic acid (Sia) on cancer cell surface is a distinctive feature associated with malignant properties including invasiveness and metastatic potential. However, the biological function of sialidase (also termed as Neuraminidase, NEU), but not sialyltransferases, were less documented in cancer research. In preliminary study, higher expression of Sias, but lower expression of neuraminidase 1 (NEU1) in bladder cancer cells than in normal epithelial bladder cells were observed. In NEU1 expressed bladder cancer cells, expression of fibronectin (FN) and integrinα5β1 were down-regulated, cell proliferation was inhibited and cell apoptosis was enhanced. Furthermore, overexpression of NEU1 in bladder cancer cells suppressed tumor growth in vivo. However, the mechanism of NEU1 on the suppression of bladder cancer remains unclear. Based on above observations, we proposed that NEU1 could enhance cell apoptosis and decrease cell proliferation by disrupting the interaction of FN and integrinα5β1 and attenuating downstream signaling pathway. This proposal will explore the correlation of NEU1 expression and the progression of bladder cancer, explain the mechanism how NEU1 modulates the interaction of FN and integrinα5β1 and its signaling pathway, evaluate the efficiency and safety of NEU1 in vivo, and make a contribution to the clinical therapy of bladder cancer.

唾液酸的高表达显著影响肿瘤细胞的侵袭及迁移，但肿瘤中唾液酸酶的生物学功能很少受到关注。申请人在预实验中发现膀胱癌细胞中唾液酸酶NEU1的低表达是唾液酸升高的主要因素；在膀胱癌细胞过表达NEU1，纤连蛋白（FN）和整合素α5β1表达显著下降；细胞增殖受到抑制，凋亡升高；过表达NEU1的膀胱癌细胞在小鼠体内成瘤能力显著下降。但是NEU1抑制膀胱癌的分子机制尚不清楚。基于预实验结果，本项目推测：NEU1通过调节FN和整合素α5β1的相互作用及其下游的信号通路，影响膀胱癌细胞的增殖与凋亡。本项目将评估NEU1表达与膀胱癌病情发展的相关性，阐明NEU1如何调节FN和整合素α5β1互相作用及相关的信号通路，并通过小鼠模型对NEU1抑制膀胱癌的有效性及安全性进行论证，以期为膀胱癌的临床诊疗开拓新思路并提供理论依据。

唾液酸是一种以九个碳原子作为骨架的酸性糖，常以单体或多聚形式存在于糖链末端。已证实唾液酸在肿瘤的发生发展过程中具有重要的生物学功能。唾液酸修饰受唾液酸转移酶和唾液酸酶共同调控，唾液酸转移酶在肿瘤组织中高表达且有助于癌细胞累积唾液酸修饰，但唾液酸酶在肿瘤中的作用仍存在争议。.本研究以膀胱癌为研究模型，结合基于质谱的糖组学和蛋白组学技术，研究了唾液酸修饰和膀胱癌发生发展的关联及分子机理。主要结论如下：.（1）发现膀胱癌唾液酸修饰的异常增加与唾液酸酶NEU1低表达有关。经高通量N-糖组学分析发现恶性膀胱癌细胞含有较多唾液酸修饰。经唾液酸酶活性检测以及高通量蛋白质谱分析，发现膀胱癌唾液酸修饰增加与唾液酸酶NEU1低表达有关。分析临床样本发现膀胱癌患者癌组织中NEU1的表达水平显著低于癌旁组织，且NEU1的低表达预示着患者有更短的生存期。.（2）唾液酸酶NEU1 催化纤维连接蛋白（FN）的去唾液酸修饰，抑制由整合素α5β1介导的Akt 磷酸化和细胞增殖。发现NEU1定位于细胞膜且能够催化FN去唾液酸化修饰。而FN唾液酸修饰影响其与整合素α5β1的亲和力，致使由整合素介导的细胞黏附状态信号传递减弱，细胞内吞并降解整合素α5β1和FN，导致细胞增殖减少，凋亡增加小鼠皮下成瘤模型证实YTS-1/NEU1组肿瘤生长显著慢于对照组，且肿瘤组织切片染色显示NEU1高表达降低FN、整合素α5和β1的含量，并抑制细胞增殖，促进细胞凋亡。.总之，本研究从膀胱癌唾液酸修饰与唾液酸酶的关联等方面展开研究，发现了唾液酸酶NEU1在原位催化FN去唾液酸修饰的现象，并分析FN的唾液酸化对整和蛋白亲和力的影响，证实NEU1与膀胱癌细胞唾液酸修饰的负相关性，并揭示了NEU1对细胞生长和凋亡调控的分子机理，为膀胱癌的临床诊疗提供了科学依据。