甲羟戊酸代谢物介导RhoB异戊二烯化在膀胱癌中作用及其机制的研究

Lipid metabolism is closely related to bladder cancer, however, its exact mechanism in cancer remains unclear. Our previous study suggested that the mevalonate pathway was activated in bladder cancer. Inhibition of the activity or interference with expression of the mevalonate pathway related enzyme could reduce the content of intracellular FPP and GGPP, inhibit the proliferation and metastasis of bladder cancer and increase the expression of RhoB. We also found that compared with paracancer tissues, the expression of RhoB was down-regulated in bladder cancer, and the down-regulated RhoB was closely related to tumor stage and prognosis. In addition, the prenytylated RhoB protein is more susceptible to degradation by lysosomes. According to our previous study, we hypothesis that due to the activation of the mevalonate pathway, the content of FPP and GGPP is increased in bladder cancer, leading RhoB more prone to prenylation and degradation. In this study, clinical samples will been collected to detect the clinical relevance of RhoB expression, FPP, GGPP in bladder cancer. We plan to establish cell and animal models to elucidate the regulation mechanism of mevalonate metabolites on RhoB expression and isoprenylation, and to investigate the effects of RhoB and its prenylation modification in bladder cancer and its mechanism. The aim of our study is to provide the new ideas of diagnosis and treatment of bladder cancer together with the metabolites and clarify the regulatory role.

脂类代谢与膀胱癌密切相关，但其调控机制尚未阐明。我们前期发现甲羟戊酸途径在膀胱癌中处于激活状态。降低该途径相关酶的活性，可抑制膀胱癌的增殖和转移、降低胞内甲羟戊酸代谢物FPP和GGPP的含量、促进抑癌基因RhoB的表达。而RhoB在膀胱癌中低表达，并与肿瘤分期和预后相关，但机制不明。文献提示FPP和GGPP能够参与异戊二烯化修饰调控过程，且RhoB蛋白经过异戊二烯化修饰后更易被溶酶体降解。据此推测：膀胱癌中激活的甲羟戊酸途径产生较多FPP和GGPP，导致RhoB更倾向于异戊二烯化修饰而被降解，从而无法发挥抑癌作用。本项目拟结合临床样本，探讨RhoB表达、甲羟戊酸代谢产物含量与膀胱癌的临床相关性；应用细胞和动物模型，探究甲羟戊酸代谢物对RhoB及其异戊二烯化的调控机制、阐明RhoB及其异戊二烯化修饰对膀胱癌的作用机制；从代谢物及其调控作用出发为膀胱癌诊疗新思路提供相关依据。

代谢异常与肿瘤发生发展密切相关，从“肿瘤代谢”的角度出发，探究膀胱癌发生发展的机制，有望为膀胱癌的诊疗提供潜在新靶点。本项目从团队前期研究发现的膀胱癌中甲羟戊酸途径代谢异常出发，利用膀胱癌及癌旁组织样本证实了RhoB在膀胱癌组织中低表达，并与患者预后呈正相关；发现膀胱癌组织中胆固醇的含量更高；应用TCGA数据库中膀胱癌测序数据分析了甲羟戊酸途径相关酶（HMGCR、MVK、MVD、FDPS、HMGCS1、HMGCS2、PMVK、GGPS1）和RhoB表达与膀胱癌的临床相关性，进一步证实了甲羟戊酸途径在膀胱癌中被激活，且RhoB在膀胱癌以及转移性膀胱癌组织中的表达更高。通过敲低FDPS（甲羟戊酸途径酶）或者唑来膦酸（FDPS抑制剂）来抑制甲羟戊酸途径后，膀胱癌细胞的迁移和增殖能力明显受到抑制，并伴随着RhoB蛋白的表达上调。在培养基中添加甲羟戊酸途径中间代谢产物GGPP（而非FPP）能够逆转甲羟戊酸途径抑制导致的膀胱癌细胞迁移能力的抑制，并降低RhoB蛋白的表达。同时，我们还证实了GGPP能够通过参与RhoB蛋白的香叶基香叶基化修饰和泛素化修饰来影响RhoB蛋白的表达。此外，我们应用细胞实验和动物实验证实了RhoB对膀胱癌细胞增殖和迁移能力的抑制作用，并发现其能够影响膀胱癌细胞上皮间质化转变（EMT）过程及整合素β1蛋白在细胞质和膜间的分布调控。上述结果表明，膀胱癌细胞中被激活的甲羟戊酸途径，能够使细胞中GGPP等甲羟戊酸代谢产物的含量增加，从而通过参与RhoB蛋白香叶基香叶基化修饰和泛素化修饰的调控而导致其蛋白降解，抑制其抑癌功能，进而通过EMT及整合素β1的质膜分布等调控来促进膀胱癌细胞的增殖和转移。综上可知，抑制膀胱癌细胞中激活的甲羟戊酸途径有望为当前膀胱癌诊疗困境提供新的解决策略或方案。