CAB39介导上皮间质转化在膀胱癌侵袭转移中的作用及机制研究

Epithelial-mesenchymal transition (EMT) plays a significant role in the invasion and metastasis of cancer cells, the molecular mechanism of which remains to be elucidated. We previously found that CAB39 gene was highly expressed in invasive and metastatic bladder cancer. Expression level of CAB39 was closely related to invasive and metastatic potential of bladder cancer cells, while positively and negatively related to the two major EMT markers, Vimentin and E-cadherin, respectively. Therefore, we hypothesized that, CAB39 promotes invasion and metastasis of bladder cancer through regulation of EMT. In this project, we will establish a possible regulatory role of CAB39 in EMT-related proteins, and analyze their co-relations with clinical and pathological features of bladder cancer. Using gene manipulation measurements including stable transfection and knockdown techniques, we will study the effect of altering CAB39 expression on bladder cancer cell growth and EMT in cell and nude mice models, and on EMT and JNK signaling pathways by TGF-β stimulation and JNK inhibitors, respectively. In the aim of revealing the underlying molecular mechanism of CAB39 regulating EMT through JNK signaling pathway, this project will provide experimental data and theoretical basis for future development of gene targeted therapy and medicine for bladder cancer treatment.

侵袭转移是恶性肿瘤的主要生物学特征。上皮-间质转化（EMT）在肿瘤侵袭转移中起重要作用，其分子机制不明。申请人前期研究发现CAB39基因在侵袭转移性膀胱癌组织中显著高表达，其表达量与膀胱癌细胞的侵袭转移潜能和癌组织中EMT相关蛋白Vimentin表达正相关和E-cadherin表达负相关；据此提出假设:CAB39通过调控EMT促进膀胱癌侵袭转移。本课题拟在组织标本中观察CAB39和EMT相关蛋白的相互作用，并分析它们与临床病理特征的相关性；通过基因稳定转染和干扰等技术在细胞和裸鼠模型中探讨CAB39表达改变对膀胱癌细胞生长和EMT的影响；进一步结合外源性TGF-β刺激实验和利用JNK抑制剂处理，观察其对EMT和JNK信号通路的影响，从而阐明CAB39通过JNK信号通路调控EMT发生，以促进膀胱癌侵袭转移的分子机制,为探寻膀胱癌基因治疗靶点和药物开发提供理论基础和实验依据。

膀胱肿瘤是泌尿系最常见肿瘤之一；侵袭转移是恶性肿瘤的主要生物学特征；上皮-间质转化（EMT）在肿瘤侵袭转移中起重要作用，其分子机制尚不清楚。申请人前期研究发现CAB39基因在侵袭转移性膀胱癌组织中显著高表达，其表达量与膀胱癌细胞的侵袭转移潜能和癌组织中EMT相关蛋白Vimentin表达正相关和E-cadherin表达负相关；据此提出假设: CAB39通过调控EMT促进膀胱癌侵袭转移。本研究通过免疫组化及实时荧光定量PCR检测组织标本及细胞中CAB39的表达，并分析它们与膀胱癌临床病理特征的相关性；通过基因稳定转染和干扰等技术构建CAB39敲减的细胞和裸鼠模型，通过细胞划痕愈伤实验、侵袭实验及CCK8增殖实验等观察CAB39表达改变对膀胱癌细胞二维迁移、侵袭、增殖及体内成瘤能力的影响。进一步结合GSEA生物信息学分析膀胱癌组织中CAB39密切相关的关键分子，并采用Western blot检测关键蛋白的表达。本研究发现CAB39在膀胱癌组织及癌细胞中异常高表达，其表达水平和膀胱癌患者预后密切相关；CAB39基因被敲减后膀胱癌细胞的二维迁移、侵袭及体内成瘤能力明显受抑制；其可能是通过激活NF-kB信号通路，抑制E-cadherin并促进N-cadherin的表达，以此诱导EMT的发生，从而促进膀胱癌侵袭与转移。本研究发现可为膀胱癌基因治疗靶点和药物开发提供理论基础和实验依据。