调控AMPK-HNF4α-WNT致瘤信号轴：小檗碱抗胃癌作用的关键靶向通路？

Previous evidence has demonstrated the efficacy of berberine on type 2 diabetes mellitus (T2DM) targeting HNF4α. Recent researches have shown that Metformin, a widely used anti-diabetes drug, was found to inhibit cancer cell lines and xenograft growth through regulating ‘metabolic switch’. Given that the abnormal energy metabolism of cells contributes to T2DM and cancer，we hypothesize that regulating ‘metabolic switch’ AMPK-HNF4α-WNT signaling axis: a potentially targetable pathway for Berberine on Gastric Cancer? To test our hypothesis, we will establish xenograft (NCI-N87 and MKN45) mouse models and GC cell models. Experiments have been designed to examine the activity and expression of AMPK-HNF4α signaling axis in GC tumors models. The modern techniques including pathophysiology, biochemistry and cell molecular biology such as ELISA, flow cytometry, Real time PCR, immunohistochemistry, western blot, report (Luciferase) assay, and siRNA transfection will be performed in vivo and in vitro. The successful completion if the proposed research will illustrate the mechanisms of the anti-gastric cancer actions of berberine, based on which we will be able to develop a new treatment for cancers in view of cell energy metabolism. Additionally, our study will provide scientific rationale for the potential utility of new anti-diabetes and anti-carcinoma agents which possess molecular structure as berberine and the feature of “mono-target point, multi-pathway, multi-actions” in the clinical practice. At the same time, the proposed research will explain scientific connotation of traditional Chinese medicine at “treating different diseases with the same method or the same target”.

本项目在前期研究证实小檗碱具有靶向肝细胞核因子HNF4α抗糖尿病功效的基础上，以“细胞能量代谢异常是2型糖尿病与恶性肿瘤共同的病理生理学基础”为切入点，结合抗2型糖尿病药物通过调控“代谢开关”而发挥抗肿瘤功效这一最新研究动态，提出“调控AMPK-HNF4α-WNT致瘤信号轴可能是小檗碱抗胃癌作用的关键靶向通路之一”的全新假说。本项目拟采用现代病理生理，生物化学，细胞分子生物学等技术，整体与离体实验相结合，机能与代谢检测相结合，深入系列研究AMPK-HNF4α-WNT信号轴与胃癌发生的相关性以及小檗碱抗胃癌作用的相关机制，为从细胞能量代谢角度寻找恶性肿瘤的治疗靶点提供新的思路，进而为研发以小檗碱为母核分子结构，具有“单靶点，多途径，多功效”的新型治疗特色且兼具抗糖尿病与抗肿瘤双重功效的天然植物类新药奠定理论基础，同时也初步揭示出中药“异病同治”的科学内涵。

细胞能量代谢异常是2型糖尿病和恶性肿瘤发生共同的病理生理学基础，肝细胞核因子(HNF4α)是肝脏最重要的转录因子之一，一方面参与糖脂代谢及胰岛素分泌的基因调控，与代谢性疾病如2型糖尿病，肥胖，代谢综合征密切相关；另一方面，HNF4α是与肿瘤分化相关性最高的基因之一，在转录水平上调控着细胞的成熟与分化，也是胃癌的一个通道癌蛋白。HNF4α是小檗碱抗糖尿病作用的关键靶向分子之一，因此，我们设想以HNF4α为关键分子靶点的小檗碱可能正是通过调控“代谢开关”AMPK-HNF4α-WNT致瘤信号轴而发挥抗肿瘤功效!.我们从临床探讨HNF4α异常表达与胃癌的临床病理特征关系，以求证HNF4α是否参与胃癌的发生发展及临床意义；同时探讨小檗碱对离体胃癌细胞系和在体的胃癌皮下移植瘤小鼠模型的抗肿瘤效应，以求证小檗碱是否具有抗胃癌效应，以及AMPK-HNF4α-WNT致瘤信号轴是否为抗胃癌功效的关键靶向通路。我们的研究结果证实，相比癌旁组织，HNF4α和WNT5a在胃癌组织中表达上调，且两者之间的表达呈显著正相关；胞质中的β-catenin在胃癌组织中表达下调。体外实验证实，小檗碱具有抗胃癌作用，包括抑制胃癌细胞的增殖，诱导胃癌细胞周期停滞，减弱细胞的迁移和侵袭能力等，其作用机制与小檗碱磷酸化AMPK，靶向抑制HNF4α，进而下调其下游的WNT5a和β-catenin的表达有关。同时，体内实验也证实了小檗碱可以有效抑制胃癌皮下移植瘤的生长，其机制与小檗碱抑制HNF4α的表达并下调其下游的WNT5a和β-catenin的信号有关。.我们的研究结果为抗胃癌药物的研发提供了潜在的靶向通路，也为从细胞能量代谢角度寻找恶性肿瘤的治疗靶点提供了新的思路，进而为研发以小檗碱为母核分子结构，具有“单靶点，多途径，多功效”的新型治疗特色且兼具抗糖尿病与抗肿瘤双重功效的天然植物类新药奠定理论基础，同时也初步揭示出中药“异病同治”的科学内涵。