Targeting crosstalk between insulin and Gq signaling systems in pancreatic cancer

描述(申请人提供)：胰腺癌是美国男性和女性癌症死亡的第四大原因，总体5年存活率仅为3-5%。由于目前的治疗方法提供的生存益处非常有限，迫切需要新的治疗策略来治疗这种侵袭性疾病。鉴于胰腺内分泌和外分泌之间的密切关系，胰腺微环境的一个独特特征是局部产生的胰岛素对胰腺癌细胞生长的潜在影响。胰岛素受体和G蛋白偶联受体信号系统之间的串扰在许多生理功能的调节和包括癌细胞增殖在内的各种异常过程的发病机制中起着关键作用。尽管这种受体串扰具有重要的生物学和临床意义，但其涉及的分子机制(S)仍未得到很好的描述。R21的应用(针对RFA，题为“胰腺癌的初步研究”)是基于我们最近的发现，即人胰腺癌细胞暴露于生理浓度的胰岛素会迅速增强早期细胞事件，例如GQ偶联受体激动剂，包括血管紧张素II、缓激肽、蛙皮素和神经降压素引起的[钙]i增加。通过对单细胞中PtdIns(4，5)P2水解和Ins(1，4，5)P3生成的变化进行实时成像，我们发现胰岛素增强了这些第二信使生成反应的速度和幅度。胰岛素诱导的GPCR信号的增强是通过雷帕霉素敏感的PI3-激酶/mTOR依赖的途径来介导的。我们的初步结果还表明，胰岛素和GPCR激动剂协同刺激DNA合成和锚定非依赖性细胞增殖。我们推测，胰岛素通过mTOR依赖的途径增强GPCR信号通路，从而提供了一种机制，通过胰岛素增强胰腺细胞对GPCR激动剂的反应性，广泛用于治疗2型糖尿病的药物二甲双胍通过激活AMPK来负向调节mTOR功能。我们假设二甲双胍优先消除胰岛素受体和gpr信号之间的串扰。的确，初步结果表明，二甲双胍抑制了胰岛素受体和GPCR信号系统在人胰腺癌细胞中钙信号和DNA合成方面的串扰。因此，我们建议在GRANT应用中追求两个特定的目标：1)在一组人胰腺癌细胞系中表征胰岛素受体和gpr信号之间的串扰的机制(S)和促进生长的作用；2)表征二甲双胍在体外和体内对胰岛素受体和gpr信号之间的串扰的抑制作用。鉴于二甲双胍和雷帕霉素衍生物是FDA批准的药物，这些研究可能为激动人心的快速翻译研究铺平道路。与公共卫生相关：增加胰腺胰岛素生成的疾病，包括肥胖和早期II型糖尿病，会增加患胰腺癌的风险。这一应用中一个令人兴奋的新结果是，广泛用于治疗II型糖尿病的药物二甲双胍抑制了胰岛素受体和GQ偶联受体信号之间的一种新的串扰，阻止了人胰腺癌细胞的生长，从而为胰腺癌提供了一种新的治疗策略。

DESCRIPTION (provided by applicant): Pancreatic cancer is the 4th leading cause of cancer fatalities in the United States both in men and women, with an overall 5-year survival rate of only 3-5%. As the current therapies offer very limited survival benefits, novel therapeutic strategies are urgently needed to treat this aggressive disease. Given the close relationship between the endocrine and exocrine pancreas, a unique feature of the pancreatic microenvironment is the potential influence of locally produced insulin on pancreatic cancer cell development. Crosstalk between the insulin receptor and G protein-coupled receptor (GPCR) signaling systems plays a critical role in the regulation of many physiological functions and in the pathogenesis of a variety of abnormal processes, including cancer cell proliferation. Despite the biological and clinical importance of this receptor crosstalk, the molecular mechanism(s) involved remain poorly characterized. This R21 application (in response to RFA, entitled "Pilot Studies in Pancreatic Cancer") is based on our recent findings showing that exposure of human pancreatic cancer cells to physiological concentrations of insulin rapidly enhances early cellular events, such as increase in [Ca2+]i elicited by Gq-coupled receptor agonists, including angiotensin II, bradykinin, bombesin and neurotensin. Using real-time imaging of changes in PtdIns(4,5)P2 hydrolysis and generation of Ins(1,4,5)P3 in single cells, we found that insulin enhances the rate and magnitude of these second messenger-generating responses. The insulin-induced potentiation of GPCR signaling is mediated through a rapamycin-sensitive, PI3- kinase/mTOR-dependent pathway. Our preliminary results also show that insulin and GPCR agonists synergistically stimulate DNA synthesis and anchorage-independent cell proliferation. We hypothesize that the potentiation of GPCR signaling by insulin through a mTOR-dependent pathway provides a mechanism by which insulin enhances the responsiveness of pancreatic cells to GPCR agonists, Metformin, a widely used drug for treatment of type 2 diabetes mellitus, negatively regulates mTOR function via AMPK activation. We hypothesize that metformin preferentially abolishes the crosstalk between insulin receptor and GPCR signaling. Indeed, preliminary results show that metformin inhibits the crosstalk between insulin receptor and GPCR signaling systems in Ca2+ signaling and DNA synthesis in human pancreatic cancer cells. Consequently, we propose to pursue two Specific Aims in this Grant application: 1) Characterize the mechanism(s) and growth-promoting effects of the crosstalk between insulin receptor and GPCR signaling in a panel of human pancreatic cancer cell lines; 2) Characterize the inhibitory effects of metformin on the crosstalk between insulin receptor and GPCR signaling in vitro and in vivo. Given that metformin and rapamycin derivatives are FDA-approved drugs, these studies may pave the way for exciting and rapid translational research. PUBLIC HEALTH RELEVANCE: Conditions that enhance insulin production in the pancreas, including obesity and early stages of Type II diabetes, increase the risk of pancreatic cancer. An exciting new result shown in this application is that metformin, a drug widely used in the treatment of type II diabetes, inhibits a novel crosstalk between insulin receptor and Gq-coupled receptor signaling, blocks the growth of human pancreatic cancer cells and thus, can provide a novel therapeutic strategy for pancreatic cancer.