Role of EPAC Signaling in Melanoma Progression

摘要 皮肤黑色素瘤源于皮肤中的黑素细胞。暴露于太阳紫外线（UV）辐射是 重大风险因素。因此，美国在世界各地的高紫外线指数地区部署军事人员， 在服兵役期间和之后增加患黑色素瘤的风险。如果早期诊断，切除原发肿瘤， 皮肤可以治愈。然而，随着黑色素瘤的转移， 局部和区域淋巴结以及远端器官。尽管最近的成功和有针对性的承诺， 早期诊断和局部原发癌的治疗仍然是最好的 降低癌症死亡率的战略。因此，详细了解所涉及的分子机制 在黑色素瘤中，肿瘤进展对于侵袭性黑色素瘤的准确和可靠的诊断和治疗是至关重要的。 皮肤黑素瘤环磷酸腺苷（cAMP）信号通路在黑素细胞增殖和黑素细胞分化中起着关键作用。 分化这一途径是在腺苷酸的催化活性引起细胞内cAMP升高时启动的。 环化酶，并通过蛋白激酶A和转录因子CREB转导，最终表达 cAMP反应基因，包括黑素细胞主调节因子MITF。cAMP信号通路的作用 在黑色素瘤中的作用还不完全清楚。采用人原发性黑色素瘤组织微阵列（TMA）， 原发性和转移性黑色素瘤细胞系以及黑色素瘤的BrafV 600 E/Pten-/-小鼠模型，我们最近 证明了cAMP信号在原发性和转移性黑色素瘤细胞中发挥相反的作用，即。例如，作为一个亲， 在原发性细胞中作为增殖信号，在转移性细胞中作为抗增殖信号。我们还发现这个开关 cAMP的作用是由涉及EPAC-RAP 1（交换因子）的替代cAMP信号轴介导的 直接由CAMP-Ras相关蛋白1激活）。其他研究表明，EPAC促进细胞周期 进展，激活mTORC 1信号传导，并调节细胞代谢和线粒体ROS产生 选择性地在原发性黑素瘤细胞中。此外，使用一组原发性和转移性黑色素瘤的研究 来自同一患者的细胞系显示EPAC功能的转换是渐进性的 表明EPAC是原发性黑色素瘤生长的关键成分 肿瘤进展过程中信号网络的重新布线消除了转移性肿瘤中的EPAC依赖性， 黑素瘤在这里，我们提出测试的假设，a）致癌转化的黑素细胞激活 EPAC信号传导和EPAC作为一个关键的信号节点，抑制应激诱导的ROS产生，促进 衰老逃逸和原发性黑素瘤发展，和B）基因表达变化和代谢 在转移过程中发生的适应逐渐消除EPAC依赖性。具体目标是 应用是：1）建立EPAC在黑色素瘤肿瘤发生和进展中的作用，2）确定EPAC在黑色素瘤中的作用。 EPAC作用的生物化学和分子机制; 3）研究EPAC在代谢中的作用 黑色素瘤进展期间的适应。成功完成这里提出的实验可能会导致 可靶向抑制黑素瘤进展和治疗转移性黑素瘤的途径和蛋白的鉴定 黑素瘤因此，我们相信该项目将在黑色素瘤领域产生持续的影响。

ABSTRACT Cutaneous melanoma arises from melanocytes in the skin. Exposure to solar ultraviolet (UV) radiation is a significant risk factor. Deployment of US military personnel around the world in areas of high UV index thus increases risk of melanoma during and after military service. If diagnosed early, excision of the primary tumor in the skin can be curative. However, the five-year survival rates decrease precipitously as melanoma metastasizes to local and regional lymph nodes and distant organs. Despite the recent successes and promise of targeted therapies and immunotherapy, early diagnosis and treatment of the localized primary cancer remain the best strategies to reduce cancer mortality. Therefore, a detailed understanding of the molecular mechanisms involved in melanoma tumor progression is critical for accurate and reliable diagnosis and treatment of aggressive cutaneous melanoma. Cyclic AMP (cAMP) signaling pathway plays a critical role in melanocyte proliferation and differentiation. This pathway is initiated upon elevation of intracellular cAMP by the catalytic activity of adenylyl cyclases and transduced through protein kinase A and transcription factor CREB culminating in expression of cAMP-responsive genes including the melanocyte master regulator, MITF. The role for cAMP signaling pathway in melanoma is not fully understood. Employing a human primary melanoma tissue microarray (TMA), a panel primary and metastatic melanoma cell lines and the BrafV600E/Pten-/- mouse model for melanoma, we recently demonstrated that cAMP signaling plays opposite roles in primary and metastatic melanoma cells, i. e., as a pro- proliferative signal in primary and as anti-proliferative signal in metastatic cells. We also showed that this switch in the cAMP of action is mediated by the alternative cAMP signaling axis involving EPAC-RAP1 (Exchange factor directly activated by cAMP- Ras-related protein 1). Additional studies showed that EPAC promotes cell cycle progression, activates mTORC1 signaling, and regulates cellular metabolism and mitochondrial ROS production selectively in primary melanoma cells. Furthermore, studies using a panel of primary and metastatic melanoma cell lines derived from the same patient revealed that the switch in the functions of EPAC occurs progressively during melanoma metastasis suggesting that EPAC is a critical component for the growth of primary melanoma and that rewiring of signaling networks during tumor progression abolish EPAC dependency in metastatic melanoma. Here, we propose to test the hypotheses that a) oncogenic transformation of melanocytes activates EPAC signaling and EPAC acts as a critical signaling node to suppress stress induced ROS generation, promote senescence escape and primary melanoma development, and b) gene expression changes and metabolic adaptions that occur during metastasis progressively abolish EPAC dependency. The specific aims of this application are: 1) Establish the role of EPAC in melanoma tumorigenesis and progression, 2) Determine the biochemical and molecular mechanisms of action of EPAC and 3) Investigate the role of EPAC in metabolic adaptation during melanoma progression. Successful completion of experiments proposed here could lead to identification of pathways and proteins that can be targeted to inhibit melanoma progression and treat metastatic melanoma. Therefore, we believe this project will have sustained impact in the field of melanoma.