Analysis of Redox Modulated Signaling Networks in Response to Ionizing Radiation

描述(由申请人提供)： 预测性、预防性和个性化医学是患者和医疗保健提供者的共同目标。在过去的几年里，人类基因组计划的完成和对数百种人类癌症的基因筛查，导致在不同类型的癌症中发现了数百种“驱动”激酶突变。为了利用从这些研究中获得的信息，我们的长期努力旨在应用先进的系统生物学方法，以更好地定义头颈部鳞状细胞癌(HNSCC)联合突变的后果。这种方法将允许在未来为患者量身定做治疗，从而实现更高的治愈率和更低的毒性。除了致癌基因突变外，细胞信号转导的另一个重要调节成分是由生长因子启动的信号转导、放射治疗、药物/外源物质代谢等因素所产生的活性氧物种(ROS)。在HNSCC或其他癌症中，致癌基因突变和信号蛋白氧化的共同作用几乎没有受到关注。信号网络的氧化还原调节在放射治疗中尤其重要，因为人们对辐射诱导的ROS与促进细胞死亡或细胞增殖的信号通路之间的相互作用知之甚少。为了研究控制肿瘤生长和对放射和药物治疗反应的信号网络的氧化还原调节，我们在这里描述了一种跨学科的翻译方法，基于i)蛋白质组学方法，ii)以毫秒时间分辨率的生长因子刺激细胞的特定仪器，iii)首次使用高度特异的分子探针来检测作为氧化还原信号关键中间体的含磺酸的蛋白质，以及iv]计算方法来整合和评估蛋白质组学方法产生的大量数据。我们建议使用HNSCC的同源抗辐射模型来定义正常(EGF刺激)和治疗条件(辐射加或不加Erlotinib)下蛋白质氧化和蛋白质磷酸化的靶点。此外，我们还描述了一系列后续研究，以评估氧化修饰对蛋白质功能和辐射抗性表型的影响，这些研究选定了一些被蛋白质组学研究确定为氧化的信号蛋白。该项目的成果将对辐射后敏感和耐药细胞系中的磷酸和氧化信号产生系统水平的理解。该建议结合了多组实验数据、模型和理论，以发展对这些扰动的特性和生物学后果的系统级理解。

DESCRIPTION (provided by applicant): Predictive, preventive and personalized medicine is the common goal of patients and health-care providers. The completion of the human genome project and genetic screenings of several hundreds of human cancers over the last years, have led to the identification of hundreds of "driver" kinase mutations in different types of cancers. To take advantage of the information emerging from these studies, our long-term efforts are directed towards applying advanced systems biology methodologies to better define the consequences of combined mutations in head and neck squamous cell carcinomas (HNSCC). Such an approach will allow patient tailored therapies to be prescribed in the future, resulting in higher cure rates and lower toxicity. In addition to oncogenic mutations, another important regulatory component of cell signaling is represented by the generation of reactive oxygen species (ROS) in response to growth factors initiated signaling, radiation therapy, drugs/xenobiotics metabolism and other factors. The combined contribution of oncogenic mutations and oxidation of signaling proteins in HNSCC or other cancers has received little attention. The redox regulation of signaling networks is particularly important in the context of radiation therapies as little is known about the interaction of radiation induced ROS and signaling pathways that promote cell death or cell proliferation. To investigate the redox regulation of signaling networks that control tumor growth and the response to radiation and drug therapies, we describe here a cross-disciplinary, translational approach based on i] proteomics methodologies, ii] specific instrumentation for cellular stimulation with growth factors with millisecond time resolution, iii] first time use of highly specific molecular probes for the detection of sulfenic acid containing proteins as key intermediates in redox signaling, and iv] computational methods to integrate and evaluate the massive amount of data generated by the proteomics approach. We propose to define targets of protein oxidation and protein phosphorylation under normal (EGF stimulation) and therapeutic conditions (radiation plus or minus Erlotinib) using an isogenic radiation-resistant model of HNSCC. Also, we describe a series of follow-up studies to assess the consequences of oxidative modification on protein function and radiation resistance phenotype for a selected number of signaling proteins identified as oxidized by the proteomics studies. The outcome of this project will yield a systems-level understanding of phospho- and oxidative signaling following radiation, both in sensitive and resistant cell lines. This proposal combines multiple sets of experimental data, modeling, and theory for developing a systems-level understanding of properties and biological consequences of these perturbations.