Human models of radioprotection

项目3将分析放射调节剂（rma）对人体细胞的体外影响——来自正常个体（目标1），来自已知DMA修复障碍患者（目标2），以及来自一组未确诊疾病的放射敏感患者（目标3）。待测试的rma将通过高通量筛选（Core D）确定，然后在酵母和小鼠模型中进行额外的表征（项目2和3）。我们有独特的条件来进行项目3中概述的研究，因为我们实验室拥有大量的细胞库和长期的功能和诊断性DNA修复分析经验，例如通过菌落存活进行的辐射敏感性测试，耐辐射DNA合成，IRIF形成，丝氨酸激酶活性和彗星分析。光谱核型将识别染色体畸变特征，并有助于量化候选RPAs的影响（Core E）。在这些分析的最后阶段，细胞内酪氨酸激酶信号和全局蛋白表达微阵列将描绘这些化合物的下游效应（Core D）。通过使用已知在1)atm相关通路（A-T， NBS, Mre11）， 2) smc1独立放射敏感性（Fanconi通路）和3)非同源（非法）末端连接修复（LIG4）中发生突变的放射敏感（RS）细胞，候选rma将被分组为操作类别和共同的作用机制。随着对人类放射敏感性机制的新见解的发现，bbbb40淋巴母细胞系的RS组将使这些类别得到改进，并应用于特定分子缺陷的治疗和辐射调节化合物的潜在作用机制。这些协调中心研究的附加价值将包括确定a)用于评估人类辐射暴露的新双标志物和b)用于减轻和预防辐射损伤全面影响的新辐射调节剂。

Project 3 will analyze the in vitro effects of radiomodulatory agents (RMAs) on human cells - from normal individuals (Aim 1), from patients with known DMA repair disorders (Aim 2), and from a panel of radiosensitive patients with undiagnosed disorders (Aim 3). The RMAs to be tested will have been identified by high throughput screening (Core D), followed by additional characterization in yeast and mouse models (Projects 2 and 3). We are uniquely poised to perform the studies outlined in Project 3 because pur laboratory has a large cell repository and long-standing experience with functional and diagnostic DNA repair assays, such as radiosensitivity testing by colony survival, radioresistant DNA synthesis, IRIF formation, serine kinase activity, and comet assays. Spectral karyotyping will identify characteristic chromosomal aberrations and help to quantify the effects of candidate RPAs (Core E). In the final stages of these analyses, intracellular tyrosine kinase signaling and global protein expression microarrays will profile the downstream effects of these compounds (Core D). Through the use of radiosensitive (RS) cells with known mutations in 1) ATM-related pathways (A-T, NBS, Mre11), 2) SMC1-independent radiosensitivity (Fanconi pathway), and 3) non-homologous (illegitimate) end joining repair (LIG4), candidate RMAs will be grouped into operational categories and common mechanisms of action. As new insights into the mechanisms of human radiosensitivity are discovered, a RS panel of >40 lymphoblastpid cell lines will allow these categories to be refined and applied towards the treatment of specific molecular defects and the underlying mechansims of action of the radiomodulatory compounds. The added value of these coordinated center studies will include the identification of a) new bipmarkers for evaluating human radiation exposure and b) novel radiomodulatory agents for mitigating and preventing the full effects of radiation damage.