Quantitative RT/PCR-based radiation dosimetry

基于 RT/PCR 的定量辐射剂量测定

• 批准号: 7486748
• 负责人: DEREK L STIREWALT
• 金额: $ 48.99万
• 依托单位: FRED HUTCHINSON CANCER RESEARCH CENTER
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7486748
• 关键词: Adverse effects; Affect; Animal Model; Animals; Biological Assay; Blinded; Blood Cells; Bone Marrow; CD4 Positive T Lymphocytes; Candidate Disease Gene; Canis familiaris; Caring; Cells; Cessation of life; Classification; Clinical; DNA Microarray Chip; DNA Microarray format; Development; Dose; Emergency Situation; Ensure; Ethical Issues; Event; Extracellular Space; Genes; Genetic; Genetic Transcription; Goals; Hematopoietic; Human; In Vitro; Lead; Length; Leukocytes; Licensing; Life; Lymphocyte; Modeling; Nuclear; Organ failure; Patients; Physiologic pulse; Plasma; Pulse taking; RNA; Radiation; Radioactive; Radiometry; Rate; Reagent; Risk; Sampling; Source; Stratification; System; Testing; Time; Tissues; Transplant Recipients; Whole-Body Irradiation; base; biodosimetry; clinically relevant; day; design; dosimetry; in vivo; injured; irradiation; novel; radiation effect; response

In the event of a terrorist attack using radioactive material, risk-stratification of patients based on exposure doses will be critical to ensure appropriate care. Current dosimetry assays take days to perform, which would delay life-saving therapies. This proposal seeks to develop RNA-based dosimetry assays using the GeneXpert platform that will risk-stratify patients immediately after radiation exposures. Specific Aim 1. Discover genes that display dose-dependent expression changes after in vitro irradiation of human peripheral blood cells. Radiation exposures cause dose-dependent expression changes in peripheral blood cells. We hypothesize that these expression changes can be utilized to develop RNA-based assays that discriminate between clinically relevant doses of irradiation. Therefore, DNA microarrays will be used to identify genes with dose-dependent expression changes in hematopoietic cells after in vitro irradiation. Quantitative RT/PCR assays will validate these dose-dependent expression changes, and candidate genes will be examined in Specific Aims 2 and 3. Specific Aim 2. Determine whether RNA-based dosimetry assays using plasma or cells best correlate with radiation doses in the canine model. Cells injured from radiation die, leaking RNA into the extracellular space, and this RNA can be detected using quantitative RT/PCR assays. We hypothesize that RNA-based assays using plasma will be more specific in discriminating between radiation doses. Ethical issues preclude systematic examinations of radiation responses in humans, but dogs offer an optimal animal model to examine the in vivo effects of irradiation. Therefore, quantitative RT/PCR assays for candidate genes from Specific Aim 1 will be examined in the canine model to determine which genes display in vivo dose-dependent expression changes and whether plasma or blood cells should be used for the development of dosimetry assays. Dosimetry assays will then be developed using the GeneXpert platform, and these GeneXpert dosimetry assays will be validated in additional dogs. Specific Aim 3. Determine if in vitro dose-dependent genes correlate with radiation exposure in transplant patients. There may be subtle differences in radiation-induced responses between dogs and humans. In addition, exposure over more extended periods of time may invoke different radiation responses. We hypothesize that RNA expression changes may be useful in determining the cumulative dose of irradiation after prolonged exposures. Therefore, quantitative RT/PCR assays for candidate genes from Specific Aim 1 will be examined in transplant patients in order to determine which candidate genes display dose-dependent responses in humans receiving irradiation over several days and whether plasma or blood cells are optimal for the development of dosimetry assays in humans.

如果发生使用放射性材料的恐怖袭击，根据暴露剂量对患者进行风险分层对于确保适当护理至关重要。目前的剂量测定需要数天才能完成，这将延误挽救生命的治疗。该提案旨在使用GeneXpert平台开发基于RNA的剂量测定法，该平台将在辐射暴露后立即对患者进行风险分层。具体目标1.发现在体外照射人外周血细胞后显示剂量依赖性表达变化的基因。辐射暴露引起外周血细胞中剂量依赖性表达变化。我们假设这些表达变化可用于开发基于RNA的检测方法，以区分临床相关剂量的辐射。因此，DNA微阵列将用于鉴定体外照射后造血细胞中具有剂量依赖性表达变化的基因。定量RT/PCR试验将验证这些剂量依赖性表达变化，并将在特定目的2和3中检查候选基因。具体目标2。确定使用血浆或细胞的基于RNA的剂量测定法是否与犬模型中的辐射剂量最相关。辐射损伤的细胞死亡，RNA泄漏到细胞外空间，这种RNA可以使用定量RT/PCR检测。我们假设使用血浆的基于RNA的测定在区分辐射剂量方面将更特异。伦理 这些问题阻碍了对人体辐射反应的系统检查，但狗提供了检查辐射体内效应的最佳动物模型。因此，将在犬模型中检查Specific Aim 1候选基因的定量RT/PCR测定，以确定哪些基因表现出体内剂量依赖性表达变化，以及是否应使用血浆或血细胞来开发剂量测定法。然后将使用GeneXpert平台开发剂量测定试验，并在其他犬中对这些GeneXpert剂量测定试验进行确认。具体目标3。确定体外剂量依赖性基因是否与移植患者的辐射暴露相关。狗和人对辐射的反应可能有细微的差别。此外，长时间暴露可能引起不同的辐射反应。我们假设RNA表达的变化可能有助于确定长期暴露后的累积辐射剂量。因此，定量RT/PCR检测候选基因从 将在移植患者中检查特异性目标1，以确定哪些候选基因在接受辐射数天的人体中显示剂量依赖性反应，以及血浆或血细胞是否最适合用于开发人体剂量测定法。