Rusalatide Acetate (TP508) Mitigation of Genotoxic Radiation Damage in Human Lens Epithelial Cells

醋酸鲁沙拉肽 (TP508) 减轻人晶状体上皮细胞的基因毒性辐射损伤

• 批准号: 10384634
• 负责人: USHA P ANDLEY
• 金额: $ 32.75万
• 依托单位: AFFIRMED PHARMA, LLC
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-02-01 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10384634
• 关键词: Acetates; Amyloid beta-Protein; Animal Model; Animals; Apoptosis; BCL2 gene; Biological Assay; Blindness; CASP3 gene; CHEK1 gene; CHEK2 gene; Cancer Patient; Cataract; Cell Cycle; Cell Cycle Regulation; Cell Death; Cell Line; Cell Proliferation; Cell Survival; Cells; Complex; Cornea; Crystallins; Cyclins; DNA Double Strand Break; DNA Repair; DNA-PKcs; Data; Disease; Dose; Double Strand Break Repair; Drug Delivery Systems; Endothelial Cells; Epithelial; Epithelial Cells; Event; Family; Functional disorder; Future; Genes; Health; Hour; Human; Individual; Investigation; Laboratories; Lead; Lens Fiber; Life; Medical Economics; Methods; Mitotic; Molecular; Necrosis; Occupational Exposure; Oral mucous membrane structure; Pathway interactions; Peptides; Persons; Pharmaceutical Preparations; Phase; Population; Procedures; Proteins; Proton Radiation; Radiation; Radiation Dose Unit; Radiation Induced DNA Damage; Radiation Protection; Radiation Tolerance; Radiation exposure; Radiation induced damage; Radiation therapy; Radio; Retina; Retinal Diseases; Risk; Roentgen Rays; Schedule; Signal Pathway; Signal Transduction Pathway; Signaling Protein; Social Impacts; Societies; Stains; TP53 gene; Techniques; Testing; Therapeutic; Time; Tissues; Treatment Protocols; Western Blotting; Xerostomia; dosage; economic impact; eye dryness; fiber cell; genotoxicity; health related quality of life; improved outcome; irradiation; lens; optimal treatments; p53-binding protein 1; prevent; promoter; protective effect; protein aggregation; protein expression; radiation adverse effect; regenerative; response; senescence; side effect; single fraction radiation

Cancer patients benefit from radiation therapy but can incur side effects to non-targeted tissues including cataracts. Although not directly life threatening, cataract disease has major medical, economic, and social impacts on individuals, families, and society as a whole. Radiation-induced lens opacification is a complex event and has been attributed to DNA double strand breaks (DSB) in the germinative epithelium, leading to defective differentiation of lens fiber cells and subsequent abnormal folding of lens proteins. Rusalatide acetate (TP508) is a radio-modulating peptide that has been shown to increase survival of irradiated animals via activation of signal transduction pathways in endothelial cells, initiating repair of DSB, increasing NO levels and reversing of endothelial cell dysfunction. This investigation will determine if, in the absence of endothelial cells, TP508 will have a similar effect on human lens epithelial cells (HLEC) and mitigate radiation induced pathophysiological pathways that lead to DSB. The hypothesis is that through the direct activation of molecular pathways in irradiated HLEC, TP508 treatment will mitigate or repair DSB. In contrast to other investigative approaches that focus on a single downstream mechanism, this investigation will examine molecular activity of TP508 across multiple pathophysiological pathways associated with the health of HLEC. Study aims are to establish the molecular activity and optimum dosage thresholds, and timing of treatments of TP508 in mitigating X-ray or proton radiation damage with single fraction exposures of 0.5, 1.0, 2.0, or 4 Gy in HLEC (CRL-11421 [B3] and SRA01/04 lines). Aim (1) is to determine the most optimized concentration and administration schedule for TP508 effects on radiation induced HLEC viability using a clonogenic survival assay, MTT assay, and cell doubling time to assess the effects of various concentrations of TP508 on the sensitivity of HLECs applied before and after irradiation. Aim (2) is to further identify the effects of TP508 on specific HLEC molecular responses, using the most optimized concentration and administration schedule of TP508 comparing the single fraction radiation exposures applied with or without TP508, and analyzed with assays for apoptosis, necrosis, senescence, mitotic catastrophe and protein expression; 53BP1 foci staining for dynamics of DSB; and western blot assays for related signaling pathways and protein profiles including amyloid beta, an early marker of cataract formation. Studies are expected to provide the following: (i) determine if TP508 provides a survival effect on irradiated HLEC at different doses applied before and after radiation; (ii) identify if the molecular mechanisms and protein profiles underlying the protective effect of TP508 in HLEC at different doses of radiation are attributed to DSB repair; and (iii) determine the most effective dosage and timing (before or after radiation) of TP508 application. Successfully developed, future investigations of TP508 could expand its application to mitigate additional radiation side effects including dry eye, retinopathy, and xerostomia. .

癌症患者受益于放射治疗，但可能对非靶组织产生副作用，包括 白内障白内障疾病虽然不直接威胁生命，但在医疗、经济和社会方面具有重大意义 对个人、家庭和整个社会的影响。辐射诱导的透镜混浊是一种复杂的 事件，并已被归因于DNA双链断裂（DSB）在生发上皮，导致 透镜纤维细胞的分化缺陷和随后的透镜蛋白质的异常折叠。鲁萨尔茨基 乙酸酯（TP 508）是一种放射性调节肽，已被证明可以增加受辐射的 动物通过激活内皮细胞中的信号转导途径，启动DSB的修复，增加 NO水平和逆转内皮细胞功能障碍。这项调查将确定，如果没有 因此，TP 508对人透镜上皮细胞（HLEC）具有类似的作用，并且减轻了对人晶状体上皮细胞（HLEC）的刺激。 辐射诱导的病理生理途径导致DSB。假设是，通过直接 激活照射后HLEC中的分子通路，TP 508治疗将减轻或修复DSB。在 与关注单一下游机制的其他调查方法相比， 将检查TP 508在与糖尿病相关的多种病理生理学途径中的分子活性。 HLEC的健康研究目的是确定分子活性和最佳剂量阈值， TP 508治疗减轻X射线或质子辐射损伤的时间选择 HLEC中的剂量为0.5、1.0、2.0或4戈伊（CRL-11421 [B3]和SRA 01/04系）。目的（1）是确定最 TP 508对辐射诱导的HLEC活力影响的优化浓度和给药方案 使用克隆形成存活测定、MTT测定和细胞倍增时间来评估各种 TP 508浓度对照射前后应用的HLEC的敏感性的影响。目标（2）是 进一步确定TP 508对特异性HLEC分子反应的影响，使用最优化的 TP 508的浓度和给药方案，比较单次辐射暴露 在有或没有TP 508的情况下施用，并用细胞凋亡、坏死、衰老、有丝分裂 突变和蛋白质表达; 53 BP 1焦点染色用于DSB的动力学;和蛋白质印迹测定用于DSB的免疫印迹分析。 相关的信号通路和蛋白质谱，包括淀粉样蛋白β，白内障的早期标志物 阵研究预期提供以下内容：（i）确定TP 508是否提供了对受试者的存活效果。 在辐射之前和之后施加不同剂量的经辐射的HLEC;（ii）确定分子是否 TP 508在不同剂量下对HLEC的保护作用的机制和蛋白质谱 辐射归因于DSB修复;以及（iii）确定最有效的剂量和时间（之前或之后）。 辐射后）的TP 508应用。成功开发，TP 508的未来研究可以扩大 其用于减轻包括干眼症、视网膜病和口干症的额外辐射副作用。 .