Mechanisms of UV-induced DNA damage responses and carcinogenesis in skin

紫外线诱导的皮肤 DNA 损伤反应和致癌机制

• 批准号: 9038985
• 负责人: PAUL NGHIEM
• 金额: $ 38.28万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-04-01 至 2021-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9038985
• 关键词: ATR gene; Address; Apoptosis; Beverages; Biological; Biology; Bypass; Caffeine; Cancer Burden; Cell Cycle Progression; Cell Survival; Cells; Chromatin; Chronic; Complex; DNA Damage; DNA Sequence Alteration; DNA biosynthesis; DNA damage checkpoint; DNA lesion; Data; Development; Dissociation; Dose; Employee Strikes; Epidemiologic Studies; Epidemiology; Flow Cytometry; Genetic; Goals; Health; Human; In Vitro; Incidence; Intake; Lead; Lesion; Malignant Neoplasms; Mediating; Mus; Mutation; Names; Pathway interactions; Phosphotransferases; Polymerase; Preventive; Proteins; Public Health; Pyrimidine Dimers; Recruitment Activity; Regulation; Risk; Role; S Phase; Scientist; Signal Transduction; Site; Skin; Skin Cancer; Source; Stress; System; TP53 gene; Testing; Time; Translating; UV Radiation Exposure; UV induced; base; cancer type; carcinogenesis; combat; high throughput screening; improved; in vivo; inhibitor/antagonist; innovation; insight; kinase inhibitor; mutant; novel; novel strategies; prevent; repaired; response; skin cancer prevention; small molecule; small molecule inhibitor; tool; ultraviolet; ultraviolet irradiation; ultraviolet lesions

 DESCRIPTION (provided by applicant): The number of solar UV-induced skin cancers is greater than all other types of cancers combined. A better understanding of the underlying biology is needed to develop novel approaches to address this problem. UV irradiation instantaneously generates two structurally distinct types of DNA lesion: cyclobutane pyrimidine dimers (CPD) and 6-4 photoproducts (6-4PP). Replicating cells rapidly respond to these deleterious lesions by activating ATR kinase and its downstream target Chk1. However, it is not clear whether these lesions have different effects on DNA replication and/or ATR-Chk1 activation. To address these questions, in Aim 1, we will generate cells that have one, both, or neither type of DNA lesion by combining lesion-specific photolyases with multi-parameter flow cytometry. The proposed studies will test the hypothesis, supported by preliminary data, that there are striking differences in the mechanisms and impact of these two lesion types on DNA replication and UV-DNA damage responses. Human epidemiological studies and mouse in vivo data demonstrate that intake of caffeine (a non- selective ATR inhibitor) prevents UV-associated skin cancers. Indeed, we estimate that 260,000 skin cancers are prevented annually in the U.S. by caffeinated beverage intake. This cancer-preventive effect of caffeine could be further optimized by an improved understanding of its mechanism including whether ATR should be inhibited immediately after UV or long after UV. In Aim 2, we will determine the mechanism by which ATR inhibition suppresses UV carcinogenesis. We will test the hypothesis that immediate (not delayed) ATR inhibition reduces the mutation burden in skin by blocking error-prone lesion bypass. ATR pathway activation requires recruiting more than 10 proteins to a site of DNA damage. Theoretically, it is possible to target any of these proteins to inhibit the ATR pathway, with likely differences in the ability to sensitize to DNA damage. However, current approaches to block this pathway are limited to ATP-competitive inhibitors that target the kinase activity of ATR or Chk1. To identify other druggable mechanisms in this pathway, we performed a cell-based high-throughput screen and identified several small molecules that inhibit the ATR pathway through mechanisms other than ATP competition. One compound ("ARPIN") displayed an important mechanism not shared by any other inhibitor: blockage of Chk1 release from chromatin following damage. In Aim 3, we will use this unique biological effect of ARPIN as a tool to dissect the fundamental mechanism by which Chk1 is normally released from chromatin in order to carry out its effector functions. The three proposed Aims will markedly advance our fundamental understanding of how the replication checkpoint functions, characterize a mechanistically novel inhibitor of the pathway, and provide insight that can be directly translated to further augment an already extensive public health impact. The studies will be highly collaborative with five scientists whose expertise will facilitate critical innovative aspects of te proposed Aims.

 描述(由申请人提供)：太阳紫外线诱发的皮肤癌的数量比所有其他类型的癌症的总和还要多。需要更好地了解潜在的生物学，以开发新的方法来解决这个问题。紫外线照射会瞬间产生两种不同结构的DNA损伤类型：环丁烷嘧啶二聚体(CPD)和6-4光产物(6-4PP)。复制细胞通过激活ATR激酶及其下游靶标Chk1，对这些有害的损伤迅速做出反应。然而，目前尚不清楚这些损伤是否对DNA复制和/或ATR-Chk1激活有不同的影响。为了解决这些问题，在目标1中，我们将通过结合病变特异性光解酶和多参数流式细胞术来产生具有一种、两种或两种类型DNA损伤的细胞。拟议的研究将检验这一假说，并得到初步数据的支持，即这两种损伤类型在DNA复制和UV-DNA损伤反应的机制和影响方面存在显著差异。人类流行病学研究和小鼠体内数据表明，摄入咖啡因(一种非选择性ATR抑制剂)可以预防紫外线相关的皮肤癌。事实上，我们估计，在美国，每年有26万人通过摄入含咖啡因的饮料来预防皮肤癌。咖啡因的这种癌症预防作用可以通过对其机制的进一步了解来进一步优化，包括是否应该在紫外线后立即抑制ATR或在紫外线后很长时间才抑制ATR。在目标2中，我们将确定ATR抑制抑制紫外线致癌的机制。我们将测试这样一种假设，即立即(而不是延迟)抑制ATR通过阻断容易出错的病变旁路来减少皮肤中的突变负担。ATR途径的激活需要招募10个以上的蛋白质到DNA损伤的位置。从理论上讲，靶向这些蛋白质中的任何一种来抑制ATR途径是可能的，但对DNA损伤的敏感度可能有所不同。然而，目前阻断这一途径的方法仅限于以atr的激酶活性为靶点的atp竞争性抑制剂。 或Chk1。为了确定这一途径中的其他可用药机制，我们进行了基于细胞的高通量筛选，并确定了几个通过ATP竞争以外的机制抑制ATR途径的小分子。一种化合物(“精氨酸”)显示了一种其他任何抑制剂都没有的重要机制：损伤后阻止染色质释放Chk1。在目标3中，我们将利用Arpin这一独特的生物学效应作为工具来剖析Chk1正常地从染色质中释放出来以实现其效应功能的基本机制。提出的三个目标将显著促进我们对复制检查点如何发挥作用的基本理解，表征一种机械上新颖的途径抑制物，并提供可以直接翻译的见解 进一步扩大本已广泛的公共卫生影响。这些研究将与五名科学家高度合作，他们的专业知识将促进TE拟议目标的关键创新方面。