DNA Repair Phenotype the Missing Link in Breast Cancer Risk Assessment

摘要 DNA修复是维持细胞基因组稳定性的重要机制。DNA修复机制中的缺陷 增加细胞对DNA损伤剂的脆弱性和基因组中突变的积累，并导致 包括癌症在内的各种疾病的发展。已测量DNA修复能力(DRC)的研究， 包括我们自己在内，估计患乳腺癌(BC)的风险比大多数其他人高得多(3-15倍 除了BRCA1和BRCA2等基因的高度穿透性突变外，已确定的BC危险因素， 对DNA修复至关重要的基因。尽管这种关联性很强，但没有关于卑诗省的大规模前瞻性研究 是存在的。尽管一些BC风险模型包括已知的DNA修复基因突变，但基因分型只是部分 解释了表型，而BC风险模型目前不包括表型DNA修复措施。缺乏 包括一个主要的风险因素--DRC--可能是临床上的BC风险模型只有适度的主要原因 绩效--这使得以有效的初级预防方案为目标非常具有挑战性(例如， 化学预防)，用于大多数不是已知突变携带者的女性。此外，二级预防 选项(例如，通过乳房X光检查或其他补充方法进行BC筛查的开始时间、频率和方法) 如果有更准确的风险评估，可以更有效地瞄准目标。DRC使用的主要限制 对于靶向预防，一直缺乏高通量的DRC分析，特别是表型DRC分析， 整合到癌症风险评估中。我们通过调整我们的高吞吐量，克服了这一重大差距， 全自动ɣ-H_2AX分析系统，最初设计用于检测DNA双链断裂 (Dsb)新鲜抽出的血液，与档案血样一起使用。我们提出了最大的潜在客户之一 使用一组丰富的队列(n=12,563)评估DSB修复效果的研究 绝对的BC风险。在这个队列中使用嵌套病例对照设计(699例，1：1匹配)，我们将测量 基线收集的档案生物标本中的DSB-DRC(目标1a)。我们将优化检测方案，以满足 新鲜指血测定DSB-DRC及青少年DSB-DRC纵向变化 妇女(年龄40岁)(目标1b)(n=100，间隔1-2岁)。然后我们将全面评估 DSB-DRC对DSB修复基因的遗传和表观遗传改变的独立贡献，并评估 以及遗传和表观遗传变化是否与DSB-DRC相互作用增加BC风险(目标2)。我们会 通过量化标准BC风险模型性能的改善来研究DSB-DRC的临床应用 DSB-DRC与BC术后5年存活率之间的关系 诊断(目标3b)。我们的研究将提供必要的经验证据，将功能分析整合到 人口研究，以加快与DNA损伤异常反应相关的有针对性的预防选择。这 研究将由BC流行病学、分子生物学领域的一组老牌调查人员领导 流行病学、高通量DNA修复能力评估和生物统计学。

ABSTRACT DNA repair is a crucial mechanism for maintaining genomic stability in cells. Defects in the DNA repair machinery increase cell vulnerability to DNA-damaging agents and accumulation of mutations in the genome, and lead to the development of various disorders including cancers. Studies that have measured DNA repair capacity (DRC), including our own, have estimated a much higher risk of breast cancer (BC) (3-15-fold) than most other established risk factors for BC, with the exception of highly penetrant mutations in genes like BRCA1 and BRCA2, genes critical to DNA repair. Despite the strength of this association, no large-scale prospective studies of BC exist. Even though some BC risk models include known mutations in DNA repair genes, genotype only partially explains phenotype, and BC risk models currently do not include phenotypic DNA repair measures. The lack of inclusion of a major risk factor – DRC – is likely the major reason that clinical BC risk models have only modest performance - which makes it very challenging to target effective primary prevention options (e.g., chemoprevention) for the majority of women who are not known mutation carriers. Further, secondary prevention options (e.g., onset, frequency, and method of BC screening by mammography or other supplemental methods) could be targeted more efficiently if more accurate risk assessment existed. The main limitation of use of DRC for targeted prevention has been the lack of a high-throughput DRC assay, in particular a phenotypic DRC assay, for integration into cancer risk assessment. We have overcome this major gap by adapting our high-throughput, fully-automated ɣ-H2AX assay system which was originally designed for assaying DNA double strand breaks (DSB) in freshly-drawn blood for use with archival blood samples. We propose one of the largest prospective studies estimating the effect of DSB repair using an enriched cohort (n=12,563) that spans the spectrum of absolute BC risk. Using a nested case-control design within this cohort (699 cases, 1:1 match), we will measure DSB-DRC in archival biospecimens collected at baseline (Aim 1a). We will optimize the assay protocol for measuring DSB-DRC using fresh fingerstick blood and measure longitudinal changes in DSB-DRC in young women (age <40 years) (Aim 1b) (n=100, 1-2 years apart). We will then comprehensively assess the independent contribution of DSB-DRC over genetic and epigenetic alterations in DSB repair genes, and assess and whether genetic and epigenetic changes interact with DSB-DRC in increasing BC risk (Aim 2). We will investigate the clinical utility of DSB-DRC by quantifying the improvement in standard BC risk model performance from its inclusion (Aim 3a), and evaluating the association between DSB-DRC and 5 year survival after BC diagnosis (Aim 3b). Our study will provide essential empirical evidence from integrating functional assays into population studies to accelerate targeted prevention options linked to aberrant responses to DNA damage. This research will be led by a team of established investigators in the fields of BC epidemiology, molecular epidemiology, high-throughput DNA repair capacity assessment, and biostatistics.