Development of Cell-based Functional Tests for Rare Germline ATM Gene Variants in Hereditary Ovarian Cancer Families

遗传性卵巢癌家族中罕见种系 ATM 基因变异的基于细胞的功能测试的开发

基本信息

  • 批准号:
    9307327
  • 负责人:
  • 金额:
    $ 7.7万
  • 依托单位:
  • 依托单位国家:
    美国
  • 项目类别:
  • 财政年份:
    2017
  • 资助国家:
    美国
  • 起止时间:
    2017-08-15 至 2019-07-31
  • 项目状态:
    已结题

项目摘要

Whole genome sequencing of DNA from subjects at high risk for cancer predisposition has led to the identification of numerous variants of unknown significance, VUSs. We evaluated 48 women from high-risk families diagnosed with ovarian cancer for germline mutations by whole exome sequencing of their germline DNA. These subjects were previously found to be negative for any known pathogenic mutations in either BRCA1 or BRCA2. Numerous genetic variants with a moderate to high effect in relevant pathways were found to be present in this cohort. In particular, we identified 2 deleterious and 5 potentially damaging variants in the ATM gene in the normal DNA from these high-risk women with ovarian cancer. The role of ATM in ovarian cancer has been underestimated because of a lack of functional assays for the effects of these genetic changes. In this project we will pilot the development of a general approach to determine the nature of private and rare mutations in DNA repair associated genes using cell-based functional assays. The functional analysis of novel germline variants in the gene ataxia-telangiectasia mutated (ATM) will be studied in immortalized normal ovarian and mammary epithelial cells. This approach is superior to using established tumor cells that already have DNA repair defects. The PI's lab is experienced in studying the genetics of cancer and the ramifications of mutant hereditary cancer genes on normal cell biology and genomic instability focusing previously on Li-Fraumeni Syndrome. ATM codes for a protein kinase that regulates the DNA damage response initiating the signaling cascades that activate cell cycle checkpoints and the repair of DNA double-strand breaks. We will develop cell-based assays that can be used for the functional analysis of genetic variants of unknown significance (VUSs) in the ATM gene. ATM mutations are associated with the risk of breast cancer; however little information is available on ovarian cancer. Therefore, we will compare the biological and biochemical impact on immortalized normal mammary and ovarian epithelial cells that have had their ATM genes edited to contain the SNPs that we have identified in patients' DNA. We will determine whether these clearly deleterious and potentially damaging variants have significant impact on cell cycle or DNA repair mechanisms in a cell type specific fashion. Deleterious and potentially damaging VUSs in ATM were statistically overrepresented in our cohort (Table 1) compared to the MAF in the 1000 genomes database. However each VUS will require some functional analysis to provide clinically useful information to patients. This project will be pursued in two aims: AIM 1: We will employ gene editing of the endogenous ATM gene in hTERT immortalized normal mammary and ovarian epithelial cells using CRISPR/Cas9 to produce potentially deleterious ATM variants. AIM 2: Using functional assays in pathways known to be associated with inherited risk of ovarian cancer we will determine the impact of these VUSs on cell cycle control and DNA repair mechanisms.
对来自癌症易感性高风险受试者的DNA进行全基因组测序, 鉴定了许多意义不明的变异体,VUS。我们评估了48名高风险女性, 通过全外显子组测序诊断为卵巢癌的生殖系突变的家庭, 生殖系DNA这些受试者先前被发现对任何已知的致病性突变呈阴性, BRCA 1或BRCA 2。在相关途径中具有中度至高度影响的多种遗传变异 在这个群体中也有。特别是,我们确定了2种有害物质和5种潜在的破坏性物质, 这些高危卵巢癌妇女正常DNA中ATM基因的变异。的作用 ATM在卵巢癌中的作用一直被低估,因为缺乏对ATM作用的功能测定。 这些基因变化。在本项目中,我们将试点开发一种通用方法,以确定 使用基于细胞的功能测定,研究DNA修复相关基因中私人和罕见突变的性质。 将对共济失调-毛细血管扩张突变(ATM)基因中新的种系变异体进行功能分析。 在永生化的正常卵巢和乳腺上皮细胞中研究。这种方法上级使用 已经存在DNA修复缺陷的肿瘤细胞。PI的实验室在研究 癌症遗传学和突变遗传性癌症基因对正常细胞生物学的影响, 基因组不稳定性之前集中在李-弗劳梅尼综合征。ATM编码一种蛋白激酶 调节DNA损伤反应,启动激活细胞周期检查点的信号级联反应 和DNA双链断裂的修复。我们将开发基于细胞的检测方法, ATM基因中未知意义的遗传变体(VUS)的功能分析。ATM突变 与乳腺癌的风险有关;然而,关于卵巢癌的信息很少。 因此,我们将比较对永生化正常乳腺癌的生物学和生化影响, 卵巢上皮细胞的ATM基因经过编辑,包含我们在 病人的DNA我们将确定这些明显有害和潜在破坏性的变异是否具有 以细胞类型特异性方式对细胞周期或DNA修复机制产生显著影响。有害和 在我们的队列中,ATM中潜在的损害性VUS在统计学上占比过高(表1), 千人基因组数据库里的MAF但是,每个VUS都需要进行一些功能分析, 为患者提供临床有用的信息。这一项目的目标有两个: 目的1:我们将在hTERT永生化的正常人中使用内源性ATM基因的基因编辑, 乳腺和卵巢上皮细胞使用CRISPR/Cas9产生潜在有害的ATM变体。 目的2:在已知与卵巢癌遗传风险相关的途径中使用功能测定 我们将确定这些VUS对细胞周期控制和DNA修复机制的影响。

项目成果

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MICHAEL A TAINSKY其他文献

MICHAEL A TAINSKY的其他文献

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{{ truncateString('MICHAEL A TAINSKY', 18)}}的其他基金

Diagnostic Assays for OVCA Recurrence using Paraneoplastic Antigens and Epitopes
使用副肿瘤抗原和表位诊断 OVCA 复发
  • 批准号:
    8887317
  • 财政年份:
    2014
  • 资助金额:
    $ 7.7万
  • 项目类别:
Diagnostic Assays for OVCA Recurrence using Paraneoplastic Antigens and Epitopes
使用副肿瘤抗原和表位诊断 OVCA 复发
  • 批准号:
    8753213
  • 财政年份:
    2014
  • 资助金额:
    $ 7.7万
  • 项目类别:
Validation of an Antibody Test for Early Diagnosis of Ovarian Cancer
卵巢癌早期诊断抗体测试的验证
  • 批准号:
    8154030
  • 财政年份:
    2011
  • 资助金额:
    $ 7.7万
  • 项目类别:
Validation of an Antibody Test for Early Diagnosis of Ovarian Cancer
卵巢癌早期诊断抗体测试的验证
  • 批准号:
    8509625
  • 财政年份:
    2011
  • 资助金额:
    $ 7.7万
  • 项目类别:
Serum Biomarkers for Colorectal Cancer Detection
用于结直肠癌检测的血清生物标志物
  • 批准号:
    7586435
  • 财政年份:
    2009
  • 资助金额:
    $ 7.7万
  • 项目类别:
Serum Biomarkers for Colorectal Cancer Detection
用于结直肠癌检测的血清生物标志物
  • 批准号:
    7826919
  • 财政年份:
    2009
  • 资助金额:
    $ 7.7万
  • 项目类别:
Early Detection of Breast Cancer Using Autoantibody Mar*
使用自身抗体 Mar* 早期检测乳腺癌
  • 批准号:
    7105117
  • 财政年份:
    2005
  • 资助金额:
    $ 7.7万
  • 项目类别:
Early Detection of Breast Cancer Using Autoantibody Mar*
使用自身抗体 Mar* 早期检测乳腺癌
  • 批准号:
    7003625
  • 财政年份:
    2005
  • 资助金额:
    $ 7.7万
  • 项目类别:
Early Detection of Breast Cancer Using Autoantibody Mar*
使用自身抗体 Mar* 早期检测乳腺癌
  • 批准号:
    7287417
  • 财政年份:
    2005
  • 资助金额:
    $ 7.7万
  • 项目类别:
Early Detection of Breast Cancer Using Autoantibody Mar*
使用自身抗体 Mar* 早期检测乳腺癌
  • 批准号:
    7681241
  • 财政年份:
    2005
  • 资助金额:
    $ 7.7万
  • 项目类别:

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非洲人群中 HIV 氨基酸变异与 CHD1L 和 HLA I 类基因座的保护性宿主等位基因的关联
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