Discovery of New Genes for Inherited Predisposition to Breast Cancer by Exome Seq

描述（由申请人提供）：该项目的目标是识别乳腺癌遗传易感性的新基因。DNA捕获和大规模并行测序技术将被用来在1822个家庭的队列中识别这些基因，每个家庭至少有四个患有乳腺癌的亲属。其中1500多个家庭中乳腺癌的遗传原因仍未得到解决。在这些家族中发现新的乳腺癌基因涉及三个目标。在AIM 1中，所有已知的乳腺癌遗传易感性基因将在150个家庭的所有受影响亲属中进行完全测序。所有未解决家族的先证者在BRCA1、BRCA2、CHEK 2、PALB 2、CHEK 2、BRIP 1、p53和PTEN处具有野生型序列。然而，其他受影响的家庭成员可能携带这些基因的突变，先证者可能是散发病例。受影响亲属的组成DNA将用于制备配对末端文库，其将与定制寡核苷酸池杂交以捕获所有已知的乳腺癌易感基因，然后将富集的文库在Illumina GAIIx基因组分析仪上以多重设计进行条形码编码和测序。突变将通过桑格测序进行验证。在AIM 2中，将再次使用Illumina平台，对所有已知乳腺癌基因的野生型序列进行筛选中的所有女性的整个外显子组进行完全测序。将过滤变体以鉴定破坏基因的罕见无义突变、移码、剪接位点突变以及基因组缺失和重复。筛选的突变将被验证并测试与乳腺癌的共遗传。多种有害突变是所有已知遗传性乳腺癌基因的标志，也是新乳腺癌基因功能后果的最有力证据。在Aim 3A中，最有希望的候选基因将在来自一系列独立的家族性乳腺癌患者的DNA样本中进行完全测序，以揭示其他突变。在Aim 3B中，将在大型随访系列中通过TaqMan检测对这些基因中的潜在功能突变进行基因分型，以估计个体和全基因突变频率和相对风险。新的乳腺癌易感基因的发现将使预防策略扩展到目前尚不清楚的致病基因的家庭，以更好地识别处于风险中的妇女，允许对这些妇女进行更密切的医疗监督，刺激基于遗传证据的新预防策略的设计，并提供更好地了解乳腺癌发展所涉及的生物学途径。

DESCRIPTION (provided by applicant): The goal of this project is to identify new genes for inherited susceptibility to breast cancer. DNA capture and massively parallel sequencing technologies will be exploited to identify these genes in a cohort of 1822 families, each with at least four relatives with breast cancer. The genetic causes of breast cancer in more than 1500 of these families remain unresolved. Discovery of new breast cancer genes in these families involves three aims. In AIM 1, all known genes for inherited predisposition to breast cancer will be fully sequenced in all affected relatives from 150 of the families. The probands of all unresolved families have wildtype sequences at BRCA1, BRCA2, CHEK2, PALB2, CHEK2, BRIP1, p53, and PTEN. However, other affected family members may carry mutations in these genes, and the proband may be a sporadic case. Constitutional DNA of affected relatives will be used to prepare paired-end libraries, which will be hybridized to custom oligonucleotide pools to capture all known breast cancer susceptibility genes, then the enriched libraries will be bar-coded and sequenced in a multiplex design on an Illumina GAIIx genome analyzer. Mutations will be validated by Sanger sequencing. In AIM 2, the Illumina platform will be used again, to fully sequence the entire exomes of all women in the screen with wildtype sequences at all known breast cancer genes. Variants will be filtered to identify rare nonsense mutations, frameshifts, splice site mutations, and genomic deletions and duplications that disrupt genes. Filtered mutations will be validated and tested for co-inheritance with breast cancer. Multiple deleterious mutations are a hallmark of all known genes for inherited breast cancer and are the strongest proof of functional consequence of new breast cancer genes. In Aim 3A, the most promising candidate genes will be fully sequenced in DNA samples from an independent series of familial breast cancer patients to reveal additional mutations. In Aim 3B, potentially functional mutations in these genes will be genotyped by TaqMan assays in large follow-up series to estimate individual and gene-wide mutation frequencies and relative risks. Discovery of new breast cancer susceptibility genes will allow prevention strategies to be extended to families for which causal genes are currently unknown, to better identify women at risk, to allow closer medical surveillance of these women, to stimulate design of new prevention strategies based on genetic evidence, and to offer a better understanding of the biological pathways involved in breast cancer development.