New Familial Colon Cancer Gene Discovery Via Combined Linkage and SNP Association

描述(申请人提供)：我们的研究小组已经发现并确认了一个新的染色体区域，9q22.2-31.2，它证明了(P=0.00016)与家族性结肠肿瘤连锁的证据，该模式与该区域的常染色体显性疾病等位基因一致，约占我们研究的结肠肿瘤家族中结肠癌和晚期结肠腺瘤的40%。我们最初的发现是对同胞配对研究的连锁分析的结果，该研究旨在基于无偏见的全基因组扫描识别新的人类结肠癌易感基因，该研究采用来自53个“研究-4”家庭的同胞配对，其中两个或更多的兄弟姐妹在65岁或更年轻时受到结肠癌的影响，或者受到晚期结肠腺瘤(大小1厘米或组织学显示高度不典型增生)的影响，这些腺瘤是癌症的直接先兆。我们的初步发现现已在120个“研究-4”家庭的扩大队列中得到证实，英国和斯堪的纳维亚半岛的独立调查人员也证实了这一点。经过精细定位，我们目前精细化的连锁区域跨度为8.8Mb。这一新建议的主要目的是使用基于非连锁的技术来进一步缩小该候选区域，例如i)SNP关联，以及ii)定位家族性肿瘤目标丢失或获得的基因区域；然后在9q22.2-31.2区间内识别实际的新疾病基因和致病等位基因。我们现在建议的具体步骤是：i)分析9q22.2-31.2区间，通过分别检查该区间是否杂合性缺失(LOH)(或基因拷贝数增加)来分析9q22.2-31.2区间是否存在肿瘤抑制基因或癌基因。然后通过定义这些家族性结肠肿瘤共有的最小LOH(或基因拷贝数增加)区域来进一步缩小这个基因组间隔。Ii)检查疾病之间的关联{5147个SNP多态的综合小组，平均间距为2.7kb，捕捉到9q22.2-31.2连锁区间中常见和罕见的遗传变异；}从而直接或间接地定位任何潜在的创始人疾病等位基因的位置，这些等位基因将解释许多或所有这些结肠肿瘤家族的肿瘤发展。Iii)通过对我们最窄的9q22.2-31.2区间内的所有基因进行高通量测序，研究我们最好的连锁家系，检查目前缩小的8.8Mb连锁区间内最多包含102个基因，从而直接寻找潜在的疾病基因和致病等位基因。IV)在我们最好的连锁家庭中构建“转化克隆”人-鼠体细胞杂交，以捕获单个亲本人类染色体9个拷贝。利用这些捕获的单倍体双亲染色体来测试我们最大限度地缩小区间内的基因，以检测标准方法无法检测到的致病突变，包括检测外显子缺失和导致外显子跳过的突变。这一项目的成功将对公共卫生产生重大影响。公共卫生相关性：结肠癌是美国癌症死亡的第二大原因，20%的结肠癌患者报告有结肠癌阳性家族史，也影响到父母或兄弟姐妹。识别一种新的基因，这是家族性结肠癌的常见原因，将有助于识别这种疾病的高危人群，对他们来说，积极的筛查和早期检测可能会挽救他们的生命。

DESCRIPTION (provided by applicant): Our group of investigators has identified and confirmed a novel chromosomal region, 9q22.2-31.2, that demonstrates evidence for linkage (P=0.00016) to familial colon neoplasia in a pattern consistent with dominant autosomal disease alleles in this region accounting for an estimated 40% of colon cancers and advanced colon adenomas in the colon neoplasia families we studied. Our initial finding was the result of linkage analysis of a sibling pair study that was designed to identify novel human colon cancer susceptibility loci based on an unbiased whole genome scan employing sibling pairs drawn from 53 "study-4" kindreds in which 2 or more siblings had by age 65 or younger been affected by colon cancer, or affected by advanced colon adenomas (of size >1cm or with histology demonstrating high grade dysplasia) that are direct cancer precursors. Our initial discovery has now been confirmed by us in an enlarged cohort of 120 "study-4" kindreds, as well by independent investigators in Britain and in Scandinavia. After fine mapping, our currently refined linkage region spans 8.8Mb. The main purpose of this new proposal is to further narrow this candidate region using non-linkage based techniques, such as i) SNP association, and ii) mapping gene regions that familial tumors target for loss {or for gains}; followed by then identifying the actual novel disease gene and pathogenetic disease alleles within the 9q22.2-31.2 interval. The specific steps we thus now propose are: i) To analyze the 9q22.2-31.2 interval for evidence of either a tumor suppressor gene {or an oncogene} by examining this interval for, respectively, either loss of heterozygosity (LOH) {or for increased gene copy number} in the tumors arising in kindreds that show linkage to the region. To then further narrow this genomic interval by defining the minimal region of LOH {or of increased gene copy number} that is shared in common among these familial colon tumors. ii) To examine disease association among {a comprehensive panel of 5147 SNP polymorphisms, with average spacing of average 2.7kb, that capture both the common and the rare genetic variation across the 9q22.2-31.2 linkage interval;} thereby directly or indirectly mapping the location of any potential founder disease allele that would account for tumor development in many or all of these colon neoplasia families. iii) To directly look for potential disease genes and pathogenetic disease alleles by studying our best linked kindreds via high throughput sequencing of all genes within our maximally narrowed 9q22.2-31.2 interval, examining a maximum of 102 genes contained within our currently narrowed 8.8Mb linkage interval. iv) To in our best linked families construct "converted clone" human-mouse somatic cell hybrids to capture individual parental human chromsome 9 copies. To employ these captured haploid parental chromsomes to test the genes in our maximally narrowed interval for disease causing mutations that would not be detected by standard approaches, including testing for exon deletions and testing for mutations that cause exon skipping. Success of this project would have substantial implications for public health. PUBLIC HEALTH RELEVANCE: Colon cancer is the second leading cause of cancer death in the United States, with 20% of colon cancer patients reporting a positive family history of colon cancer also affecting either a parent or sibling. Identifying a new gene that is a common cause of familial colon cancer would allow identification of individuals at high risk for this disease, for whom aggressive screening and early detection could be life saving.