A new approach to identify rare genetic variants influencing melanoma risk

描述（由申请人提供）：许多全基因组关联研究（GWAS）已经成功地在复杂性状的病因学中发现了共同的snp。到目前为止，发现的大多数变异导致的风险增量相对较小（1.1-1.5倍），并且只能解释一小部分家族聚类。最近的研究表明，常见疾病可能是由于具有广泛等位基因频率的功能失调变异。到目前为止，罕见变异的研究仅限于少数表型和基因，但测序技术的进步应该会导致候选基因和基因组的广泛关联研究。如果罕见的变异比常见的变异有更大的影响，这应该有助于它们的检测。此外，它们的识别应对风险评估、疾病预防和治疗产生更大的影响。然而，罕见变异的分析是具有挑战性的，因为用于常见变异的方法能力不足。先前，我们使用来自乳腺癌患者和对照组的突变筛选数据来证明从截断和剪接变异体以及罕见的错义替换中检测致病性证据的能力。该方法包括将病例和/或对照组中观察到的罕见错义替换按从最小到最可能对进化有害的一系列等级进行分层，然后进行逻辑回归趋势检验，比较病例和对照组中变异等级的频率分布。最初的模型是为了评估乳腺癌相关基因错义替换的致病性而开发的。在这里，我们提出测试我们的分析策略对皮肤恶性黑色素瘤（CMM）的效率。为了确定新的风险等位基因，我们将对来自10个欧洲国家的1300多例患者和1300名匹配对照患者的色素沉着途径最强候选基因进行突变筛选。由于主要环境因素（暴露于太阳紫外线辐射）与已知易感基因之间的关系已经相当清楚，CMM为研究多因子疾病发展过程中的基因-基因和基因-环境相互作用提供了一个独特的模型。大约20-25%的黑色素瘤易发家族携带CDKN2A和CDK4的高风险突变。相比之下，色素沉着基因MC1R中的一些错义替换已被证明是中等风险或中等风险的易感等位基因，并且也增加了CDKN2A突变的外显率。最后，两个最近的GWAS在MC1R或相同途径的基因中发现了低外显率snp。相关snp占家族相对风险的比例不超过12%。因此，对CMM的大部分遗传易感性仍有待解释。在我们的方法在强有力的候选者上得到验证后，我们设想对整个生化途径的基因进行大规模平行测序，以产生与黑色素瘤易感性相关的致病序列变异的风险频谱的全面图景。

DESCRIPTION (provided by applicant): A number of genome-wide association studies (GWAS) have successfully implicated common SNPs in the etiology of complex traits. Most variants identified so far confer relatively small increments in risk (1.1-1.5-fold), and explain oly a small proportion of familial clustering. Recent studies have demonstrated that common diseases can be due to dysfunctional variants with a wide spectrum of allele frequencies. So far, rare variants studies have been limited to a handful of phenotypes and genes, but the advancement of sequencing technologies should lead to widespread association studies of candidate genes and genomes. If rare variants have larger effects than common variants, this should aid in their detection. Also their identification should have a greater impact on risk assessment, disease prevention and treatment. However, the analysis of rare variants is challenging since methods used for common variants are underpowered. Previously, we used data from mutation screening of breast cancer patients and controls to demonstrate the ability to detect evidence of pathogenicity from both truncating and splice junction variants and rare missense substitutions. The method involves stratifying rare missense substitutions observed in cases and/or in controls into a series of grades ordered from least to most likely to be evolutionarily deleterious, followed by a logistic regression trend test to compare the frequency distributions of the grades of variants in cases versus controls. The original model was developed to assess the pathogenicity of missense substitutions in breast cancer-related genes. Here we propose to test the efficiency of our analysis strategy to cutaneous malignant melanoma (CMM). To identify novel risk alleles, we will mutation screen the strongest candidate genes of the pigmentation pathway in over 1,300 cases and 1,300 matched controls from 10 European countries enrolled in the EPIC cohort. CMM provides a unique model for studies of gene-gene and gene-environmental interactions in the development of multifactorial diseases, since relationships between the major environmental factor (exposure to solar UV radiation) and known susceptibility genes are reasonably well understood. High-risk mutations in CDKN2A and CDK4 are carried by about 20-25% of melanoma-prone families. In contrast, some missense substitutions in the pigmentation gene MC1R have been proven to be modest-risk or intermediate-risk susceptibility alleles, and also to increase the penetrance of CDKN2A mutations. Finally, two recent GWAS identified low- penetrance SNPs in MC1R or in genes of the same pathway. Associated SNPs will account for no more than 12% of the Familial Relative Risk. Thus, the majority of the genetic susceptibility to CMM remains to be explained. After validation of our method on strong candidates, massive parallel sequencing of genes of entire biochemical pathways is envisaged to generate a comprehensive picture of the risk-frequency spectrum for pathogenic sequence variants involved in susceptibility to melanoma.