Design Considerations for Massively Parallel Sequencing Studies of Complex Human Disease
Design Considerations for Massively Parallel Sequencing Studies of Complex Human Disease
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DOI：
10.1371/journal.pone.0023221
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发表时间：
2011-08-05
期刊：
影响因子：
3.7
通讯作者：
Goldgar, David E.
中科院分区：
文献类型：
--
作者：
Feng, Bing-Jian;Tavtigian, Sean V.;Goldgar, David E.
大规模并行测序（MPS）现在可以以合理的成本对整个外显子组和基因组进行测序，并且其在识别罕见孟德尔疾病基因方面的效用已得到证明。然而，对于复杂的疾病，研究设计需要适应相当程度的基因座、等位基因和表型异质性，以及基因型和表型之间的复杂关系。这些考虑因素包括仔细选择用于测序的样本，以及制定完善的策略，从许多不相关的基因中识别出少数“真正的”疾病易感性基因，这些基因将被发现含有罕见的变异。为了研究这些问题，我们进行了基于模拟的分析，以比较复杂疾病中 MPS 测序的几种策略。检查的因素包括遗传结构、样本大小、选择进行测序的个体的数量和关系，以及基于变异类型的各种过滤器、基因的多重观察以及谱系内遗传变异的一致性。假设采用两阶段设计，其中来自高风险家庭 MPS 分析的基因在第二次筛选阶段对更多具有较温和家族史的先证者进行评估。使用成本函数评估设计，该函数假设对整个外显子组进行测序的成本是对单个候选基因进行测序的 400 倍。结果表明，虽然要求在多个谱系和/或同一谱系的多个个体中鉴定变异是减少假阳性的有效策略，但存在过度过滤的危险，导致大多数真正的易感基因被遗漏。在大多数情况下，对每个谱系的两个以上个体进行测序会导致功效降低，并且在降低总体成本方面没有任何好处。此外，我们的结果表明，尽管没有单一策略是最佳的，但模拟可以为研究设计提供重要指导。
Massively Parallel Sequencing (MPS) allows sequencing of entire exomes and genomes to now be done at reasonable cost, and its utility for identifying genes responsible for rare Mendelian disorders has been demonstrated. However, for a complex disease, study designs need to accommodate substantial degrees of locus, allelic, and phenotypic heterogeneity, as well as complex relationships between genotype and phenotype. Such considerations include careful selection of samples for sequencing and a well-developed strategy for identifying the few "true" disease susceptibility genes from among the many irrelevant genes that will be found to harbor rare variants. To examine these issues we have performed simulation-based analyses in order to compare several strategies for MPS sequencing in complex disease. Factors examined include genetic architecture, sample size, number and relationship of individuals selected for sequencing, and a variety of filters based on variant type, multiple observations of genes and concordance of genetic variants within pedigrees. A two-stage design was assumed where genes from the MPS analysis of high-risk families are evaluated in a secondary screening phase of a larger set of probands with more modest family histories. Designs were evaluated using a cost function that assumes the cost of sequencing the whole exome is 400 times that of sequencing a single candidate gene. Results indicate that while requiring variants to be identified in multiple pedigrees and/or in multiple individuals in the same pedigree are effective strategies for reducing false positives, there is a danger of over-filtering so that most true susceptibility genes are missed. In most cases, sequencing more than two individuals per pedigree results in reduced power without any benefit in terms of reduced overall cost. Further, our results suggest that although no single strategy is optimal, simulations can provide important guidelines for study design.