The potential for next generation sequencing to characterise the genetic variation underlying nonsyndromic cleft lip and palate phenotypes

Abstract

Genetics of Complex DiseasesAbstract Introduction Next-generation sequencing is revolutionising the study of genetic variation and its role in disease. Individual DNA samples can now be sequenced cost-effectively enabling analysis of the complete spectrum of genetic variation. This technology has the potential to contribute significantly to the understanding of non-syndromic cleft lip and/or palate. This condition occurs with relatively high frequency and only a proportion of the underlying genetic causal factors have been identified. Many of the genes implicated have been found through genome-wide association studies but further progress is limited because these approaches consider only common genetic variants and neglect rarer variations. Because many of the causal genetic variants remain unknown, the role of gene-environment and genegene interaction is difficult to characterise. The identification of novel, low frequency, variants will provide new insights into the biological mechanisms and pathways involved in the condition. Sequence-based analysis will also be invaluable for fine mapping causal variants in the larger regions already identified by linkage and association studies for which positive identification of causal genetic variants has proven difficult. This review considers the available evidence for the genes involved and current understanding of how genetic variation interacts with environmental factors known to influence risk. Only by characterising the underlying genetic factors will the effort to understand gene-environment interaction and underlying functional processes be successful. Conclusion Success with next-generation sequencing will lead to improvements in prediction, prevention, and treatment for cleft lip and palate patients. IntroductionNext-generation sequencing (NGS) is revolutionising genomics 1 and this trend of increasingly significant impact is likely to continue. Most importantly, NGS provides a route to characterising and understanding the role of genetic variation in disease. This is important because evidence from genome-wide association studies (GWAS) suggests that much of the heritability underlying complex disease phenotypes is not explained by common deleterious genetic variants with small effect sizes 2 . NGS enables new analytical strategies, which are not achievable through genome-wide association studies. These include: the identification of the complete complement of DNA variants in samples (across the allele frequency spectrum); tests on the burden of rare variation(do specific genes contain many rare variants which collectively impair gene function?); the identification of de novo mutations and the genes underlying rare Mendelian forms of disease; fine mapping of causal variants within broader regions identified by linkage and/or association and the characterisation of important structural variation, such as differences in copy number, which may contribute to disease.Orofacial cleft lip and/or palate (CLP) represents a complex phenotype for which NGS offers the po...

Deleterious coding variants in multi-case families with non-syndromic cleft lip and/or palate phenotypes

“…For investigations aiming to resolve the genetic factors underlying NSCLP in multiple case families, exome sequencing presents a relatively cost-effective approach in which sequencing a small number of affected family members can identify candidate underlying genetic variation. NSCLP provides a particular challenge for genetic studies, with incomplete penetrance and environmental factors hindering the identification of aetiological variance 2 39 . We have aimed to minimise this effect by careful selection of pedigrees exhibiting clefting in multiple individuals, where we would expect a stronger genetic component.…”

“…The list is derived in part (363 genes out of the 865) from the professional Human Gene Mutation Database 12 , using search terms related to clefts and clefting syndromes. The remaining genes in the list were included after corresponding interrogation of OMIM 13 , and a small number of additional CLP-related genes from the review by Collins et al 2 .…”

“…Genetic studies including linkage analysis, genome-wide association (GWAS), and GWAS-based meta-analysis, have yielded reproducible evidence for the involvement of several genes and gene regions. Collins et al 2 , listed 16 genes and gene regions which have been firmly implicated in NSCLP through linkage and association analysis. Several of these are broad regions where the underlying causal variant(s) have yet to be pinpointed, however, polymorphisms in genes such as IRF6 are strongly associated with NSCLP 3 and more minor roles have been established for MSX1 4 5 , PVRL1 , FGFR2 , PAX7 , NOG and SPRY2 among others 6 .…”

Resolving clinical diagnoses for syndromic cleft lip and/or palate phenotypes using whole‐exome sequencing

“…This list comprised 363 genes. Additional genes were included after a corresponding interrogation of OMIM (http://omim.org/, accessed July 2014), and a small number of additional CLP‐related genes from the review were also included by Collins et al . The complete list of 865 genes considered in variant filtering is given in Table S2.…”

“…Samples from six individuals with syndromic phenotypes from three families were exome-sequenced. DNA derived from peripheral blood was sequenced for five I1 I2 II1 II3 II2 III1 III10 III9 I V1 I V2 I V3 III11 III12 III13 P III14 III15 III2 III3 III4 III5 III6 III7 III8 II4 II5 II6 II7 II8 II9 I3 II10 were also included by Collins et al (7). The complete list of 865 genes considered in variant filtering is given in Table S2.…”