Exome sequencing can improve diagnosis and alter patient management

Abstract

The translation of "next-generation" sequencing directly to the clinic is still being assessed but has the potential for genetic diseases to reduce costs, advance accuracy, and point to unsuspected yet treatable conditions. To study its capability in the clinic, we performed whole-exome sequencing in 118 probands with a diagnosis of a pediatric-onset neurodevelopmental disease in which most known causes had been excluded. Twenty-two genes not previously identified as disease-causing were identified in this study (19% of cohort), further establishing exome sequencing as a useful tool for gene discovery. New genes identified included EXOC8 in Joubert syndrome and GFM2 in a patient with microcephaly, simplified gyral pattern, and insulin-dependent diabetes. Exome sequencing uncovered 10 probands (8% of cohort) with mutations in genes known to cause a disease different from the initial diagnosis. Upon further medical evaluation, these mutations were found to account for each proband's disease, leading to a change in diagnosis, some of which led to changes in patient management. Our data provide proof of principle that genomic strategies are useful in clarifying diagnosis in a proportion of patients with neurodevelopmental disorders.

Fig. 1

Summary of probands prioritized for whole-exome sequencing analysis. A total of 188 probands, each from a unique family, were priori-tized for analysis in this study, and of these, 40 (21%) had mutations in known disease-causing genes as determined by candidate gene screening. Thirty demonstrated a single linkage peak (16%) and were thus not candidates for whole-exome sequencing. The remaining 118 were analyzed by whole-exome sequencing, and of these, 10 were found to have mutations in genes known to cause a disease different from their initial diagnosis (8%). In all 10 cases, the whole-exome sequencing information led to a modification of the diagnosis.

Fig. 2

Variant filter, prioritization, and validation pipelines. All whole-exome sequencing variants were filtered to include coding variants, splice variants or INDELs (insertions-deletions), nonsynonymous amino acid changes, homozygous genotype calls, variants not found in homozygous form in population studies, and those found in linkage intervals. The remaining variants were then prioritized on the basis of type of mutation (INDELs > nonsense > missense), conservation of the amino acid across species, predicted damage to the protein, and relevance to neurodevelopmental disorders. Variant validation pipeline: Variants from the filtering pipeline were then checked for segregation according to a recessive model of disease and their absence in 200 ethnically matched control individuals.

Fig. 3

Genetic data for family 650. (A) Pedigree and MRI scans from the living affected members of family 650. In the pedigree, an asterisk denotes that individual's DNA was used in linkage analysis. The brain MRI (axial, midline sagittal, and coronal views) of one affected member shows a thickened cortex with a paucity of white matter, simplified gyral pattern, and thickened cerebral mantle, compatible with pachygyria. (B) Linkage plot for family 650 shows LOD score plotted against chromosome number and reveals one major linkage block on chromosome 5. (C) Final variant table after whole-exome sequencing analysis reveals four candidate genes, only one of which segregates in the family. Columns in table denote chromo-some (Chr), nucleotide position (Pos; hg19 build), reference nucleotide (RefNT), alternate nucleotide (AltNT), amino acid change (AA change), POLYPHEN damage prediction, conservation as reported by GERP score (GERP >5 is highly conserved), gene name, whether the variant segregates in the family, and whether the variant is present in 200 healthy control individuals. (D) Conservation analysis for the GFM2 variant shows that this amino acid (p.D576) is fully conserved from fish to human. This variant occurs at the exon/intron boundary, and shading denotes the exonic region.

Fig. 4

Genetic data for family 982. (A) Pedigree and an MRI scan for an affected male member of family 982 showing the hallmark molar tooth malformation characteristic of Joubert syndrome (circled region). (B) Linkage plot of LOD score against chromosome number reveals multiple link age blocks for this family. (C) Final variant table after whole-exome sequencing analysis reveals five potential candidates, only one of which segregated in the family. (D) Conservation analysis for the EXOC8 variant shows that amino acid E²⁶⁵ (p.E265) is fully conserved from fish to human.