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Whole-Exome Sequencing in the Differential Diagnosis of Primary Adrenal Insufficiency in Children.

Chan LF, Campbell DC, Novoselova TV, Clark AJ, Metherell LA - Front Endocrinol (Lausanne) (2015)

Bottom Line: However, with the availability, and increasing cost effectiveness of whole-exome sequencing, there is the potential for this to become a powerful diagnostic tool.Here, we report the results of whole-exome sequencing of 43 patients referred to us with a diagnosis of familial glucocorticoid deficiency (FGD) who were mutation negative for MC2R, MRAP, and STAR the most commonly mutated genes in FGD.The diagnosis of isolated or familial glucocorticoid deficiency was only confirmed in 3 of the 17 patients, other genetic diagnoses were adrenal hypo- and hyperplasia, Triple A, and autoimmune polyendocrinopathy syndrome type I, emphasizing both the difficulty of phenotypically distinguishing between disorders of PAI and the utility of WES as a tool to achieve this.

View Article: PubMed Central - PubMed

Affiliation: Centre for Endocrinology, William Harvey Research Institute, Barts and The London School of Medicine and Dentistry, Queen Mary University of London , London , UK.

ABSTRACT
Adrenal insufficiency is a rare, but potentially fatal medical condition. In children, the cause is most commonly congenital and in recent years a growing number of causative gene mutations have been identified resulting in a myriad of syndromes that share adrenal insufficiency as one of the main characteristics. The evolution of adrenal insufficiency is dependent on the variant and the particular gene affected, meaning that rapid and accurate diagnosis is imperative for effective treatment of the patient. Common practice is for candidate genes to be sequenced individually, which is a time-consuming process and complicated by overlapping clinical phenotypes. However, with the availability, and increasing cost effectiveness of whole-exome sequencing, there is the potential for this to become a powerful diagnostic tool. Here, we report the results of whole-exome sequencing of 43 patients referred to us with a diagnosis of familial glucocorticoid deficiency (FGD) who were mutation negative for MC2R, MRAP, and STAR the most commonly mutated genes in FGD. WES provided a rapid genetic diagnosis in 17/43 sequenced patients, for the remaining 60% the gene defect may be within intronic/regulatory regions not covered by WES or may be in gene(s) representing novel etiologies. The diagnosis of isolated or familial glucocorticoid deficiency was only confirmed in 3 of the 17 patients, other genetic diagnoses were adrenal hypo- and hyperplasia, Triple A, and autoimmune polyendocrinopathy syndrome type I, emphasizing both the difficulty of phenotypically distinguishing between disorders of PAI and the utility of WES as a tool to achieve this.

No MeSH data available.


Related in: MedlinePlus

Filtration strategy to screen variants from WES data.
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Figure 2: Filtration strategy to screen variants from WES data.

Mentions: Exome sequencing was performed using the Agilent SureSelect all exon V4 capture and paired-end (2 × 100) sequencing on an Illumina HiSeq 2000 at Otogenetics (Norcross, GA). First analysis pipeline: initially sequencing read alignment, variant calling, and annotation were performed by DNAnexus (DNAnexus Inc.1, Mountain View, CA, USA), their Nucleotide-Level Variation analysis outputs were then screened with our list of genes via the DNAnexus Classic platform which permitted variants table viewing and filtering functionality. This platform no longer exists and more recently these data were reanalyzed uploading the vcf files to Ingenuity variant analysis2. Second analysis pipeline: the raw data were also reanalyzed, aligning to the H. Sapiens GRCh37–b37 (1000genomes Phase 1) reference genome with BWA-MEM FastQ Readmapper VCF files, generated by Vendor Human Exome GATK-Lite Variant Caller (Unified Genotyper) and uploaded to Ingenuity variant analysis. Single Nucleotide Polymorphisms, with threshold coverage of at least 10 reads on the respective nucleotide, were included in the analysis. The variant files from both analyses were screened for causal variants using Ingenuity variant analysis with the filtering strategy outlined in Figure 2. Sequence changes in PAI causal genes were confirmed by PCR designed to cover the affected region followed by Sanger sequencing (primer sequences available on request).


Whole-Exome Sequencing in the Differential Diagnosis of Primary Adrenal Insufficiency in Children.

Chan LF, Campbell DC, Novoselova TV, Clark AJ, Metherell LA - Front Endocrinol (Lausanne) (2015)

Filtration strategy to screen variants from WES data.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4525066&req=5

Figure 2: Filtration strategy to screen variants from WES data.
Mentions: Exome sequencing was performed using the Agilent SureSelect all exon V4 capture and paired-end (2 × 100) sequencing on an Illumina HiSeq 2000 at Otogenetics (Norcross, GA). First analysis pipeline: initially sequencing read alignment, variant calling, and annotation were performed by DNAnexus (DNAnexus Inc.1, Mountain View, CA, USA), their Nucleotide-Level Variation analysis outputs were then screened with our list of genes via the DNAnexus Classic platform which permitted variants table viewing and filtering functionality. This platform no longer exists and more recently these data were reanalyzed uploading the vcf files to Ingenuity variant analysis2. Second analysis pipeline: the raw data were also reanalyzed, aligning to the H. Sapiens GRCh37–b37 (1000genomes Phase 1) reference genome with BWA-MEM FastQ Readmapper VCF files, generated by Vendor Human Exome GATK-Lite Variant Caller (Unified Genotyper) and uploaded to Ingenuity variant analysis. Single Nucleotide Polymorphisms, with threshold coverage of at least 10 reads on the respective nucleotide, were included in the analysis. The variant files from both analyses were screened for causal variants using Ingenuity variant analysis with the filtering strategy outlined in Figure 2. Sequence changes in PAI causal genes were confirmed by PCR designed to cover the affected region followed by Sanger sequencing (primer sequences available on request).

Bottom Line: However, with the availability, and increasing cost effectiveness of whole-exome sequencing, there is the potential for this to become a powerful diagnostic tool.Here, we report the results of whole-exome sequencing of 43 patients referred to us with a diagnosis of familial glucocorticoid deficiency (FGD) who were mutation negative for MC2R, MRAP, and STAR the most commonly mutated genes in FGD.The diagnosis of isolated or familial glucocorticoid deficiency was only confirmed in 3 of the 17 patients, other genetic diagnoses were adrenal hypo- and hyperplasia, Triple A, and autoimmune polyendocrinopathy syndrome type I, emphasizing both the difficulty of phenotypically distinguishing between disorders of PAI and the utility of WES as a tool to achieve this.

View Article: PubMed Central - PubMed

Affiliation: Centre for Endocrinology, William Harvey Research Institute, Barts and The London School of Medicine and Dentistry, Queen Mary University of London , London , UK.

ABSTRACT
Adrenal insufficiency is a rare, but potentially fatal medical condition. In children, the cause is most commonly congenital and in recent years a growing number of causative gene mutations have been identified resulting in a myriad of syndromes that share adrenal insufficiency as one of the main characteristics. The evolution of adrenal insufficiency is dependent on the variant and the particular gene affected, meaning that rapid and accurate diagnosis is imperative for effective treatment of the patient. Common practice is for candidate genes to be sequenced individually, which is a time-consuming process and complicated by overlapping clinical phenotypes. However, with the availability, and increasing cost effectiveness of whole-exome sequencing, there is the potential for this to become a powerful diagnostic tool. Here, we report the results of whole-exome sequencing of 43 patients referred to us with a diagnosis of familial glucocorticoid deficiency (FGD) who were mutation negative for MC2R, MRAP, and STAR the most commonly mutated genes in FGD. WES provided a rapid genetic diagnosis in 17/43 sequenced patients, for the remaining 60% the gene defect may be within intronic/regulatory regions not covered by WES or may be in gene(s) representing novel etiologies. The diagnosis of isolated or familial glucocorticoid deficiency was only confirmed in 3 of the 17 patients, other genetic diagnoses were adrenal hypo- and hyperplasia, Triple A, and autoimmune polyendocrinopathy syndrome type I, emphasizing both the difficulty of phenotypically distinguishing between disorders of PAI and the utility of WES as a tool to achieve this.

No MeSH data available.


Related in: MedlinePlus