Targeted next-generation sequencing: a novel diagnostic tool for primary immunodeficiencies

TY - JOUR

T1 - Targeted next-generation sequencing

T2 - a novel diagnostic tool for primary immunodeficiencies

AU - Nijman, Isaac J

AU - van Montfrans, Joris M

AU - Hoogstraat, Marlous

AU - Boes, Marianne L

AU - van de Corput, Lisette

AU - Renner, Ellen D

AU - van Zon, Patrick

AU - van Lieshout, Stef

AU - Elferink, Martin G

AU - van der Burg, Mirjam

AU - Vermont, Clementien L

AU - van der Zwaag, Bert

AU - Janson, Esther

AU - Cuppen, Edwin

AU - Ploos van Amstel, Johannes K

AU - van Gijn, Marielle E

N1 - Copyright © 2014 American Academy of Allergy, Asthma & Immunology. Published by Mosby, Inc. All rights reserved.

PY - 2014/2

Y1 - 2014/2

N2 - BACKGROUND: Primary immunodeficiency (PID) disorders are a heterogeneous group of inherited disorders caused by a variety of monogenetic immune defects. Thus far, mutations in more than 170 different genes causing PIDs have been described. A clear genotype-phenotype correlation is often not available, which makes a genetic diagnosis in patients with PIDs complex and laborious.OBJECTIVE: We sought to develop a robust, time-effective, and cost-effective diagnostic method to facilitate a genetic diagnosis in any of 170 known PID-related genes by using next-generation sequencing (NGS).METHODS: We used both targeted array-based and in-solution enrichment combined with a SOLiD sequencing platform and a bioinformatic pipeline developed in house to analyze genetic changes in the DNA of 41 patients with PIDs with known mutations and 26 patients with undiagnosed PIDs.RESULTS: This novel NGS-based method accurately detected point mutations (sensitivity and specificity >99% in covered regions) and exonic deletions (100% sensitivity and specificity). For the 170 genes of interest, the DNA coverage was greater than 20× in 90% to 95%. Nine PID-related genes proved not eligible for evaluation by using this NGS-based method because of inadequate coverage. The NGS method allowed us to make a genetic diagnosis in 4 of 26 patients who lacked a genetic diagnosis despite routine functional and genetic testing. Three of these patients proved to have an atypical presentation of previously described PIDs.CONCLUSION: This novel NGS tool facilitates accurate simultaneous detection of mutations in 161 of 170 known PID-related genes. In addition, these analyses will generate more insight into genotype-phenotype correlations for the different PID disorders.

AB - BACKGROUND: Primary immunodeficiency (PID) disorders are a heterogeneous group of inherited disorders caused by a variety of monogenetic immune defects. Thus far, mutations in more than 170 different genes causing PIDs have been described. A clear genotype-phenotype correlation is often not available, which makes a genetic diagnosis in patients with PIDs complex and laborious.OBJECTIVE: We sought to develop a robust, time-effective, and cost-effective diagnostic method to facilitate a genetic diagnosis in any of 170 known PID-related genes by using next-generation sequencing (NGS).METHODS: We used both targeted array-based and in-solution enrichment combined with a SOLiD sequencing platform and a bioinformatic pipeline developed in house to analyze genetic changes in the DNA of 41 patients with PIDs with known mutations and 26 patients with undiagnosed PIDs.RESULTS: This novel NGS-based method accurately detected point mutations (sensitivity and specificity >99% in covered regions) and exonic deletions (100% sensitivity and specificity). For the 170 genes of interest, the DNA coverage was greater than 20× in 90% to 95%. Nine PID-related genes proved not eligible for evaluation by using this NGS-based method because of inadequate coverage. The NGS method allowed us to make a genetic diagnosis in 4 of 26 patients who lacked a genetic diagnosis despite routine functional and genetic testing. Three of these patients proved to have an atypical presentation of previously described PIDs.CONCLUSION: This novel NGS tool facilitates accurate simultaneous detection of mutations in 161 of 170 known PID-related genes. In addition, these analyses will generate more insight into genotype-phenotype correlations for the different PID disorders.

KW - Adolescent

KW - Adult

KW - Child

KW - Genetic Predisposition to Disease

KW - High-Throughput Nucleotide Sequencing

KW - Humans

KW - Immunologic Deficiency Syndromes

KW - Male

KW - Mutation

KW - Sequence Analysis, DNA

U2 - 10.1016/j.jaci.2013.08.032

DO - 10.1016/j.jaci.2013.08.032

M3 - Article

C2 - 24139496

SN - 0091-6749

VL - 133

SP - 529

EP - 534

JO - Journal of Allergy and Clinical Immunology

JF - Journal of Allergy and Clinical Immunology

IS - 2

ER -