Targeted NGS Platforms for Genetic Screening and Gene Discovery in Primary Immunodeficiencies

Cristina Cifaldi, Immacolata Brigida, Federica Barzaghi, Matteo Zoccolillo, Valentina Ferradini, Davide Petricone, Maria Pia Cicalese, Dejan Lazarevic, Davide Cittaro, Maryam Omrani, Enrico Attardi, Francesca Conti, Alessia Scarselli, Maria Chiriaco, Silvia Di Cesare, Francesco Licciardi, Montin Davide, Francesca Ferrua, Clementina Canessa, Claudio PignataSilvia Giliani, Simona Ferrari, Georgia Fousteri, Graziano Barera, Pietro Merli, Paolo Palma, Simone Cesaro, Marco Gattorno, Antonio Trizzino, Viviana Moschese, Loredana Chini, Anna Villa, Chiara Azzari, Andrea Finocchi, Franco Locatelli, Paolo Rossi, Federica Sangiuolo, Alessandro Aiuti, Caterina Cancrini, Gigliola Di Matteo

Research output: Contribution to journalArticle

Abstract

Background: Primary Immunodeficiencies (PIDs) are a heterogeneous group of genetic immune disorders. While some PIDs can manifest with more than one phenotype, signs, and symptoms of various PIDs overlap considerably. Recently, novel defects in immune-related genes and additional variants in previously reported genes responsible for PIDs have been successfully identified by Next Generation Sequencing (NGS), allowing the recognition of a broad spectrum of disorders. Objective: To evaluate the strength and weakness of targeted NGS sequencing using custom-made Ion Torrent and Haloplex (Agilent) panels for diagnostics and research purposes. Methods: Five different panels including known and candidate genes were used to screen 105 patients with distinct PID features divided in three main PID categories: T cell defects, Humoral defects and Other PIDs. The Ion Torrent sequencing platform was used in 73 patients. Among these, 18 selected patients without a molecular diagnosis and 32 additional patients were analyzed by Haloplex enrichment technology. Results: The complementary use of the two custom-made targeted sequencing approaches allowed the identification of causative variants in 28.6% (n = 30) of patients. Twenty-two out of 73 (34.6%) patients were diagnosed by Ion Torrent. In this group 20 were included in the SCID/CID category. Eight out of 50 (16%) patients were diagnosed by Haloplex workflow. Ion Torrent method was highly successful for those cases with well-defined phenotypes for immunological and clinical presentation. The Haloplex approach was able to diagnose 4 SCID/CID patients and 4 additional patients with complex and extended phenotypes, embracing all three PID categories in which this approach was more efficient. Both technologies showed good gene coverage. Conclusions: NGS technology represents a powerful approach in the complex field of rare disorders but its different application should be weighted. A relatively small NGS target panel can be successfully applied for a robust diagnostic suspicion, while when the spectrum of clinical phenotypes overlaps more than one PID an in-depth NGS analysis is required, including also whole exome/genome sequencing to identify the causative gene.

Original languageEnglish
Pages (from-to)316
JournalFrontiers in Immunology
Volume10
DOIs
Publication statusPublished - 2019

Fingerprint

Genetic Testing
Genetic Association Studies
Ions
Phenotype
Genes
Technology
Exome
Inborn Genetic Diseases
Workflow
Immune System Diseases
Signs and Symptoms
Genome
Research

Cite this

Targeted NGS Platforms for Genetic Screening and Gene Discovery in Primary Immunodeficiencies. / Cifaldi, Cristina; Brigida, Immacolata; Barzaghi, Federica; Zoccolillo, Matteo; Ferradini, Valentina; Petricone, Davide; Cicalese, Maria Pia; Lazarevic, Dejan; Cittaro, Davide; Omrani, Maryam; Attardi, Enrico; Conti, Francesca; Scarselli, Alessia; Chiriaco, Maria; Di Cesare, Silvia; Licciardi, Francesco; Davide, Montin; Ferrua, Francesca; Canessa, Clementina; Pignata, Claudio; Giliani, Silvia; Ferrari, Simona; Fousteri, Georgia; Barera, Graziano; Merli, Pietro; Palma, Paolo; Cesaro, Simone; Gattorno, Marco; Trizzino, Antonio; Moschese, Viviana; Chini, Loredana; Villa, Anna; Azzari, Chiara; Finocchi, Andrea; Locatelli, Franco; Rossi, Paolo; Sangiuolo, Federica; Aiuti, Alessandro; Cancrini, Caterina; Di Matteo, Gigliola.

In: Frontiers in Immunology, Vol. 10, 2019, p. 316.

Research output: Contribution to journalArticle

Cifaldi, C, Brigida, I, Barzaghi, F, Zoccolillo, M, Ferradini, V, Petricone, D, Cicalese, MP, Lazarevic, D, Cittaro, D, Omrani, M, Attardi, E, Conti, F, Scarselli, A, Chiriaco, M, Di Cesare, S, Licciardi, F, Davide, M, Ferrua, F, Canessa, C, Pignata, C, Giliani, S, Ferrari, S, Fousteri, G, Barera, G, Merli, P, Palma, P, Cesaro, S, Gattorno, M, Trizzino, A, Moschese, V, Chini, L, Villa, A, Azzari, C, Finocchi, A, Locatelli, F, Rossi, P, Sangiuolo, F, Aiuti, A, Cancrini, C & Di Matteo, G 2019, 'Targeted NGS Platforms for Genetic Screening and Gene Discovery in Primary Immunodeficiencies', Frontiers in Immunology, vol. 10, pp. 316. https://doi.org/10.3389/fimmu.2019.00316
Cifaldi, Cristina ; Brigida, Immacolata ; Barzaghi, Federica ; Zoccolillo, Matteo ; Ferradini, Valentina ; Petricone, Davide ; Cicalese, Maria Pia ; Lazarevic, Dejan ; Cittaro, Davide ; Omrani, Maryam ; Attardi, Enrico ; Conti, Francesca ; Scarselli, Alessia ; Chiriaco, Maria ; Di Cesare, Silvia ; Licciardi, Francesco ; Davide, Montin ; Ferrua, Francesca ; Canessa, Clementina ; Pignata, Claudio ; Giliani, Silvia ; Ferrari, Simona ; Fousteri, Georgia ; Barera, Graziano ; Merli, Pietro ; Palma, Paolo ; Cesaro, Simone ; Gattorno, Marco ; Trizzino, Antonio ; Moschese, Viviana ; Chini, Loredana ; Villa, Anna ; Azzari, Chiara ; Finocchi, Andrea ; Locatelli, Franco ; Rossi, Paolo ; Sangiuolo, Federica ; Aiuti, Alessandro ; Cancrini, Caterina ; Di Matteo, Gigliola. / Targeted NGS Platforms for Genetic Screening and Gene Discovery in Primary Immunodeficiencies. In: Frontiers in Immunology. 2019 ; Vol. 10. pp. 316.
@article{1fa06e3d6f444d3dad74077b13f7d3ff,
title = "Targeted NGS Platforms for Genetic Screening and Gene Discovery in Primary Immunodeficiencies",
abstract = "Background: Primary Immunodeficiencies (PIDs) are a heterogeneous group of genetic immune disorders. While some PIDs can manifest with more than one phenotype, signs, and symptoms of various PIDs overlap considerably. Recently, novel defects in immune-related genes and additional variants in previously reported genes responsible for PIDs have been successfully identified by Next Generation Sequencing (NGS), allowing the recognition of a broad spectrum of disorders. Objective: To evaluate the strength and weakness of targeted NGS sequencing using custom-made Ion Torrent and Haloplex (Agilent) panels for diagnostics and research purposes. Methods: Five different panels including known and candidate genes were used to screen 105 patients with distinct PID features divided in three main PID categories: T cell defects, Humoral defects and Other PIDs. The Ion Torrent sequencing platform was used in 73 patients. Among these, 18 selected patients without a molecular diagnosis and 32 additional patients were analyzed by Haloplex enrichment technology. Results: The complementary use of the two custom-made targeted sequencing approaches allowed the identification of causative variants in 28.6{\%} (n = 30) of patients. Twenty-two out of 73 (34.6{\%}) patients were diagnosed by Ion Torrent. In this group 20 were included in the SCID/CID category. Eight out of 50 (16{\%}) patients were diagnosed by Haloplex workflow. Ion Torrent method was highly successful for those cases with well-defined phenotypes for immunological and clinical presentation. The Haloplex approach was able to diagnose 4 SCID/CID patients and 4 additional patients with complex and extended phenotypes, embracing all three PID categories in which this approach was more efficient. Both technologies showed good gene coverage. Conclusions: NGS technology represents a powerful approach in the complex field of rare disorders but its different application should be weighted. A relatively small NGS target panel can be successfully applied for a robust diagnostic suspicion, while when the spectrum of clinical phenotypes overlaps more than one PID an in-depth NGS analysis is required, including also whole exome/genome sequencing to identify the causative gene.",
author = "Cristina Cifaldi and Immacolata Brigida and Federica Barzaghi and Matteo Zoccolillo and Valentina Ferradini and Davide Petricone and Cicalese, {Maria Pia} and Dejan Lazarevic and Davide Cittaro and Maryam Omrani and Enrico Attardi and Francesca Conti and Alessia Scarselli and Maria Chiriaco and {Di Cesare}, Silvia and Francesco Licciardi and Montin Davide and Francesca Ferrua and Clementina Canessa and Claudio Pignata and Silvia Giliani and Simona Ferrari and Georgia Fousteri and Graziano Barera and Pietro Merli and Paolo Palma and Simone Cesaro and Marco Gattorno and Antonio Trizzino and Viviana Moschese and Loredana Chini and Anna Villa and Chiara Azzari and Andrea Finocchi and Franco Locatelli and Paolo Rossi and Federica Sangiuolo and Alessandro Aiuti and Caterina Cancrini and {Di Matteo}, Gigliola",
year = "2019",
doi = "10.3389/fimmu.2019.00316",
language = "English",
volume = "10",
pages = "316",
journal = "Frontiers in Immunology",
issn = "1664-3224",
publisher = "Frontiers Media S.A.",

}

TY - JOUR

T1 - Targeted NGS Platforms for Genetic Screening and Gene Discovery in Primary Immunodeficiencies

AU - Cifaldi, Cristina

AU - Brigida, Immacolata

AU - Barzaghi, Federica

AU - Zoccolillo, Matteo

AU - Ferradini, Valentina

AU - Petricone, Davide

AU - Cicalese, Maria Pia

AU - Lazarevic, Dejan

AU - Cittaro, Davide

AU - Omrani, Maryam

AU - Attardi, Enrico

AU - Conti, Francesca

AU - Scarselli, Alessia

AU - Chiriaco, Maria

AU - Di Cesare, Silvia

AU - Licciardi, Francesco

AU - Davide, Montin

AU - Ferrua, Francesca

AU - Canessa, Clementina

AU - Pignata, Claudio

AU - Giliani, Silvia

AU - Ferrari, Simona

AU - Fousteri, Georgia

AU - Barera, Graziano

AU - Merli, Pietro

AU - Palma, Paolo

AU - Cesaro, Simone

AU - Gattorno, Marco

AU - Trizzino, Antonio

AU - Moschese, Viviana

AU - Chini, Loredana

AU - Villa, Anna

AU - Azzari, Chiara

AU - Finocchi, Andrea

AU - Locatelli, Franco

AU - Rossi, Paolo

AU - Sangiuolo, Federica

AU - Aiuti, Alessandro

AU - Cancrini, Caterina

AU - Di Matteo, Gigliola

PY - 2019

Y1 - 2019

N2 - Background: Primary Immunodeficiencies (PIDs) are a heterogeneous group of genetic immune disorders. While some PIDs can manifest with more than one phenotype, signs, and symptoms of various PIDs overlap considerably. Recently, novel defects in immune-related genes and additional variants in previously reported genes responsible for PIDs have been successfully identified by Next Generation Sequencing (NGS), allowing the recognition of a broad spectrum of disorders. Objective: To evaluate the strength and weakness of targeted NGS sequencing using custom-made Ion Torrent and Haloplex (Agilent) panels for diagnostics and research purposes. Methods: Five different panels including known and candidate genes were used to screen 105 patients with distinct PID features divided in three main PID categories: T cell defects, Humoral defects and Other PIDs. The Ion Torrent sequencing platform was used in 73 patients. Among these, 18 selected patients without a molecular diagnosis and 32 additional patients were analyzed by Haloplex enrichment technology. Results: The complementary use of the two custom-made targeted sequencing approaches allowed the identification of causative variants in 28.6% (n = 30) of patients. Twenty-two out of 73 (34.6%) patients were diagnosed by Ion Torrent. In this group 20 were included in the SCID/CID category. Eight out of 50 (16%) patients were diagnosed by Haloplex workflow. Ion Torrent method was highly successful for those cases with well-defined phenotypes for immunological and clinical presentation. The Haloplex approach was able to diagnose 4 SCID/CID patients and 4 additional patients with complex and extended phenotypes, embracing all three PID categories in which this approach was more efficient. Both technologies showed good gene coverage. Conclusions: NGS technology represents a powerful approach in the complex field of rare disorders but its different application should be weighted. A relatively small NGS target panel can be successfully applied for a robust diagnostic suspicion, while when the spectrum of clinical phenotypes overlaps more than one PID an in-depth NGS analysis is required, including also whole exome/genome sequencing to identify the causative gene.

AB - Background: Primary Immunodeficiencies (PIDs) are a heterogeneous group of genetic immune disorders. While some PIDs can manifest with more than one phenotype, signs, and symptoms of various PIDs overlap considerably. Recently, novel defects in immune-related genes and additional variants in previously reported genes responsible for PIDs have been successfully identified by Next Generation Sequencing (NGS), allowing the recognition of a broad spectrum of disorders. Objective: To evaluate the strength and weakness of targeted NGS sequencing using custom-made Ion Torrent and Haloplex (Agilent) panels for diagnostics and research purposes. Methods: Five different panels including known and candidate genes were used to screen 105 patients with distinct PID features divided in three main PID categories: T cell defects, Humoral defects and Other PIDs. The Ion Torrent sequencing platform was used in 73 patients. Among these, 18 selected patients without a molecular diagnosis and 32 additional patients were analyzed by Haloplex enrichment technology. Results: The complementary use of the two custom-made targeted sequencing approaches allowed the identification of causative variants in 28.6% (n = 30) of patients. Twenty-two out of 73 (34.6%) patients were diagnosed by Ion Torrent. In this group 20 were included in the SCID/CID category. Eight out of 50 (16%) patients were diagnosed by Haloplex workflow. Ion Torrent method was highly successful for those cases with well-defined phenotypes for immunological and clinical presentation. The Haloplex approach was able to diagnose 4 SCID/CID patients and 4 additional patients with complex and extended phenotypes, embracing all three PID categories in which this approach was more efficient. Both technologies showed good gene coverage. Conclusions: NGS technology represents a powerful approach in the complex field of rare disorders but its different application should be weighted. A relatively small NGS target panel can be successfully applied for a robust diagnostic suspicion, while when the spectrum of clinical phenotypes overlaps more than one PID an in-depth NGS analysis is required, including also whole exome/genome sequencing to identify the causative gene.

U2 - 10.3389/fimmu.2019.00316

DO - 10.3389/fimmu.2019.00316

M3 - Article

C2 - 31031743

VL - 10

SP - 316

JO - Frontiers in Immunology

JF - Frontiers in Immunology

SN - 1664-3224

ER -