Frontiers in Immunology (Apr 2019)

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

  • Cristina Cifaldi,
  • Immacolata Brigida,
  • Federica Barzaghi,
  • Federica Barzaghi,
  • Federica Barzaghi,
  • Matteo Zoccolillo,
  • Matteo Zoccolillo,
  • Valentina Ferradini,
  • Davide Petricone,
  • Maria Pia Cicalese,
  • Maria Pia Cicalese,
  • 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,
  • Francesca Ferrua,
  • Francesca Ferrua,
  • Clementina Canessa,
  • Clementina Canessa,
  • Claudio Pignata,
  • Silvia Giliani,
  • Simona Ferrari,
  • Georgia Fousteri,
  • Graziano Barera,
  • Pietro Merli,
  • Paolo Palma,
  • Simone Cesaro,
  • Marco Gattorno,
  • Antonio Trizzino,
  • Viviana Moschese,
  • Viviana Moschese,
  • Loredana Chini,
  • Loredana Chini,
  • Anna Villa,
  • Anna Villa,
  • Chiara Azzari,
  • Chiara Azzari,
  • Andrea Finocchi,
  • Andrea Finocchi,
  • Franco Locatelli,
  • Paolo Rossi,
  • Paolo Rossi,
  • Federica Sangiuolo,
  • Alessandro Aiuti,
  • Alessandro Aiuti,
  • Alessandro Aiuti,
  • Caterina Cancrini,
  • Caterina Cancrini,
  • Gigliola Di Matteo,
  • Gigliola Di Matteo

DOI
https://doi.org/10.3389/fimmu.2019.00316
Journal volume & issue
Vol. 10

Abstract

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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.

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