A novel 33-Gene targeted resequencing panel provides accurate, clinical-grade diagnosis and improves patient management for rare inherited anaemias

Noémi B.A. Roy, Edward A. Wilson, Shirley Henderson, Katherine Wray, Christian Babbs, Steven Okoli, Wale Atoyebi, Avery Mixon, Mary R. Cahill, Peter Carey, Jonathan Cullis, Julie Curtin, Helene Dreau, David J.P. Ferguson, Brenda Gibson, Georgina Hall, Joanne Mason, Mary Morgan, Melanie Proven, Amrana QureshiJoaquin Sanchez Garcia, Nongnuch Sirachainan, Juliana Teo, Ulf Tedgård, Doug Higgs, David Roberts, Irene Roberts*, Anna Schuh

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

67 Citations (Scopus)

Abstract

Accurate diagnosis of rare inherited anaemias is challenging, requiring a series of complex and expensive laboratory tests. Targeted next-generation-sequencing (NGS) has been used to investigate these disorders, but the selection of genes on individual panels has been narrow and the validation strategies used have fallen short of the standards required for clinical use. Clinical-grade validation of negative results requires the test to distinguish between lack of adequate sequencing reads at the locations of known mutations and a real absence of mutations. To achieve a clinically-reliable diagnostic test and minimize false-negative results we developed an open-source tool (CoverMi) to accurately determine base-coverage and the ‘discoverability’ of known mutations for every sample. We validated our 33-gene panel using Sanger sequencing and microarray. Our panel demonstrated 100% specificity and 99·7% sensitivity. We then analysed 57 clinical samples: molecular diagnoses were made in 22/57 (38·6%), corresponding to 32 mutations of which 16 were new. In all cases, accurate molecular diagnosis had a positive impact on clinical management. Using a validated NGS-based platform for routine molecular diagnosis of previously undiagnosed congenital anaemias is feasible in a clinical diagnostic setting, improves precise diagnosis and enhances management and counselling of the patient and their family.

Original languageEnglish
Pages (from-to)318-330
Number of pages13
JournalBritish Journal of Haematology
Volume175
Issue number2
DOIs
Publication statusPublished - 1 Oct 2016
Externally publishedYes

Bibliographical note

Publisher Copyright:
© 2016 John Wiley & Sons Ltd

Keywords

  • congenital dyserythropoietic anaemia
  • inherited anaemia
  • molecular genetics
  • next-generation sequencing
  • pyruvate kinase deficiency

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