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

Roy, Noémi B A; Wilson, Edward A.; Henderson, Shirley; Wray, Katherine; Babbs, Christian; Okoli, Steven; Atoyebi, Wale; Mixon, Avery; Cahill, Mary R. and Carey, Peter, et al. (2016) In British Journal of Haematology 175(2). p.318-330
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... (More)

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.

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publication status
published
subject
keywords
Congenital dyserythropoietic anaemia, Inherited anaemia, Molecular genetics, Next-generation sequencing, Pyruvate kinase deficiency
in
British Journal of Haematology
volume
175
issue
2
pages
318 - 330
publisher
Federation of European Neuroscience Societies and Blackwell Publishing Ltd
external identifiers
  • scopus:84978634344
ISSN
0007-1048
DOI
10.1111/bjh.14221
language
English
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no
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d5b871db-3c8d-4620-825a-a4df074f67c2
date added to LUP
2016-08-15 11:55:07
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2017-10-01 05:22:09
@article{d5b871db-3c8d-4620-825a-a4df074f67c2,
  abstract     = {<p>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.</p>},
  author       = {Roy, Noémi B A and Wilson, Edward A. and Henderson, Shirley and Wray, Katherine and Babbs, Christian and Okoli, Steven and Atoyebi, Wale and Mixon, Avery and Cahill, Mary R. and Carey, Peter and Cullis, Jonathan and Curtin, Julie and Dreau, Helene and Ferguson, David J P and Gibson, Brenda and Hall, Georgina and Mason, Joanne and Morgan, Mary and Proven, Melanie and Qureshi, Amrana and Sanchez Garcia, Joaquin and Sirachainan, Nongnuch and Teo, Juliana and Tedgård, Ulf and Higgs, Doug and Roberts, David and Roberts, Irene and Schuh, Anna},
  issn         = {0007-1048},
  keyword      = {Congenital dyserythropoietic anaemia,Inherited anaemia,Molecular genetics,Next-generation sequencing,Pyruvate kinase deficiency},
  language     = {eng},
  number       = {2},
  pages        = {318--330},
  publisher    = {Federation of European Neuroscience Societies and Blackwell Publishing Ltd},
  series       = {British Journal of Haematology},
  title        = {A novel 33-Gene targeted resequencing panel provides accurate, clinical-grade diagnosis and improves patient management for rare inherited anaemias},
  url          = {http://dx.doi.org/10.1111/bjh.14221},
  volume       = {175},
  year         = {2016},
}