Clinical evaluation of panel testing by next-generation sequencing (NGS) for gene mutations in myeloid neoplasms

Au, Chun Hang; Wa, Anna; Ho, Dona N.; Chan, Tsun Leung; S.K., Edmond

doi:10.1186/s13000-016-0456-8

Cited by 84 publications

(89 citation statements)

References 45 publications

(46 reference statements)

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“…INDELseek detected all three complex indels (Table 2; Additional file 2: Figure S1-S3), demonstrating 100% sensitivity and 100% specificity. INDELseek was also applied to a myeloid neoplasm (MN) panel dataset of 23 samples [16]. The 54-gene panel targeted somatic mutations (Illumina MiSeq).…”

Section: Resultsmentioning

confidence: 99%

“…Five of the DNA samples carried somatic pathogenic CALR or JAK2 complex indels and were regarded as positive controls. Remaining seventeen DNA samples of healthy adults with normal complete blood profile were regarded as negative controls as described [16]. Ten core-binding factor leukemia DNA samples were retrieved from Queen Mary Hospital, Hong Kong from January 2003 to December 2014 as a discovery cohort of KIT exon 8 mutations.…”

Section: Methodsmentioning

confidence: 99%

“…Ten core-binding factor leukemia DNA samples were retrieved from Queen Mary Hospital, Hong Kong from January 2003 to December 2014 as a discovery cohort of KIT exon 8 mutations. A total of 32 DNA samples were screened by MiSeq sequencing of a 54-gene myeloid NGS gene panel as described [16, 17]. Complex indel detection by INDELseek was performed on BWA-MEM (version 0.7.7) alignments of MiSeq NGS data of CALR, JAK2 and KIT (exon 8 only).…”

Section: Methodsmentioning

confidence: 99%

“…CALR and JAK2 complex indels were confirmed by conventional PCR and Sanger sequencing or conventional PCR fragment analysis [5, 16]. KIT exon 8 complex indels were confirmed by conventional PCR fragment analysis [16] or microfluidic PCR followed by MiSeq sequencing [25]. The primers used in these validation studies were different from those used in the original NGS datasets (Additional file 1: Table S3).…”

Section: Methodsmentioning

confidence: 99%

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INDELseek: detection of complex insertions and deletions from next-generation sequencing data

Leung

Kwong

et al. 2017

BMC Genomics

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BackgroundComplex insertions and deletions (indels) from next-generation sequencing (NGS) data were prone to escape detection by currently available variant callers as shown by large-scale human genomics studies. Somatic and germline complex indels in key disease driver genes could be missed in NGS-based genomics studies.ResultsINDELseek is an open-source complex indel caller designed for NGS data of random fragments and PCR amplicons. The key differentiating factor of INDELseek is that each NGS read alignment was examined as a whole instead of “pileup” of each reference position across multiple alignments. In benchmarking against the reference material NA12878 genome (n = 160 derived from high-confidence variant calls), GATK, SAMtools and INDELseek showed complex indel detection sensitivities of 0%, 0% and 100%, respectively. INDELseek also detected all known germline (BRCA1 and BRCA2) and somatic (CALR and JAK2) complex indels in human clinical samples (n = 8). Further experiments validated all 10 detected KIT complex indels in a discovery cohort of clinical samples. In silico semi-simulation showed sensitivities of 93.7–96.2% based on 8671 unique complex indels in >5000 genes from dbSNP and COSMIC. We also demonstrated the importance of complex indel detection in accurately annotating BRCA1, BRCA2 and TP53 mutations with gained or rescued protein-truncating effects.ConclusionsINDELseek is an accurate and versatile tool for complex indel detection in NGS data. It complements other variant callers in NGS-based genomics studies targeting a wide spectrum of genetic variations.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-3449-9) contains supplementary material, which is available to authorized users.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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INDELseek: detection of complex insertions and deletions from next-generation sequencing data

Leung

Kwong

et al. 2017

BMC Genomics

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“…Так, одно из исследований касалось миелоидных неоплазий [5]. Наблюдали группу из 46 пациентов среднего возраста.…”

Section: фундаментальные исследования в практической медицине на соврunclassified

Next-Generation Gene Sequencing and Its Applications in Oncohematology

Barkhatov¹,

Predeus²,

Чухловин³

2016

Onkogematologiâ

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Key words: next generation sequencing, technologies, leukemia, gene mutations, bioinformaticsВведение Общеизвестно, что развитие злокачественных опухолевых заболеваний системы крови связано с воз-никновением ряда последовательных мутаций генов, ведущих к развитию злокачественного клеточного клона.Известно также, что в процессе терапии первичная популяция злокачественных клеток может приобретать вторичные мутации, изменяясь под влиянием различных факторов отбора в организме больного (иммунологиче-ская селекция, действие цитостатических препаратов и др.). Эти злокачественные субклоны с сочетаниями генных мутаций могут впоследствии приводить к ре-цидивам заболевания. Разнообразие генной патологии в организме больного существенно влияет на прогноз и исходы лечения больных с лейкозами и лимфомами. Для оценки клинического риска в каждом конкретном случае требуется детальная характеристика генетических аберраций, а для этого необходимо определение по-следовательности нуклеотидов целевого гена в злока-чественных клетках.Современная молекулярная диагностика при лей-козах основана на выявлении конкретных генных мутаций, характерных для того или иного вида лейко-за. Основным методом является молекулярно-генети-ческая диагностика на базе полимеразной цепной реакции (ПЦР) с анализом целевых участков генов, которая проводится в лабораториях крупных онкоге-матологических клиник. Кроме того, с начала 1990-х годов для задач дешифровки генов, в первую очередь смысловых последовательностей (сиквенсов) молекул матричной РНК (мРНК), применяли главным образом классическое секвенирование по Сэнгеру. Проект «Ге-ном человека», основной задачей которого была сбор-ка референсной последовательности человеческого

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FLT3 Mutation Testing in Acute Myeloid Leukemia

2017

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A previously healthy man in his 50s presented with headache and blurry vision. Laboratory data are summarized in the Table . He was diagnosed with acute myeloid leukemia (AML) complicated by leukostasis. Bone marrow karyotyping revealed a normal karyotype; 46 XX [20]. Multigene polymerase chain reaction (PCR) testing of the bone marrow identified a 44-base pair (bp) internal tandem duplication in the FLT3 gene (OMIM 136351), with an allelic ratio of 0.57 (range, 0.01 to >100). No other mutations on the panel (CEBPA, NPM1, KIT, IDH1, and IDH2) were detected. AnswerA. The patient is more likely to have a worse prognosis.

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Clinical evaluation of panel testing by next-generation sequencing (NGS) for gene mutations in myeloid neoplasms

Cited by 84 publications

References 45 publications

INDELseek: detection of complex insertions and deletions from next-generation sequencing data

INDELseek: detection of complex insertions and deletions from next-generation sequencing data

Next-Generation Gene Sequencing and Its Applications in Oncohematology

FLT3 Mutation Testing in Acute Myeloid Leukemia

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