Detection of minimal residual disease in NPM1-mutated acute myeloid leukemia by next-generation sequencing

Detection of minimal residual disease predicts adverse outcome in patients with acute myeloid leukemia. Currently, minimal residual disease may be detected by RQ-PCR or flow cytometry, both of which have practical and diagnostic limitations. Here, we describe a next-generation sequencing assay for m...

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Main Authors:	Salipante, Stephen J., Fromm, Jonathan R., Shendure, Jay, Wood, Brent L., Wu, David
Format:	Online
Language:	English
Published:	2014
Online Access:	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4201902/

id	pubmed-4201902
recordtype	oai_dc
spelling	pubmed-42019022015-05-01 Detection of minimal residual disease in NPM1-mutated acute myeloid leukemia by next-generation sequencing Salipante, Stephen J. Fromm, Jonathan R. Shendure, Jay Wood, Brent L. Wu, David Article Detection of minimal residual disease predicts adverse outcome in patients with acute myeloid leukemia. Currently, minimal residual disease may be detected by RQ-PCR or flow cytometry, both of which have practical and diagnostic limitations. Here, we describe a next-generation sequencing assay for minimal residual disease detection in NPM1-mutated acute myeloid leukemia, which encompasses ∼60% of patients with normal karyotype acute myeloid leukemia. Exon 12 of NPM1 was PCR amplified using sequencing adaptor-linked primers and deep sequenced to enable detection of low-prevalence, acute myeloid leukemia-specific activating mutations. We benchmarked our results against flow cytometry, the standard-of-care for acute myeloid leukemia minimal residual disease diagnosis at our institution. The performance of both approaches was evaluated using defined dilutions of an NPM1-mutation positive cell line and longitudinal clinical samples from acute myeloid leukemia patients. Using defined control material, we found this assay sensitive to approximately 0.001% mutant cells, outperforming flow cytometry by an order-of-magnitude. Next-generation sequencing was precise and semi-quantitative over 4 orders-of-magnitude. In 22 longitudinal samples from 6 acute myeloid leukemia patients, next-generation sequencing detected minimal residual disease in all samples deemed negative by flow cytometry. Further, in one-third of patients, sequencing detected alternate NPM1-mutations in addition to the patient's index mutation, consistent with tumor heterogeneity. Next-generation sequencing provides information without prior knowledge of NPM1 mutation subtype or validation of allele-specific probes as required for RQ-PCR assays, and without generation and interpretation of complex multi-dimensional flow cytometry data. This approach may complement current technologies to enhance patient-specific clinical decision-making. 2014-04-18 2014-11 /pmc/articles/PMC4201902/ /pubmed/24743218 http://dx.doi.org/10.1038/modpathol.2014.57 Text en http://www.nature.com/authors/editorial_policies/license.html#terms Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use:http://www.nature.com/authors/editorial_policies/license.html#terms
repository_type	Open Access Journal
institution_category	Foreign Institution
institution	US National Center for Biotechnology Information
building	NCBI PubMed
collection	Online Access
language	English
format	Online
author	Salipante, Stephen J. Fromm, Jonathan R. Shendure, Jay Wood, Brent L. Wu, David
spellingShingle	Salipante, Stephen J. Fromm, Jonathan R. Shendure, Jay Wood, Brent L. Wu, David Detection of minimal residual disease in NPM1-mutated acute myeloid leukemia by next-generation sequencing
author_facet	Salipante, Stephen J. Fromm, Jonathan R. Shendure, Jay Wood, Brent L. Wu, David
author_sort	Salipante, Stephen J.
title	Detection of minimal residual disease in NPM1-mutated acute myeloid leukemia by next-generation sequencing
title_short	Detection of minimal residual disease in NPM1-mutated acute myeloid leukemia by next-generation sequencing
title_full	Detection of minimal residual disease in NPM1-mutated acute myeloid leukemia by next-generation sequencing
title_fullStr	Detection of minimal residual disease in NPM1-mutated acute myeloid leukemia by next-generation sequencing
title_full_unstemmed	Detection of minimal residual disease in NPM1-mutated acute myeloid leukemia by next-generation sequencing
title_sort	detection of minimal residual disease in npm1-mutated acute myeloid leukemia by next-generation sequencing
description	Detection of minimal residual disease predicts adverse outcome in patients with acute myeloid leukemia. Currently, minimal residual disease may be detected by RQ-PCR or flow cytometry, both of which have practical and diagnostic limitations. Here, we describe a next-generation sequencing assay for minimal residual disease detection in NPM1-mutated acute myeloid leukemia, which encompasses ∼60% of patients with normal karyotype acute myeloid leukemia. Exon 12 of NPM1 was PCR amplified using sequencing adaptor-linked primers and deep sequenced to enable detection of low-prevalence, acute myeloid leukemia-specific activating mutations. We benchmarked our results against flow cytometry, the standard-of-care for acute myeloid leukemia minimal residual disease diagnosis at our institution. The performance of both approaches was evaluated using defined dilutions of an NPM1-mutation positive cell line and longitudinal clinical samples from acute myeloid leukemia patients. Using defined control material, we found this assay sensitive to approximately 0.001% mutant cells, outperforming flow cytometry by an order-of-magnitude. Next-generation sequencing was precise and semi-quantitative over 4 orders-of-magnitude. In 22 longitudinal samples from 6 acute myeloid leukemia patients, next-generation sequencing detected minimal residual disease in all samples deemed negative by flow cytometry. Further, in one-third of patients, sequencing detected alternate NPM1-mutations in addition to the patient's index mutation, consistent with tumor heterogeneity. Next-generation sequencing provides information without prior knowledge of NPM1 mutation subtype or validation of allele-specific probes as required for RQ-PCR assays, and without generation and interpretation of complex multi-dimensional flow cytometry data. This approach may complement current technologies to enhance patient-specific clinical decision-making.
publishDate	2014
url	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4201902/
_version_	1613146348038651904

Detection of minimal residual disease in NPM1-mutated acute myeloid leukemia by next-generation sequencing

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