A Method for Next-Generation Sequencing of Paired Diagnostic and Remission Samples to Detect Mitochondrial DNA Mutations Associated with Leukemia

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

Somatic mitochondrial DNA (mtDNA) mutations have been identified in many human cancers, including leukemia. To identify somatic mutations, it is necessary to have a control tissue from the same individual for comparison. When patients with leukemia achieve remission, the remission peripheral blood may be a suitable and easily accessible control tissue, but this approach has not previously been applied to the study of mtDNA mutations. We have developed and validated a next-generation sequencing approach for the identification of leukemia-associated mtDNA mutations in 26 chronic myeloid leukemia patients at diagnosis using either nonhematopoietic or remission blood samples as the control. The entire mt genome was amplified by long-range PCR and sequenced using Illumina technology. Variant caller software was used to detect mtDNA somatic mutations, and an empirically determined threshold of 2% was applied to minimize false-positive results because of sequencing errors. Mutations were called against both nonhematopoietic and remission controls: the overall concordance between the two approaches was 81% (73/90 mutations). Some discordant results were because of the presence of somatic mutations in remission samples, because of either minimal residual disease or nonleukemic hematopoietic clones. This method could be applied to study somatic mtDNA mutations in leukemia patients who achieve minimal residual disease, and in patients with nonhematopoietic cancers who have a matched uninvolved tissue available.

LanguageEnglish
Pages711-721
Number of pages11
JournalJournal of Molecular Diagnostics
Volume19
Issue number5
DOIs
Publication statusPublished - Sep 2017

Keywords

  • Journal Article

Cite this

@article{4fc097b762c84146a8c80bb27344ff23,
title = "A Method for Next-Generation Sequencing of Paired Diagnostic and Remission Samples to Detect Mitochondrial DNA Mutations Associated with Leukemia",
abstract = "Somatic mitochondrial DNA (mtDNA) mutations have been identified in many human cancers, including leukemia. To identify somatic mutations, it is necessary to have a control tissue from the same individual for comparison. When patients with leukemia achieve remission, the remission peripheral blood may be a suitable and easily accessible control tissue, but this approach has not previously been applied to the study of mtDNA mutations. We have developed and validated a next-generation sequencing approach for the identification of leukemia-associated mtDNA mutations in 26 chronic myeloid leukemia patients at diagnosis using either nonhematopoietic or remission blood samples as the control. The entire mt genome was amplified by long-range PCR and sequenced using Illumina technology. Variant caller software was used to detect mtDNA somatic mutations, and an empirically determined threshold of 2{\%} was applied to minimize false-positive results because of sequencing errors. Mutations were called against both nonhematopoietic and remission controls: the overall concordance between the two approaches was 81{\%} (73/90 mutations). Some discordant results were because of the presence of somatic mutations in remission samples, because of either minimal residual disease or nonleukemic hematopoietic clones. This method could be applied to study somatic mtDNA mutations in leukemia patients who achieve minimal residual disease, and in patients with nonhematopoietic cancers who have a matched uninvolved tissue available.",
keywords = "Journal Article",
author = "Pagani, {Ilaria S} and Kok, {Chung H} and Saunders, {Verity A} and {Van der Hoek}, {Mark B} and Heatley, {Susan L} and Schwarer, {Anthony P} and Hahn, {Christopher N.} and Hughes, {Timothy P} and White, {Deborah L} and Ross, {David M.}",
note = "Copyright {\circledC} 2017 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.",
year = "2017",
month = "9",
doi = "10.1016/j.jmoldx.2017.05.009",
language = "English",
volume = "19",
pages = "711--721",
journal = "Journal of Molecular Diagnostics",
issn = "1525-1578",
publisher = "Association of Molecular Pathology",
number = "5",

}

TY - JOUR

T1 - A Method for Next-Generation Sequencing of Paired Diagnostic and Remission Samples to Detect Mitochondrial DNA Mutations Associated with Leukemia

AU - Pagani, Ilaria S

AU - Kok, Chung H

AU - Saunders, Verity A

AU - Van der Hoek, Mark B

AU - Heatley, Susan L

AU - Schwarer, Anthony P

AU - Hahn, Christopher N.

AU - Hughes, Timothy P

AU - White, Deborah L

AU - Ross, David M.

N1 - Copyright © 2017 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.

PY - 2017/9

Y1 - 2017/9

N2 - Somatic mitochondrial DNA (mtDNA) mutations have been identified in many human cancers, including leukemia. To identify somatic mutations, it is necessary to have a control tissue from the same individual for comparison. When patients with leukemia achieve remission, the remission peripheral blood may be a suitable and easily accessible control tissue, but this approach has not previously been applied to the study of mtDNA mutations. We have developed and validated a next-generation sequencing approach for the identification of leukemia-associated mtDNA mutations in 26 chronic myeloid leukemia patients at diagnosis using either nonhematopoietic or remission blood samples as the control. The entire mt genome was amplified by long-range PCR and sequenced using Illumina technology. Variant caller software was used to detect mtDNA somatic mutations, and an empirically determined threshold of 2% was applied to minimize false-positive results because of sequencing errors. Mutations were called against both nonhematopoietic and remission controls: the overall concordance between the two approaches was 81% (73/90 mutations). Some discordant results were because of the presence of somatic mutations in remission samples, because of either minimal residual disease or nonleukemic hematopoietic clones. This method could be applied to study somatic mtDNA mutations in leukemia patients who achieve minimal residual disease, and in patients with nonhematopoietic cancers who have a matched uninvolved tissue available.

AB - Somatic mitochondrial DNA (mtDNA) mutations have been identified in many human cancers, including leukemia. To identify somatic mutations, it is necessary to have a control tissue from the same individual for comparison. When patients with leukemia achieve remission, the remission peripheral blood may be a suitable and easily accessible control tissue, but this approach has not previously been applied to the study of mtDNA mutations. We have developed and validated a next-generation sequencing approach for the identification of leukemia-associated mtDNA mutations in 26 chronic myeloid leukemia patients at diagnosis using either nonhematopoietic or remission blood samples as the control. The entire mt genome was amplified by long-range PCR and sequenced using Illumina technology. Variant caller software was used to detect mtDNA somatic mutations, and an empirically determined threshold of 2% was applied to minimize false-positive results because of sequencing errors. Mutations were called against both nonhematopoietic and remission controls: the overall concordance between the two approaches was 81% (73/90 mutations). Some discordant results were because of the presence of somatic mutations in remission samples, because of either minimal residual disease or nonleukemic hematopoietic clones. This method could be applied to study somatic mtDNA mutations in leukemia patients who achieve minimal residual disease, and in patients with nonhematopoietic cancers who have a matched uninvolved tissue available.

KW - Journal Article

U2 - 10.1016/j.jmoldx.2017.05.009

DO - 10.1016/j.jmoldx.2017.05.009

M3 - Article

VL - 19

SP - 711

EP - 721

JO - Journal of Molecular Diagnostics

T2 - Journal of Molecular Diagnostics

JF - Journal of Molecular Diagnostics

SN - 1525-1578

IS - 5

ER -