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
AU - Kok, Chung
AU - Saunders, Verity
AU - Van der Hoek, Mark
AU - Heatley, Susan
AU - Schwarer, Anthony
AU - Hahn, Christopher
AU - Hughes, Timothy
AU - White, Deborah
AU - Ross, David
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.
UR - http://www.scopus.com/inward/record.url?scp=85027730594&partnerID=8YFLogxK
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
JF - Journal of Molecular Diagnostics
SN - 1525-1578
IS - 5
ER -