Targeted sequencing aids in identifying clonality in chronic myelomonocytic leukemia

Sang Mee Hwang, Sung Min Kim, Youngwon Nam, Jinhyun Kim, Sungsik Kim, Yong Oon Ahn, Yong Park, Sung-Soo Yoon, Sue Shin, Sunghoon Kwon, Dong Soon Lee

Research output: Contribution to journalArticleResearchpeer-review

Abstract

Chronic myelomonocytic leukemia (CMML) typically shows monocytosis in the peripheral blood (PB), which must be differentiated from reactive monocytosis. To determine the clonality of CMML, we performed molecular and cytogenetic analysis in Korean patients. To investigate whether monocytes in the PB harbored clonal mutational changes, we performed single-cell sequencing after selecting monocytes, neutrophils, and lymphocytes by morphology-aided laser microdissection. Targeted sequencing was performed in 35 patients with CMML with 41 bone marrow samples. Single-cell analysis was performed in two cases. Most (94.3%) patients harbored at least one variant, in genes considered as potential therapeutic targets, while cytogenetic aberrations occurred in only 28.6% of cases. ASXL1 (54.3%), SRSF2 (37.1%), NRAS (31.4%), and TET2 (25.7%) were frequently mutated, with lower frequencies of TET2 mutation and higher frequencies of NRAS, DNMT3A (17.1%), and NPM1 (11.4%) mutations compared to in previous studies of Caucasians. Patients with SETBP1 mutation and those with more than two variants showed poorer survival than those without mutation (P < 0.001 and P = 0.007, respectively). Most (70.8%) variants were detected at diagnosis and follow-up with no significant differences in variant allele frequency, warranting sequencing during follow-up if diagnostic samples were unavailable. Single-cell analysis revealed clonal monocytes with mutations, and the same mutations were also identified in lymphocytes and neutrophils. Targeted sequencing aided in clonality detection in most patients with CMML and single-cell sequencing facilitated identification of clonal monocytes and the co-existence of mutations in non-myeloid cells, suggesting that certain mutations are acquired by pluripotent stem cells.

Original languageEnglish
Article number106190
JournalLeukemia Research
Volume84
DOIs
StatePublished - 1 Sep 2019

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Leukemia, Myelomonocytic, Chronic
Mutation
Monocytes
Single-Cell Analysis
Neutrophils
Lymphocytes
Microdissection
Pluripotent Stem Cells
Cytogenetic Analysis
Mutation Rate
Gene Frequency
Chromosome Aberrations
Lasers
Bone Marrow
Survival

Keywords

  • Chronic myelomonocytic leukemia
  • Monocytosis
  • Morphology-aided laser microdissection
  • Single cell analysis
  • Targeted sequencing

Cite this

Hwang, Sang Mee ; Kim, Sung Min ; Nam, Youngwon ; Kim, Jinhyun ; Kim, Sungsik ; Ahn, Yong Oon ; Park, Yong ; Yoon, Sung-Soo ; Shin, Sue ; Kwon, Sunghoon ; Lee, Dong Soon. / Targeted sequencing aids in identifying clonality in chronic myelomonocytic leukemia. In: Leukemia Research. 2019 ; Vol. 84.
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title = "Targeted sequencing aids in identifying clonality in chronic myelomonocytic leukemia",
abstract = "Chronic myelomonocytic leukemia (CMML) typically shows monocytosis in the peripheral blood (PB), which must be differentiated from reactive monocytosis. To determine the clonality of CMML, we performed molecular and cytogenetic analysis in Korean patients. To investigate whether monocytes in the PB harbored clonal mutational changes, we performed single-cell sequencing after selecting monocytes, neutrophils, and lymphocytes by morphology-aided laser microdissection. Targeted sequencing was performed in 35 patients with CMML with 41 bone marrow samples. Single-cell analysis was performed in two cases. Most (94.3{\%}) patients harbored at least one variant, in genes considered as potential therapeutic targets, while cytogenetic aberrations occurred in only 28.6{\%} of cases. ASXL1 (54.3{\%}), SRSF2 (37.1{\%}), NRAS (31.4{\%}), and TET2 (25.7{\%}) were frequently mutated, with lower frequencies of TET2 mutation and higher frequencies of NRAS, DNMT3A (17.1{\%}), and NPM1 (11.4{\%}) mutations compared to in previous studies of Caucasians. Patients with SETBP1 mutation and those with more than two variants showed poorer survival than those without mutation (P < 0.001 and P = 0.007, respectively). Most (70.8{\%}) variants were detected at diagnosis and follow-up with no significant differences in variant allele frequency, warranting sequencing during follow-up if diagnostic samples were unavailable. Single-cell analysis revealed clonal monocytes with mutations, and the same mutations were also identified in lymphocytes and neutrophils. Targeted sequencing aided in clonality detection in most patients with CMML and single-cell sequencing facilitated identification of clonal monocytes and the co-existence of mutations in non-myeloid cells, suggesting that certain mutations are acquired by pluripotent stem cells.",
keywords = "Chronic myelomonocytic leukemia, Monocytosis, Morphology-aided laser microdissection, Single cell analysis, Targeted sequencing",
author = "Hwang, {Sang Mee} and Kim, {Sung Min} and Youngwon Nam and Jinhyun Kim and Sungsik Kim and Ahn, {Yong Oon} and Yong Park and Sung-Soo Yoon and Sue Shin and Sunghoon Kwon and Lee, {Dong Soon}",
year = "2019",
month = "9",
day = "1",
doi = "10.1016/j.leukres.2019.106190",
language = "English",
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Targeted sequencing aids in identifying clonality in chronic myelomonocytic leukemia. / Hwang, Sang Mee; Kim, Sung Min; Nam, Youngwon; Kim, Jinhyun; Kim, Sungsik; Ahn, Yong Oon; Park, Yong; Yoon, Sung-Soo; Shin, Sue; Kwon, Sunghoon; Lee, Dong Soon.

In: Leukemia Research, Vol. 84, 106190, 01.09.2019.

Research output: Contribution to journalArticleResearchpeer-review

TY - JOUR

T1 - Targeted sequencing aids in identifying clonality in chronic myelomonocytic leukemia

AU - Hwang, Sang Mee

AU - Kim, Sung Min

AU - Nam, Youngwon

AU - Kim, Jinhyun

AU - Kim, Sungsik

AU - Ahn, Yong Oon

AU - Park, Yong

AU - Yoon, Sung-Soo

AU - Shin, Sue

AU - Kwon, Sunghoon

AU - Lee, Dong Soon

PY - 2019/9/1

Y1 - 2019/9/1

N2 - Chronic myelomonocytic leukemia (CMML) typically shows monocytosis in the peripheral blood (PB), which must be differentiated from reactive monocytosis. To determine the clonality of CMML, we performed molecular and cytogenetic analysis in Korean patients. To investigate whether monocytes in the PB harbored clonal mutational changes, we performed single-cell sequencing after selecting monocytes, neutrophils, and lymphocytes by morphology-aided laser microdissection. Targeted sequencing was performed in 35 patients with CMML with 41 bone marrow samples. Single-cell analysis was performed in two cases. Most (94.3%) patients harbored at least one variant, in genes considered as potential therapeutic targets, while cytogenetic aberrations occurred in only 28.6% of cases. ASXL1 (54.3%), SRSF2 (37.1%), NRAS (31.4%), and TET2 (25.7%) were frequently mutated, with lower frequencies of TET2 mutation and higher frequencies of NRAS, DNMT3A (17.1%), and NPM1 (11.4%) mutations compared to in previous studies of Caucasians. Patients with SETBP1 mutation and those with more than two variants showed poorer survival than those without mutation (P < 0.001 and P = 0.007, respectively). Most (70.8%) variants were detected at diagnosis and follow-up with no significant differences in variant allele frequency, warranting sequencing during follow-up if diagnostic samples were unavailable. Single-cell analysis revealed clonal monocytes with mutations, and the same mutations were also identified in lymphocytes and neutrophils. Targeted sequencing aided in clonality detection in most patients with CMML and single-cell sequencing facilitated identification of clonal monocytes and the co-existence of mutations in non-myeloid cells, suggesting that certain mutations are acquired by pluripotent stem cells.

AB - Chronic myelomonocytic leukemia (CMML) typically shows monocytosis in the peripheral blood (PB), which must be differentiated from reactive monocytosis. To determine the clonality of CMML, we performed molecular and cytogenetic analysis in Korean patients. To investigate whether monocytes in the PB harbored clonal mutational changes, we performed single-cell sequencing after selecting monocytes, neutrophils, and lymphocytes by morphology-aided laser microdissection. Targeted sequencing was performed in 35 patients with CMML with 41 bone marrow samples. Single-cell analysis was performed in two cases. Most (94.3%) patients harbored at least one variant, in genes considered as potential therapeutic targets, while cytogenetic aberrations occurred in only 28.6% of cases. ASXL1 (54.3%), SRSF2 (37.1%), NRAS (31.4%), and TET2 (25.7%) were frequently mutated, with lower frequencies of TET2 mutation and higher frequencies of NRAS, DNMT3A (17.1%), and NPM1 (11.4%) mutations compared to in previous studies of Caucasians. Patients with SETBP1 mutation and those with more than two variants showed poorer survival than those without mutation (P < 0.001 and P = 0.007, respectively). Most (70.8%) variants were detected at diagnosis and follow-up with no significant differences in variant allele frequency, warranting sequencing during follow-up if diagnostic samples were unavailable. Single-cell analysis revealed clonal monocytes with mutations, and the same mutations were also identified in lymphocytes and neutrophils. Targeted sequencing aided in clonality detection in most patients with CMML and single-cell sequencing facilitated identification of clonal monocytes and the co-existence of mutations in non-myeloid cells, suggesting that certain mutations are acquired by pluripotent stem cells.

KW - Chronic myelomonocytic leukemia

KW - Monocytosis

KW - Morphology-aided laser microdissection

KW - Single cell analysis

KW - Targeted sequencing

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U2 - 10.1016/j.leukres.2019.106190

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M3 - Article

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