Department of Laboratory Medicine, Seoul National University Bundang Hospital, Seongnam, Republic of Korea; Department of Laboratory Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea.
Cancer Research Institute, Seoul National University College of Medicine, Seoul, Republic of Korea.
Leuk Res. 2019 Sep;84:106190. doi: 10.1016/j.leukres.2019.106190. Epub 2019 Jul 12.
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.
慢性髓单核细胞白血病（CMML）通常在外周血（PB）中表现为单核细胞增多症，必须与反应性单核细胞增多症区分开来。为了确定 CMML 的克隆性，我们对韩国患者进行了分子和细胞遗传学分析。为了研究 PB 中的单核细胞是否存在克隆突变，我们通过形态辅助激光微切割选择单核细胞、中性粒细胞和淋巴细胞后进行单细胞测序。对 35 例 CMML 患者的 41 个骨髓样本进行了靶向测序。对两例进行了单细胞分析。大多数（94.3%）患者至少存在一个潜在治疗靶点的基因变异，而细胞遗传学异常仅发生在 28.6%的病例中。ASXL1（54.3%）、SRSF2（37.1%）、NRAS（31.4%）和 TET2（25.7%）突变频率较高，而 TET2 突变频率较低，NRAS、DNMT3A（17.1%）和 NPM1（11.4%）突变频率较高与之前的高加索人群研究相比。SETBP1 突变患者和有两个以上变异的患者的生存情况比无突变患者差（P<0.001 和 P=0.007）。大多数（70.8%）变异在诊断和随访时均能检测到，且变异等位基因频率无显著差异，如果无法获得诊断样本，则在随访期间需要进行测序。单细胞分析显示存在突变的克隆性单核细胞，淋巴细胞和中性粒细胞中也存在相同的突变。靶向测序有助于检测大多数 CMML 患者的克隆性，单细胞测序有助于识别克隆性单核细胞和非髓系细胞突变的共存，提示某些突变是由多能干细胞获得的。
