State Institution "National Research Center for Radiation Medicine, National Academy of Medical Sciences of Ukraine", Kyiv, Ukraine.
Exp Oncol. 2023 Dec 28;45(3):322-327. doi: 10.15407/exp-oncology.2023.03.322.
The typical chronic lymphocytic leukemia (CLL) immunophenotype is vital for diagnosis, but the expression of some antigens varies and has prognostic value. There are data that reduced CD20 expression is associated with NOTCH1 and SF3B1 gene mutations.
To determine a high-risk group of CLL patients for prediction of unfavorable NOTCH1 and SF3B1 gene mutations based on immunophenotyping of leukemic cells.
Flow cytometric and molecular-genetic analysis (mutations of NOTCH1, SF3B1, and TP53 genes using the polymerase chain reaction followed by direct sequencing) was performed in a group of 86 previously untreated CLL patients.
The immunophenotype of leukemic cells of all examined patients met the criteria of CLL diagnosis. NOTCH1 gene mutations were found in 21 patients (24.4%), and SF3B1 gene mutations - in 7 patients (8.1%). There were no TP53 gene mutations among the examined patients. A decreased number of CD20+CD5+ cells and a downward trend in the relative index of mean fluorescence intensity (iMFI) of CD20+ cells were found in patients with NOTCH1 and SF3B1 gene mutations. Based on the iMFI level (higher and/or lower than 3.0) and the number of CD20+CD5+ cells among all B-cells (higher and/or lower than 50%), we distinguished CLL cases with low and relatively high levels of CD20 antigen expression. Using ROC analysis and the parameter of low CD20 antigen expression, we could predict the presence of NOTCH1 and SF3B1 gene mutations in 73.3 ± 0.06% of patients (p = 0.001). The risk of NOTCH1 and SF3B1 gene mutations in cases with low CD20 antigen expression was 6.96 (95% CI = 2.53-19.18; p = 0.0001). The revealed regularities were statistically significant for patients in whom the diagnosis was established in all Binet - Rai stages except A0-AI.
Our data confirmed a reduced CD20 expression in CLL patients with NOTCH1 and SF3B1 mutations. In addition, an approach was proposed to identify high-risk CLL patients for prediction of such mutations: previously untreated CLL patients at advanced Binet - Rai stages (BII, CIII, CIV) with a reduced number of double-positive CD20+CD5+ cells in peripheral blood and/or low iMFI of CD20+ cells.
典型的慢性淋巴细胞白血病 (CLL) 免疫表型对诊断至关重要，但某些抗原的表达存在差异，且具有预后价值。有数据表明，CD20 表达降低与 NOTCH1 和 SF3B1 基因突变有关。
基于白血病细胞的免疫表型，确定 CLL 患者的高危组，以预测不利的 NOTCH1 和 SF3B1 基因突变。
对 86 例未经治疗的 CLL 患者进行流式细胞术和分子遗传学分析（使用聚合酶链反应直接测序检测 NOTCH1、SF3B1 和 TP53 基因突变）。
所有检查患者的白血病细胞免疫表型均符合 CLL 诊断标准。21 例患者（24.4%）存在 NOTCH1 基因突变，7 例患者（8.1%）存在 SF3B1 基因突变。检查患者中均未发现 TP53 基因突变。NOTCH1 和 SF3B1 基因突变患者的 CD20+CD5+细胞数量减少，CD20+细胞相对平均荧光强度指数（iMFI）呈下降趋势。基于 iMFI 水平（高于和/或低于 3.0）和所有 B 细胞中 CD20+细胞的数量（高于和/或低于 50%），我们区分了具有低和相对高 CD20 抗原表达水平的 CLL 病例。使用 ROC 分析和低 CD20 抗原表达参数，我们可以在 73.3±0.06%的患者中预测存在 NOTCH1 和 SF3B1 基因突变（p=0.001）。CD20 抗原低表达病例中 NOTCH1 和 SF3B1 基因突变的风险为 6.96（95%CI=2.53-19.18;p=0.0001）。除了 A0-AI 期，在所有 Binet-Rai 分期确诊的患者中，这些规律均具有统计学意义。
我们的数据证实了 CLL 患者中存在 NOTCH1 和 SF3B1 突变时 CD20 表达降低。此外，我们提出了一种识别高危 CLL 患者以预测这些突变的方法：未经治疗的处于晚期 Binet-Rai 分期（BII、CIII、CIV）的 CLL 患者，外周血双阳性 CD20+CD5+细胞数量减少和/或 CD20+细胞 iMFI 降低。
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