Single-cell sequencing reveals shared clonal signatures in nonmalignant B and tumor cells in T-prolymphocytic leukemia

Blood Neoplasia Pub Date : 2025-02-16 DOI:10.1016/j.bneo.2025.100076

Caroline Hesselager , Ingrid Thörn , Millaray Marincevic , Claes Ladenvall , Jonas Almlöf , Sara Löfgren , Simone Weström , Helena Nord , Lesley-Ann Sutton , Lucia Cavelier , Panagiotis Baliakas , Rose-Marie Amini

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Abstract

This study aimed to elucidate the clonal origin and evolutionary dynamics of T-cell prolymphocytic leukemia (T-PLL) using targeted next generation sequencing (NGS) of paired samples from diagnosis and relapse. DNA from both nonmalignant and tumor cells was extracted from sorted cell fractions obtained from 16 patients with T-PLL. NGS was performed using a customized Haloplex gene panel comprising 19 genes recurrently mutated in T-PLL (ATM and JAK/STAT pathway). Droplet digital polymerase chain reaction was performed to confirm mutations detected by NGS with low variant allele frequencies. Single-cell analysis of genomic DNA combined with cell surface protein markers was performed using the Mission Bio Tapestri Platform. The most frequently mutated gene was ATM (n = 10) followed by STAT5B (n = 7), JAK3 (n = 3), EZH2 (n = 3), BCOR (n = 1), and STAT6 (n = 1). Relapse samples were available for 9 of the 16 patients. Varying patterns of clonal shifts were observed between diagnosis and relapse (increase, decrease, both increase and decrease, and no change). The presence of pathogenic variants in ATM, EZH2, STAT5B, and JAK3 in both normal sorted B cells and clonal T cells was confirmed. Single-cell analysis revealed shared mutations in both nonmalignant B and clonal T cells in 1 case. A pathogenic variant within the ATM gene of potential germ line origin was observed in 1 case. T-PLL exhibits variable patterns of clonal evolution between diagnosis and relapse. Single-cell multiomics analysis reveals shared mutational signatures in both nonmalignant B cells and clonal T cells. The role of germ line ATM mutations in the pathogenesis of T-PLL should be further explored.

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单细胞测序显示t -原淋巴细胞白血病的非恶性B细胞和肿瘤细胞具有共同的克隆特征

摘要本研究旨在利用诊断和复发配对样本的靶向下一代测序（targeted next generation sequencing， NGS）研究t细胞原淋巴细胞白血病（T-PLL）的克隆起源和进化动力学。从16例T-PLL患者的分选细胞中提取非恶性细胞和肿瘤细胞的DNA。NGS采用定制的Haloplex基因面板，该面板包含T-PLL （ATM和JAK/STAT通路）中19个反复突变的基因。采用微滴数字聚合酶链反应对NGS检测到的低变异等位基因频率突变进行确证。使用Mission Bio Tapestri平台进行基因组DNA与细胞表面蛋白标记的单细胞分析。最常见的突变基因是ATM (n = 10)，其次是STAT5B （n = 7）、JAK3 （n = 3）、EZH2 （n = 3）、BCOR （n = 1）和STAT6 （n = 1）。16例患者中有9例有复发样本。在诊断和复发之间观察到不同的克隆转移模式（增加，减少，增加和减少，无变化）。证实ATM、EZH2、STAT5B和JAK3在正常分选B细胞和克隆T细胞中均存在致病性变异。单细胞分析显示1例非恶性B细胞和克隆T细胞共有突变。在1例中观察到潜在种系起源的ATM基因内的致病变异。T-PLL在诊断和复发之间表现出不同的克隆进化模式。单细胞多组学分析揭示了在非恶性B细胞和克隆T细胞中共享的突变特征。种系ATM突变在T-PLL发病机制中的作用有待进一步探讨。

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Blood Neoplasia

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