文章：

拓扑异构酶II抑制剂诱导白血病T细胞系及人外周血单个核细胞中DNA双链断裂与游离DNA的异质性形成

Heterogeneous Formation of DNA Double-Strand Breaks and Cell-Free DNA in Leukemia T-Cell Line and Human Peripheral Blood Mononuclear Cells in Response to Topoisomerase II Inhibitors

原文发布日期：12 November 2024

DOI: 10.3390/cancers16223798

类型: Article

开放获取: 是

英文摘要：

Background: Improving precision medicine in chemotherapy requires highly sensitive and easily applicable diagnostic tools. In addition, non-invasive molecular real-time monitoring of cytotoxic response is highly desirable. Here, we employed the kinetics of DNA double-strand breaks (DSB) and cell-free DNA (cfDNA) in a cell model of topoisomerase II-inhibitors in T cell leukemia (Jurkat cells) compared to normal cells (peripheral blood mononuclear cells, PBMCs). Methods: We applied automated microscopy to quantify immuno-stained phosphorylated H2AX (γH2AX) as a marker for either DNA damage response (DDR) or cell death and quantitative PCR-based analysis of nuclear and mitochondrial cfDNA concentrations. Results: Jurkat cells displayed a DDR to cytotoxic drug treatment significantly earlier than PBMCs, and etoposide (ETP) induced DSB formation faster than doxorubicin (DOX) in both Jurkat and PBMCs. Jurkat cells exhibited an earlier cytotoxic response compared to PBMC, with a significantly increased mitochondrial cfDNA formation after 2 h of DOX application. In PBMCs, increased cell death was detected after 4 h of incubation with ETP, whereas DOX treatment was less effective. Conclusions: Both automated microscopy and mitochondrial cfDNA quantification analysis indicate that (malignant) Jurkat cells are more sensitive to DOX than (healthy) PBMC. Our real-time approach can improve DDR inducing drug selection and adaptation in cancer therapy and aids in decisions for optimal patient biosampling.

摘要翻译： 

背景：提升化疗精准医疗水平需要高灵敏度且易于应用的诊断工具。此外，对细胞毒性反应进行无创分子实时监测具有重要临床价值。本研究通过对比拓扑异构酶II抑制剂在T细胞白血病（Jurkat细胞）与正常细胞（外周血单个核细胞，PBMCs）中的效应，系统分析了DNA双链断裂动力学与游离DNA的动态变化。方法：采用自动化显微成像技术定量免疫染色的磷酸化H2AX（γH2AX）作为DNA损伤应答或细胞死亡标志物，同时通过定量PCR技术检测核DNA与线粒体cfDNA浓度。结果：Jurkat细胞对细胞毒性药物的DDR响应显著早于PBMCs；在两种细胞系中，依托泊苷诱导DSB形成的速度均快于阿霉素。与PBMCs相比，Jurkat细胞表现出更早的细胞毒性反应，阿霉素处理2小时后线粒体cfDNA生成显著增加。PBMCs经依托泊苷孵育4小时后可检测到细胞死亡增加，而阿霉素处理效果较弱。结论：自动化显微成像与线粒体cfDNA定量分析均表明（恶性）Jurkat细胞比（健康）PBMCs对阿霉素更敏感。本研究所建立的实时监测方法可优化癌症治疗中DDR诱导药物的选择与调整策略，并为患者最佳生物样本采集时机的决策提供依据。

原文链接：

Heterogeneous Formation of DNA Double-Strand Breaks and Cell-Free DNA in Leukemia T-Cell Line and Human Peripheral Blood Mononuclear Cells in Response to Topoisomerase II Inhibitors