文章：

通过多模态成像优化离体CAR-T细胞介导的细胞毒性检测方法

Optimizing Ex Vivo CAR-T Cell-Mediated Cytotoxicity Assay through Multimodality Imaging

原文发布日期：9 July 2024

DOI: 10.3390/cancers16142497

类型: Article

开放获取: 是

英文摘要：

CAR-T cell-based therapies have demonstrated remarkable efficacy in treating malignant cancers, especially liquid tumors, and are increasingly being evaluated in clinical trials for solid tumors. With the FDA’s initiative to advance alternative methods for drug discovery and development, full human ex vivo assays are increasingly essential for precision CAR-T development. However, prevailing ex vivo CAR-T cell-mediated cytotoxicity assays are limited by their use of radioactive materials, lack of real-time measurement, low throughput, and inability to automate, among others. To address these limitations, we optimized the assay using multimodality imaging methods, including bioluminescence, impedance tracking, phase contrast, and fluorescence, to track CAR-T cells co-cultured with CD19, CD20, and HER2 luciferase reporter cancer cells in real-time. Additionally, we varied the ratio of CAR-T cells to cancer cells to determine optimal cytotoxicity readouts. Our findings demonstrated that the CAR-T cell group effectively attacked cancer cells, and the optimized assay provided superior temporal and spatial precision measurements of ex vivo CAR-T killing of cancer cells, confirming the reliability, consistency, and high throughput of the optimized assay.

摘要翻译： 

基于CAR-T细胞的疗法在治疗恶性肿瘤，尤其是血液肿瘤方面已展现出显著疗效，并正越来越多地在实体瘤临床试验中得到评估。随着美国食品药品监督管理局（FDA）推动药物发现与开发的替代方法，全人源体外检测在精准CAR-T开发中日益重要。然而，当前主流的体外CAR-T细胞介导的细胞毒性检测方法存在诸多局限，包括使用放射性材料、缺乏实时测量能力、通量低以及无法自动化等。为克服这些不足，我们通过多模态成像方法（包括生物发光、阻抗追踪、相差成像及荧光成像）对检测体系进行优化，实现了对CAR-T细胞与CD19、CD20及HER2荧光素酶报告基因癌细胞共培养过程的实时追踪。此外，我们通过调整CAR-T细胞与癌细胞的比例，确定了最优的细胞毒性检测读数。研究结果表明，CAR-T细胞组能有效攻击癌细胞，优化后的检测方法可提供更优的时空精度测量体外CAR-T杀伤癌细胞的效果，证实了该优化检测方法的可靠性、一致性和高通量特性。

原文链接：

Optimizing Ex Vivo CAR-T Cell-Mediated Cytotoxicity Assay through Multimodality Imaging