Maria Alieva, Mario Barrera Román, Sam de Blank, Diana Petcu, Amber L. Zeeman, Noël M. M. Dautzenberg, Annelisa M. Cornel, Cesca van de Ven, Rob Pieters, Monique L. den Boer, Stefan Nierkens, Friso G. J. Calkoen, Hans Clevers, Jürgen Kuball, Zsolt Sebestyén, Ellen J. Wehrens, Johanna F. Dekkers, Anne C. Rios
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Here, we describe the application of BEHAV3D, a platform that implements multi-color live 3D imaging and computational tools for: (i) analyzing tumor death dynamics at both single-organoid or cell and population levels, (ii) classifying T cell behavior and (iii) producing data-informed 3D images and videos for visual inspection and further insight into obtained results. Together, this enables a refined assessment of how solid and liquid tumors respond to cellular immunotherapy, critically capturing both inter- and intratumoral heterogeneity in treatment response. In addition, BEHAV3D uncovers T cell behavior involved in tumor targeting, offering insight into their mode of action. Our pipeline thereby has strong implications for comparing, prioritizing and improving immunotherapy products by highlighting the behavioral differences between individual tumor donors, distinct T cell therapy concepts or subpopulations. The protocol describes critical wet lab steps, including co-culture preparations and fast 3D imaging with live cell dyes, a segmentation-based image processing tool to track individual organoids, tumor and immune cells and an analytical pipeline for behavioral profiling. This 1-week protocol, accessible to users with basic cell culture, imaging and programming expertise, can easily be adapted to any type of co-culture to visualize and exploit cell behavior, having far-reaching implications for the immuno-oncology field and beyond. BEHAV3D is a 3D live imaging platform for analyzing engineered T cell behavior and tumor response. 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引用次数: 0
摘要
通过使用源自患者的材料和免疫细胞共培养物建立免疫肿瘤学模型,可以推进我们对免疫细胞以患者特异性方式靶向肿瘤的理解,为改进细胞免疫疗法提供线索。然而,充分利用这些活体培养物需要分析建模的动态细胞特征,而目前这方面的方案还很有限。在这里,我们介绍了 BEHAV3D 的应用,这是一个实现多色实时三维成像和计算工具的平台,可用于(i)在单器官或细胞和群体水平上分析肿瘤死亡动态,(ii)对 T 细胞行为进行分类,(iii)生成数据信息三维图像和视频,用于视觉检查和进一步深入了解所获得的结果。这样就能对实体瘤和液体瘤如何对细胞免疫疗法做出反应进行精细评估,并准确捕捉治疗反应中瘤间和瘤内的异质性。此外,BEHAV3D 还能发现参与肿瘤靶向的 T 细胞行为,从而深入了解它们的作用模式。因此,我们的研究方法通过突出单个肿瘤供体、不同 T 细胞疗法概念或亚群之间的行为差异,对比较、优先考虑和改进免疫疗法产品具有重要意义。该方案描述了关键的湿实验室步骤,包括共培养制备和活细胞染料快速三维成像,基于分割的图像处理工具跟踪单个器官组织、肿瘤和免疫细胞,以及行为分析管道。这个为期一周的方案可供具备基本细胞培养、成像和编程专业知识的用户使用,可轻松适用于任何类型的共培养,以可视化和利用细胞行为,对免疫肿瘤学领域及其他领域具有深远影响。
BEHAV3D: a 3D live imaging platform for comprehensive analysis of engineered T cell behavior and tumor response
Modeling immuno-oncology by using patient-derived material and immune cell co-cultures can advance our understanding of immune cell tumor targeting in a patient-specific manner, offering leads to improve cellular immunotherapy. However, fully exploiting these living cultures requires analysis of the dynamic cellular features modeled, for which protocols are currently limited. Here, we describe the application of BEHAV3D, a platform that implements multi-color live 3D imaging and computational tools for: (i) analyzing tumor death dynamics at both single-organoid or cell and population levels, (ii) classifying T cell behavior and (iii) producing data-informed 3D images and videos for visual inspection and further insight into obtained results. Together, this enables a refined assessment of how solid and liquid tumors respond to cellular immunotherapy, critically capturing both inter- and intratumoral heterogeneity in treatment response. In addition, BEHAV3D uncovers T cell behavior involved in tumor targeting, offering insight into their mode of action. Our pipeline thereby has strong implications for comparing, prioritizing and improving immunotherapy products by highlighting the behavioral differences between individual tumor donors, distinct T cell therapy concepts or subpopulations. The protocol describes critical wet lab steps, including co-culture preparations and fast 3D imaging with live cell dyes, a segmentation-based image processing tool to track individual organoids, tumor and immune cells and an analytical pipeline for behavioral profiling. This 1-week protocol, accessible to users with basic cell culture, imaging and programming expertise, can easily be adapted to any type of co-culture to visualize and exploit cell behavior, having far-reaching implications for the immuno-oncology field and beyond. BEHAV3D is a 3D live imaging platform for analyzing engineered T cell behavior and tumor response. This provides insights into the mode of action of cellular immunotherapy, capturing heterogeneity within and between tumors during treatment response.
期刊介绍:
Nature Protocols focuses on publishing protocols used to address significant biological and biomedical science research questions, including methods grounded in physics and chemistry with practical applications to biological problems. The journal caters to a primary audience of research scientists and, as such, exclusively publishes protocols with research applications. Protocols primarily aimed at influencing patient management and treatment decisions are not featured.
The specific techniques covered encompass a wide range, including but not limited to: Biochemistry, Cell biology, Cell culture, Chemical modification, Computational biology, Developmental biology, Epigenomics, Genetic analysis, Genetic modification, Genomics, Imaging, Immunology, Isolation, purification, and separation, Lipidomics, Metabolomics, Microbiology, Model organisms, Nanotechnology, Neuroscience, Nucleic-acid-based molecular biology, Pharmacology, Plant biology, Protein analysis, Proteomics, Spectroscopy, Structural biology, Synthetic chemistry, Tissue culture, Toxicology, and Virology.