Objectives: Developing ex vivo models that replicate immune–tumor interactions with high fidelity is essential for advancing immunotherapy research, as traditional two-dimensional in vitro systems often lack the complexity required to fully represent these interactions.Methods: In this study, we establish a comprehensive 3D redirect lysis (3D-RDL) assay using colorectal cancer spheroids and adult stem cell-derived, healthy human organoids to evaluate the efficacy and safety profile ofCibisatamab, a bispecific antibody targeting carcinoembryonic antigens (CEAs) on cancer cells and CD3 on T cells. This model allows us to assess cytotoxic activity and immune responses, capturing variations in therapeutic response not observable in simpler systems. Our model integrates live imaging and cytotoxicity analyses to enable precise, real-time tracking of antibody effects on CEA-expressing tumor cells compared to healthy cells. Additionally, by standardizing effector-to-target cell ratios in each co-culture, we establish a reproducible workflow that enhances data accuracy and comparability across assays. Flow cytometry and Granzyme B release profiling further allow us to characterize immune cell activation, revealing distinct T cell activation markers and Granzyme B release patterns tied toCibisatamabtreatment.Results: Our results show thatCibisatamabeffectively induces cell death in cancer spheroids with high CEA expression while being dose-dependent on target, off-tumor binding and killing on non-cancerous cells of healthy organoids with intermediate CEA levels. This highlights our model’s potential to predict clinical immunotherapy outcomes, capturing complex responses like immune activation, therapeutic selectivity, and potential resistance mechanisms.Conclusions: These findings underscore the utility of our model as a reliable, physiologically relevant tool for screening new immunotherapies and advancing our understanding of tumor-immune dynamics.
目的：开发能够高保真模拟免疫-肿瘤相互作用的离体模型对于推进免疫治疗研究至关重要，因为传统的二维体外系统通常缺乏充分展现这些相互作用所需的复杂性。方法：在本研究中，我们利用结直肠癌球体和成人干细胞来源的健康人类类器官，建立了一种全面的三维重定向裂解（3D-RDL）检测方法，以评估靶向癌细胞癌胚抗原（CEA）和T细胞CD3的双特异性抗体Cibisatamab的疗效与安全性特征。该模型使我们能够评估细胞毒活性与免疫反应，捕捉在更简单系统中无法观察到的治疗反应差异。我们的模型整合了活体成像与细胞毒性分析，能够精确、实时地追踪抗体对表达CEA的肿瘤细胞相较于健康细胞的影响。此外，通过标准化每个共培养体系中的效应细胞与靶细胞比例，我们建立了一个可重复的工作流程，从而提高了数据准确性及不同检测间的可比性。流式细胞术和颗粒酶B释放谱分析进一步使我们能够表征免疫细胞活化情况，揭示了与Cibisatamab治疗相关的独特T细胞活化标志物及颗粒酶B释放模式。结果：我们的结果显示，Cibisatamab能有效诱导高表达CEA的癌球体细胞死亡，同时对中等CEA水平的健康类器官非癌细胞表现出剂量依赖性的脱靶结合与杀伤作用。这凸显了我们模型在预测临床免疫治疗结果方面的潜力，能够捕捉免疫激活、治疗选择性及潜在耐药机制等复杂反应。结论：这些发现强调了我们的模型作为一种可靠且具有生理相关性的工具，可用于筛选新型免疫疗法并增进我们对肿瘤-免疫动力学的理解。
肿瘤(癌症)患者之家