Zeming Kerwin Kwek, Quek Kai Yun, Sin Wei-Xiang, Teo Denise Bei Lin, Cheung Ka-Wai, Goh Chin Ren, Kairi Faris, Lee Elizabeth, Lim Francesca Lorraine Wei Inng, Seng Michaela Su-Fern, Soh Shui Yen, Birnbaum Michael E, Han Jongyoon
Critical Analytics for Manufacturing of Personalized Medicine, Singapore-MIT Alliance for Research and Technology (SMART), Singapore, Singapore.
Department of Haematology, Singapore General Hospital, Singapore, Singapore.
Nat Commun. 2025 May 22;16(1):4775. doi: 10.1038/s41467-025-59789-w.
Chimeric Antigen Receptor (CAR) T cell therapy is a pivotal treatment for hematological malignancies. However, CAR T cell products exhibit batch-to-batch variability in cell number, quality, and in vivo efficacy due to donor-to-donor heterogeneity, and pre/post-manufacturing processes, and the manufacturing of such products necessitates careful testing, both post-manufacturing and pre-infusion. Here, we introduce the Cell Trajectory Modulation (CTM) assay, a microfluidic, label-free approach for the rapid evaluation of the functional attributes of CAR T cells based on biophysical features (i.e., size, deformability). CTM assay correlates with phenotypic metrics, including CD4:CD8 ratio, memory subtypes, and cytotoxic activity. Validated across multiple donors and culture platforms, the CTM assay requires fewer than 10,000 cells and delivers results within 10 minutes. Compared to labeled flow cytometry processing, the CTM assay offers real-time data to guide adaptive manufacturing workflows. Thus, the CTM assay offers an improvement over existing phenotypic assessments, marking a step forward in advancing CAR T cell therapy manufacturing.
嵌合抗原受体（CAR）T细胞疗法是血液系统恶性肿瘤的关键治疗方法。然而，由于供体间的异质性以及制造前/后的过程，CAR T细胞产品在细胞数量、质量和体内疗效方面表现出批次间的差异，并且此类产品的制造需要在制造后和输注前进行仔细检测。在此，我们介绍细胞轨迹调制（CTM）检测法，这是一种基于生物物理特征（即大小、可变形性）对CAR T细胞功能属性进行快速评估的微流控、无标记方法。CTM检测法与包括CD4:CD8比率、记忆亚型和细胞毒性活性在内的表型指标相关。该CTM检测法在多个供体和培养平台上得到验证，所需细胞少于10000个，且能在10分钟内给出结果。与标记流式细胞术处理相比，CTM检测法可提供实时数据以指导适应性制造工作流程。因此，CTM检测法相较于现代表型评估有了改进，标志着CAR T细胞疗法制造向前迈进了一步。
