Complex In-Vitro Models, GSK, Collegeville, PA, United States of America.
Genome Biology, GSK, Collegeville, PA, United States of America.
Biofabrication. 2024 Oct 24;17(1). doi: 10.1088/1758-5090/ad847f.
Tumors in patients non-responsive to immunotherapy harbor a series of barriers that impede the efficacy of effector T-cells. Consequently, therapeutically modulating the chemotaxis machinery to enable effector T cell infiltration and function in the tumor could result in more successful therapeutic outcomes. Complexmodels allow re-creation oftumor complexities in ansetting, allowing improved translatability to patient biology at the laboratory scale. We identified a gap in available industrial scale microphysiological (MPS) assays for faster validation of targets and strategies that enable T-cell chemotaxis and effector function within tumor microenvironments. Using a commercially available, 96-chip 2-lane microfluidic assay system, we present a novel, scalable, complexMPS assay to study 3D T-cell chemotaxis and function within native, extracellular matrix (ECM)-rich multicellular tumor environments. Activated or naïve CD3+ T-cells stained with far-red nuclear stain responded to the chemokine gradients generated within the matrigel-collagen ECM by migrating into the microfluidic channel (∼5 mm horizontal window), in a concentration- and cell type-dependent manner. Furthermore, we observed and tracked chemotaxis and cancer cell killing function of antigen-specific CD4.CD8. chimeric antigen receptor (CAR)-T cells that responded to CXCR3 agonist gradient built through the expansive 5 mm of cancer cell colony containing stroma. The 2-lane assay system yielded useful information regarding donor and dose-dependent differences in CAR-T cell chemotaxis and tumor killing. The scalable assay system allows a granular window into immune cell migration and function in tissue spaces beyond endothelium, addressing a missing gap in studying tissue-specific immune cell chemotaxis and function to bring forward advancements in cancer immunotherapy.
在对免疫疗法无反应的患者中，肿瘤存在一系列阻碍效应 T 细胞疗效的障碍。因此，通过治疗性调节趋化机制，使效应 T 细胞能够浸润肿瘤并发挥功能，可能会产生更成功的治疗效果。复杂模型允许在设置中重新创建肿瘤的复杂性，从而在实验室规模上提高对患者生物学的可转化性。我们发现现有的工业规模微生理（MPS）检测方法存在空白，无法快速验证能够使 T 细胞趋化和在肿瘤微环境中发挥效应功能的靶点和策略。我们使用一种商业上可用的、96 芯片 2 通道微流控检测系统，提出了一种新颖的、可扩展的复杂 MPS 检测方法，用于研究天然细胞外基质（ECM）丰富的多细胞肿瘤环境中 3D T 细胞的趋化性和功能。用远红核染料染色的激活或幼稚 CD3+T 细胞对基质胶-胶原 ECM 中产生的趋化因子梯度做出响应，以浓度和细胞类型依赖的方式迁移到微流控通道（约 5 毫米水平窗口）。此外，我们观察并跟踪了特异性针对抗原的 CD4.CD8.嵌合抗原受体（CAR）-T 细胞的趋化性和杀伤肿瘤细胞的功能，这些细胞对通过含有基质的 5 毫米肿瘤细胞集落构建的 CXCR3 激动剂梯度做出反应。2 通道检测系统提供了有关供体和剂量依赖性差异的有用信息，这些差异与 CAR-T 细胞的趋化性和肿瘤杀伤有关。该可扩展的检测系统允许深入了解免疫细胞在超越内皮细胞的组织空间中的迁移和功能，填补了研究组织特异性免疫细胞趋化性和功能以推动癌症免疫治疗进展的空白。
