PO.TB04.01 · 肿瘤生物学
NAMs时代:将肿瘤异质性和免疫背景纳入与人体相关的患者化身癌症模型
The era of NAMS, incorporating tumor heterogeneity and immune context into human-relevant patient avatar cancer models
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
患者来源微肿瘤化身处于NAMs(新方法学)的前沿,它们能够重现肿瘤内异质性并纳入完整的免疫细胞肿瘤微环境。为解决转化性癌症建模中持续存在的挑战,我们开发了一个平台,能够将肿瘤浸润淋巴细胞(TIL)、CAR T细胞、PBMC、检查点阻断剂(aPD1)、抗体药物偶联物和常规化疗药物二次添加到这些微肿瘤系统中。利用来自卵巢癌、胰腺癌、膀胱癌、结直肠癌和子宫内膜癌的经验证模型,我们证明了对治疗反应和免疫动态的高保真评估。高分辨率延时成像以单细胞分辨率揭示了免疫效应细胞清除肿瘤的时空动力学,更多影像展示了免疫介导的肿瘤根除机制。这些发现验证了微肿瘤化身在临床前免疫肿瘤学研究和机制检测方面的稳健性和转化潜力。
目的:评估微肿瘤化身在癌症研究中的应用,重点关注在NAMs中整合肿瘤异质性和免疫背景以构建忠实的转化模型。
方法:通过使用LIVE/DEAD细胞成像以及配备NSPARC超分辨率检测器的Nikon AXR和多光子共聚焦显微镜进行免疫荧光成像,定期评估NAMs的活力。使用多种针对细胞核、Ki67活性、I型胶原、EpCAM、E-Cadherin、N-Cadherin、VE-Cadherin、Vimentin、CD45、CD3、Trop-2、Nectin-4等的染色对患者微肿瘤进行表征。在长时间延时实验之后,采用我们常规的固定和透化方案进行超分辨率显微成像。
结果:患者来源微组织在培养中维持了较长时间,并表现出持续的活力和多样的细胞组成,这一点通过上述成像技术得到证实。免疫荧光在所有微肿瘤中揭示了增殖活性、动态的组织结构和可变的标志物表达。使用靶向药物、免疫检查点抑制剂和广谱细胞毒药物的药物反应研究表明,体外结果与患者的治疗结果一致,包括实体瘤模型中的敏感和耐药模式。此外,使用CAR T细胞和TIL等工程化免疫细胞的功能检测证实了这些微肿瘤平台内稳健的免疫介导肿瘤杀伤。
结论:这些结果支持,先进的患者来源微组织NAMs现在能够为提高转化性癌症研究和治疗发现的精准度提供稳健的机会。
查看英文原文 English abstract
Patient-derived microtumor avatars are at the forefront of NAMs (New Approach Methodologies) that recapitulate intratumoral heterogeneity and incorporate the full immune cell tumor microenvironment. To address ongoing challenges in translational cancer modeling, we developed a platform enabling secondary addition of tumor-infiltrating lymphocytes (TILs), CAR T-cells, PBMCs, checkpoint blockade agents (aPD1), antibody-drug conjugates, and conventional chemotherapies to these microtumor systems. Using validated models from ovarian, pancreatic, bladder, colorectal, and endometrial cancers, we have demonstrated high-fidelity assessment of therapeutic response and immune dynamics. High-resolution time-lapse imaging reveals spatiotemporal kinetics of tumor clearance by immune effectors at single-cell resolution, and additional movies demonstrate mechanisms of immune-mediated tumor eradication. These findings benchmark the robustness and translational potential of microtumor avatars for preclinical immuno-oncology research and mechanistic testing.
Purpose: To evaluate the use of microtumor avatars in cancer research, focusing on the integration of tumor heterogeneity and immune context for faithful translational models in NAMs.
Methods: NAMs viability was regularly assessed via immunofluorescence imaging using LIVE/DEAD cell imaging and a Nikon AXR and Multiphoton confocal microscope outfitted with the NSPARC super-resolution detector. Characterization of the patient microtumors was performed using a variety of stains for nuclei, Ki67 activity, Collagen Type I, EpCAM, E-Cadherin, N-Cadherin, VE-Cadherin, Vimentin, CD45, CD3, Trop-2, Nectin-4, and others. Following long duration time-lapse experiments, super-resolution microscopy was performed using our typical fixation and permeabilization protocols.
Results: Patient-derived microtissues were maintained in culture for extended periods and exhibited sustained viability and diverse cellular composition, as confirmed by the above imaging techniques. Proliferative activity, dynamic tissue architecture, and variable marker expression were revealed using immunofluorescence across all microtumors. Drug response studies using targeted agents, immune checkpoint inhibitors, and broad-spectrum cytotoxics demonstrated in vitro outcomes concordant with patient therapies, including both sensitivity and resistance patterns in solid tumor models. Furthermore, functional assays with engineered immune cells such as CAR T-cells and TILs confirmed robust immune-mediated tumor killing within these microtumor platforms.
Conclusions: These results support that advanced patient-derived microtissue NAMs now enable robust opportunities for increased precision in translational cancer research and therapeutic discovery.
利益披露 Disclosure
W. Sawyer,
Merck ).
R. Smolchek,
Merck ).
D. Nguyen,
Merck ).
A. Pepe,
Merck ).
D. Pedro,
Merck ).
E. George,
Merck ).
R. Li,
Merck ).