PO.TB02.02 · 肿瘤生物学
增强的体内与离体分析可对用于免疫肿瘤学研究的人源化小鼠模型进行更深入的表征
Enhanced in vivo and ex vivo analysis enables deeper characterization of humanized mouse models for immuno-oncology research
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
背景
我们此前已证明，可根据研究目的，使用外周血单个核细胞（PBMC）、PBMC亚群（如T细胞和NK细胞）或CD34⁺造血干细胞（HSC）来构建人源化小鼠模型。随着新一代免疫疗法的推进——包括检查点抑制剂（CPI）、工程化免疫细胞及免疫细胞衔接器——建立合适的体内与离体分析平台变得至关重要。同时含有人类免疫系统和人类肿瘤的全人源化小鼠模型，为评估这些疗法提供了更符合生理学的环境。
方法
通过将CD34⁺细胞、PBMC或纯化的NK细胞或T细胞静脉移植到免疫缺陷小鼠体内，建立人源化小鼠。细胞来源异种移植（CDX）和患者来源异种移植（PDX）肿瘤分别以皮下（s.c.）或原位（静脉注射或注入乳腺脂肪垫）方式接种。肿瘤进展通过卡尺测量（皮下）或生物发光成像（BLI，用于原位模型）进行评估。
血液、骨髓、脾脏和肿瘤组织中免疫细胞组成的定量分析采用流式细胞术。建立了基于Ultramicroscope Blaze的3D光片荧光显微技术，以实现对肿瘤和脾脏中免疫细胞浸润的空间分析与定位。
结果
来自多种癌症类型的CDX和PDX肿瘤均成功植入人源化小鼠体内，植入成功率>70%，且与非人源化对照相比，肿瘤生长动力学无显著差异。免疫治疗揭示了不同的应答者与非应答者特征，表现为免疫细胞浸润模式的差异。
流式细胞术可可靠地监测人类免疫重建，并随时间定量免疫细胞和肿瘤细胞群体。流式细胞术还被用于追踪治疗效果。BLI为原位模型的纵向肿瘤评估提供了非侵入性方法。
基于Ultramicroscope Blaze的3D光片成像能够验证基于FACS的发现，并可视化肿瘤浸润淋巴细胞（TIL），实现对肿瘤及淋巴器官内免疫细胞定位的空间映射。
结论
对我们人源化小鼠模型的持续优化，使得对新兴免疫疗法进行稳健的临床前评估成为可能。整合空间生物学通过对肿瘤微环境内免疫细胞行为提供高分辨率可视化，增强了机制层面的洞察，强化了这些平台的转化价值。
查看英文原文 English abstract
Background
We previously demonstrated that humanized mouse models can be generated using peripheral blood mononuclear cells (PBMCs), PBMC subpopulations such as T and NK cells, or CD34⁺ hematopoietic stem cells (HSCs), depending on the research objective. As next-generation immunotherapies advance-including checkpoint inhibitors (CPIs), engineered immune cells, and immune-cell engagers-establishing suitable in vivo and ex vivo analytical platforms becomes essential. Fully humanized mouse models containing both a human immune system and human tumors provide a more physiologically relevant setting for evaluating these therapies.
Methods
Humanized mice were established through intravenous transplantation of CD34⁺ cells, PBMCs, or purified NK or T cells into immunodeficient mice. Cell-derived xenograft (CDX) and patient-derived xenograft (PDX) tumors were engrafted either subcutaneously (s.c.) or orthotopically (intravenously or into the mammary fat pad). Tumor progression was assessed using caliper measurements (s.c.) or bioluminescence imaging (BLI) for orthotopic models.
Quantitative immune cell composition in blood, bone marrow, spleen, and tumor tissue was analyzed by flow cytometry. 3D-Light-sheet fluorescence microscopy with the Ultramicroscope Blaze was established to enable spatial analysis and localization of immune-cell infiltration in tumors and spleen.
Results
Both CDX and PDX tumors from multiple cancer types successfully engrafted in humanized mice, with >70% engraftment success and no significant differences in tumor growth kinetics compared to non-humanized controls. Immunotherapy treatment revealed distinct responder and non-responder profiles, characterized by differential immune-cell infiltration patterns.
Flow cytometry reliably monitored human immune reconstitution and quantified immune and tumor cell populations over time. Flow cytometry has also been used to follow therapeutic treatment effects. BLI provided a non-invasive method for longitudinal tumor assessment in orthotopic models.
3D-Light-sheet imaging with the Ultramicroscope Blaze enables the confirmation of FACS-based findings and visualized tumor-infiltrating lymphocytes (TILs), unlocking spatial mapping of immune-cell localization within tumors and lymphoid organs.
Conclusions
Continuous refinement of our humanized mouse models enables robust preclinical evaluation of emerging immunotherapies. Integration of spatial biology enhances mechanistic insight by providing high-resolution visualization of immune-cell behavior within the tumor microenvironment, strengthening the translational value of these platforms.
利益披露 Disclosure
M. Stecklum,
EPO GmbH Employment.
J. Alcaniz,
EPO GmbH Employment.
L. Bornemann,
Miltenyi Biotec B.V. & Co. KG Employment.
J. Hoffmann,
EPO GmbH Employment, Stock.