Department of Molecular Genetics and Cell Biology, The University of Chicago, Chicago, IL, USA.
Department of Biopharmaceutical Sciences, The University of Illinois at Chicago, Chicago, IL, USA.
Lab Invest. 2019 Sep;99(9):1400-1413. doi: 10.1038/s41374-018-0156-y. Epub 2018 Nov 6.
Enumeration of tumor-infiltrating lymphocytes (TILs) in H&E stained tissue sections has demonstrated limited value in predicting immune responses to cancer immunotherapy, likely reflecting the diversity of cell types and immune activation states among tumor infiltrates. Multiparametric flow cytometry enables robust phenotypic and functional analysis to distinguish suppression from activation, but tissue dissociation eliminates spatial context. Multiplex methods for immunohistochemistry (IHC) are emerging, but these interrogate only a single tissue section at a time. Here, we report transparent tissue tomography (T3) as a tool for three-dimensional (3D) imaging cytometry in the complex architecture of the tumor microenvironment, demonstrating multiplexed immunofluorescent analysis in core needle biopsies. Using T3 imaging, image processing and machine learning to map CD3CD8 cytotoxic T cells (CTLs) in whole core needle biopsies from Her2 murine mammary tumors and human head and neck surgical specimens revealed marked inhomogeneity within single needle cores, confirmed by serial section IHC. Applying T3 imaging cytometry, we discovered a strong spatial correlation between CD3CD8 CTLs and microvasculature in the EGFR parenchyma, revealing significant differences among head and neck cancer patients. These results show that T3 offers simple and rapid access to three-dimensional and quantitative maps of the tumor microenvironment and immune infiltrate, offering a new diagnostic tool for personalized cancer immunotherapy.
在 H&E 染色组织切片中对肿瘤浸润淋巴细胞（TILs）进行计数，其预测癌症免疫治疗反应的价值有限，这可能反映了肿瘤浸润物中细胞类型和免疫激活状态的多样性。多参数流式细胞术能够进行强大的表型和功能分析，以区分抑制和激活，但组织解离消除了空间背景。免疫组化（IHC）的多重方法正在出现，但这些方法一次只能检测单个组织切片。在这里，我们报告透明组织断层成像术（T3）作为肿瘤微环境复杂结构中三维（3D）成像细胞术的工具，在核心针活检中证明了多重免疫荧光分析。使用 T3 成像、图像处理和机器学习来绘制 Her2 鼠乳腺肿瘤和人类头颈部手术标本的整个核心针活检中的 CD3CD8 细胞毒性 T 细胞（CTLs），发现单个针芯内存在明显的不均匀性，通过连续切片 IHC 得到证实。应用 T3 成像细胞术，我们发现 CD3CD8 CTLs 与 EGFR 实质中的微血管之间存在很强的空间相关性，揭示了头颈部癌症患者之间的显著差异。这些结果表明，T3 为肿瘤微环境和免疫浸润的三维和定量图谱提供了简单快速的获取途径，为个性化癌症免疫治疗提供了一种新的诊断工具。
