PO.BCS01.08 · 生物信息与计算
通过一种新型17重免疫荧光方法补充H&E对实体瘤进行空间免疫分析
Spatial immune profiling of solid tumors by complementing H&E with a novel 17-plex immunofluorescence approach
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
背景：空间生物学越来越多地利用“空间邻域”来整合细胞的组织和相互作用方式，但定义和识别这些邻域的标准仍在发展中。多重荧光免疫组化（fIHC）提供了对免疫和肿瘤细胞状态的详细分析，而苏木精和伊红（H&E）染色仍然是可视化组织和细胞形态的金标准。我们在Orion™（RareCyte）平台上验证了一种17重fIHC检测，并将fIHC与同一切片的H&E图像对齐，以提高乳腺癌、肺癌、前列腺癌、黑色素瘤和结直肠癌中细胞邻域识别的保真度。
方法：我们开发了一个可推广的工作流程，用于对齐同一切片的H&E和17重fIHC Orion图像，通过整合细胞表型分析（通过fIHC）与组织结构（通过H&E）来评估细胞邻域。17重fIHC检测包括靶向Granzyme B、Ki67、CD3e、CD20、CD4、CD163、CD8a、PD-L1、NKp46、PD-1、LAG3、FOXP3、CD11b、CD138、SOX10、Cytokeratin的抗体-荧光团偶联物以及Hoechst核染色。切片首先通过fIHC染色和成像，随后在Orion上进行H&E染色和成像。细胞表型分析和切片配准在HALO®（Indica Labs）上进行。
结果：将H&E与多重fIHC整合，支持了空间主题的识别，包括富集肿瘤浸润淋巴细胞的区域、免疫荒漠以及占据肿瘤/间质界面的混合/过渡区。这种方法通过将细胞表型与组织结构形态相连接，改善了对这些模式的可解释性，支持以更高的一致性识别微环境。
结论：将H&E与多重fIHC结合，能够在多种肿瘤类型中实现对细胞邻域的解剖学信息化映射。这种发现导向的工作流程改善了空间模式的可解释性，可适应多样化的数据集和流程，并可能为免疫微环境提供新见解，以指导下一代治疗。
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查看英文原文 English abstract
Background: Spatial biology has increasingly leveraged “spatial neighborhoods” to integrate how cells are organized and interact, yet standards for defining and identifying these neighborhoods are still developing. Multiplex fluorescence immunohistochemistry (fIHC) offers detailed profiling of immune and tumor cell states, while hematoxylin and eosin (H&E) staining remains a gold standard for visualizing tissue and cellular morphology. We validated a 17-plex fIHC assay on the Orion™ (RareCyte) platform and aligned the fIHC with same slide H&E images to improve fidelity of cellular neighborhoods identification across breast, lung, prostate, melanoma, and colorectal cancers.
Methods: We developed a generalizable workflow to align same-slide H&E and 17-plex fIHC Orion images for assessment of cellular neighborhoods through integration cell phenotyping (via fIHC) with tissue architecture (via H&E). The 17-plex fIHC assay includes antibody-fluorophore conjugates targeting Granzyme B, Ki67, CD3e, CD20, CD4, CD163, CD8a, PD-L1, NKp46, PD-1, LAG3, FOXP3, CD11b, CD138, SOX10, Cytokeratin, and Hoechst nuclear stain. Slides were first stained and imaged by fIHC, followed by H&E staining and imaging on the Orion. Cell phenotyping and slide registration were performed on HALO® (Indica Labs).
Results: Integrating H&E with multiplex fIHC supported identification of spatial themes, including regions with enriched tumor-infiltrating lymphocytes, immune deserts, and mixed/transitional zones populating the tumor/stroma interface. This approach improves interpretability of these patterns by connecting cell phenotypes with tissue architecture morphology, supporting identification of microenvironments with greater consistency.
Conclusions: Combining H&E with multiplex fIHC enables anatomically informed mapping of cellular neighborhoods across multiple tumor types. This discovery-oriented workflow improves interpretability of spatial patterns, is adaptable to diverse datasets and pipelines, and may yield new insights into the immune microenvironment to inform next-generation therapies.
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利益披露 Disclosure
R. Van Krieken,
Navigate BioPharma Services, Inc. Employment.
Novartis Stock.
J. Kaur,
Navigate BioPharma Services, Inc. Employment.
X. Li,
Navigate BioPharma Services, Inc. Employment.
N. Dakappagari,
Navigate BioPharma Services, Inc. Employment.
J. Bordeaux,
Navigate BioPharma Services, Inc. Employment.