PO.CL01.13 · 临床研究
在临床试验中使用 17 重免疫荧光检测对实体瘤进行全面的免疫图谱分析
Comprehensive immune landscape analysis of solid tumors using a 17-Plex immunofluorescence assay in clinical trials
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
背景：肿瘤微环境高度复杂；越来越多的证据表明，免疫细胞组成和空间分布可促进疾病进展并抑制抗肿瘤免疫。目前人们正日益努力利用新技术深入分析肿瘤免疫微环境的复杂性。我们验证了一个 17 重、亚细胞分辨率的泛免疫细胞组合，用于在全组织切片中探究免疫细胞类型及其空间组织。
方法：我们与 RareCyte® 合作，利用 Orion™ 平台共同开发了一种 17 重泛免疫细胞免疫荧光（IF）检测，用于表征来自黑色素瘤、乳腺癌、肺癌、前列腺癌和结直肠癌的福尔马林固定石蜡包埋（FFPE）组织中的免疫细胞群体及其功能状态。使用 intelliPATH™ FLX（Biocare Medical），采用靶向 Granzyme B、Ki67、CD3e、CD20、CD4、CD163、CD8a、PDL1、NKp46、PD-1、LAG3、FOXP3、CD11b、CD138、SOX10、Cytokeratin 的抗体-荧光基团偶联物混合液以及 Hoechst 核染料，对单张 4 微米 FFPE 切片进行一步法染色。成像在 Orion™ 平台上进行，数据使用 HALO®（Indica Labs）进行分析。
结果：该检测能够全面地对肿瘤和基质区室内的 T 细胞、B 细胞、NK 细胞和髓系细胞分布进行空间图谱分析。基于免疫检查点和活化标志物的表达，区分了 T 细胞的功能状态（细胞毒性、调节性、耗竭）以及巨噬细胞/B 细胞生态位。邻近分析识别了免疫突触和有组织的淋巴结构，有助于稳健的微环境表征。
结论：Orion™ 17 重泛免疫细胞 IF 检测将能够对临床试验标本中的肿瘤微环境进行全面的定性和定量分析。我们预期该方法将通过实现对多种癌症类型中免疫细胞和肿瘤细胞空间生物学的详细表征，支持下一代疗法的开发。
查看英文原文 English abstract
Background: The tumor micro-environment is highly complex; accumulating evidence indicates that immune cell composition and spatial distribution can contribute to disease progression and inhibition of antitumor immunity. There is a growing effort to leverage new technologies for in-depth analysis of the complexities within the tumor immune microenvironment. We validated a 17-plex, subcellular resolution, pan-immune cell panel to interrogate the immune cell types and their spatial organization in whole tissue sections.
Methods: We co-developed a 17-plex pan-immune cell immunofluorescence (IF) assay with RareCyte® utilizing the Orion™ platform to characterize immune cell populations and their functional states in formalin-fixed paraffin-embedded (FFPE) tissues from melanoma, breast, lung, prostate, and colorectal cancers. A single, 4-micron FFPE section was stained in one-step, using the intelliPATH™ FLX (Biocare Medical) with a cocktail of antibody-fluorophore conjugates targeting Granzyme B, Ki67, CD3e, CD20, CD4, CD163, CD8a, PDL1, NKp46, PD-1, LAG3, FOXP3, CD11b, CD138, SOX10, Cytokeratin, and Hoechst nuclear stain. Imaging was performed on the Orion™ platform, and data were analyzed using HALO® (Indica Labs).
Results: The assay enabled comprehensive spatial profiling of T, B, NK, and myeloid cell distributions within tumor and stromal compartments. Functional states of T cells (cytotoxic, regulatory, exhausted) and macrophage/B-cell niches were distinguished based on immune checkpoint and activation marker expression. Proximity analyses identified immune synapses and organized lymphoid structures, facilitating robust microenvironment characterization.
Conclusions: The Orion™ 17-plex pan-immune cell IF assay will enable comprehensive qualitative and quantitative analysis of the tumor microenvironment in clinical trial specimens. We anticipate that this approach will support the development of next-generation therapies by enabling detailed characterization of immune cell and tumor cell spatial biology across multiple cancer types.
利益披露 Disclosure
J. Kaur,
Navigate BioPharma Services, Inc. Employment.
X. Li,
Navigate BioPharma Services, Inc. Employment.
Y. Huang,
Novartis BioMedical Research Employment.
L. Kattenhorn,
Novartis BioMedical Research Employment.
N. Dakappagari,
Navigate BioPharma Services, Inc. Employment.
M. E. McLaughlin,
Novartis BioMedical Research Employment.
J. Bordeaux,
Navigate BioPharma Services, Inc. Employment.