PO.CL01.12 · 临床研究
用于空间解析免疫检查点分析的邻近连接-成像质谱流式细胞术平台的开发
Development of a proximity ligation-imaging mass cytometry platform for spatially resolved immune checkpoint analysis
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
引言：免疫检查点抑制剂（ICI）已经变革了癌症治疗，但持久的临床获益仍局限于一部分患者。当前的生物标志物，如PD-L1免疫组织化学（IHC），反映的是蛋白丰度而非功能性受体-配体结合，因此仅提供有限的预测价值。为解决这一问题，我们此前证明，使用邻近连接分析（PLA）绘制PD1-PD-L1相互作用图谱在预测非小细胞肺癌（NSCLC）免疫治疗应答方面优于PD-L1 IHC[1]。
方法：基于这些发现，我们开发了一种三重PLA，可检测PD1-PD-L1、PD1-PD-L2和CD8-MHC I相互作用，并将其与成像质谱流式细胞术（IMC）整合。该方法能够在详细免疫表型分析的同时对活跃免疫信号通路进行高多重（≈40个标志物）空间定量。该平台应用于作为生物学对照的人扁桃体组织，以及治疗前的NSCLC和三阴性乳腺癌（TNBC）活检组织，以绘制肿瘤微环境中检查点结合、抗原识别和T细胞功能状态的图谱。
结果：在扁桃体中，PD1-PD-L1和PD1-PD-L2相互作用主要观察到发生在CD8⁺ T细胞与滤泡B细胞之间，而额外的PD1-PD-L1相互作用发生在CD8⁺ T细胞与CD68⁺巨噬细胞之间。这些模式提示淋巴组织中PD-L1和PD-L2结合存在部分不同的细胞背景。与此一致，初步的肿瘤数据表明，与NSCLC相比，PD1-PD-L2相互作用在TNBC中相对罕见，提示PD-L2参与因组织类型和免疫环境而异。
结论：PLA-IMC提供了一个功能性、空间解析的平台，可直接在组织中筛查和定量活跃的免疫检查点信号。该方法有望识别功能性生物标志物，用于优化患者分层和绘制背景特异性检查点活性图谱，以指导实体瘤中合理的免疫治疗联合方案。
参考文献：[1] Lindberg A, Muhl L, Yu H, et al. In situ detection of programmed cell death protein 1 and programmed death ligand 1 interactions as a functional predictor for response to immune checkpoint inhibition in NSCLC. J Thorac Oncol. 2025; 20:625-640.
查看英文原文 English abstract
Introduction: Immune checkpoint inhibitors (ICIs) have transformed cancer therapy, yet durable clinical benefit remains limited to a subset of patients. Current biomarkers, such as PD-L1 immunohistochemistry (IHC), reflect protein abundance rather than functional receptor-ligand engagement and therefore provide only modest predictive value. To address this, we previously demonstrated that mapping PD1-PD-L1 interactions using a proximity ligation assay (PLA) outperforms PD-L1 IHC in predicting immunotherapy response in non-small cell lung cancer (NSCLC) [1].
Methods: Building on these findings, we developed a triplex PLA detecting PD1-PD-L1, PD1-PD-L2, and CD8-MHC I interactions and integrated it with Imaging Mass Cytometry (IMC). This approach enables high-plex (≈40 markers) spatial quantification of active immune signaling pathways alongside detailed immune phenotyping. The platform was applied to human tonsil tissue as a biological control and to pre-treatment NSCLC and triple-negative breast cancer (TNBC) biopsies to map checkpoint engagement, antigen recognition, and T-cell functional states within the tumor microenvironment.
Results: In tonsil, PD1-PD-L1 and PD1-PD-L2 interactions were observed primarily between CD8⁺ T cells and follicular B cells, while additional PD1-PD-L1 interactions occurred between CD8⁺ T cells and CD68⁺ macrophages. These patterns suggest partly distinct cellular contexts for PD-L1 and PD-L2 engagement in lymphoid tissue. Consistent with this, preliminary tumor data indicate that PD1-PD-L2 interactions are relatively rare in TNBC compared with NSCLC, suggesting that PD-L2 involvement varies across tissue types and immune environments.
Conclusion: PLA-IMC provides a functional, spatially resolved platform for screening and quantifying active immune checkpoint signaling directly in tissue. This approach holds promise to identify functional biomarkers for refined patient stratification and mapping of context-specific checkpoint activity to guide rational immunotherapy combinations in solid tumors.
References: [1] Lindberg A, Muhl L, Yu H, et al. In situ detection of programmed cell death protein 1 and programmed death ligand 1 interactions as a functional predictor for response to immune checkpoint inhibition in NSCLC. J Thorac Oncol. 2025; 20:625-640.
利益披露 Disclosure
G. Lessan Toussi, None..
L. Muhl, None..
A. Gorbunova, None.
A. Rayford,
Bergen Bio (BGB) Employment.
A. Lindberg, None..
N. Hekmati, None..
V. Thurfjell, None..
A. Schiza, None..
A. Zieba Wicher, None..
P. Micke, None..
C. Strell, None.