PO.TB10.08 · 肿瘤生物学
空间和血管微环境特征预测胶质母细胞瘤中肿瘤浸润淋巴细胞的扩增
Spatial and vascular microenvironmental features predict tumor-infiltrating lymphocyte expansion in glioblastoma
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摘要 Abstract
中文摘要
引言/理论依据：肿瘤浸润淋巴细胞（TIL）的过继转移已在多种实体瘤中显示出有意义的临床活性，但其在胶质母细胞瘤中的应用受到深度免疫功能障碍、稀疏的T细胞浸润以及限制肿瘤反应性淋巴细胞体外增殖的空间受限免疫微龛的挑战。更深入地理解区分能够产生TIL培养物的肿瘤与失败肿瘤的空间、细胞和血管特征，可能识别成功扩增的微环境决定因素，并指导患者选择或微环境修饰策略，用于未来基于TIL的治疗。
方法：在40例肿瘤中尝试TIL扩增，其中24例（60%）在优化的快速扩增方案（REP）中获得成功产量。通过光谱流式细胞术和TCRseq对扩增产物进行表征。对TIL阳性（TIL⁺；n = 5）和TIL阴性（TIL⁻；n = 6）肿瘤的一个子集进行了bulk RNA和TCRseq、单细胞RNAseq、CODEX多重空间蛋白质组学以及Xenium原位空间转录组学分析，以界定免疫细胞状态、空间组织的微龛、受体-配体通讯网络和血管结构。
结果：扩增的TIL从REP前到REP增殖了100-1000倍，并富集CD4⁺细胞，CD4⁺和CD8⁺区室均以效应记忆表型为主，且含有极少的初始或调节性亚群。尽管肿瘤中总体T细胞浸润较低（<所有细胞的3%），但TIL⁺肿瘤含有更多干细胞样TCF7⁺ CD8⁺ T细胞，且HAVCR2和TOX的表达较低，与终末耗竭减少一致。空间分析显示，TIL⁺肿瘤中CD31⁺内皮密度高出2.3倍，且T细胞-内皮距离显著缩短（p < 0.05），表明血管微龛更易接近。相比之下，TIL⁻肿瘤表现出强1.6倍的抑制性TAM-小胶质细胞信号传导，而TIL⁺区域则富集T细胞-内皮和TAM-巨噬细胞相互作用，反映出更为许可的免疫结构。
结论：我们的数据强调血管可及性、免疫空间组织和抑制性髓系信号传导减少是胶质母细胞瘤中TIL成功扩增的关键决定因素。空间转录组学、多重蛋白质组学和单细胞测序的整合识别出与TIL增殖相关的微环境标志，并提示调控血管或髓系微龛可能提高基于TIL的治疗在胶质母细胞瘤中的可行性和有效性。这些见解支持微环境引导的患者选择，并为开发旨在重塑胶质母细胞瘤肿瘤微环境以改善对基于细胞的免疫治疗反应性的联合方法提供依据。
查看英文原文 English abstract
Introduction/Rationale: Adoptive transfer of tumor-infiltrating lymphocytes (TILs) has demonstrated meaningful clinical activity in several solid tumors, yet its application in glioblastoma is challenged by profound immune dysfunction, sparse T-cell infiltration, and spatially restricted immune niches that constrain the outgrowth of tumor-reactive lymphocytes ex vivo. A deeper understanding of the spatial, cellular, and vascular features that distinguish tumors capable of yielding TIL cultures from those that fail may identify microenvironmental determinants of successful expansion and guide patient selection or microenvironment-modifying strategies for future TIL-based therapies.
Methods: TIL expansion was attempted in 40 tumors, 24 (60%) yielded successful yield in an optimized rapid expansion protocol (REP). Expanded products were characterized by spectral flow cytometry and TCRseq. A subset of TIL-positive (TIL⁺; n = 5) and TIL-negative (TIL⁻; n = 6) tumors underwent bulk RNA and TCRseq, single-cell RNAseq, CODEX multiplex spatial proteomics, and Xenium in situ spatial transcriptomics to define immune cell states, spatially organized niches, receptor-ligand communication networks, and vascular architecture.
Results: Expanded TILs proliferated 100-1000-fold from pre-REP to REP and were enriched for CD4⁺ cells, with both CD4⁺ and CD8⁺ compartments dominated by effector-memory phenotypes and containing minimal naïve or regulatory subsets. Although overall T-cell infiltration in tumors was low (<3% of all cells), TIL⁺ tumors contained more stem-like TCF7⁺ CD8⁺ T cells and exhibited lower expression of HAVCR2 and TOX, consistent with reduced terminal exhaustion. Spatial analysis demonstrated 2.3-fold higher CD31⁺ endothelial density and significantly shorter T cell-endothelium distances in TIL⁺ tumors (p < 0.05), indicating more accessible vascular niches. In contrast, TIL⁻ tumors exhibited 1.6-fold stronger inhibitory TAM-microglia signaling, whereas TIL⁺ regions were enriched for T cell-endothelial and TAM-macrophage interactions, reflecting a more permissive immune architecture.
Conclusion: Our data highlight vascular accessibility, immune spatial organization, and reduced inhibitory myeloid signaling as key determinants of successful TIL expansion in glioblastoma. The integration of spatial transcriptomics, multiplex proteomics, and single-cell sequencing identifies microenvironmental hallmarks associated with TIL outgrowth and suggests that modulating vascular or myeloid niches may enhance the feasibility and effectiveness of TIL-based therapies in glioblastoma. These insights support microenvironment-guided patient selection and inform the development of combination approaches aimed at reconditioning the glioblastoma tumor microenvironment to improve responsiveness to cell-based immunotherapy.
利益披露 Disclosure
J. Jepson, None..
K. Van Batavia, None.