Spatial single-cell proteotyping reveals immunotherapy-resistant features within the complex tumor microenvironment of metastatic NSCLC
Kohsuke Isomoto, Koji Haratani, Takahiro Tsujikawa, Shuta Tomida, Yusuke Makutani, Masayuki Takeda, Kimio Yonesaka, Kaoru Tanaka, Tsutomu Iwasa, Kazuko Sakai, Kazuto Nishio, Akihiko Ito, Kazuhiko Nakagawa, Hidetoshi Hayashi
Kohsuke Isomoto, Koji Haratani, Takahiro Tsujikawa, Shuta Tomida, Yusuke Makutani, Masayuki Takeda, Kimio Yonesaka, Kaoru Tanaka, Tsutomu Iwasa, Kazuko Sakai, Kazuto Nishio, Akihiko Ito, Kazuhiko Nakagawa, Hidetoshi Hayashi
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Clinical Research and Public Health Clinical Research Immunology Oncology

Spatial single-cell proteotyping reveals immunotherapy-resistant features within the complex tumor microenvironment of metastatic NSCLC

Abstract

BACKGROUND Immune checkpoint inhibitors (ICIs) targeting the programmed cell death 1 axis have revolutionized metastatic non–small cell lung cancer (mNSCLC) treatment. However, disease progression remains a concern, and the role of the complex tumor microenvironment (TME) in treatment failure is not fully understood.METHODS In this biomarker study involving 103 patients with mNSCLC, including 81 patients who received ICI treatment, we evaluated the association between heterogeneous immune cell subsets and ICI efficacy through single-cell spatial profiling of pretreatment tumor tissue, using a 29-marker multiplex IHC platform built for in-depth dissection of the TME.RESULTS Among various types of intratumoral lymphocytes, including Th1, Treg, and NK cells, only CD8+ T cells (tumor-infiltrating lymphocytes [TILs]) were associated with ICI efficacy. Computational tissue segmentation underscored the importance of direct physical interactions between CD8+ TILs and cancer cells for ICI efficacy. TIL phenotyping identified CD39/CD103/Ki-67 positivity as a hallmark of exhausted yet functional tumor-reactive CD8+ TILs. Immunosuppressive tumor-associated macrophages (TAMs) and cancer-associated fibroblasts were independent unfavorable adversaries. High CD73 expression on cancer cells was suggested to confer tolerance to ICI in EGFR/ALK-oncogene+ NSCLC, potentially through M2-TAM accumulation and aberrant angiogenesis.CONCLUSION Our study delineates the clinical relevance of heterogeneous immune cell subsets in ICI-treated mNSCLC, aiding the development of targeted therapeutic strategies.FUNDING Osaka Cancer Society, KANAE Foundation for the Promotion of Medical Science, SGH Foundation, and YOKOYAMA Foundation for Clinical Pharmacology.

Authors

Kohsuke Isomoto, Koji Haratani, Takahiro Tsujikawa, Shuta Tomida, Yusuke Makutani, Masayuki Takeda, Kimio Yonesaka, Kaoru Tanaka, Tsutomu Iwasa, Kazuko Sakai, Kazuto Nishio, Akihiko Ito, Kazuhiko Nakagawa, Hidetoshi Hayashi

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Figure 7

Association of intratumoral immune–stromal contexture with ICI efficacy.

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Association of intratumoral immune–stromal contexture with ICI efficacy....
(A) Unsupervised hierarchical clustering results of ICI cohort according to Ki-67+ Trm-like CD8+ TIL density, CD206+ TAM density, and FAP positivity in stromal cells are visualized as a heatmap. Each tile represents the z score for key TME features (yellow, highest; black, median; blue, lowest). Additional mIHC-derived TME features are shown in a second heatmap (red, highest; black, median; green, lowest). Clinicopathologic characteristics and PFS for each patient are shown below the heatmaps. (B) Ki-67+ Trm-like CD8+ TIL density, CD206+ M2-TAM density, and FAP positivity in stromal cells compared between patients in clusters 2 and 6 (N = 11) and those in other clusters (N = 70). (C) The z scores for Ki-67+ Trm-like CD8+ TIL density (top), CD206+ M2-TAM density (middle), and FAP positivity in stromal cells (bottom) across all 10 clusters. (D) KM curves for PFS of ICI treatment comparing favorable (clusters 2 and 6) and unfavorable (other clusters) TME. Data are presented in violin plots in C. Each dot represents 1 patient in B and C. Data are presented as mean ± SEM in dot plots. P values for dot plots and survival analyses were determined by Mann-Whitney U test and log-rank test, respectively. Vertical bars on KM curves indicate censoring. Mono, monotherapy; chemo, chemotherapy; Adeno, adenocarcinoma; Sq, squamous cell carcinoma; Adeno-sq, adenosquamous.