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 4

Association of CD8+ T cell features within TN with ICI efficacy.

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Association of CD8+ T cell features within TN with ICI efficacy. (A) KM ...
(A) KM curves for PFS of ICI treatment stratified by Trm-like (CD39+CD103+) CD8+ TIL density in TN. (B) Forest plot showing HRs for PFS estimated by multivariable Cox proportional hazards regression models evaluating the association between ICI efficacy and the indicated biomarkers. Other covariates not shown here are provided in the supplemental material. (C) Correlation between exhaustion marker positivity in CD8+ TILs in TN (LAG-3 vs. CD103 [left]; TIM-3 vs. CD39 [right]) (N = 80). One case was excluded because CD8+ T cells were absent, precluding positivity calculation. Pairwise correlations were calculated using Spearman’s correlation analysis. (D) LAG-3 (left) and TIM-3 (right) positivity in CD8+ TILs in TN compared between the DCB (N = 30) and non-DCB (N = 48) groups. Two cases were excluded because DCB/non-DCB status could not be determined due to early censoring (<1 year) of PFS data. Another case in the non-DCB group was excluded because CD8+ T cells were absent in the tumor, precluding positivity calculation. (E) KM curves for PFS stratified by LAG-3 or TIM-3 positivity in Trm-like CD8+ TILs in TN. Data are presented in violin plots in D, with each dot representing 1 patient; P values were determined by Mann-Whitney U test. P values for the survival analyses were determined by log-rank test. Vertical bars on KM curves indicate censoring.