Abstract 3980: Exploring the diversity and dynamics of the macrophage compartment in non-small cell lung cancer

Abstract Introduction: Although immune checkpoint inhibitors (ICIs) has improved the survival rate for some cancer, EGFR mutant (MT) non-small cell lung cancer (NSCLC) patients exhibit a poor response to ICIs than those with EGFR wild-type (WT). EGFR-MT cells contribute to immunosuppressive tumor microenvironment (TME) by secreting inhibitory cytokines and recruiting pro-tumor immunogenic cells, thereby diminishing the efficacy of ICIs. Tumor associated macrophages (TAMs) represent a major component of the TME, playing complex roles in tumor progression and metastasis. While single-cell RNA (scRNA) analyses have been performed in various cell types, studies on macrophage (MØ) diversity between EGFR-WT and -MT NSCLCs are limited. Therefore, we aimed to reveal the presence of multifaceted MØ populations, using scRNA sequencing. Methods: Using the 10x Genomics, we generated scRNA expression data from 904, 174 cells in primary lung tissues of 95 patients: EGFR-WT (n=35), treatment-naïve EGFR-MT (n=55), EGFR-TKI post-treatment (n=5), and matched normal (n=22). We annotated five distinct cell lineages—lymphocytes, epithelial cells, endothelial cells, fibroblasts, and myeloid cells—using canonical marker gene expression. We focused on MØs (n=158, 640 cells) in myeloid, including markers such as APOE, FCGR1A, and MARCO. Results: We characterized 13 transcriptional states of MØs: Four clusters of alveolar MØs (AMs; PPARG+), two clusters of interstitial MØs (IMs; MERTK+, LYVE1+) and eight clusters of MØs expressing canonical markers (CD68+, C1QA+) with high monocyte derived MØ (MDM) signatures. To define significant clusters, we analyzed their proportions based on sample type, tumor stage, and recurrence status. MØ cluster compartment varied among normal, EGFR-WT, -MT, and -TKI samples. AMs were primarily found in normal tissues, while IMs and MDMs predominate in tumors. Some IM and MDM clusters showed opposing trends in proportion between EGFR-WT and -MT across tumor stages, with major recurrence-associated clusters differing between the two types. Next, we defined high proportion of four major TAM clusters identified in EGFR-WT, -MT, and -TKI, expressing the representative genes SPP1, TREM2, IL1B, and FOLR2, respectively. Gene ontology pathway analyses revealed the complexity of the TME. SPP1 TAMs, predominant in EGFR-MT, were associated with cellular stress response, while IL1B TAMs, mainly in EGFR-WT, were associated with response to external stimuli. FOLR2 TAMs were associated with chemoattraction and were prevalent in both EGFR-WT and -TKI. TREM2 TAMs, associated with lipid metabolism, were evenly distributed across all sample types. Conclusions: We characterized the heterogeneity of MØ compartments between EGFR-WT, -MT and -TKI samples. These results highlight the need for selective treatment strategies based on EGFR mutation and treatment status in NSCLCs for successful immunotherapy. Citation Format: Eun Ji LEE, You Won Lee, Minyeop Kim, Seung Yeon Oh, YoungJoon Park, Su-Jin Choi, JuHyeon Lee, Kyoung-Ho Pyo, Jae Hwan Kim, Jii Bum Lee, Min Hee Hong, Sun Min Lim, Mi Ran Yun, Byoung Chul Cho. Exploring the diversity and dynamics of the macrophage compartment in non-small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 3980.