Abstract 2664: Spatial and single-cell transcriptomics reveal molecular heterogeneity in pulmonary pleomorphic carcinoma

Abstract Background: Pulmonary pleomorphic carcinoma (PPC) is a rare and aggressive subtype of non-small cell lung cancer characterized by the presence of both epithelial and sarcomatoid components. We conducted multi-omics analyses on surgically resected specimens from nine primary PPC cases to explore characteristics of this tumor at molecular level. Materials and Methods: Whole-exome sequencing and RNA sequencing on bulk tumor specimens to identify mutational profiles. Spatial transcriptomics using the Digital Spatial Profiler was employed to analyze gene expression separately for epithelial and sarcomatoid tumor components. Single-cell RNA sequencing (scRNA-seq) was conducted on three cases to explore intratumoral heterogeneity and clonal evolution. Gene signature based on extracellular matrix (ECM) remodeling was developed and validated in public lung cancer datasets to assess its association with tumor aggressiveness and prognosis. Results: Mutational analyses revealed MET exon 14 skipping in two cases and an ALK-EML4 fusion in one case, while no EGFR mutations were observed. We found various differential expressed genes (DEGs) between components. Further analysis (GSEA and Transcriptional factor scoring) revealed there was a common enrichment trend observed among pathways related to MYC, such as those involved in cell proliferation, division, and the cell cycle. On the other hand, significant differences were observed in pathways related to cancer invasion and cell-cell interactions, such as extracellular matrix (ECM) remodeling and hypoxia. identified an atypical cell population bridging epithelial cells and fibroblasts, which we defined as sarcomatoid cells. The clusters defined as epithelial tumor cells and sarcomatoid cells showed upregulation of DEGs corresponding to the respective component identified through spatial transcriptomics. Trajectory analysis visualized tumor evolution, suggesting a transition from epithelial tumor cells to sarcomatoid cells. A consistent upregulation of seven ECM remodeling-related genes was observed in both tumor components compared to normal lung tissue. When applied to a GEO lung adenocarcinoma cohort, this gene signature was associated with more aggressive histological subtypes (P < 0.01). Survival analysis using TCGA demonstrated that high ECM remodeling signature scores correlated with significantly poorer prognosis (P < 0.01). Conclusion: This study provides a comprehensive molecular analysis of PPC, revealing both shared and distinct features between its components. The trajectory analysis supports the hypothesis of a common cellular origin for these components and highlights the transition to a more aggressive sarcomatoid phenotype. These findings provide new insights into the biology of PPC and identify potential therapeutic targets to address its intratumoral heterogeneity and clonal evolution. Citation Format: Atsushi Matsuoka, Kazuhiko Shien, Shuta Tomida, Hirofumi Inoue, Kazuya Hisamatsu, Ryota Fujiwara, Kousei Ishimura, Ryunosuke Fujii, Ryo Yoshichika, Tomoaki Higashihara, Naohiro Hayashi, Kazuhiro Okada, Fumiaki Mukohara, Mao Yoshikawa, Ken Suzawa, Hidetaka Yamamoto, Shinichi Toyooka. Spatial and single-cell transcriptomics reveal molecular heterogeneity in pulmonary pleomorphic carcinoma. [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 2664.