Abstract 5126: Time-series single-cell RNA sequencing uncovers PLAU enhancing early lung metastatic seeding of esophageal squamous cell carcinoma

Abstract Background: Metastasis continues to be the primary cause of cancer death even with recent advancements in cancer treatment. The initiating ability of metastatic growth bottlenecked in cancer development and could be a promising target. We have successfully captured the distinctive features of early metastatic cells through time-series scRNA-seq using the pulmonary metastatic model. Methods: To characterize the early metastatic tumor microenvironment (TME), we looked into the highly expressed genes encoding the secreted protein in early metastasis performing GO and KEGG pathway analysis. Clinical implication of PLAU overexpression was studied in ESCC patients. Multiplex immunohistochemistry staining of PLAU and previously identified metastatic initiating genes in ESCC patient samples were completed. Functional assays, including cell migration, invasion, wound healing, and foci formation were undertaken to evaluate the metastatic ability. PLAU knockdown ESCC cells were injected intravenously and subsequent in vivo lung imaging was obtained. Results: Through GO and KEGG pathway analysis of the single-cell profiling, the genes highly expressed in early metastatic lung exhibited a significant enrichment in processes related to coagulation and fibrinolysis (F3, THBS1, PLAU, SEPRINE1 and ANXA2). Among these genes, PLAU increased greatly in 6 hours and was maintained at a relatively high level during the early metastatic phase. We further explored the implications of PLAU. The analysis of the TCGA database reveals that PLAU is significantly upregulated in tumor tissues compared to non-tumor tissues .Multiplex immunohistochemistry staining of both ESCC cell lines and clinical samples demonstrated co-expression of PLAU and the previously identified metastatic initiating signature CD44 and TACSTD2. Knockdown of PLAU greatly impaired the ability of migration, invasion, wound healing and foci formation (p<0.05). In addition, the knockdown of PLAU led to significantly fewer surviving cancer cells in mice lungs in the early phase of metastasis (p<0.05). Conclusions: In all, the finding indicated that PLAU co-expressed with previously identified metastatic initiating genes and increased early dissemination of ESCC lung metastasis. Targeting PLAU could offer a promising approach to limit early metastatic progression in ESCC. Citation Format: Yingchen Lyu, Ching Ngar Wong, Hongyu Zhou, Songna Wang, Xinyuan Guan. Time-series single-cell RNA sequencing uncovers PLAU enhancing early lung metastatic seeding of esophageal squamous cell 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 5126.