Single‐cell transcriptome sequencing analysis reveals intra‐tumor heterogeneity in esophageal squamous cell carcinoma

Abstract Esophageal squamous cell carcinoma (ESCC) is a prevalent malignant tumor of the digestive system that poses a significant threat to human life and health. It is crucial to thoroughly investigate the mechanisms of esophageal carcinogenesis and identify potential key molecular events in its carcinogenesis. Single‐cell transcriptome sequencing is an emerging technology that has gained prominence in recent years for studying molecular mechanisms, which may help to further explore the underlying mechanisms of the ESCC tumor microenvironment in depth. The single‐cell dataset was obtained from GSE160269 in the Gene Expression Omnibus database, including 60 tumor samples and four paracancer samples. The single‐cell data underwent dimensional reduction clustering analysis to identify clusters and annotate expression profiles. Subcluster analysis was conducted for each cellular taxon. Copy number variation analysis of tumor cell subpopulations was performed to primarily identify malignant cells within them. A proposed chronological analysis was performed to obtain the process of cell differentiation. In addition, cell communication, transcription factor analysis, and tumor pathway analysis were also performed. Relevant risk models and key genes were established by univariate COX regression and LASSO analysis. The key genes obtained from the screen were subjected to appropriate silencing and cellular assays, including CCK‐8, 5‐ethynyl‐2′‐deoxyuridine, colony formation, and western blot. Single‐cell analysis revealed that normal samples contained a large number of fibroblasts, T cells, and B cells, with fewer other cell types, whereas tumor samples exhibited a relatively balanced distribution of cell types. Subclassification analysis of immune cells, fibroblasts, endothelial cells, and epithelial cells revealed their specific spatial characteristics. The prognostic risk model, we constructed successfully, achieved accurate prognostic stratification for ESCC patients. The screened key gene, UPF3A, was found to be significantly associated with the development of ESCC by cellular assays. This process might be linked to the phosphorylation of ERK and P38. Single‐cell transcriptome analysis successfully revealed the distribution of cell types and major expressed factors in ESCC patients, which could facilitate future in‐depth studies on the therapeutic mechanisms of ESCC.