Dissecting the single-cell transcriptome network underlying esophagus non-malignant tissues and esophageal squamous cell carcinoma

Abstract

Background: Esophageal squamous cell carcinoma (ESCC) is among the most prevalent causes of cancer-related death in adults. Tumor microenvironment (TME) has been associated with therapeutic failure and lethal outcomes for patients. However, published reports on the heterogeneity and TME in ESCC are scanty.

Methods: Five tumor samples and five corresponding non-malignant samples were subjected to scRNA-seq analysis. Bulk RNA sequencing data were retrieved in publicly available databases.

Findings: From the scRNA-seq data, a total of 128,688 cells were enrolled for subsequent analyses. Gene expression and CNV status exhibited high heterogeneity of tumor cells. We further identified a list of tumor-specific genes and four malignant signatures, which are potential new markers for ESCC. Metabolic analysis revealed that energy supply-related pathways are pivotal in cancer metabolic reprogramming. Moreover, significant differences were found in stromal and immune cells between the esophagus normal and tumor tissues, which promoted carcinogenesis at both cellular and molecular levels in ESCC. Immune checkpoints, regarded as potential targets for immunotherapy in ESCC were significantly highly expressed in ESCC, including LAG3 and HAVCR2. Eventually, we constructed a cell-to-cell communication atlas based on cancer cells and immune cells and performed the flow cytometry, qRT-PCR, immunofluorescence, and immunohistochemistry analyses to validate the results.

Interpretation: This study demonstrates a widespread reprogramming across multiple cellular elements within the TME in ESCC, particularly in transcriptional states, cellular functions, and cell-to-cell interactions. The findings offer an insight into the exploration of TME and heterogeneity in the ESCC and provide new therapeutic targets for its clinical management in the future.

Funding: The work was supported by the Shanghai Pujiang Program (2020PJD009) and Research Development Fund of Zhongshan Hospital, Fudan University (2019ZSFZ002 and 2019ZSFZ19).

Keywords: Esophageal squamous cell carcinoma; Heterogeneity; Immunotherapy; Single-cell RNA-seq; Tumor microenvironment.