# Single-Cell Analysis Reveals Epithelial Heterogeneity and Tumor Microenvironment Characteristics During the Malignant Progression of Colorectal Cancer

## TL;DR

This study uses single-cell sequencing to explore epithelial cell diversity and tumor microenvironment changes in colorectal cancer progression.

## Contribution

Identifies 11 epithelial cell subtypes and their roles in CRC progression, highlighting KCNMA1+ and MKI67+ cells as key indicators.

## Key findings

- Epithelial cells were divided into 11 subgroups marked by genes like MKI67 and KCNMA1.
- Downregulation of KCNMA1 and upregulation of MKI67 correlate with poor CRC prognosis.
- Cell–cell communication suggests bidirectional regulation between epithelial and fibroblast subsets.

## Abstract

Background/Objectives: To mine single-cell sequencing data for colorectal cancer (CRC), identify CRC epithelial cell subtypes, and explore the heterogeneity of epithelial cells and their impact on the tumor microenvironment (TME). Methods: The GSE201348 dataset, including normal, colorectal adenoma, high-grade colorectal intraepithelial neoplasia, and CRC tumor tissue samples, was downloaded from the Gene Expression Omnibus. The Seurat package of R software was used for data quality control, data integration, normalization, and clustering. The Feature Plot and the Recode function were executed to annotate and group the epithelial cells. Finally, genetic differences, copy number variant heterogeneity, pseudotime, cell–cell communication, and Gene Set Variation Analysis (GSVA) were further conducted. Results: In total, 26,335 gene matrices from 263,872 cells were obtained for subsequent analyses. Four cell clusters, including immune cells, fibroblasts, endothelial cells, and epithelial cells, were identified. Epithelial cells were further divided into 11 subgroups characterized by MKI67, SLC27A6, PLCE1, NKD1, KCNMA1, GDA, CLCA4, BEST4, LRMP, ACTG2, and ASPM. GSVA enrichment analysis suggested a role of the “P53 pathway,” “Wnt–β-catenin signaling,” and “MYC targets V1” pathways in epithelial cells during the malignant progression of tumors. Survival analysis indicated that downregulation of KCNMA1 and upregulation of MKI67 were associated with poor prognosis. Cell–cell communication analysis suggested a bidirectional regulatory role between epithelial and fibroblast subsets. Conclusions: This study analyzed the gene expression characteristics of 11 types of epithelial cells during the malignant progression of CRC. KCNMA1+ and MKI67+ epithelial subpopulations are important indicators for the malignant progression of CRC.