Integrative epitranscriptomic and transcriptomic characterization in human colorectal cancer

Abstract

Introduction: Despite significant progress in understanding the molecular basis of colorectal cancer (CRC), the precise mechanisms driving its development and progression remain poorly defined. This gap limits the identification of novel therapeutic targets and the development of effective early detection methods. N6-methyladenosine (m6A) has merged as a key role in CRC pathogenesis. But research on mRNA methylation in CRC remains sparse.

Objectives: We aimed to address the roles of m6A modifications in CRC and to understand how mRNA methylation contributes to CRC development and progression.

Methods: We obtained a comprehensive mapping of altered m6 peaks within mRNAs using Methylated RNA Immunoprecipitation Sequencing (MeRIP-seq) from 34 CRC samples and 34 adjacent normal tissue samples and dissect molecular mechanisms of identified key CRC gene(s). By integrating these data, we identified differentially expressed mRNAs with altered m6A levels. Further analysis identified 119 overlapping peaks with both significantly altered RNA methylation and expression levels. Finally, we analyzed the relationship between m6A-regulated gene expression and Immune infiltrates using CRC patient-derived PBMC samples. Additionally, we established a subcutaneous xenograft tumor model to explore the role of SIM2 in CRC progression.

Results: Our comprehensive analysis of 68 fresh-frozen CRC samples identified 119 overlapping m6A peaks across 77 genes, classified based on m6A and mRNA expression changes. Survival analyses revealed a signature of m6A-modified genes with prognostic potential. These methylated genes were significant associated with immune cell profiles in the tumor microenvironment and immune checkpoints regulation, highlighting m6A as a promising immunotherapeutic target for CRC. SIM2 emerged as a key candidate, exhibiting elevated m6A and RNA expression levels in tumors and Macrophage M2 cells. NTMT1 (a writer) and YTHDF1 (a reader) were identified as pivotal regulators of m6A modifications in the 5'-UTR of SIM2 mRNA, emphasizing their role in CRC progression. Silencing methylated SIM2 significantly suppressed tumor growth, suggesting its potential as a therapeutic target.

Conclusion: Our integrative analyses provide a valuable resource for unraveling the molecular landscape of CRC. These finding offer new insight for advancing diagnostic precision and refining prognostic and therapeutic strategies for CRC.

Keywords: Colorectal cancer; Diagnosis; Immunotherapy; Single-minded 2, NTMT1; Survival outcome; m6A modification.