Comprehensive Analysis of N6-Methyladenosine (m6A) RNA Methylation Regulators in Soft Tissue Leiomyosarcoma

Abstract

N6-methyladenosine (m6A), a widespread RNA modification, plays a vital role in various biological processes, including carcinogenesis, tumor progression, and immune regulation. We conducted this study to investigate the relationship between m6A regulators, such as METTL3, METTL14, WTAP, FTO, ALKBH5, and YTHDF1-3, and their association with c-Myc and programmed death ligand 1 (PD-L1) expression in leiomyosarcoma (LMS). The expression of these epitranscriptome regulator genes was evaluated using the next-generation sequencing data of 53 patients with LMS obtained from an online public database. We next determined the relationship between m6A regulators and c-Myc and CD274 (PD-L1) mRNA expression in an LMS cell line. We also performed immunohistochemical staining of 69 LMS cases. Immunohistochemical staining showed that cases with higher expression of METTL3, METTL14, ALKBH5, FTO, and WTAP exhibited higher c-Myc expression, and cases with higher expression of ALKBH5, YTHDF2, and WTAP exhibited higher mitotic activity. Gene set enrichment analysis revealed that METTL3, METTL14, and FTO knockdown significantly suppressed c-Myc target gene expression. Knockdown of METTL3, ALKBH5, YTHDF1, WTAP, and FTO in LMS cell lines reduced cell proliferation. These results suggest the relationship between m6A modifications and c-Myc-driven oncogenesis. Moreover, knockdown of YTHDF2 inhibited interferon gamma-induced PD-L1 expression, suggesting its role in immune evasion through PD-L1 regulation. Multivariate Cox proportional hazards analysis revealed that lower YTHDF2 expression and higher WTAP expression are unfavorable prognostic factors. These findings provide potential therapeutic targets for LMS, particularly in combination with immune checkpoint inhibitors. Further investigation into the molecular mechanisms of m6A-mediated regulation of PD-L1 and c-Myc expression is required to develop more effective treatments for LMS.

Keywords: N6-methyladenosine; RNA modification; carcinogenesis; immune regulation; leiomyosarcoma; tumor progression.