Subtype cluster analysis unveiled the correlation between m6Aand cuproptosis-related lncRNAs and the prognosis, immune microenvironment, and treatment sensitivity of esophageal cancer

Ming Zhang Yani Su Pengfei Wen Xiaolong Shao Peng Yang Peng An Wensen Jing Lin Liu Yang Zhi Mingyi Yang ^*

• Xi'an Honghui Hospital, Xi'an, China

Objective: Esophageal cancer (EC) is characterized by a high degree of malignancy and poor prognosis. N6-methyladenosine (m6A), a prominent post-transcriptional modification of mRNA in mammalian cells. Similarly, cuproptosis has garnered attention for its potential implications in cancer biology. This study seeks to elucidate the impact of m6A- and cuproptosis-related long non-coding RNAs (m6aCRLncs) on the prognosis of patients with EC.Methods: The EC transcriptional data and corresponding clinical information were retrieved from The Cancer Genome Atlas (TCGA) database. Data on 23 m6A regulators and 25 cuproptosis-related genes were sourced from the latest literature. Differentially expressed m6aCRLncs associated with EC prognosis were screened using the limma package in R and univariate Cox regression analysis. Subtype clustering was performed to classify EC patients, enabling the investigation of differences in clinical outcomes and immune microenvironment across patient clusters. A risk prognostic model was constructed using LASSO regression. Single-sample gene set enrichment analysis (ssGSEA), immune cell infiltration analysis, and immune checkpoint differential expression analysis were conducted. Drug sensitivity analysis was performed to identify potential therapeutic agents for EC. Finally, the mRNA expression levels of m6aCRLncs in EC cell lines were validated using reverse transcription quantitative polymerase chain reaction (RT-qPCR).Results: We developed a prognostic risk model based on five m6aCRLncs, namely ELF3-AS1, HNF1A-AS1, LINC00942, LINC01389, and MIR181A2HG, to predict survival outcomes and characterize the immune microenvironment in EC patients. Immune profiling further identified distinct immune cell populations and functional pathways associated with risk scores. Additionally, we identified key immune checkpoint-related genes with significant differential expression between risk groups, including TNFRSF14, TNFSF15, TNFRSF18, LGALS9, CD44, HHLA2, and CD40. Furthermore, nine candidate drugs with potential therapeutic efficacy in EC were identified: Bleomycin, Cisplatin, Cyclopamine, PLX4720, Erlotinib, Gefitinib, RO.3306, XMD8.85, and WH.4.023. Finally, RT-qPCR validation of the mRNA expression levels of m6aCRLncs in EC cell lines demonstrated that ELF3-AS1 expression was significantly upregulated in the EC cell lines KYSE-30 and KYSE-180 compared to normal esophageal epithelial cells.Conclusion: This study elucidates the role of m6aCRLncs in shaping the prognostic outcomes and immune microenvironment of EC. Furthermore, it identifies potential therapeutic agents with efficacy against EC.