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Background <p>Platinum-based chemotherapy resistance, particularly to carboplatin, is a major challenge in the treatment of non-small cell lung cancer (NSCLC). N6-methyladenosine (m6A) methylation, which regulates gene and protein expression by influencing numerous aspects of mRNA metabolism, has been implicated in NSCLC pathogenesis, but its role in treatment resistance remains unclear.</p> Methods <p>We performed an innovative integrated analysis by combining genome-wide association study (GWAS) summary statistics, methylation quantitative trait loci (mQTL) data, and single-cell RNA sequencing (scRNA-seq) data from NSCLC patients. First, a transcriptome-wide association study (TWAS) using FUSION analysis was conducted to identify associations between m6A methylation sites, genes, and NSCLC by integrating m6A methylation profiles with GWAS summary statistics. Subsequently, scRNA-seq data were used to characterize NSCLC epithelial cell subpopulations and dynamically delineate the transition from treatment-sensitive to treatment-resistant states. Finally, a cross-omics correlation network was constructed by integrating the m6A and scRNA-seq datasets to identify associations between dynamic m6A modifications and carboplatin resistance.</p> Results <p>Our integrated analysis identified 6 m6A sites and 9 genes significantly associated with NSCLC. ScRNA-seq analysis revealed a significant increase in the proportion of epithelial cells in the carboplatin-resistant group. Marker genes of these carboplatin-resistant epithelial cells were enriched in pathways related to glutathione metabolism, ferroptosis, and endoplasmic reticulum protein processing. The RAS oncogene family member RAB7A was identified as a critical driver of treatment resistance. MeRIP-qPCR confirmed that RAB7A mRNA carries m6A modifications, and RIP-qPCR demonstrated direct binding of the m6A reader IGF2BP2 to RAB7A transcripts. Functional assays showed that RAB7A overexpression significantly increased intracellular glutathione levels and reduced lipid ROS, indicating enhanced antioxidant capacity and ferroptosis suppression. RAB7A expression was significantly upregulated in NSCLC patients with a poor response to neoadjuvant chemotherapy. Functional validation in vitro confirmed that overexpression of RAB7A conferred carboplatin resistance in NSCLC cells.</p> Conclusions <p>This study establishes a novel paradigm in which m6A-mediated gene regulation is a key contributor to chemoresistance. Furthermore, we identify RAB7A as a functionally relevant gene associated with carboplatin resistance, warranting further investigation as a candidate therapeutic target.</p>

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Integration of genome-wide association study and single-cell transcriptome analysis reveals the role of N6-methyladenosine in treatment resistance in lung cancer

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2026年8月4日

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  • Huikai Miao,
  • Chunmei Xu,
  • Dongni Chen,
  • Zhesheng Wen,
  • Jiajun Du,
  • Youfang Chen

摘要

Background

Platinum-based chemotherapy resistance, particularly to carboplatin, is a major challenge in the treatment of non-small cell lung cancer (NSCLC). N6-methyladenosine (m6A) methylation, which regulates gene and protein expression by influencing numerous aspects of mRNA metabolism, has been implicated in NSCLC pathogenesis, but its role in treatment resistance remains unclear.

Methods

We performed an innovative integrated analysis by combining genome-wide association study (GWAS) summary statistics, methylation quantitative trait loci (mQTL) data, and single-cell RNA sequencing (scRNA-seq) data from NSCLC patients. First, a transcriptome-wide association study (TWAS) using FUSION analysis was conducted to identify associations between m6A methylation sites, genes, and NSCLC by integrating m6A methylation profiles with GWAS summary statistics. Subsequently, scRNA-seq data were used to characterize NSCLC epithelial cell subpopulations and dynamically delineate the transition from treatment-sensitive to treatment-resistant states. Finally, a cross-omics correlation network was constructed by integrating the m6A and scRNA-seq datasets to identify associations between dynamic m6A modifications and carboplatin resistance.

Results

Our integrated analysis identified 6 m6A sites and 9 genes significantly associated with NSCLC. ScRNA-seq analysis revealed a significant increase in the proportion of epithelial cells in the carboplatin-resistant group. Marker genes of these carboplatin-resistant epithelial cells were enriched in pathways related to glutathione metabolism, ferroptosis, and endoplasmic reticulum protein processing. The RAS oncogene family member RAB7A was identified as a critical driver of treatment resistance. MeRIP-qPCR confirmed that RAB7A mRNA carries m6A modifications, and RIP-qPCR demonstrated direct binding of the m6A reader IGF2BP2 to RAB7A transcripts. Functional assays showed that RAB7A overexpression significantly increased intracellular glutathione levels and reduced lipid ROS, indicating enhanced antioxidant capacity and ferroptosis suppression. RAB7A expression was significantly upregulated in NSCLC patients with a poor response to neoadjuvant chemotherapy. Functional validation in vitro confirmed that overexpression of RAB7A conferred carboplatin resistance in NSCLC cells.

Conclusions

This study establishes a novel paradigm in which m6A-mediated gene regulation is a key contributor to chemoresistance. Furthermore, we identify RAB7A as a functionally relevant gene associated with carboplatin resistance, warranting further investigation as a candidate therapeutic target.