High-resolution detection of RPS24 microexon variations reveals novel splicing patterns in response to KRAS-targeted therapy in lung adenocarcinoma.

Abstract

Alternative splicing (AS) dysregulation is increasingly recognized as a critical factor in cancer progression and drug response. However, precisely detecting and characterizing complex splicing events, particularly those involving microexons, remains technically challenging. Ribosomal Protein 24 (RPS24), which contains three microexons (3, 18, and 22 bp), serves as an ideal model for studying complex AS regulation in cancer. We developed a high-resolution detection method for RPS24 microexon variations and investigate their relationship with KRAS proto-oncogene, GTPase (KRAS) inhibition in lung adenocarcinoma (LUAD) to identify potential biomarkers for KRAS-targeted inhibitors. We established an integrated methodological approach combining RNA-seq analysis with fragment analysis to detect RPS24 AS patterns. Using this method, we analyzed RPS24 AS across a panel of lung cancer cell lines and examined AS changes in KRAS-mutant cell lines following treatment with KRAS inhibitors. Our method successfully characterized distinct RPS24 AS isoform compositions across lung cancer cell lines, demonstrating high accuracy in detecting 3 bp variations. In KRASmutant cell lines, we observed a consistent upregulation of the 3 bp-containing isoform following KRAS inhibition, indicating a specific correlation with treatment response. This study provides a robust methodology for analyzing complex AS events and supports the RPS24 3 bp-containing isoform as a potential biomarker for KRAS inhibitor response in LUAD. These findings offer new insights into the molecular mechanisms of KRAS inhibitor therapy and strategies for monitoring treatment response.