CMTR2 deficiency defines a novel RNA splicing-driven subtype of lung adenocarcinoma with therapeutic vulnerabilities

Background: RNA splicing is frequently perturbed in cancer, yet upstream genomic drivers and tractable vulnerabilities remain incompletely defined in lung adenocarcinoma (LUAD). CMTR2 catalyzes 2′-O-methylation at the mRNA cap (Cap2), a modification implicated in mRNA maturation. The oncologic significance of CMTR2 alterations and their impact on splicing have not been systematically characterized. We performed comprehensive splicing analysis to identify novel therapeutic targets in LUAD. Methods: We analyzed 1,017 primary lung cancer specimens with integrated DNA- and RNA-sequencing to map alternative splicing (AS) events (rMATS) to identify alternative splicing events. t-SNE visualization revealed distinct splicing patterns. CMTR2 variants were functionally profiled using CRISPR knockout models. Splicing changes were validated by orthogonal assays, including long-read direct RNA sequencing. Physical links between CMTR2 and the spliceosome were assessed by proteomics/Co-IP. Therapeutic dependencies were tested using the RBM39-degrader splicing modulator indisulam in vitro and in vivo, and by evaluating response to anti-PD-1 immune checkpoint blockade (ICB) and combination regimens in immunocompetent models. Results: Genomic profiling identified 3.8 % of LUAD harboring truncating or functionally impaired CMTR2 alterations, which associated with a distinct AS signature across multiple event classes. CMTR2 physically interacted with spliceosomal components; these interactions were compromised by truncating variants or loss of CMTR2 , coinciding with widespread shifts in exon usage and intron retention detected by both short- and long-read sequencing. CMTR2 -deficient cells exhibited heightened sensitivity to splicing modulation with indisulam, producing robust anti-tumor effects in xenograft models. In immunocompetent settings, CMTR2 loss increased tumor susceptibility to ICB; combining indisulam with anti-PD-1 further enhanced tumor control. Together, these data establish a mechanistic link between cap modification, spliceosome engagement, and therapeutic vulnerability. Conclusions: CMTR2 alterations constitute an upstream genomic driver of splicing dysregulation in LUAD and unveil an exploitable liability to splicing modulation and ICB therapy. Cap-dependent RNA maturation and spliceosome coupling represent a convergent axis of vulnerability with immediate translational potential for patient stratification and combination strategies. Clinical Relevance: CMTR2 status may serve as a biomarker to identify patients likely to benefit from splicing modulators and/or ICB, and provides a rationale for clinical exploration of CMTR2-targeted combinations in lung cancer.