Abstract 94: Tumor RNA isoforms are associated with recurrence in resected non-small cell lung cancer

Abstract Introduction: Surgical resection remains a mainstay of treatment for individuals with non-small cell lung cancer (NSCLC). However, disease recurrence rates remain a major concern, standing at 5-19% for stage I disease, 11-27% for stage II disease, and 24-40% for stage III disease. This study focused on identifying tumor transcript variants that allow patients to be risk-stratified for recurrence/disease progression. Methods: Patients with stage I-IIIA NSCLC tumors (T1-2N0-2M0) who did not receive neoadjuvant therapy and had follow-up clinical data available were included. The primary outcome was event free survival (EFS), defined as time until recurrence or death. Tumor RNAseq data was interrogated for RNA isoforms at the splice variant (n=645 subjects) and fusion transcript (n=555 subjects) levels, identifying 225, 896 unique splice variants via FRASER and 1, 382 unique fusion transcripts via STAR-Fusion, respectively. Differential analyses were performed via edgeR based on nodal and recurrence status (i.e. N0 Non-recurrent, N0 Recurrent, N1-2 Non-recurrent, and N1-2 Recurrent) at the individual isoform level using mean feature importances of a supervised learning model (3 random forest ensemble) to rank isoforms on prognostic value. Aggregate analyses were also performed, assessing the relationship between recurrence and tumor fusion burden in two ways (sum of normalized counts and sum of unique transcripts). Results: From the intersection of top features and differential expression analysis comparing the N0 Non-recurrent group (n=325) to those that recurred/progressed (N0 Recurrent, N1-2 Non-recurrent, and N1-2 Recurrent; n=230) we identified differential utilization at multiple splice site loci, including at chr 7: 16, 861, 429 - 16, 861, 430 (RGA3 exon 6) that was associated with progression (q = 2.145·10¯ 7; log fold change = 0.374). Total normalized fusion transcript fragments (as FFPM) per subject pass Kruskal-Wallis test for differences between the four individual groups (p = 0.3126). Tumors with greater than 10 unique fusion transcript variants detected were associated with a superior 5-year EFS (HR = 0.59 (95%CI: 0.4-0.86), p = 0.0069). Conclusions: These data suggest multiple loci producing splice variants associated with recurrence exist, and a fusion burden >10 unique transcripts correlates with a superior EFS in patients with resected stage I-IIIA NSCLC, which may be reflective of increased neoantigen load and immune activation. Current efforts are focused on validation of these findings in the TracerX cohort and mass spectrometry. Citation Format: Kelly M. Cagin, Arsalan Khan, Savan Shah, Wara Naeem, Minha Ansari, Oluwamuyiwa W. Adebayo, Hita Moudgalya, Michael J. Liptay, Christopher W. Seder, Jeffrey A. Borgia. Tumor RNA isoforms are associated with recurrence in resected non-small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 94.