ESTRO 2025 - Abstract Book

S3996

Radiobiology - Tumour radiobiology

ESTRO 2025

Material/Methods: Four MIBC cell lines (HT1376, T24, J82, UMUC3) were exposed to a clinical radiotherapy protocol (2.75 Gy daily, 5 days per week, totalling 55 Gy). Cells were allowed to recover after treatment to generate radioresistant populations. Radioresistance was confirmed through clonogenic assays. RNAseq was used to identify differentially expressed genes (DEGs) in radioresistant cells. DEGs consistently significant (fold change ≥2 or ≤-2 and p adj <0.05) across all models were validated for in vitro stratification using hierarchical, K-means, and PCA clustering. DEGs were retrospectively validated in two clinical cohorts (e.g., BCON [n=151] and GemX [n=184]) and three publically available non-radiotherapy cohorts (e.g., GSE [n=831], IMVigor210 [n=261] and TCGA [n=407]) using Cox proportional hazards models. Clonogenic assays, seen in Figure 1 , confirmed that cells surviving radiotherapy acquire radioresistance for HT1376 (p<0.001), J82 (p<0.001) and T24 (p<0.001). Pathway analysis highlighted extracellular matrix (ECM) remodelling, Rho GTPase signalling, and mTOR pathway inhibition as mechanisms supporting radioresistance in all models. 19,666 DEGs were identified, of which 29 were identified in all radioresistant lines and selected as biomarker candidates (Figure 2) . Seven of the candidates were retrospectively validated in clinical cohorts as potential biomarkers: CYF1P2 ([IMVigor210, HR 0.81, p=0.03]), DEPTOR (TCGA, HR=1.15, p=0.04), MAGEA11 (IMVigor210, HR=1.16, p=0.03), MYL9 ([GSE13507, HR=0.7, p=0.007]), PPP1R9A (GSE13507, HR=1.5, p=0.001), PSG6 (GSE5287, HR=3.2, p=0.002), and ZSWIM5 (GemX, HR=1.7, p=0.02). Five of the genes ( MYL9 , PPP1R9A , CYF1P2 , PSG6 ) were associated with ECM processes. Conclusion: In this study, 29 genes were consistently associated with in vitro MIBC radioresistance . Seven genes were significantly prognostic in clinical cohorts and should be further investigated as radiosensitivity biomarkers. Some DEGs in radioresistant cells were associated with ECM-related pathways, suggesting ECM remodelling plays a role in radioresistance. Future work should aim to use the identified genes to establish and validate a radiosensitivity signature, which could improve patient stratification and MIBC treatment outcomes. Results: