ESTRO 2025 - Abstract Book

S3903

Radiobiology - Microenvironment

ESTRO 2025

3914

Proffered Paper Fractionated radiation alters the extracellular matrix produced by muscle-invasive bladder cancer cells Luisa Vanesa Biolatti 1 , Conrado Guerrero Quiles 1 , S. Fahy 1 , M. Bartak 1 , Julia Gonzalez 2 , Emilia Powel 3 , Rachel Reed 1 , Kimberley Reeves 1 , Alex Baker 1 , Peter Hoskin 1 , Catharine M. West 1 , Ananya Choudhury 1 1 Division of Cancer Sciences, The University of Manchester, Manchester, United Kingdom. 2 Immunity and Cancer Unit, Institute Curie, Paris, France. 3 Cancer Research UK Manchester Institute, The University of Manchester, Manchester, United Kingdom Purpose/Objective: Radiotherapy is standard-of-care in muscle-invasive bladder cancer (MIBC). Radiation-induced extracellular matrix (ECM) remodelling and fibrosis can induce tumour resistance and recurrence, but it has not been studied in MIBC. We characterised the impact of radiation on the ECM composition of MIBC. Material/Methods: Bladder cancer cell lines (T24, UMUC3, J82) were cultured in McCoy's 5A medium with 10% FBS at 37°C, 5% CO 2 . Cells were irradiated (2.75 Gy/day, Monday-Friday) over two weeks. One month post radiation, cell-derived ECMs were extracted and analysed by mass spectrometry. Immunofluorescence staining for FN, COL1, and COL5 was performed by confocal imaging. ECM genes were tested for prognostic significance in one cystectomy (TCGA-BLCA, n=397) and one radiotherapy (BCON, n=151) cohort. Gene expression correlation among genes of interest was evaluated using Spearman statistics. GraphPad Prism (v. 9.3.1) and RStudio (v. 4.3.3) were used in statistical analyses. Results: We identified 613 proteins significantly affected by radiation (p.adj<0.05, fc >2 or <-2), of which 68 were ECM proteins (Fig 1A). We found a general increase in proteases and protease regulators, suggesting radiation promotes ECM degradation. Immunofluorescence confirmed radiation affected ECM structure by consistently reducing FN1 and COL5 fibre numbers (Fig 1 B-C). Fibre morphology was also affected, with a significant decrease in COL5 (p<0.05) and COL1 (p<0.0001) width across all cell lines. High expression levels of FN1, COL5A1 and COL1A1 mRNA expression were adverse prognostic markers in both cohorts (Fig. 2). Significant prognostic value was only found in the radiotherapy cohort for patients without hypoxia-modifying treatment. Finally, we found significant correlations between the expression of FN1, COL5A1 and COL1A1 (r>0.84, p<0.001), suggesting alterations in the ECM structure are associated.