341 - Targeting Angiotensin and Inflammation to Prevent Radiotherapy-Induced Bladder Toxicity

A. Groves¹, N. Paris¹, E. Hernady¹, C. J. Johnston², Y. F. Lee³, S. L. Kerns⁴, and B. Marples¹; ¹Department of Radiation Oncology, University of Rochester, Rochester, NY, ²Department of Pediatrics, University of Rochester, Rochester, NY, ³Department of Urology, University of Rochester, Rochester, NY, ⁴Medical College of Wisconsin, Milwaukee, WI

Purpose/Objective(s): Radiotherapy (RT) dose-volume constraints help to limit bladder radiotoxicity for prostate cancer (PCa) patients, but the incidence and severity of injury varies. This suggests that patient-specific biologic differences alter the response of each patient. A large genome-wide association study (GWAS) in PCa patients treated with RT provided clues to the etiology of gross hematuria. The top associated SNPs correlated with hematuria were in AGT (angiotensinogen), part of the renin-angiotensin system (RAS). To investigate this association, we evaluated the ability of the angiotensin-converting enzyme (ACE) inhibitor lisinopril to prevent RT-induced bladder toxicity. Materials/

Methods: C57BL/6 male mice were treated with lisinopril (oral, 6mg/L) 5 days before focal bladder X-irradiation (3 fractions of 10Gy over 5 days), or sham-RT, and lisinopril maintained for 20 weeks. RT was delivered using a medical technology company SARRP Muriplan CT-image-guidance with parallel-opposed lateral beams. Bladder toxicity was assessed using a micturition assay and by histopathology. Total RNA was isolated from separated urothelium and muscle layers and expression of CCL2, CCR2, IL1ß and MMP12 determined by qRT-PCR 1wk and 20 wk after RT. Then, the signaling and recruitment of monocyte-derived macrophages to the irradiated bladder was assessed at early and late timepoints post-RT.
Results: Compared with non-irradiated animals, RT treatment resulted in an increase in urinary frequency and reduced the volume of individual voids at 20 weeks, and these changes were associated with persistent recruitment of macrophages to the bladder muscle, thickening of the bladder muscle and development of fibrosis as assessed by Trichrome staining of the bladder. These long-term changes in the pattern of micturition were reduced by lisinopril treatment (p<0.05). Irradiation significantly increased gene expression of inflammatory related genes (CCL2, CCR2, MMP12, IL1b, FRAS1) in both the bladder urothelium and muscle at early time points post RT but this was only evident in the muscle at later times. Lisinopril treatment decreased CCL2 expression in the muscle indicating an abrogation of macrophage recruitment signaling, and decreased MMP12 in the muscle showing an inhibition of processes involved in collagen and matrix progressing, a precursor for fibrosis. In addition, Lisinopril treatment reduced expression of the FRAS1 extracellular matrix protein, while FRAS1 was enriched for hematuria risk SNPs in a prior GWAS study of bladder toxicity in prostate cancer patients.
Conclusion: This murine study demonstrated that lisinopril treatment protected against late RT-induced bladder injury and provided support that macrophages are critical in the mechanism of injury.