S9-Fibronectin, EGF-R, HB-EGF:biomarkers of urothelial damage during intravesical adjuvant therapy?

Intravesical chemotherapy and immunotherapy with BCG represent the standard therapy to prevent recurrence after transurethral resection (TUR) of non-muscle invasive bladder cancer (NMI-BC). Maintenance for at least one year is considered the best regimen. Noteworthy, a relevant number of patients do not complete the planned treatment due to local toxicity of the drug given intravesically1, 2. A major challenge for the urologists is to identify an early urothelial damage biomarker to prevent severe local toxicity requiring treatment interruption and to improve patient's compliance. The preliminary purpose of our research was to verify the possible correlation between urothelial damage induced by intravesical treatment and the expression of potential biomarkers in urine or bladder washing solution. Fibronectin (FN), Epidermal Growth Factor-Receptor (EGF-R) and Heparin-binding Epidermal Growth Factor-like Growth Factor (HB-EGF) have been preliminary investigated. The urinary HB-EGF expression in patients with interstitial cystitis has been already analyzed by some studies3, 4, 5 The biomarkers trend during therapy with hyaluronic acid and chondroitin sulphate solution will be investigated. Materials and Methods: we collected 50 ml urine and bladder washing solution during intravesical therapy in 55 patients after NMI-BC TUR and in 10 healty controls for a total of 200 samples. After centrifugation, total cellular RNA was isolated from the cell pellett using miRNeasy Mini Kit (Qiagen®) according to the manufacturer's instructions. We investigated FN and EGF-R gene expression by Real Time quantitative PCR. Finally, the abundance of HB-EGF in urine samples was measured using Enzyme-linked immunosorbent assay (ELISA) (Abcam®) following manufacturer's instructions. Results: The FN gene expression levels in NMI-BC compared with controls were increased 4.7 fold while EGF-R levels were decreased 0.9 fold. In the patients in whom local toxicity due to intravesical therapy was clinically relevant, the FN gene expression levels were increased 5.82 fold, and the EGF-R one was decreased 0.88 fold. In contrast patients before starting therapy or with a good tolerance showed gene expression levels increased 1.9 fold for FN and 1.1 fold for EGF-R. In patients receiving hyaluronic acid and chondroitin sulphate solution to treat the severe vesical toxicity the average FN gene expression levels were decreased from 3 to 0.6 fold, with concomitant symptomatic improvement. HB-EGF protein levels in urine in patients receiving intravesical chemo or immunotherapy were increased 1.2 fold compared to controls and remained unchanged during therapy. No fold change was detected in patients treated with hyaluronic acid and chondroitin sulphate solution. Our preliminary data will be updated in the final presentation. Statistical analysis is ongoing. Conclusion: EGF-R gene expression in the bladder washing solution and HB-EGF levels in urine did not show significant fold change in relation to urothelial damage compared to controls. Preliminarly, FN gene expression in bladder washing solution showed an overexpression during urothelial damage and decrease after therapy with hyaluronic acid and chondroitin sulphate solution in correlation to symptoms relief.