T. Okamoto

243 Glycan modification of podoplanin enhances growth factor production through stabilization of platelet aggregation in bladder cancer cell

INTRODUCTION AND OBJECTIVES: Non-muscle invasive bladder cancer is often treated by intravesical chemotherapy following transurethral resection. Mechanism of the effect of intravesical chemotherapy has yet to be elucidated. In this study, we investigated cell death mechanism induced by high-dose chemotherapeutics using a newly-developed culture method for primary cancer cells. METHODS: We have developed a novel culture method which enables preparation and culture of primary cancer cells as multicellular spheroids which are termed cancer tissue-originated spheroids (CTOSs) (Okuyama 2013 J Urol). First we generated a bladder cancer patient-derived xenograft. We prepared CTOSs from the xenograft, and exposed to high-dose (1mg/ml) of epirubicin (e-ADM) or mitomycin C (MMC) for 2 hours, or low-dose (0.01mg/ml) of those chemotherapeutics. We investigated absorption of chemotherapeutics into CTOSs and assessed the mode of cell death. Next, we prepared CTOSs directly from surgical specimens of bladder cancer and tested the individual responses after exposure to high-dose e-ADM and MMC. RESULTS: At high dose, e-ADM was promptly and homogenously delivered into cancer cells in the CTOSs. High-dose e-ADM and MMC decreased ATP levels in CTOSs within an hour accompanied with mitochondrial swelling and reduction of mitochondrial membrane potential, while plasma membrane integrity was maintained. Some of the molecules in mitochondrial apoptotic pathway were subsequently activated, while at later time point the cells lost integrity of cell membrane without DNA laddering. In contrast, CTOSs exposed to low-dose of eADM or MMC exhibited typical DNA laddering. Intriguingly, the decrease of ATP levels was varied among CTOSs derived from 20 surgical specimens. CONCLUSIONS: High-dose chemotherapeutics used in intravesical therapy may induce necrosis-like cell death, different from typical apoptosis caused by chemotherapeutics at the dose of systemic chemotherapy. Each bladder cancer may have a different response to high-dose chemotherapeutics used in intravesical therapy, indicating necessity of chemosensitivity test aiming for personalized medicine.