Abstract. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) triggers apoptosis in a variety of tumor cells by engaging the death receptors 4 (DR4) and 5 (DR5). We investigated the effect of chemotherapeutic drugs on DR4-mediated apoptosis in human bladder cancer cells, using a human monoclonal agonistic antibody specific for DR4, mapatumumab. Cytotoxicity was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Synergy was assessed by isobolographic analysis. Treatment of human bladder cancer T24 cells with mapatumumab in combination with mitomycin C, vinblastine or gemcitabine did not overcome resistance to these agents. However, treatment with mapatumumab in combination with epirubicin (EPI) had a synergistic cytotoxic effect. Synergy was also obtained in KU7 and RT112 human bladder cancer cells. A synergistic effect was also observed with mapatumumab in combination with pirarubicin. The synergy obtained in cytotoxicity with mapatumumab and EPI was also achieved in apoptosis. EPI markedly increased DR4 expression in the bladder cancer cells at both the mRNA and protein levels. Furthermore, the combination-induced cytotoxicity was significantly suppressed by the DR4:Fc chimeric protein. The combination of EPI and mapatumumab significantly activated the caspase cascade, including caspase-8, -9 and -3, which are the downstream molecules of death receptors. These findings indicate that EPI sensitizes bladder cancer cells to DR4-mediated apoptosis through induction of DR4 and activation of caspases, suggesting that the combination therapy of EPI and mapatumumab may be effective for bladder cancer therapy.
IntroductionEach year more than 350,000 new cases of bladder cancer are diagnosed globally (1), making bladder cancer the ninth most frequent cancer worldwide (2). Epirubicin (EPI) and pirarubicin (THP) are a new generation of anthracycline drugs, widely used as anticancer chemotherapeutic agents in various types of cancers including bladder cancer (2,3). Furthermore, these anthracycline drugs are less cardiotoxic than doxorubicin. However intrinsic and acquired resistance to chemotherapeutic drugs are considered as major problems. Therefore, it is important to develop new treatment strategies for bladder cancer patients to prevent recurrence and reduce the risk of tumor progression.Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is a pro-apoptotic member of the TNF superfamily and a potentially effective anticancer agent. TRAIL has been identified as a powerful activator of programmed cell death or apoptosis in various tumor cells, yet is considered relatively nontoxic against most normal cells (4-6). TRAIL induces apoptosis by interacting with two death-inducing receptors TRAIL-R1 (DR4) and TRAIL-R2 (DR5) following activation of the caspase cascade that initiates both extrinsic and intrinsic apoptotic pathways (4,7-9). In addition, TRAIL binds to two other receptors, TRAIL-R3 (DcR1) and TRAIL-R4 (DcR2), which lack a functional cytoplasmic death dom...