Abstract
Carnosine, a naturally occurring dipeptide, was recently reported to exhibit anticancer activity; however, the molecular mechanisms and regulators underlying its activity against tumor-associated angiogenesis remain unidentified. In this study, we evaluated the in vitro and in vivo antitumor effects of carnosine in EJ bladder cancer cells and EJ-xenografted BALB/c nude mice, respectively. In addition, in vitro capillary tube formation of HUVECs, ex vivo aortic ring and in vivo Matrigel plug assays were employed to examine the antiangiogenic potential of carnosine. Carnosine significantly inhibited EJ cell proliferation. Flow cytometric and immunoblot analyses indicated that carnosine modulated regulators of the G1 cell cycle phase, including cyclin D1, CDK4 and p21WAF1. The mitogen-activated protein kinases, ERK and p38, but not JNK or AKT, responded to carnosine. Carnosine inhibited the migratory and invasive potential of EJ cells by inhibiting MMP-9 activity, which was associated with suppression of binding activity of NF-κB, SP-1 and AP-1. In xenograft tumors, carnosine exhibited antitumor activity equivalent to cisplatin, but no weight loss occurred in carnosine-treated mice. In HUVECs, carnosine inhibited VEGF-mediated proliferation, colony tube formation, migration and invasion. The antiangiogenic activity of carnosine was partially due to the suppression of VEGFR-2-mediated ERK/AKT/eNOS signaling and MMP-2. Furthermore, using aortic ring and Matrigel plug assays, we confirmed the antiangiogenic activity of carnosine. Given that targeting tumor-associated angiogenesis is a proven effective therapeutic strategy, our results may provide valuable information for the development of preventive or therapeutic agents for bladder cancer patients.