Some bladder disease therapies can benefit from intravesical drug delivery, which involves direct instillation of drug into the bladder via a catheter, to attain high local concentrations of the drug with minimal systemic effects. Deguelin is a potential anticancer agent, however, its poor water solubility and neurotoxicity restrict its clinical application. To address these challenges, we investigated the promising application of deguelin in the intravesical therapy of bladder cancer by designing a novel intravesical drug delivery system for deguelin. It was found that deguelin could efficiently kill bladder cancer cells and inhibit angiogenesis. Intravesically administrated deguelin had better tolerance than systemically applied deguelin. Encapsulation of deguelin in cationic DOTAP and monomethoxy poly(ethylene glycol)-poly(ε-caprolactone) (MPEG-PCL) hybrid nanoparticles (DMP) created the deguelin loaded DMP nanoparticles (D/DMP). They had a mean particle size of 35 nm and zeta potential of 21 mV, rendering deguelin completely dispersible in aqueous media. Encapsulation of deguelin in cationic DMP nanoparticles enhanced the anticancer activity of deguelin in vitro. In addition, D/DMP nanoparticles were incorporated into a thermo-sensitive Pluronic F127 hydrogel, forming a novel D/DMP-F system, which remained in a flowing liquid state at lower than 25 °C, but underwent gelation at higher temperatures. The DMP nanoparticles in the F127 hydrogel system (DMP-F) could significantly extend the hydrophobic drug residence time and increase the drug concentration within the bladder. These results suggested that DMP-F was a good intravesical drug delivery system and D/DMP-F may have promising applications in intravesical therapy of bladder cancer.