In the present study, a novel nanocarrier was developed for the delivery of anticancer drug to the cancer tissues. For this purpose, 5-FU loaded methoxy poly(ethylene glycol)-poly(lactide) (mPEG-PLA) (5-FU NP) based polymeric nanoparticles was designed to increase the chemotherapeutic efficacy against breast cancers. Nanoprecipitation method was used to prepare the drug-loaded nanoparticles. The nanoparticle was evaluated in terms of DLS, TEM, in vitro release kinetics, and in vivo parameters. The average particle diameter of drug loaded NP was similar to 110 nm and exhibited a sustained drug release pattern for up to 120 h. The NP exhibited a pH-response drug release pattern with accelerated release at acidic media. The in vitro cytotoxicity assay showed the enhanced cytotoxicity effect of NP formulations in comparison to free drug. The NP system showed remarkable G2/M phase cell cycle arrest with significant amount of apoptosis cells in late and early phase of flow cytometer analysis. Consistently, NP formulation greatly decreased the tumor burden of mice with no sign of adverse effect. TUNEL assay further confirmed the superior anticancer effect of NP formulations which showed a high number of apoptotic cells. The favorable results obtained from this study highlights the potential application of encapsulated 5-FU NP in the treatment of breast cancers. The remarkable anticancer therapeutic efficacy with negligible toxicity profile of 5-FU NP makes it one of the possible alternative for the successful breast cancer therapy. (C) 2015 Elsevier B.V. All rights reserved.