The present investigation explores the use of folic acid (FA)-conjugated chitosan (CS)-functionalized poly (D,L-lactide-co-glycolide) (PLGA) nanocarriers for treatment of prostate cancer. A folic acid-chitosan conjugate was prepared to coat nanoparticles (NPs). Bicalutamide (BCL)-loaded nanoparticles were prepared using the nanoprecipitation method under a Box-Behnken-RSM design and characterized through MPS, PDI, ZP, SEM, PXRD, DSC, in vitro release, in vitro cytotoxicity, protein adsorption, hemolysis and stability studies. The MPS, ZP, %EE and %DL of the BCL-loaded FA conjugated CS-functionalized PLGA NPs (CPN) formulation were 206.9 nm, +21.7 mV, 87.11% and 9.37%, respectively. The drug release of the optimized BCL-loaded CPN was found to be 101.27 +/- 1.61% at 120 h. The IC50 value of the optimized coated batch was <80 mu g/ml, as compared with a value of >80 mu g/mlfor the BCL suspension, determined through a cytotoxicity assay. DSC, FTIR and PXRD studies confirmed sufficient drug entrapment along with amorphous behavior of the drug in optimized formulations. Hemolytic studies revealed that the BCL-loaded CPN was stable in blood. The stability data revealed that the CPN was stable in a phosphate buffer (pH 7.4). The CPN was also stable in short term studies carried out according to ICH Q1A (R2) guidelines. It can be concluded from all these studies that FA conjugated CS-functionalized PLGA NPs are safe and stable, so may useful in cancer therapy. (C) 2015 Elsevier B.V. All rights reserved.