Nanoparticle preparation

For preparation of nanoparticles, nanoprecipitation method was used [18]. Briefly, the appropriate amount of polymer (PLGA or PLGA-PEG-Folate) and disulfiram was dissolved in a DMSO to form a diffusing phase. In synthesis of disulfiram encapsulated PLGA-PEG-folate nanoparticle, a combination of PLGA-PEG-folate and PLGA ranging from (1:1) to (1:10) was chosen. The ratio of drug (disulfiram) to polymer (PLGA or PLGE-PEG-Folate) was 1:10 (w/w). The mixture was then added into the dispersing phase (PVA 0.5 % in water) using a syringe that positioned directly in the medium under moderate magnetic stirring (300 rpm, 10 min). The ratio of diffusing phase to dispersing phase was 1:20 (v/v). The freshly formed nanoparticles were obtained by dialyzing against water for 24 h. The nanoparticles were centrifuged at 20,000g for 15 min to remove DMSO and free disulfiram followed by several washing steps with distilled water. The purity of NPs was analyzed using spectrophotometry. The absence of DMSO in nanoparticle solution (in PBS) was confirmed at 265 nm, the absence of un-capsulated disulfiram was confirmed at 433 nm. The nanoparticles were then freeze-dried and kept at 4 °C.