Synthesis, Characterization and In–vitro Evaluation of Anticancer Potential of Chitosan–coated Polyoxometalates Nanoparticles

Abstract

Synthesis, Characterization and In–vitro Evaluation of Anticancer Potential of Chitosan–coated Polyoxometalates Nanoparticles Polyoxometalates (POMs) are discrete anions and have become significant in biomedical research due to their structural diversity which renders them highly active against bacterial, viral, cancer and HIV infection. In this study six different POMs were resynthesized and encapsulated within chitosan (CTS) through inotropic gelation technique. The synthesized nanoparticles were characterized in terms of their surface morphology, particle size and zeta potential. All nanoparticles were observed non–spherical with hollow surface having particle diameter below 200 nm. For each formulation the observed zeta potential value was in acceptable limits (> 25 mV). The lowest particle diameter (91 ± 4 nm) was recorded for CTS–TiW11Co with zeta potential 52.0 ± 5.21 mV. The entrapment efficiency, dissolution studies and release kinetics were estimated for all nano formulations. The CTS–P5W30 nanoparticles showed the maximum entrapment efficiency (92 ± 9 %) while the release pattern of POMs from nanoparticles was observed as diffusion and polymer surface erosion. Enzyme inhibition study on tissue non–specific alkaline phosphatase was determined where free POMs and their nanoparticles were analyzed and compared with the standard inhibitor. The inhibition constant (Ki) value for CTS–TiW11Co (10.2 ± 9 ng/mL) was ten–fold lower than the Ki of levamisole (137.5 ± 29 ng/mL). The anticancer potential of free POMs and CTS conjugated nanoparticles were studied on two different cancer cell lines including human cervical cancer cells (HeLa cells) and human breast cancer cells (MCF–7 cells). Furthermore, the toxicity of these compounds was studied on normal cells (vero cells). The compounds CTS–TiW11Co and CTS–P5W30 were very effective on HeLa cells with IC50 of 8.94 ± 2.33 and 7.26 ± 2.55 μg/mL respectively. While these compounds showed the minimum toxicity on vero cells. The CTS–TiW11Co and CTS–P5W30 compounds also showed the lowest IC50 values when tested on MCF–7 cells with 4.55 ± 1.98 and 6.36 ± 1.22 (μg/mL) correspondingly. Based on maximum potential of cytotoxicity on x cancer cells and low toxicity toward normal cells, CTS–TiW11Co and CTS–P5W30 were selected for further experiments. The CTS–TiW11Co and CTS–P5W30 were morphologically analyzed for any signs of apoptosis with DAPI staining. The treated cells (HeLa and MCF–7) were characterized by chromatin condensation, cell shrinkage and formation of apoptotic bodies. A microscopic analysis of the production of reactive oxygen species (ROS) was carried out with the help of fluorescent dye DCFH–DA. The dye was incubated with HeLa and MCF–7 cells after treatment with CTS–TiW11Co and CTS–P5W30 nanoparticles. The treated cells were characterized by glowing cells showing signs of lipid peroxidation and chromatin condensation. Furthermore, a DNA fragmentation analysis was carried out on HeLa and MCF–7 cells separately. The existence of DNA fragments had confirmed apoptosis in HeLa and MCF–7 cells treated with CTS–TiW11Co and CTS–P5W30 nanoparticles.