Hyaluronic Acid-Coated Chitosan/Gelatin Nanoparticles as a New Strategy for Topical Delivery of Metformin in Melanoma

Abstract

Metformin is a multipotential compound for treating diabetes II and controlling hormonal acne and skin cancer. This study was designed to enhance metformin skin penetration in melanoma using nanoparticles containing biocompatible polymers. Formulations with various concentrations of chitosan, hyaluronic acid, and sodium tripolyphosphate were fabricated using an ionic gelation technique tailored by the Box-Behnken design. The optimal formulation was selected based on the smallest particle size and the highest entrapment efficiency (EE%) and used in ex vivo skin penetration study. In vitro antiproliferation activity and apoptotic effects of formulations were evaluated using MTT and flow cytometric assays, respectively. The optimized formulation had an average size, zeta potential, EE%, and polydispersity index of $329\pm 6.30$  nm, $21.94\pm 0.05$  mV, $64.71\pm 6.12\%$ , and $0.272\pm 0.010$ , respectively. The release profile of the optimized formulation displayed a biphasic trend, characterized by an early burst release, continued by a slow and sustained release compared to free metformin. The ex vivo skin absorption exhibited $1142.5\pm 156.3$  μg/cm2 of metformin deposited in the skin layers for the optimized formulation compared to $603.2\pm 93.1$  μg/cm2 for the free metformin. Differential scanning calorimetry confirmed the deformation of the drug from the crystal structure to an amorphous state. The attenuated total reflection Fourier transform infrared results approved no chemical interaction between the drug and other ingredients of the formulations. According to the MTT assay, metformin in nanoformulation exhibited a higher cytotoxic effect against melanoma cancer cells than free metformin (IC50: $3.94\pm 0.57$  mM vs. $7.63\pm 0.26$  mM, respectively, $P<0.001$ ). The results proved that the optimized formulation of metformin could efficiently decrease cell proliferation by promoting apoptosis, thus providing a promising strategy for melanoma therapy.