Enhanced Cytotoxicity of 5-Fluorouracil Against Skin Cancer Cell Lines and 3D Spheroid Tumor Model Using Solid Lipid Nanoparticles

This study aimed to develop solid lipid nanoparticles containing 5-fluorouracil (5-FU loaded PGSLNs) to achieve desired physicochemical properties and to enhance in vitro cytotoxicity against monolayered and 3D spheroid skin tumor models. 5-FU loaded PGSLNs were prepared via cold homogenization using Glyceryl Palmitostearate (GP) and Geleol® as solid lipids and Poloxamer 188 and Tween 80 as surfactant and co-surfactant, respectively. The developed nanoparticles were then physicochemically characterized, and cytotoxicity was evaluated against monolayered and 3D spheroid skin tumor models using the Cell TiterBlue® Assay and Cell-Titre Glo assays, respectively. The cellular uptake behavior of nanoparticles against monolayered and 3D spheroid tumor models was studied through flow cytometric analysis and fluorescence microscopic analysis. Nanoparticle sizes ranged from 116.3 ± 1.19 to 235.13 ± 1.15 nm, with a PDI < 0.45, and zeta potential was found to be between − 33.67 ± 0.40 and − 19.37 ± 0.25. FTIR analysis revealed no chemical interactions among components, and Differential Scanning Calorimetry showed the thermal stability of the drug in nanoparticles. The in vitro drug release profile depicted biphasic release behavior, with 30–35% burst release within 3 h and sustained release for 48 h. Enhanced cytotoxicity was observed against monolayered skin tumor models (B16F10, A375, and A431) and a 3D spheroid skin tumor model (A431). Rhodamine-labeled PGSLNs showed increased uptake in monolayered models via fluorescence microscopy and flow cytometry, with enhanced uptake in 3D spheroid models confirmed by fluorescence microscopy. The hemocompatibility assay indicated the safety of nanoparticles at physiological levels. The designed solid lipid nanoparticles significantly enhanced the cytotoxic effect of 5-FU against monolayered and 3D spheroid skin tumor models and can be used as promising approach for skin cancer.