Enhancing the Cytotoxic Activity of Encorafenib Using Lipid–Polymer Hybrid Nanoparticles: An Integrated In Silico and In Vitro Evaluation

Glioblastoma multiforme (GBM) is one of the most aggressive cancers of brain that remains highly therapy resistant due to the presence of the blood-brain barrier and the complexity of cancer biology. Encorafenib, a strong inhibitor of BRAF, could be considered a good drug candidate for repositioning into GBM, but the poor brain permeability and efflux due to P-glycoprotein could limit its use in the clinical setting. This study aimed to evaluate the potential of encorafenib through an integrated in silico approach and enhance its in vitro cytotoxic activity using lipid–polymer hybrid nanoparticles (LPHNPs). The interaction of the compound with selected GBM targets (EPHA2, PTPRZ1, LSD1, and ALDH1A3) was analysed via molecular docking, followed by drug-likeness, ADMET, and toxicity prediction. The HTCC-LPHNPs, containing lipid core and quaternized chitosan (HTCC) shell, were developed via the nanoprecipitation technique. The cytotoxicity profile of the encorafenib-loaded nanoparticles was determined in the U87 glioblastoma cells using MTT assay, flow cytometry-based Annexin V-FITC/PI apoptosis test, and cell-cycle analysis. Encorafenib showed promising binding capabilities with the GBM-associated targets, and the strongest docking affinities were shown by LSD1 (−9.8 kcal/mol) and ALDH1A3 (−9.4 kcal/mol) targets. In silico prediction studies confirmed poor BBB penetration and P-glycoprotein substrate properties, which further emphasised the importance of a suitable drug delivery method. The optimised HTCC-LPHNPs formulation had an average particle size of 141.4 ± 2.3 nm, PDI of 0.445 ± 0.02, positive zeta potential, and 73.6 ± 1.8% entrapment efficiency with enhanced drug release in comparison with free encorafenib. More importantly, the IC₅₀ value reduced significantly from 45.71 ± 1.51 µg/mL for free encorafenib to 9.63 ± 1.27 µg/mL for HTCC-LPHNPs (p < 0.0001). Moreover, nanoparticle treatment induced apoptosis by increasing the apoptotic cells and the Sub-G1 cell population. Thus, the combined results reveal the significant increase in in vitro cytotoxic effect of encorafenib when used in combination with HTCC-LPHNPs. Keywords: Glioblastoma; Encorafenib; Lipid–polymer hybrid nanoparticles; Quaternized chitosan; Molecular docking; Apoptosis; Cell-cycle analysis