Application of Fluconazole-Loaded pH-Sensitive Lipid Nanoparticles for Enhanced Antifungal Therapy

Cryptococcus neoformans is a yeast-like fungus that can cause the life-threatening disease cryptococcal meningitis. Numerous reports have shown increased resistance of this fungus against antifungal treatments, such as fluconazole (Fluc), contributing to an 80% global mortality rate. This work presents a novel approach to improve the delivery of the antifungal agent Fluc and increase the drug's targetability and availability at the infection site. Exploiting the acidic environment surrounding a C. neoformans infected site, we have developed pH-sensitive lipid nanoparticles (LNP) encapsulating Fluc to inhibit the growth of resistant C. neoformans. The LNP-Fluc delivery system consists of a neutral lipid monoolein (MO) and a novel synthetic ionizable lipid 2-morpholinoethyl oleate (O2ME). At neutral pH, because of the presence of O2ME, the nanoparticles are neutral and exhibit a liquid crystalline hexagonal nanostructure (hexosomes). At an acidic pH, they are positively charged with a cubic nanostructure (cubosomes), which facilitates the interaction with the negatively charged fungal cell wall. This interaction results in the MIC50 and MIC90 values of the LNP-Fluc being significantly lower than that of the free-Fluc control. Confocal laser scanning microscopy and scanning electron microscopy further support the MIC values, showing fungal cells exposed to LNP-Fluc at acidic pH were heavily distorted, demonstrating efflux of cytoplasmic molecules. In contrast, fungal cells exposed to Fluc alone showed cell walls mostly intact. This current study represents a significant advancement in delivering targeted antifungal therapy to combat fungal antimicrobial resistance.

Automated summary

This study presents a novel approach to improve the delivery of antifungal agent and combat the resistance of Cryptococcus neoformans. By encapsulating the agent in pH-sensitive lipid nanoparticles, the growth of resistant C. neoformans is inhibited. The nanoparticles exhibit different structures and charges at neutral and acidic pH, facilitating interaction with the fungal cell wall and leading to a significant decrease in the minimum inhibitory concentration of the agent. Microscopic observations support these findings, showing distortion and efflux of cytoplasmic molecules in fungal cells exposed to the nanoparticles. This represents a significant advancement in targeted antifungal therapy for combating resistance.