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

Abstract

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 pHsensitive 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.

PEGylation of Phosphatidylglycerol/Docosahexaenoic Acid Hexosomes with d-α-Tocopheryl Succinate Poly(ethylene glycol)2000 Induces Morphological Transformation into Vesicles with Prolonged Circulation Times

“…14,56,58−60 For instance, it was reported on pH-mediated hexosome-cubosome colloidal transformation in nanodispersions stabilized with Pluronic F127 that can be used to enhance antifungal therapy. 14 Notably in this study, PEGylation at pH 6.0 led to a dramatic increase in zeta potential, which we attribute to the steric shielding effect provided by nonionic PEG chains on the nanoparticle surface charge. 61,62 In addition to SAXS analysis, cryo-TEM imaging was conducted to study the effect of PEGylation on the morphological features of DOPG/DHA nanoparticles at pH 6.0 (Figure 2A−F).…”

“…This is consistent with our recent studies, and presumably arising from the presence of ionized carboxylic groups of DHA at relatively higher local concentrations at the DOPG–water interface. There are also different recent examples in the literature on pH-mediated structural transitions. ,,− For instance, it was reported on pH-mediated hexosome-cubosome colloidal transformation in nanodispersions stabilized with Pluronic F127 that can be used to enhance antifungal therapy . Notably in this study, PEGylation at pH 6.0 led to a dramatic increase in zeta potential, which we attribute to the steric shielding effect provided by nonionic PEG chains on the nanoparticle surface charge. , …”

“…This is in line with our recent findings and indicates that changes in the acyl chain length and unsaturation degree of ω-3 PUFA are not associated with a significant impact on the internal nanostructures of this family of hexosomes. It is also worth noting that the lattice parameters of the H 2 phases in these nanodispersions are greater than those reported for other hexosomal preparations (in the range of 5.50–6.80 nm), which are prepared using different lipid types and compositions and temperatures. ,, This is most likely attributed to the enhanced electrostatic repulsions at the lipid–water interface among the embedded DOPG and DHA molecules with a simultaneous increase in the water uptake by the H 2 interiors …”

“…It is also worth noting that the lattice parameters of the H 2 phases in these nanodispersions are greater than those reported for other hexosomal preparations (in the range of 5.50−6.80 nm), which are prepared using different lipid types and compositions and temperatures. 14,44,45 This is most likely attributed to the enhanced electrostatic repulsions at the lipid−water interface among the embedded DOPG and DHA molecules with a simultaneous increase in the water uptake by the H 2 interiors. 6 At pH 6.0, incorporating the PEGylated lipid (TPGS-mPEG 2000 or DSPE-mPEG 2000 ) at a concentration of 1.5 wt % into the DOPG/DHA nanodispersions was associated with a direct hexosome-liposome transition (Figure 1C).…”

“…Lyotropic nonlamellar liquid crystalline (LLC) nanoparticles are attracting increasing interest as alternative vehicles to conventional drug carriers (e.g., emulsions, liposomes, and solid lipid nanoparticles) for developing nanomedicines. − LLC nanoparticles are typically produced from biologically relevant amphiphilic lipids (including single unsaturated monoglycerides and their combinations with unsaturated fatty acids) and show a diverse range of internal structural features. − ,− Among this family of nanoparticles, also known as ISAsomes ( Internally Self-Assembled somes ), cubosomes and hexosomes with unique nanostructures are the most investigated vehicles (inverse bicontinuous cubic (Q 2 ) and discontinuous hexagonal (H 2 ) phases, respectively). ,, These nano-self-assemblies can efficiently encapsulate various bioactive compounds (including drugs, peptides, and nucleic acids). ,,− …”