Background. Itraconazole (ITZ) targets Candida spp. infections in hypoxic chronic wounds. ITZ efficacy is compromised by limited solubility, short half-life, slow adsorption. These snags may be overcome by topical administration following proper encapsulation in nanodroplets (NDs). Oxygen-loaded nanodroplets (OLNDs), cored with O2-storing 2H,3H-decafluoropentane and shelled with chitosan, a promising anti-fungal polysaccharyde, proved effective in delivering oxygen to hypoxic tissues. Here, low molecular weight chitosan-shelled nanodroplets were functionalised by coupling ITZ/O2 into the core. ITZ-free/oxygen-free, ITZ-free/oxygen-loaded, ITZ-loaded/oxygen-free, and ITZ-loaded/oxygen-loaded nanodroplets (ITZF-OFNDs, ITZF-OLNDs, ITZL-OFNDs, and ITZL-OLNDs) were challenged for their anti-fungal activity against azole resistant Candida albicans and C.glabrata. ITZF-OFNDs/ITZF-OLNDs were investigated for their physical interaction with yeasts and toxicity to human skin cells, under hypoxic conditions. Materials/Methods. Nanodroplet formulations were characterised for morphology and physico-chemical parameters by microscopy and light scattering. ITZ/O2 release up to 8h was measured by HPLC chromatography or oxymetry. Yeast growth was monitored by colony counts upon incubation with 10%v/v ITZF-OFNDs, ITZF-OLNDs, ITZL-OFNDs, and ITZL-OLNDs up to 72h. The interaction between NDs and yeasts after incubation for 3 and 24h was checked by confocal microscopy. HDF, HaCAT, and HMEC-1 cell lines, employed for studies on human fibroblasts, keratinocytes, and dermal microvascular endothelium, were incubated for 24h with 10%v/v ITZF-OFNDs and ITZF-OLNDs either in normoxic (20%,O2) or hypoxic (1%,O2) conditions. Cell viability and ND cytotoxicity were evaluated by MTT and LDH assays. Results. NDs displayed spherical shapes and cationic surfaces, with average diameters from 200 nm (OFNDs) to 400 nm (OLNDs). ITZ loading did not significantly alter ND sizes. NDs showed prolonged release of O2/ITZ over time. ITZF-OLNDs/OFNDs significantly inhibited yeast growth up to 72h, confirming long-term anti-fungal properties of chitosan. ITZ addition displayed better antifungal activity, in comparison with ITZ-free nanocarriers. NDs were internalised by yeasts after 3h of incubation, without being influenced by O2. As expected, hypoxia mildly compromised human cell viability. Nevertheless, NDs were well tolerated by fibroblasts and keratinocytes, whereas they showed slightly toxicity to endothelium. Conclusions. ITZ/O2 topical administration through ND formulations may represent an innovative, nonconventional strategy to counteract fungal infections and hypoxia and to promote tissue repair processes in chronic wounds.

ITRACONAZOLE-LOADED CHITOSAN-SHELLED NANODROPLETS AS NEW NONCONVENTIONAL ANTIFUNGAL STRATEGY AGAINST CANDIDA ALBICANS AND C.GLABRATA INFECTIONS IN HYPOXIC CHRONIC WOUNDS.

Mandras N;Roana J;Scalas D;Argenziano M;Finesso N;Luganini A;Giribaldi G;Banche G;Allizond V;Cavalli R;Prato M;Cuffini AM;Tullio V
2018

Abstract

Background. Itraconazole (ITZ) targets Candida spp. infections in hypoxic chronic wounds. ITZ efficacy is compromised by limited solubility, short half-life, slow adsorption. These snags may be overcome by topical administration following proper encapsulation in nanodroplets (NDs). Oxygen-loaded nanodroplets (OLNDs), cored with O2-storing 2H,3H-decafluoropentane and shelled with chitosan, a promising anti-fungal polysaccharyde, proved effective in delivering oxygen to hypoxic tissues. Here, low molecular weight chitosan-shelled nanodroplets were functionalised by coupling ITZ/O2 into the core. ITZ-free/oxygen-free, ITZ-free/oxygen-loaded, ITZ-loaded/oxygen-free, and ITZ-loaded/oxygen-loaded nanodroplets (ITZF-OFNDs, ITZF-OLNDs, ITZL-OFNDs, and ITZL-OLNDs) were challenged for their anti-fungal activity against azole resistant Candida albicans and C.glabrata. ITZF-OFNDs/ITZF-OLNDs were investigated for their physical interaction with yeasts and toxicity to human skin cells, under hypoxic conditions. Materials/Methods. Nanodroplet formulations were characterised for morphology and physico-chemical parameters by microscopy and light scattering. ITZ/O2 release up to 8h was measured by HPLC chromatography or oxymetry. Yeast growth was monitored by colony counts upon incubation with 10%v/v ITZF-OFNDs, ITZF-OLNDs, ITZL-OFNDs, and ITZL-OLNDs up to 72h. The interaction between NDs and yeasts after incubation for 3 and 24h was checked by confocal microscopy. HDF, HaCAT, and HMEC-1 cell lines, employed for studies on human fibroblasts, keratinocytes, and dermal microvascular endothelium, were incubated for 24h with 10%v/v ITZF-OFNDs and ITZF-OLNDs either in normoxic (20%,O2) or hypoxic (1%,O2) conditions. Cell viability and ND cytotoxicity were evaluated by MTT and LDH assays. Results. NDs displayed spherical shapes and cationic surfaces, with average diameters from 200 nm (OFNDs) to 400 nm (OLNDs). ITZ loading did not significantly alter ND sizes. NDs showed prolonged release of O2/ITZ over time. ITZF-OLNDs/OFNDs significantly inhibited yeast growth up to 72h, confirming long-term anti-fungal properties of chitosan. ITZ addition displayed better antifungal activity, in comparison with ITZ-free nanocarriers. NDs were internalised by yeasts after 3h of incubation, without being influenced by O2. As expected, hypoxia mildly compromised human cell viability. Nevertheless, NDs were well tolerated by fibroblasts and keratinocytes, whereas they showed slightly toxicity to endothelium. Conclusions. ITZ/O2 topical administration through ND formulations may represent an innovative, nonconventional strategy to counteract fungal infections and hypoxia and to promote tissue repair processes in chronic wounds.
28th ECCMID European Congress of Clinical Microbiology and Infectious Diseases
Madrid (Spain)
21-24 aprile 2018
ESCMID eLibrary
ESCMID
P0326-1
P0326-2
https://www.escmid.org/escmid_publications/escmid_elibrary/
Itraconazole, nanodroplets, Candida albicans, Candida glabrata, hypoxic chronic wounds
Mandras N, Roana J, Scalas D, Argenziano M, Finesso N, Luganini A, Giribaldi G, Fumagalli M, Sangiovanni E, D’Alessandro S, Basilico N, Dell’Agli M, Banche G, Allizond V, Cavalli R, Prato M, Cuffini AM, Tullio V
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/2318/1680143
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