In situ Gelling Amphotericin B Nanofibers: A New Option for the Treatment of Keratomycosis

Benedikt Göttel,Henrike Lucas,Arne Viestenz,Karsten Mäder,Frank Syrowatka,Joana Heinzelmann,Judith Kuntsche,Wolfgang Knolle

doi:10.3389/fbioe.2020.600384

Abstract

The purpose of our research was the development of Amphotericin B-loaded in situ gelling nanofibers for the treatment of keratomycosis. Different formulation strategies were applied to increase the drug load of the sparingly water-soluble Amphotericin B in electrospun Gellan Gum/Pullulan fibers. These include bile salt addition, encapsulation in poly(lactic-co-glycolic acid) (PLGA) nanoparticles and formation of a polymeric Amphotericin B polyelectrolyte complex. The Amphotericin B polyelectrolyte complex (AmpB-Eu L) performed best and was very effective against the fungal strain Issatchenkia orientalis in vitro. The complex was characterized in detail by attenuated total reflection infrared spectroscopy, X-ray powder diffraction, and differential scanning calorimetry. A heat induced stress test was carried out to ensure the stability of the polyelectrolyte complex. To gain information about the cellular tolerance of the developed polyelectrolyte complex a new, innovative multilayered-stratified human cornea cell model was used for determination of the cellular toxicity in vitro. For a safe therapy, the applied ophthalmic drug delivery system has to be sterile. Sterilization by electron irradiation caused not degradation of pure Amphotericin B and also for the bile salt complex. Furthermore, the developed Amphotericin B polyelectrolyte complex was not degraded by the irradiation process. In conclusion, a new polyelectrolyte Amphotericin B complex has been found which retains the antifungal activity of the drug with sufficient stability against irradiation-sterilization induced drug degradation. Furthermore, in comparison with the conventional used eye drop formulation, the new AmpB-complex loaded nanofibers were less toxic to cornea cells in vitro. Electrospinning of the Amphotericin B polyelectrolyte complex with Gellan Gum/ Pullulan leads to the formation of nanofibers with in situ gelling properties, which is a new and promising option for the treatment of keratomycosis.

Highlights

Keratomycosis is a severe infection of the ocular surface and anterior segment of the eye (Bourcier et al, 2017)
Our research demonstrates different opportunities to load in situ gelling nanofibers consisting of Pullulan and Gellan Gum with Amphotericin B (AmpB) prepared from aqueous solution
The drug was spun without any further solubilization ingredient (IIa), under addition of sodium cholate as solubilization angent (IIb), drug-loaded poly(lactic-co-glycolic acid) (PLGA) nanoparticles (IIc) and using a polyelectrolyte complex (IId)

Summary

Introduction

Keratomycosis is a severe infection of the ocular surface and anterior segment of the eye (Bourcier et al, 2017). Epidemiologic investigations show that 57% of the fungal keratitis patients wore contact lenses, 30% of these had an eye operation previously and 19% had jobs in the agriculture or gardener sections (Roth et al, 2019). Mainly Voriconazole and Amphotericin B (AmpB) are used solely or in combination via surface or anterior application therapy. The antimycotic therapy is characterized by high application frequency and high doses to reach complete recovery of the fungal infection (Behrens-Baumann et al, 2015; Farrell et al, 2017; Roth et al, 2019)

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