title: Characterization of a Polyethylene Glycol-Amphotericin B Conjugate Loaded with Free AMB for Improved Antifungal Efficacy.
abstract: Amphotericin B (AMB) is a highly hydrophobic antifungal, whose use is limited by its toxicity and poor solubility. To improve its solubility, AMB was reacted with a functionalized polyethylene glycol (PEG), yielding soluble complex AmB-PEG formulations that theoretically comprise of chemically conjugated AMB-PEG and free AMB that is physically associated with the conjugate. Reverse-phase chromatography and size exclusion chromatography methods using HPLC were developed to separate conjugated AMB-PEG and free AmB, enabling the further characterization of these formulations. Using HPLC and dynamic light scattering analyses, it was observed that the AMB-PEG 2 formulation, having a higher molar ratio of 2 AMB: 1 PEG, possesses more free AMB and has relatively larger particle diameters compared to the AMB-PEG 1 formulation, that consists of 1 AMB: 1 PEG. The identity of the conjugate was also verified using mass spectrometry. AMB-PEG 2 demonstrates improved antifungal efficacy relative to AMB-PEG 1, without a concurrent increase in in vitro toxicity to mammalian cells, implying that the additional loading of free AMB in the AMB-PEG formulation can potentially increase its therapeutic index. Compared to unconjugated AMB, AMB-PEG formulations are less toxic to mammalian cells in vitro, even though their MIC50 values are comparatively higher in a variety of fungal strains tested. Our in vitro results suggest that AMB-PEG 2 formulations are two times less toxic than unconjugated AMB with antifungal efficacy on Candida albicans and Cryptococcus neoformans.
authors.affiliation: Bioprocessing Technology Institute, Agency for Science, Technology and Research (A*STAR), Singapore, Singapore.
authors.forename: Tessa Rui Min
authors.initials: TR
authors.lastname: Tan
authors.affiliation: Bioprocessing Technology Institute, Agency for Science, Technology and Research (A*STAR), Singapore, Singapore.
authors.forename: Kong Meng
authors.initials: KM
authors.lastname: Hoi
authors.affiliation: Bioprocessing Technology Institute, Agency for Science, Technology and Research (A*STAR), Singapore, Singapore.|Department of Anatomy, Yong Loo Lin School of Medicine, National University of Singapore, Singapore, Singapore.
authors.forename: Peiqing
authors.initials: P
authors.lastname: Zhang
authors.affiliation: Bioprocessing Technology Institute, Agency for Science, Technology and Research (A*STAR), Singapore, Singapore.|Department of Pharmacy, Faculty of Science, National University of Singapore, Singapore, Singapore.
authors.forename: Say Kong
authors.initials: SK
authors.lastname: Ng
mesh_terms: D000666:Amphotericin B / Q000031:analogs & derivatives* / Q000493:pharmacokinetics / Q000494:pharmacology; D000935:Antifungal Agents / Q000493:pharmacokinetics* / Q000494:pharmacology; D002176:Candida albicans / Q000187:drug effects; D002460:Cell Line / Q000187:drug effects; D002626:Chemistry, Pharmaceutical; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D003455:Cryptococcus neoformans / Q000187:drug effects; D057809:HEK293 Cells / Q000187:drug effects; D006801:Humans; D008826:Microbial Sensitivity Tests; D010316:Particle Size; D011092:Polyethylene Glycols / Q000737:chemistry; D012995:Solubility
chemical_list: D000935:Antifungal Agents; D011092:Polyethylene Glycols; D000666:Amphotericin B