Molecular co-assembly creates species selectivity in a conventional antifungal drug Kenta Morita † , Yuya Nishimura ‡ , Jun Ishii ‡ and Tatsuo Maruyama † † Department of Chemical Science and Engineering, Graduate School of Engineering, Kobe University, Japan, ‡ Division of Biotechnology and Environmental Technology, Graduate School of Science, Technology and Innovation, Kobe University, Japan Background Low-molecular-weight gelators self-assemble through non-covalent interactions to form a three-dimensional network, enabling the gelation of various solvents. In recent years, it has been discovered that low-molecular- weight gelators utilize this self-assembly ability to exhibit unique functions not present in polymers. In previous studies, we found Ac-FFFGK (P1), a short peptide that is acetylated the N-terminus, which forms hydrogels at low concentrations 1 . Peptide-based low-molecular-weight gelators composed solely of amino acids exhibit high biocompatibility, making them promising for applications in the pharmaceutical field as novel biomaterials. For instance, these compounds are amphiphilic, forming self-assemblies in water, and it is envisaged that they can create hydrogels dispersing hydrophobic drugs through co-assembly. In this study, we focused on Amphotericin B (AmB), an antimicrobial agent used for microbial infection prevention and treatment, as a model drug. Due to its high hydrophobicity, AmB cannot be formulated as an aqueous solution or gel for local administration. Therefore, we attempted to solubilize or hydrogelize the antimicrobial agent using P1. We investigated the influence of the gelling agent on the properties of the antimicrobial agent during this process. Experimentals i) To prepare the AmB-P1 Gel, Ac-FFFGK (0.5 wt%) and AmB were added to YPD medium, stirred, and heated at 100°C for 5 minutes. The mixture was dispensed into a 96-well plate and left at 4°C overnight to prepare AmB-P1 gels. As a control, the AmB-Agar gel was prepared by dissolving AmB in DMSO and gelation with Agar. Saccharomyces cerevisiae was inoculated on the prepared gels, and antibacterial activity was evaluated by observing its colony formation. ii) To assess the protease-secreting microbe-specific toxicity of AmB-P1 gel, AmB-P1 gels and AmB-Agar gels were prepared using protease-high secretion medium for Aspergillus oryzae. A. oryzae or S. cerevisiae was inoculated on the gels, and their growth was observed. Results and Discussion i) When P1 and AmB were introduced into the medium at concentrations above the gelation threshold and heated, 100 µg/ml of AmB dissolved, successfully obtaining AmB-P1 gels. However, the antibacterial activity of AmB contained in AmB-P1 gels was less than one-tenth of that of free AmB contained in AmB-Agar gels. This discrepancy is attributed to AmB being strongly trapped within the self-assemblies (gels) of P1, preventing its interaction with fungal cell membranes. Consequently, we conducted the following experiment, considering whether AmB-P1 gels specifically exhibited toxicity against fungi secreting proteases that can degrade P1. ii) AmB-P1 gels demonstrated antibacterial activity equivalent to that of free AmB against A. oryzae secreting proteases. Thus, it was demonstrated that AmB-P1 gels selectively exhibit toxicity against fungi that secrete proteases capable of degrading P1. References 1. Restu,WK. et al ., Langmuir, 34 , 8065-8074 (2018).
P88
© The Author(s), 2023
Made with FlippingBook Learn more on our blog