Ultrashort Self-Assembling Peptide Hydrogel for the Treatment of Fungal Infections

Abstract

The threat of antimicrobial resistance to society is compounded by a relative lack of new clinically effective licensed therapies reaching patients over the past three decades. This has been particularly problematic within antifungal drug development leading to a rise in fungal infection rates and associated mortality. This paper highlights the potential of an ultrashort peptide, (naphthalene-2-ly)-acetyl-diphenylalanine-dilysine-OH (NapFFKK-OH), encompassing hydrogel-forming and antifungal properties within a single peptide motif, thus overcoming formulation (e.g. solubility, drug loading) issues associated with many current employed highly hydrophobic antifungals. A range of fungal susceptibility (colony counts) and cell cytotoxicity (MTS cell viability, LIVE/DEAD staining® with fluorescent microscopy, haemolysis) assays were employed. Scanning electron microscopy confirmed the nanofibrous architecture of our self-assembling peptide, existing as a hydrogel at concentrations of 1% w/v and above. Broad-spectrum activity was demonstrated against a range of fungi clinically relevant to infection (Aspergillus niger, Candida glabrata, Candida albicans, Candida parapsilosis and Candida dubliniensis) with greater than 4 log10 CFU/mL reduction at concentrations of 0.5% w/v and above. We hypothesise antifungal activity is due to targeting of anionic components present within fungal cell membranes resulting in membrane disruption and cell lysis. NapFFKK-OH demonstrated reduced toxicity against mammalian cells (NCTC 929, ARPE-19) suggesting increased selectivity for fungal cells. However, further studies relating to safety for systemic administration is required, given the challenges toxicity has presented in the wider context of antimicrobial peptide drug development. Overall this study highlights the promise of NapFFKK-OH hydrogels, particularly as a topical formulation for the treatment of fungal infections relating to the skin and eyes, or as a hydrogel coating for the prevention of biomaterial related infection.

Nucleopeptide-Coupled Injectable Bioconjugated Guanosine-Quadruplex Hydrogel with Inherent Antibacterial Activity

“…Among various fungal species, C. albicans causes a severe broad range of clinical infections named Candidasis. 67 As most of the commercially available antifungal drugs are constituted of azole derivatives, few nucleoside analogues have also been developed for treating fungal infections. 26,68 We further extend our investigation with the nucleobase-rich G4NP G-quadruplex hydrogel to investigate the antifungal activity against C. albicans which causes serious skin infections.…”

Investigating the In Vivo Antimicrobial Activity of a Self-Assembling Peptide Hydrogel Using a Galleria mellonella Infection Model

“…NapFFKK-OH hydrogel toxicity studies using in vivo models of G. mellonella expand on previously conducted in vitro studies by our group whereby a monolayer of murine fibroblast NCTC 929 cells was exposed to varying concentrations of solubilized and hydrogel forms of NapFFKK-OH. NapFFKK-OH was nontoxic in the solubilized micromolar range (≤500 μM: >80% cell metabolic activity), 28 according to the International Standard classification for biomaterial toxicity, 29 but less than 60% NCTC 929 cell metabolic activity was observed at concentrations of ≥1.5% w/v in the hydrogel form. 10 By comparison, the in vivo studies presented (Figure 1) indicate that in more complex cell environments, for example, at an organism level, toxicity is less pronounced.…”

“…Hydrogels were formulated using 0.5 M HCl, 1 M NaOH, and sterile deionized water as previously demonstrated by our group. 28 NapFFKK-OH In Vivo Toxicity. A method similar to that used by Gibreel and Upton was used to assess peptide hydrogel toxicity.…”

“…Controls were provided in the form of untreated larvae, and these were injected separately with PBS and the comprising elements of the peptide hydrogel (deionized water, 1 M NaOH, and 0.5 M HCl) in the relevant volumes for each hydrogel concentration (Figure S2). 18,28 Verifying Maximum Nonlethal Dose of Bacteria. Methods similar to those of Hornsey and Wareham were used to conduct validation and antimicrobial studies.…”

Unravelling the antimicrobial activity of peptide hydrogel systems: current and future perspectives

“…Although studied less frequently, their benefit within antifungal applications has also been postulated. 42,43 It is also important to look closely at peptide nanostructures to establish a link between primary and secondary structure; assembly; mechanical and antimicrobial properties. There appears to be different structure-activity relationships and molecular mechanisms that determine antimicrobial activity for fully solubilized and nanofibrous peptides.…”