Synthetic Multivalent Antifungal Peptides Effective against Fungi

Rajamani Lakshminarayanan,Roger W Beuerman,Zhou Lei,Jamie Ya Ting Chang,Charles Tang,Shouping Liu,Muruganantham Nandhakumar,Howard Riezman,Lim Yih Lin,Chandra S Verma,Jianguo Li,Eunice Goh,Padhmanaban Saraswathi,Thet Tun Aung,Siti Radiah Binte Safie,Rizwan H Khan

doi:10.1371/journal.pone.0087730

Abstract

Taking advantage of the cluster effect observed in multivalent peptides, this work describes antifungal activity and possible mechanism of action of tetravalent peptide (B4010) which carries 4 copies of the sequence RGRKVVRR through a branched lysine core. B4010 displayed better antifungal properties than natamycin and amphotericin B. The peptide retained significant activity in the presence of monovalent/divalent cations, trypsin and serum and tear fluid. Moreover, B4010 is non-haemolytic and non-toxic to mice by intraperitoneal (200 mg/kg) or intravenous (100 mg/kg) routes. S. cerevisiae mutant strains with altered membrane sterol structures and composition showed hyper senstivity to B4010. The peptide had no affinity for cell wall polysaccharides and caused rapid dissipation of membrane potential and release of vital ions and ATP when treated with C. albicans. We demonstrate that additives which alter the membrane potential or membrane rigidity protect C. albicans from B4010-induced lethality. Calcein release assay and molecular dynamics simulations showed that the peptide preferentially binds to mixed bilayer containing ergosterol over phophotidylcholine-cholesterol bilayers. The studies further suggested that the first arginine is important for mediating peptide-bilayer interactions. Replacing the first arginine led to a 2–4 fold decrease in antifungal activities and reduced membrane disruption properties. The combined in silico and in vitro approach should facilitate rational design of new tetravalent antifungal peptides.

Highlights

Resistance to current antifungal drugs has become common in recent years, mainly due to increased number of immunocompromised patients, and has underscored the need for new classes of antifungals [1,2]
The Minimum Inhibitory Concentration (MIC) values of B4010 (0.37 mM) for two clinical isolates of C. albicans were lower when compared to the MIC values for amphotericin B (1.4 mM) and natamycin (15 mM), the latter is the only US FDA approved antifungal for ophthalmic applications [41]
For the ATCC strains, B4010 induced,91% killing in 1 h at 16 MIC whereas,97% killing was observed as the concentration was increased to 26 MIC

Summary

Introduction

Resistance to current antifungal drugs has become common in recent years, mainly due to increased number of immunocompromised patients, and has underscored the need for new classes of antifungals [1,2]. Pathogenic fungal infections are the 7th most common cause of infection-related deaths in the USA [3]. The number of available antifungals is limited and developing new antifungal drugs is challenging since common drug targets in fungi are homologues of similar molecular types in human which might be inhibited. The extent of drug resistance varies, as resistance to 5-FC and azoles are more common when compared to polyenes, there are many reports of resistance to polyenes as well [6,7]

Methods

Results

Conclusion