Acylhydrazones as Antifungal Agents Targeting the Synthesis of Fungal Sphingolipids

Abstract

The incidence of invasive fungal infections has risen dramatically in recent decades. Current antifungal drugs are either toxic, likely to interact with other drugs, have a narrow spectrum of activity, or induce fungal resistance. Hence, there is a great need for new antifungals, possibly with novel mechanisms of action. Previously our group reported an acylhydrazone called BHBM that targeted the sphingolipid pathway and showed strong antifungal activity against several fungi. In this study, we screened 19 derivatives of BHBM. Three out of 19 derivatives were highly active against Cryptococcus neoformansin vitro and had low toxicity in mammalian cells. In particular, one of them, called D13, had a high selectivity index and showed better activity in an animal model of cryptococcosis, candidiasis, and pulmonary aspergillosis. D13 also displayed suitable pharmacokinetic properties and was able to pass through the blood-brain barrier. These results suggest that acylhydrazones are promising molecules for the research and development of new antifungal agents.

Keywords: Aspergillus fumigatus; Candida albicans; Cryptococcus neoformans; acylhydrazones; antifungals; fungal infection; fungi; infectious disease; pharmacokinetics; sphingolipids.

FIG 1

Killing activity of BHBM (A), D2 (B), D13 (C), and D17 (D). Killing activity was determined using an in vitro killing assay in which the compounds were coincubated with C. neoformans cells at 37°C, 5% CO₂, pH 7.4. The number of CFU is counted during 96 h of incubation. All of the compounds displayed antifungal activity in a dose-dependent manner.

FIG 2

Survival studies of mice infected intranasally with 5 × 10⁵ C. neoformans cells and treated through i.p. injection. (A) Treatment started the day of infection with administration of 1.2 mg/kg/day. *, D13 versus no drug, P value of 0.0018. (B) Treatment started 5 days after infection with administration of 1.2 mg/kg/day.

FIG 3

Effect of 20 mg/kg/day oral administration against cryptococcosis, candidiasis, and aspergillosis. (A) Survival of mice infected intranasally with 5 × 10⁵ C. neoformans cells and treated with fluconazole, BHBM, or D13. *, D13 versus no drug, P value of 0.0018; #, D3 versus BHBM, P value of 0.0057. (B) Survival of mice infected intravenously with 10⁴ C. albicans cells and treated with fluconazole, BHBM, or D13. *, D13 versus no drug, P value of 0.0004; #, BHBM versus no drug, P value of 0.015. (C) Survival of mice infected intranasally with 2 × 10⁴ conidia of A. fumigatus and treated with voriconazole, BHBM, or D13. *, D13 versus no drug, P value of <0.001.

FIG 4

Pharmacokinetic studies of BHBM and D13. (A) BHBM administered i.v. at 1 mg/kg. (B) D13 administered i.v. at 1 mg/kg. (C) BHBM administered p.o. at 20 mg/kg. (D) D13 administered p.o. at 20 mg/kg. (E) BHBM administered i.p. at 1.6 mg/kg. (F) D13 administered i.p. at 1.6 mg/kg.