In Vitro and In Vivo Efficacy of a Novel and Long-Acting Fungicidal Azole, PC1244, on Aspergillus fumigatus Infection

Abstract

The antifungal effects of the novel triazole PC1244, designed for topical or inhaled administration, against Aspergillus fumigatus were tested in a range of in vitro and in vivo studies. PC1244 demonstrated potent antifungal activities against clinical A. fumigatus isolates (n = 96) with a MIC range of 0.016 to 0.25 μg/ml, whereas the MIC range for voriconazole was 0.25 to 0.5 μg/ml. PC1244 was a strong tight-binding inhibitor of recombinant A. fumigatus CYP51A and CYP51B (sterol 14α-demethylase) enzymes and strongly inhibited ergosterol synthesis in A. fumigatus with a 50% inhibitory concentration of 8 nM. PC1244 was effective against a broad spectrum of pathogenic fungi (MIC range, <0.0078 to 2 μg/ml), especially Aspergillus terreus, Trichophyton rubrum, Candida albicans, Candida glabrata, Candida krusei, Cryptococcus gattii, Cryptococcus neoformans, and Rhizopus oryzae. PC1244 also proved to be quickly absorbed into both A. fumigatus hyphae and bronchial epithelial cells, producing persistent antifungal effects. In addition, PC1244 showed fungicidal activity (minimum fungicidal concentration, 2 μg/ml) which indicated that it was 8-fold more potent than voriconazole. In vivo, once-daily intranasal administration of PC1244 (3.2 to 80 μg/ml) to temporarily neutropenic, immunocompromised mice 24 h after inoculation with itraconazole-susceptible A. fumigatus substantially reduced the fungal load in the lung, the galactomannan concentration in serum, and circulating inflammatory cytokine levels. Furthermore, 7 days of extended prophylaxis with PC1244 showed in vivo effects superior to those of 1 day of prophylactic treatment, suggesting accumulation of the effects of PC1244. Thus, PC1244 has the potential to be a novel therapy for the treatment of A. fumigatus infection in the lungs of humans.

Antifungal synergy of a topical triazole, PC945, with a systemic triazole against respiratory Aspergillus fumigatus infection

“…Broth Microdilution Minimum inhibitory concentration (MIC) values against Aspergillus fumigatus NCPF2010, TR 34 /L98H, TR 46 /Y121F/T289 strains, using the 96-well formatted EUCAST E.DEP 9.3. method. 30,31 Plates were incubated for 48 h at 35 °C, and MIC was read visually, or turbidity was assessed by measuring optical density (OD) at 530 nm using a spectrophotometer. The IC 50 and IC 90 values were calculated from the concentration-response curve generated for each test compound.…”

Anti-fungal activity of a novel triazole, PC1244, against emerging azole-resistant Aspergillus fumigatus and other species of Aspergillus

“…We also determined MIC values for clinical isolates of A. fumigatus carrying WT cyp51A ( n = 17), TR 34 /L98H ( n = 73), TR 46 /Y121F/T289A ( n = 24), M220 ( n = 6), G54 ( n = 11), TR 53 ( n = 1), TR 46 3 /Y121F/T289A ( n = 2), G448S ( n = 1), G432C ( n = 1) or P216S ( n = 1) resistance alleles, originating from India and the Netherlands, 11 using the CLSI M38-A2 method 12 by visual reading, and also against A. fumigatus carrying TR 34 /L98H ( n = 8) isolated in France using the 96-well-formatted EUCAST E.Def 9.3 method 8 , 13 . ATCC 204305 and AF293 A. fumigatus strains were used for quality control.…”

“…As previously reported, 10 we tested antifungal effects of compounds on A. fumigatus -infected, temporarily neutropenic mice. Specific pathogen-free A/J mice (male, 5 weeks old) were used for A. fumigatus infection as they have been described to be more susceptible to A. fumigatus infection 8 , 16 . Animals were then injected with hydrocortisone (Sigma H4881, 125 mg/kg, subcutaneously) on days 3, 2 and 1 before infection, and with cyclophosphamide (Sigma C0768; 250 mg/kg, intraperitoneally) 2 days before infection to induce temporary neutropenia.…”

“…PC1244, which is 4-(4-(4-(((3R,5R)-5-((1H-1,2,4-triazol-1-yl)methyl)-5-(2,4-difluorophenyl)tetrahydrofuran-3-yl)methoxy)-3-methylphenyl)piperazin-1-yl)- N -((1S,2S)-2-hydroxycyclohexyl) benzamide, is a novel triazole antifungal agent specifically designed for inhaled administration, which has demonstrated potent, broad and persistent antifungal activities in vitro and in vivo. 8 We evaluated the antifungal activity of PC1244 against a large collection of itraconazole- or pan-azole-resistant strains of clinical A. fumigatus carrying TR 34 /L98H, TR 46 /Y121F/T289A, M220, G54, TR 53 , TR 46 3 /Y121F/T289A, G448S, G432C and P216S resistance alleles originating from India, the Netherlands and France.…”

In vitro and in vivo Efficacy of a Synergistic Combination of Itraconazole and Verapamil Against Aspergillus fumigatus

“…The strategies of azole tolerance adopted by fungi (Sanglard, 2016; Rybak et al, 2018) include (i) drug target alterations, (ii) reduction of effective cellular drug concentration, and (iii) stress adaptation by modifying the cellular metabolic pathway. Many researchers have searched for alternative drug targets or tried to develop safe broad-spectrum antifungals (Wiederhold et al, 2015; Arendrup et al, 2016; Colley et al, 2018). However, eukaryotic cells, including fungal cells and their eukaryotic host cells, share evolutionarily conserved molecular signaling pathways, which limit the fungal-specific candidates that can be targeted by new drugs.…”