Discovery of a Novel Potent Tetrazole Antifungal Candidate with High Selectivity and Broad Spectrum.

Thirty-one novel albaconazole derivatives were designed and synthesized based on our previous work. All compounds exhibited potent in vitro antifungal activities against seven pathogenic fungi. Among them, tetrazole compound D2 was the most potent antifungal with MIC values of <0.008, <0.008, and 2 µg/mL against Candida albicans, Cryptococcus neoformans, and Aspergillus fumigatus, respectively, the three most common and critical priority pathogenic fungi. In addition, compound D2 also exhibited potent activity against fluconazole-resistant C. auris isolates. Notably, compound D2 showed a lower inhibitory activity in vitro against human CYP450 enzymes as well as a lower inhibitory effect on the hERG K+ channel, indicating a low risk of drug-drug interactions and QT prolongation. Moreover, with improved pharmacokinetic profiles, compound D2 showed better in vivo efficacy than albaconazole at reducing fungal burden and extending the survival of C. albicans-infected mice. Taken together, compound D2 will be further investigated as a promising candidate.

From Synergy to Monotherapy: Discovery of Novel 2,4,6-Trisubstituted Triazine Hydrazone Derivatives with Potent Antifungal Potency In Vitro and In Vivo.

Invasive fungal infections pose a serious threat to public health and are associated with high mortality and incidence rates. The development of novel antifungal agents is urgently needed. Based on hit-to-lead optimization, a series of 2,4,6-trisubstituted triazine hydrazone compounds were designed, synthesized, and biological evaluation was performed, leading to the identification of compound 28 with excellent in vitro synergy (FICI range: 0.094-0.38) and improved monotherapy potency against fluconazole-resistant Candida albicans and Candida auris (MIC range: 1.0-16.0 µg/mL). Moreover, 28 exhibited broad-spectrum antifungal activity against multiple pathogenic strains. Furthermore, 28 could inhibit hyphal and biofilm formation, which may be related to its ability to disrupt the fungal cell wall. Additionally, 28 significantly reduced the CFU in a mouse model of disseminated infection with candidiasis at a dose of 10 mg/kg. Overall, the triazine-based hydrazone compound 28 with low cytotoxicity, hemolysis, and favorable ADME/T characteristics represents a promising lead to further investigation.

Design, synthesis and evaluation of novel deferasirox derivatives with high antifungal potency in vitro and in vivo.

Here we designed and synthesized 58 deferasirox derivatives with the aim of discovering novel antifungal agents. Most compounds exhibited moderate to excellent in vitro antifungal activities against Cryptococcus neoformans H99 with MIC values ranging from 0.25 µg/mL to 16 µg/mL, including ten compounds with MIC values less than 1 µg/mL that were further screened against an additional six pathogenic fungi. This class of compounds showed high potency against Candida glabrata with MIC values ranging from <0.125 µg/mL to 1 µg/mL. We identified that compound 54 has high potency against 14 strains of Candida glabrata spp. and Cryptococcus spp. with MIC values ranging from <0.125 µg/mL to 1 µg/mL. In addition, compound 54 significantly reduced the CFU in a mouse model of disseminated infection with Cryptococcus neoformans H99 at a dose of 10 mg/kg, which is comparable to FLC. Further investigations on compound 54 are currently in progress.

Design, synthesis and in vitro biological studies of novel triazoles with potent and broad-spectrum antifungal activity.

A series of novel triazole derivatives containing aryl-propanamide side chains was designed and synthesised. In vitro antifungal activity studies demonstrated that most of the compounds inhibited the growth of six human pathogenic fungi. In particular, parts of phenyl-propionamide-containing compounds had excellent, broad-spectrum antifungal activity against Candida albicans SC5314, Cryptococcus neoformans 22-21, Candida glabrata 537 and Candida parapsilosis 22-20 with MIC values in the range of ≤0.125 µg/mL-4.0 µg/mL. In addition, compounds A1, A2, A6, A12 and A15 showed inhibitory activities against fluconazole-resistant Candida albicans and Candida auris. Preliminary structure-activity relationships (SARs) are also summarised. Moreover, GC-MS analysis demonstrated that A1, A3, and A9 interfered with the C. albicans ergosterol biosynthesis pathway by inhibiting Cyp51. Molecular docking studies elucidated the binding modes of A3 and A9 with Cyp51. These compounds with low haemolytic activity and favourable ADME/T properties are promising for the development of novel antifungal agents.

Novel antifungal triazoles with alkynyl-methoxyl side chains: Design, synthesis, and biological activity evaluation.

Previous work led to the rational design, synthesis and testing of novel antifungal triazole analogues bearing alkynyl-methoxyl side chains. Tests of in vitro antifungal activity showed Candida albicans SC5314 and Candida glabrata 537 gave MIC values of ≤0.125 µg/mL for most of the compounds. Among these, compounds 16, 18, and 29 displayed broad-spectrum antifungal activity against seven human pathogenic fungal species, two fluconazole-resistant C. albicans isolates and two multi-drug resistant Candida auris isolates. Moreover, 0.5 µg/mL of 16, 18, and 29 was more effective than 2 µg/mL of fluconazole at inhibiting fungal growth of the strains tested. The most active compound (16) completely inhibited the growth of C. albicans SC5314 at 16 µg/mL for 24 h, affected biofilm formation and destroyed the mature biofilm at 64 µg/mL. Several Saccharomyces cerevisiae strains, overexpressing recombinant Cyp51s or drug efflux pumps, indicated 16, 18, and 29 targeted Cyp51 without being significantly affected by a common active site mutation, but were susceptible to target overexpression and efflux by both MFS and ABC transporters. GC-MS analysis demonstrated that 16, 18, and 29 interfered with the C. albicans ergosterol biosynthesis pathway by inhibition at Cyp51. Molecular docking studies elucidated the binding modes of 18 with Cyp51. The compounds showed low cytotoxicity, low hemolytic activity and favorable ADMT properties. Importantly, compound 16 showed potent in vivo antifungal efficacy in the G. mellonella infection model. Taken together, this study presents more effective, broad-spectrum, low toxicity triazole analogues that can contribute to the development of novel antifungal agents and help overcome antifungal resistance.

Discovery of Novel Thiosemicarbazides Containing 1,3,5-Triazines Derivatives as Potential Synergists against Fluconazole-Resistant Candida albicans.

The clinical prevalence of antifungal drug resistance has been increasing over recent years, resulting in the failure of treatments. In an attempt to overcome this critical problem, we sought novel synergistic enhancers to restore the effectiveness of fluconazole against resistant Candida albicans. Based on the structural optimization of hit compound 8 from our in-house library, a series of novel 1,3,5-triazines derivatives was designed, synthesized, and biologically evaluated for synergistic activity in combination with fluconazole. Among them, compounds 10a-o, which contain thiosemicarbazides side chains, exhibited excellent in vitro synergistic antifungal potency (MIC80 = 0.125-2.0 µg/mL, FICI range from 0.127 to 0.25). Interestingly, compound 10l exhibited moderate C. albicans activity as monotherapy with an MIC80 value of 4.0 µg/mL, and also on several Cryptococcus strains (MIC80 ranging from ≤ 0.125-0.5 µg/mL) and C. glabrata (MIC80 ≤ 0.125 µg/mL). These effects were fungal-selective, with much lower levels of cytotoxicity towards human umbilical vein endothelial cells. Here, we report a series of thiosemicarbazides containing 1,3,5-triazines derivatives as potent synergists with fluconazole, and have preliminarily validated compound 10l as a promising antifungal lead for further investigation.

Design, synthesis, and in vitro evaluation of novel antifungal triazoles containing substituted 1,2,3-triazole-methoxyl side chains.

In order to develop new triazole derivatives, we optimized the lead compound a6 by structural modifications to obtain a series of (2R,3R)-3-((1-substituted-1H-1,2,3-triazol-4-yl) methoxy)-2-(2,4-difluorophenyl)-1-(1H-1,2,4-triazol-1-yl) butan-2-ol, compounds 5-36. Most of the target compounds exhibited excellent in vitro antifungal activity against Candida albicans 10231 and Candida glabrata 537 with MIC ≤ 0.125 µg/mL. Of particular note, compounds 6, 22, 28, 30 and 36 were highly active against Candida neoformans 32609 with MIC ≤ 0.125 µg/mL and showed broad-spectrum antifungal activity including against fluconazole-resistant Candida auris 891. In addition, compounds 6 and 22 demonstrated inhibitory effects on filamentation in the azole-resistant C. albicans isolate. Moreover, compounds 6 and 22 were minimally toxic to HUVECs and possessed weak inhibitory effects on the human CYP3A4 and CYP2D6. SARs and docking study further indicated that ortho-substituted groups in the terminal phenyl ring can promote the compounds to improve their antifungal activity.

Design, Synthesis, and In Vitro and In Vivo Antifungal Activity of Novel Triazoles Containing Phenylethynyl Pyrazole Side Chains.

A series of triazole derivatives containing phenylethynyl pyrazole moiety as side chain were designed, synthesized, and most of them exhibited good in vitro antifungal activities. Especially, compounds 5k and 6c showed excellent in vitro activities against C. albicans (MIC = 0.125, 0.0625 µg/mL), C. neoformans (MIC = 0.125, 0.0625 µg/mL), and A. fumigatus (MIC = 8.0, 4.0 µg/mL). Compound 6c also exerted superior activity to compound 5k and fluconazole in inhibiting hyphae growth of C. albicans and inhibiting drug-resistant strains of C. albicans, and it could reduce fungal burdens in mice kidney at a dosage of 1.0 mg/kg. An in vivo efficacy evaluation indicated that 6c could effectively protect mice models from C. albicans infection at doses of 0.5, 1.0, and 2.0 mg/kg. These results suggested that compound 6c deserves further investigation.