In Vitro Activities of the Novel Investigational Tetrazoles VT-1161 and VT-1598 Compared to the Triazole Antifungals against Azole-Resistant Strains and Clinical Isolates of Candida albicans

Abstract

The fungal Cyp51-specific inhibitors VT-1161 and VT-1598 have emerged as promising new therapies to combat fungal infections, including Candida spp. To evaluate their in vitro activities compared to other azoles, MICs were determined by Clinical and Laboratory Standards Institute (CLSI) method for VT-1161, VT-1598, fluconazole, voriconazole, itraconazole, and posaconazole against 68 C. albicans clinical isolates well characterized for azole resistance mechanisms and mutant strains representing individual azole resistance mechanisms. VT-1161 and VT-1598 demonstrated potent activity (geometric mean MICs ≤0.15 μg/ml) against predominantly fluconazole-resistant (≥8 μg/ml) isolates. However, five of 68 isolates exhibited MICs greater than six dilutions (>2 μg/ml) to both tetrazoles compared to fluconazole-susceptible isolates. Four of these isolates likewise exhibited high MICs beyond the upper limit of the assay for all triazoles tested. A premature stop codon in ERG3 likely explained the high-level resistance in one isolate. VT-1598 was effective against strains with hyperactive Tac1, Mrr1, and Upc2 transcription factors and against most ERG11 mutant strains. VT-1161 MICs were elevated compared to the control strain SC5314 for hyperactive Tac1 strains and two strains with Erg11 substitutions (Y132F and Y132F&K143R) but showed activity against hyperactive Mrr1 and Upc2 strains. While mutations affecting Erg3 activity appear to greatly reduce susceptibility to VT-1161 and VT-1598, the elevated MICs of both tetrazoles for four isolates could not be explained by known azole resistance mechanisms, suggesting the presence of undescribed resistance mechanisms to triazole- and tetrazole-based sterol demethylase inhibitors.

Keywords: Candida albicans; antifungal resistance; antifungal susceptibility testing; azole resistance; tetrazole.

FIG 1

Comparison of the MICs of VT-1161 and VT-1598 against the MICs of fluconazole (a), voriconazole (b), itraconazole (c), and posaconazole (d) in a collection of C. albicans clinical isolates. Plotted points represent the MICs of clinical isolates, with darker points representative of multiple, superimposed points. Concentration of points to the lower right of each plot represent favorable activity (low MICs relative to susceptible isolates) for VT-1161 or VT-1598 versus the comparator azole. Conversely, points concentrated to the top left of each plot represent isolates with high MICs of VT-1161 and VT-1598 relative to the comparator azole. Solid vertical lines represent the resistant clinical breakpoint for fluconazole and voriconazole, while dotted vertical lines represent the epidemiological cutoff values for posaconazole.

FIG 2

MICs of tested azole compounds against strains with individual known azole resistance mechanisms. Tested strains include those containing the artificially activated transcription factors Tac1 and Mrr1 in strains SCTAC1GAD1A and -B and SCMRR1GAD1A and -B, respectively, as well as Δcdr1 derivatives of SCTAC1GAD1A (SCΔcdr1TAC1GAD1A and -B), Δmdr1 derivatives of SCMRR1GAD1A and -B (SCΔmdr1MRR1GAD1A and -B), and SCUPC2R14A and -B containing the G648D gain-of-function mutation in UPC2. The MICs for the strains with artificially activated Tac1, Mrr1, and for the UPC2^G648D gain-of-function mutation are displayed as the highest MIC value of both independently created A- and B- strains for each respective transcription factor. The relative fold change in expression compared to the parent strain SC5314 of CDR1 for SCTAC1GAD1A and -B (A) and MDR1 for SCMRR1GAD1A and -B (B) is shown on the left of the figure. (C) Antifungal MICs of the UPC2^G648D homozygous strains SCUPC2R14A and -B.

FIG 3

Relative fold change compared to SC5314 in MIC of various azole antifungal agents against strains containing single and double ERG11 mutations. Open blue circles represent VT-1598 MICs, while open black circles represent VT-1161. Open gray diamonds represent fluconazole. Solid green triangles represent voriconazole. Solid orange diamonds represents itraconazole, and solid inverted purple triangles represent posaconazole.