Evaluation of Fifteen 5,6-Dihydrotetrazolo[1,5- c]quinazolines Against Nakaseomyces glabrata: Integrating In Vitro Studies, Molecular Docking, QSAR, and In Silico Toxicity Assessments

Abstract

Nakaseomyces glabrata (Candida glabrata), the second most prevalent Candida pathogen globally, has emerged as a major clinical threat due to its ability to develop high-level azole resistance. In this study, two new 5,6-dihydrotetrazolo[1,5-c]quinazoline derivatives (c11 and c12) were synthesized and characterized using IR, LC-MS, ¹H, and ¹³C NMR spectra. Along with 13 previously reported analogues, these compounds underwent in vitro antifungal testing against clinical N. glabrata isolates using a serial dilution method (0.125-64 mg/L). Remarkably, compounds c5 and c1 exhibited potent antifungal activity, with minimum inhibitory concentrations of 0.37 μM and 0.47 μM, respectively-about a 20-fold improvement in μM concentration over standard drugs like amphotericin B, caspofungin, and micafungin. A detailed structure-activity relationship analysis revealed crucial molecular features enhancing antifungal potency. Extensive molecular docking studies across 18 protein targets explored potential binding pockets and affinities of the lead compounds. A robust 3D-QSAR model, incorporating molecular descriptors Mor26m and Mor29e, displayed good predictive ability for antifungal activity. In silico predictions indicated an absence of herbicidal effect, negligible environmental toxicity (to honeybees, avian species, and aquatic organisms), and mild human toxicity concerns for these compounds. This comprehensive approach aims to develop novel and effective antifungal compounds against the clinically relevant pathogen N. glabrata.

Keywords: 5,6-dihydrotetrazolo[1,5-c]quinazolines; Nakaseomyces glabrata; QSAR; antifungal activity; molecular docking; toxicity.

Figure 1

Examples of reported antifungal compounds targeting Candida species, with a focus on the studied 5,6-dihydrotetrazolo[1,5-c]quinazolines (numbering follows a previous antimicrobial study [36], and includes investigated substances with two additional novel ones, c11 and c12, for continuity).

Figure 2

Minimum inhibitory concentration (μM) of 5,6-dihydrotetrazolo[1,5-c]quinazolines (yellow color) and references (Mf: micafungin, Cf: caspofungin, AfB: amphotericin B; blue color). And their structure–activity relationship against N. glabrata. General molecular structure was optimized by HyperChem 8.0.8, and Discovery Studio v21.1.0.20298 was used for 3D visualization.

Figure 3

Visual 3D representation of the sterol uptake control protein 2 (PDB ID: 7VPR) with lead-compound c1 (Vina score −10.4 kkal/mol), showing bonds formation in its cavity of chain D. All ten formed bonds were hydrophobic: π-σ in blue color; π-π stacked in light blue color; alkyl in pink color; π-alkyl in orange color. Discovery Studio v21.1.0.20298 was used for 3D visualization.

Figure 4

Correlation graphs of predicted vs. experimental MIC (μM/mg/L) of model equations.

Figure 5

Calculated minnow toxicity (log LD₅₀, mg/kg/day, results were multiplied in 100 for the same scale; results below 30: high acute), rat acute toxicity (LD₅₀, mg/kg; results were divided in 10 for the same scale; results under 5: strong; 5–50: moderate; 50–500: slightly; over 500: safe), and rat chronic toxicity (lowest observed adverse effect level (LOAEL), mg/kg/day; results under 10: strong; 10–50: medium; over 50: weak).

Figure 6

Pearson coefficient of correlation between predicted affinity (Vina score, kcal/mol) to CYP51 (sterol 14-alpha demethylase, PDB ID: 5TZ1) [36] and toxicity (MT: minnow toxicity, log LD₅₀, mg/kg/day), RAT: rat acute toxicity (LD₅₀, mg/kg), RCT: rat chronic toxicity (LOAEL, mg/kg/day) [72].