Abstract
Azole fungicides have been found to inhibit the activity of human 3β-hydroxysteroid dehydrogenase 1 (3β-HSD1), a key enzyme involved in steroid synthesis during normal pregnancy. However, the specific azole fungicides that exert this inhibitory effect and the structure-activity relationship (SAR) involved have not been fully elucidated. This study aimed to analyze the inhibitory potency, mode of action, SAR, and docking parameters of 9 azole fungicides on human 3β-HSD1 and its rat homolog 3β-HSD4. The results revealed that bromuconazole, climbazole, cyproconazole, and etaconazole A varying inhibitory potency against human 3β-HSD1, with IC50 values ranging from 6.21 µM (climbazole) to 104.83 µM (cyproconazole). These fungicides were identified as mixed/noncompetitive inhibitors of human 3β-HSD1. Among them, climbazole and bromuconazole were found to inhibit progesterone secretion by human JAr cells at 20 and 40 µM. It was also observed that these fungicides significantly inhibited the activity of rat placental 3β-HSD4, although with reduced potency compared to their effect on human 3β-HSD1. Furthermore, docking analysis indicated that the azole fungicides bind to the NAD+/steroid binding site via hydrogen bonds, hydrophobic, and van der Waals interactions. The study demonstrated a negative correlation between LogP and IC50 value for 3β-HSD1, indicating that the lipophilicity of the chemicals plays a critical role in their inhibitory effect. Additionally, 3D-QSAR analysis highlighted the significance of hydrogen bond donor and acceptor in the binding process. In conclusion, it was shown that certain azole fungicides exhibit inhibitory effects on human 3β-HSD1, and this inhibition is dependent on their lipophilicity. These findings suggest that these fungicides could potentially act as potential endocrine disruptors in the context of steroid synthesis during pregnancy.