Lipophilicity and Steric Constraints Govern V-Shaped Inhibition of Human and Rat Placental 3β-Hydroxysteroid Dehydrogenase by Dialkyldimethylammonium Disinfectants: Implications for Progesterone Dysregulation

Abstract Dialkyldimethylammonium chemicals (DDAs) are quaternary ammonium disinfectants. However, their effects on progesterone synthesis in placentas remain unclear. This study aims to study structure–activity relationship (SAR) and mechanism of DDAs (C2–C18) as inhibitors of human (3β-HSD1) and rat (3β-HSD4) placental 3β-hydroxysteroid dehydrogenase/Δ5-Δ4 isomerase (3β-HSD), a key enzyme in progesterone synthesis. DDAs exhibited a distinct V-shaped SAR to inhibit 3β-HSD, which was dependent on alkyl chain length. The inhibitory potency peaked with the C12 analogue, demonstrating the highest potency in human (IC50 = 4.78 μM) and rat (IC50 = 2.08 μM) assays. Conversely, a significant reduction in activity was observed with the C18 analogue, as evidenced by the substantially higher IC50 values (human: 19.1 μM; rat: 18.68 μM). Kinetic analyses confirmed mixed/noncompetitive mechanisms via NAD+/steroid interface binding, supported by molecular docking and 3D-QSAR pharmacophores highlighting hydrophobic interactions. Cellular assays in JAr cells demonstrated reduced progesterone secretion. Network toxicology analysis identifies human 3β-HSD1 as a central hub within a pathogenic network for recurrent spontaneous abortion, suggesting its dysregulation to disrupted progesterone metabolism and key signaling pathways. These findings propose DDAs as novel placental 3β-HSD inhibitors, with implications for endocrine-disrupting effects.