Novel [1,2,4]-Triazolo[3,4-]-[1,3,4]thiadizoles as Potent Pyruvate Kinase Inhibitors for Fungal Control.

To discover novel target-based fungicidal candidates, a molecular design model was established with a three-dimensional (3D) structure of pyruvate kinase (RsPK) simulated with the AlphaFold 2 and as a fungicidal lead. A series of novel [1,2,4]triazolo[3,4-][1,3,4]thiadiazole derivatives were rationally designed, synthesized, evaluated for their fungicidal performance, and validated for their mode of action. The bioassays with indicated that compounds , , and with an EC value ranging from 1.01 to 1.54 μg/mL displayed higher fungicidal activity than the positive control with its EC of 3.14 μg/mL. Especially, exhibited high potency and a broad spectrum against , , , , , and with its EC value falling between 1.54 and 13.10 μg/mL. Like all positive controls, , , and showed excellent growth inhibition against at 200 μg/mL. Even though the PK enzymatic inhibition assay showed that was approximately 2.6 times less active than (IC: 29.14 11.15 μg/mL, respectively), the similar fluorescence quenching patterns of RsPK by and , and the docking results of interactions between RsPK and or implied that they might share the similar binding site in the RsPK active pocket. Our studies suggested that could be used as a potent fungicidal lead for further optimization. The results of comparative molecular field analysis (CoMFA) provided a direction for further molecular design.