Abstract The Multi‐Threat Medical Countermeasure (MTMC) technique is crucial for developing common biochemical signaling pathways, molecular mediators, and cellular processes. This study revealed that the Nod‐like receptor 3 (NLRP3) inflammasome pathway may be a significant contributor to the cytotoxicity induced by various organophosphorus pesticides (OPPs). The study demonstrated that exposure to six different types of OPPs (paraoxon, dichlorvos, fenthion, dipterex, dibrom, and dimethoate) led to significant cytotoxicity in BV2 cells, which was accompanied by increased expression of NLRP3 inflammasome complexes (NLRP3, ASC, Caspase‐1) and downstream inflammatory cytokines (IL‐1β, IL‐18), in which the order of cytotoxicity was dichlorvos > dipterex > dibrom > paraoxon > fenthion > dimethoate, based on the IC50 values of 274, 410, 551, 585, 2,158, and 1,527,566 μM, respectively. The findings suggest that targeting the NLRP3 inflammasome pathway could be a potential approach for developing broad‐spectrum antitoxic drugs to combat multi‐OPPs‐induced toxicity. Moreover, inhibition of NLRP3 efficiently protected the cells against cytotoxicity induced by these six OPPs, and the expression of NLRP3, ASC, Caspase‐1, IL‐1β, and IL‐18 decreased accordingly. The order of NLRP3 affinity for OPPs was dimethoate > paraoxon > dichlorvos > dibrom > (fenthion and dipterex) based on KD values of 89.8, 325, 1,460, and 2,690 μM, respectively. Furthermore, the common molecular mechanism of NLRP3‐OPPs was clarified by the presence of toxicity effector groups (benzene ring, nitrogen/oxygen‐containing functional group); =O, ‐O‐, or =S (active) groups; and combination residues (Gly271, Asp272). This finding provided valuable insights into exploring the common mechanisms of multiple threats and developing effective therapeutic strategies to prevent OPPs poisoning.
