An In Vivo Zebrafish Screen Identifies Organophosphate Antidotes with Diverse Mechanisms of Action

Abstract

Organophosphates are a class of highly toxic chemicals that includes many pesticides and chemical weapons. Exposure to organophosphates, either through accidents or acts of terrorism, poses a significant risk to human health and safety. Existing antidotes, in use for over 50 years, have modest efficacy and undesirable toxicities. Therefore, discovering new organophosphate antidotes is a high priority. Early life stage zebrafish exposed to organophosphates exhibit several phenotypes that parallel the human response to organophosphates, including behavioral deficits, paralysis, and eventual death. Here, we have developed a high-throughput zebrafish screen in a 96-well plate format to find new antidotes that counteract organophosphate-induced lethality. In a pilot screen of 1200 known drugs, we identified 16 compounds that suppress organophosphate toxicity in zebrafish. Several in vitro assays coupled with liquid chromatography/tandem mass spectrometry–based metabolite profiling enabled determination of mechanisms of action for several of the antidotes, including reversible acetylcholinesterase inhibition, cholinergic receptor antagonism, and inhibition of bioactivation. Therefore, the in vivo screen is capable of discovering organophosphate antidotes that intervene in distinct pathways. These findings suggest that zebrafish screens might be a broadly applicable approach for discovering compounds that counteract the toxic effects of accidental or malicious poisonous exposures.

15 years of zebrafish chemical screening

“…Chemically induced changes in metabolomic profiles, for example, could be used as phenotypic endpoints in zebrafish chemical screens [73, 74] in addition to epigenetic, gene expression or proteomic readouts.…”

“…Our group has recently used antidote screening to identify antidotes for organophosphate poisoning [74], cyanide toxicity [73], and doxorubicin-induced cardiomyopathy [50]. Others have identified compounds able to counteract the hair-damaging effects of ototoxic drugs, such as the aminoglycosidic antibiotic neomycin [75].…”

Zebrafish as a model for acetylcholinesterase‐inhibiting organophosphorus agent exposure and oxime reactivation

“…A number of publications have described the developmental consequences of exposure to subacute concentrations of AChE‐inhibiting OP pesticides in embryonic, larval, and adult zebrafish, such as chlorpyrifos/chlorpyrifos oxon, azinphos‐methyl, parathion/paraoxon, malathion, dichlorvos, and diazinon . The major observations from a number of these studies are summarized in Table .…”

Changing the scale and efficiency of chemical warfare countermeasure discovery using the zebrafish

“…Building upon several studies of AChE function and organophosphate toxicity on zebrafish models 52, 53 , conditions were identified in which the prototypical organophosphate azinphos-methyl produced consistent behavioral deficits, paralysis, and death in early life stage zebrafish. Using death as an endpoint, a pilot screen of 1200 known drugs was conducted in 96-well format 54 . Sixteen drugs exhibiting the ability to prevent death in organophosphate-exposed zebrafish were discovered.…”