ABSTRACT
Parasitic nematodes infect over 1/4 th of the human population and are a major burden on livestock and crop production. Benzimidazole class anthelmintics are widely used to treat infections, but resistance is a widespread problem. Mutation of genes encoding the benzimidazole target ß-tubulin is a well-established mechanism of resistance, but recent evidence suggests that metabolism of the drugs may also occur. Our objective was to investigate contributions of the detoxification-response transcription factor SKN-1 to anthelmintic drug resistance using C. elegans. We find that skn-1 mutations alter EC50 of the common benzimidazole albendazole in motility assays by 1.5-1.7 fold. We also identify ugt-22 as a detoxification gene associated with SKN-1 that influences albendazole efficacy. Mutation and overexpression of ugt-22 alter albendazole EC50 by 2.3-2.5-fold. The influence of a nematode UGT on albendazole efficacy is consistent with recent studies demonstrating glucose conjugation of benzimidazoles.
Subject(s)
Albendazole/pharmacology , Caenorhabditis elegans Proteins/genetics , Caenorhabditis elegans/drug effects , Caenorhabditis elegans/genetics , DNA-Binding Proteins/genetics , Inactivation, Metabolic/genetics , Transcription Factors/genetics , Albendazole/metabolism , Animals , Anthelmintics/metabolism , Anthelmintics/pharmacology , Caenorhabditis elegans Proteins/metabolism , Drug Resistance/genetics , Mutation , Tubulin/geneticsABSTRACT
Nematodes parasitize â¼1/3 of humans worldwide, and effective treatment via administration of anthelmintics is threatened by growing resistance to current therapies. The nematode transcription factor SKN-1 is essential for development of embryos and upregulates the expression of genes that result in modification, conjugation, and export of xenobiotics, which can promote resistance. Distinct differences in regulation and DNA binding relative to mammalian Nrf2 make SKN-1 a promising and selective target for the development of anthelmintics with a novel mode of action that targets stress resistance and drug detoxification. We report 17 (ML358), a first in class small molecule inhibitor of the SKN-1 pathway. Compound 17 resulted from a vanillamine-derived hit identified by high throughput screening that was advanced through analog synthesis and structure-activity studies. Compound 17 is a potent (IC50 = 0.24 µM, Emax = 100%) and selective inhibitor of the SKN-1 pathway and sensitizes the model nematode C. elegans to oxidants and anthelmintics. Compound 17 is inactive against Nrf2, the homologous mammalian detoxification pathway, and is not toxic to C. elegans (LC50 > 64 µM) and Fa2N-4 immortalized human hepatocytes (LC50 > 5.0 µM). In addition, 17 exhibits good solubility, permeability, and chemical and metabolic stability in human and mouse liver microsomes. Therefore, 17 is a valuable probe to study regulation and function of SKN-1 in vivo. By selective targeting of the SKN-1 pathway, 17 could potentially lead to drug candidates that may be used as adjuvants to increase the efficacy and useful life of current anthelmintics.