Methylindoles and methoxyindoles are agonists and antagonists of human aryl hydrocarbon receptor

Novel methylindoles were identified as endobiotic and xenobiotic ligands of the human aryl hydrocarbon receptor (AhR). We examined the effects of 22 methylated and methoxylated indoles on the transcriptional activity of AhRs. Employing reporter gene assays in AZ-AHR transgenic cells, we determined full agonist, partial agonist, or antagonist activities of tested compounds, having substantially variable EC₅₀, IC₅₀, and relative efficacies. The most effective agonists (E_MAX relative to 5 nM dioxin) of the AhR were 4-Me-indole (134%), 6-Me-indole (91%), and 7-MeO-indole (80%), respectively. The most effective antagonists of the AhR included 3-Me-indole (IC₅₀; 19 mM), 2,3-diMe-indole (IC₅₀; 11 mM), and 2,3,7-triMe-indole (IC₅₀; 12 mM). Reverse transcription polymerase chain reaction analyses of CYP1A1 mRNA in LS180 cells confirmed the data from gene reporter assays. The compound leads, 4-Me-indole and 7-MeO-indole, induced substantial nuclear translocation of the AhR and enriched binding of the AhR to the CYP1A1 promoter, as observed using fluorescent immunohistochemistry and chromatin immunoprecipitation assays, respectively. Molecular modeling and docking studies suggest the agonists and antagonists likely share the same binding pocket but have unique binding modes that code for their affinity. Binding pocket analysis further revealed that 4-methylindole and 7-methoxyindole can simultaneously bind to the pocket and produce synergistic interactions. Together, these data show a dependence on subtle and specific chemical indole structures as AhR modulators and furthermore underscore the importance of complete evaluation of indole compounds as nuclear receptor ligands.