(P03-009-23) Molecular Networking Identifies an AHR-modulating Benzothiazole From White Button Mushrooms

Objectives: This study predicted, identified, and characterized a novel Aryl hydrocarbon receptor (AHR) modulator in Agaricus bisporus, and introduced a protocol to identify more novel AHR modulators in other fungi with nutritional and medicinal significance.

Methods: Mass Spectrometry (MS) data from Agaricus bisporus methanol extract was processed through Global Natural Products Social Molecular Networking to predict the novel compound. AHR activation was measured by luciferase-based reporter assay on HepG2 40/6 and Hepa 1.1 cells. CYP1A1 gene expression was confirmed via RT-qPCR. Compound isolated by HPLC and identified by LC-MS and ¹H NMR. Unpaired t-test used to analyze in vitro experiments. Methanol extracts of Grifola frondosa, Ganoderma lucidum, Inonotus obliquus, Pleurotus ostreatus, Lentinula edodes, Hypsizygus tessellatus, and Hericium erinaceus were also screened for AHR activity with luciferase-based reporter assay.

Results: Using a molecular networking approach, a methylated analog to benzothiazole was indicated in Agaricus bisporus, which was subsequently isolated and identified as 2-amino-4-methyl-benzothiazole(2A4). Cell-based AHR transcriptional assays revealed that 2A4 possesses agonistic activity and upregulated CYP1A1 expression. Ganoderma lucidum, Pleurotus ostreatu, Lentinula edodes, and Hypsizygus tessellatus showed significant AHR modulating activity.

Conclusions: Results contrasts with previous findings of whole Agaricus bisporus extract having overall antagonistic activity in vivo, underscoring the importance of studying the roles each chemical component plays in a whole food. Molecular networking has potential to identify novel receptor modulators from natural products. Mushrooms that showed promising AHR modulating activity could now be screened for metabolites and novel compounds identified using this approach. Future results have potential to guide dietary advice in maintaining gut health and systemic health through the multitude of biological functions AHR regulates.

Funding Sources: Supported in part by the USDA’s Agricultural and Food Research Institute, USDA’s National Institute of Food and Agriculture’s Hatch Appropriations, NIH’s National Institute of General Medical Sciences and National Institute of Environmental and Health Sciences.