Investigation of CYP2B6, 3A4 and β-esterase interactions of Withania somnifera (L.) dunal in human liver microsomes and HepG2 cells

• Published in: Journal of ethnopharmacology Vol. 270; p. 113766
• Main Authors: Kumar, Saneesh, Bouic, Patrick J., Rosenkranz, Bernd
• Format: Journal Article
• Language: English
• Published: Ireland Elsevier B.V 24.04.2021
• Subjects: CYP2B6; CYP3A4; Cytochrome P450; HepG2; Human liver microsomes; LC-MS; Phytoconstituents; Rifampicin; Withania; LC-MS; CYP2B6; Rifampicin; Withania; Cytochrome P450; Phytoconstituents; HepG2; Human liver microsomes; CYP3A4
• ISSN: 0378-8741; 1872-7573
• DOI: 10.1016/j.jep.2020.113766

Withania somnifera (L.) Dunal (Solanaceae) is a traditional herb, used in African indigenous systems of medicine for the treatment of various diseases (including HIV/AIDS and tuberculosis). The relevance of clinically significant interactions of Withania with ARVs and anti-TB drugs needs to be investigated. This study evaluated the effects of its roots on cytochromes P450 (CYPs) 2B6, 3A4, and rifampicin metabolism pathway, using methanol, ethanol, aqueous, and ethyl acetate solvent extractions. The extracts were tested on human liver microsomes (HLM) for CYP inhibition, mRNA expression in HepG2 cells for CYP induction. Biochemical qualitative tests and LC-MS/MS methodology were used to determine active phytoconstituents. The methanolic and ethyl acetate extracts inhibited CYP2B6 with IC50s 79.16 and 57.96 μg/ml respectively, while none of the extracts had any effect on rifampicin metabolism or showed time-dependant inhibition (TDI). All extracts were moderate inducers of CYP3A4; the aqueous extract exhibited 38%-fold shift induction of CYP3A4 compared to the control. The methanolic extract had the lowest CTC50 (50% of cytotoxicity inhibition) (67.13 ± 0.83 μg/ml). LC-MS/MS-PDA full scans were consistent with the presence of flavone salvigenin (m/z 327), alkaloid isopelletierine (m/z 133), steroidal lactone 2,3-dihydrowithaferin-A (m/z 472), and other withanolides including withaperuvin I (m/z 533), withaferin derivative (m/z 567), some of these compounds likely being responsible for the observed CYP2B6 inhibition and CYP3A4 induction. The putative gastrointestinal tract (GIT) concentration for the active extracts was 1800 μg/ml and the hepatic circulation concentrations were estimated at about 220 μg/ml and 13.5 μg/ml for the methanolic and ethyl acetate extracts, respectively. The extrapolated in vivo percentage of inhibition was at 85% for the methanolic extract against CYP2B6. The findings reported in this study suggest that W. somnifera extracts have the potential of causing clinically significant herb-drug interactions (HDI) as moderate inducer of CYP3A4 and inhibitor of CYP2B6 metabolism pathway (methanol and ethyl acetate extracts). [Display omitted] •Withania somnifera (L.) Dunal extracts were analyzed for CYP2B6, 3A4 and β-esterase interactions in HLM and HepG2 cells.•Methanol and ethyl acetate extracts inhibited CYP2B6 (IC50s 79.16 and 57.96 μg/ml); no inhibition of rifampicin metabolism.•The methanolic extract had the lowest CTC50 (50% of cytotoxicity inhibition) (67.13 ± 0.83 μg/ml).•All extracts moderately induced CYP3A4; aqueous extract showed 38%-fold shift induction of CYP3A4 compared to the control.•Presence of salvigenin (m/z 327), isopelletierine (m/z 133), withaperuvin I (m/z 533) and withaferin derivative (m/z 567).