Apigenin And Ε-Viniferin from Dioscorea Transversa Leaves: Isolation and in Silico Evaluation Against HPV16 E6 And Inflammatory and Angiogenic Targets

Abstract

Background and objectives: Long-term infection with HPV16 is the main cause of cervical cancer, and the viral E6 protein contributes by recruiting the host ligase E6AP, which marks p53 for destruction. Leaf extracts of Dioscorea transversa R. Br. have been reported to contain more polyphenols and to show stronger anti-inflammatory and pro-apoptotic effects than tuber extracts, but the compounds responsible were not known. This study aimed to identify them and to use molecular docking to examine how well they bind two sites on HPV16 E6 (the E6AP-binding site and the p53-binding site), as well as COX-2, TNF-α and VEGFR2.

Materials and methods: Aqueous-methanolic leaf and tuber extracts were compared by disc-diffusion assay and thin-layer chromatography. Bands present only in the leaf extract were purified by column chromatography and recrystallisation, and characterised by melting point, FTIR and HPLC co-injection with authentic standards. A library of 25 flavonoids and stilbenoids was docked using AutoDock Vina against the E6 oncoprotein of HPV16 at its E6AP- and p53-binding sites, the primary targets. Paclitaxel, hypericin and anabsinthin, previously reported as E6 ligands, were docked as reference compounds. The same 25 compounds were also docked against the secondary targets COX-2, TNF-α and vascular endothelial growth factor receptor 2 (VEGFR2), proteins involved in inflammatory and angiogenic signalling, with the respective co-crystallised ligands of each target as reference compounds. Physicochemical properties and the predicted clinical toxicity (ClinTox) probability of the two-site binders and the reference compounds were calculated in silico.

Results: The leaf extract gave wider inhibition zones than the tuber extract against S. aureus (13 vs 11 mm), E. coli (10 vs 9 mm) and C. albicans (14 vs 9 mm). The two purified compounds were identified as apigenin and ε-viniferin. Both scored better than paclitaxel at the two E6 sites (E6AP site/p53 site: apigenin −7.370/−6.098; ε-viniferin −7.900/−6.207 kcal/mol), although hypericin and anabsinthin scored better at the E6AP site. Apigenin also bound COX-2, TNF-α and VEGFR2 (−9.964, −9.061 and −9.825 kcal/mol), whereas ε-viniferin was largely limited to E6. Apigenin met all four Lipinski criteria and had a low predicted ClinTox probability (0.024).

Conclusion: Apigenin and ε-viniferin from D. transversa leaf emerge as computational leads against HPV16 E6, and apigenin may in addition act on inflammatory and angiogenic targets.