Xanthorrhizol derivatives as hyaluronidase and lipoxygenase inhibitors : design, synthesis, in vitro and in silico analyses

Abstract

Hyaluronidase (Hyal) and lipoxygenase (LOX) enzymes produced inflammatory inducers in the human body, making them suitable target for the design of anti-inflammatory agents. Xanthorrhizol (XNT) is a sesquiterpenoid isolated from Curcuma xanthorrhiza that possesses anti-inflammatory properties and holds promise as therapeutic candidate. The potential of XNT as anti-inflammatory agent can be further explored through modification of its structure. The use of computational tools can optimise the structure of XNT by precisely targeting Hyal and LOX and enhance its anti-inflammatory action. The objective of this study was to virtually screen and design potential XNT derivatives as new Hyal-1 and LOX-3 inhibitors from in-house database and in silico fragment-based drug design (FBDD) approaches, respectively. Then, the promising derivatives from both approaches were synthesised and characterised, followed by biological evaluation through in vitro enzyme inhibitory assays. The most active derivative against the enzyme in in vitro studies was subjected to further analysis using molecular dynamics (MD) simulation. Thirty XNT derivatives obtained from in-house database were virtually screened against Hyal-1 enzyme via molecular docking. Benzyl xanthorrhizyl ether (76) exhibited stronger binding energy (-8.0 kcal/mol) against Hyal-1 enzyme compared to XNT (-6.8 kcal/mol) and proposed to be synthesised as Hyal-1 inhibitor. Additionally, the in silico FBDD generated five XNT derivatives as potential Hyal-1 inhibitors, specifically (2-pyridinyl)methyl xanthorrhizyl ether (102), 4-nitrobenzyl xanthorrhizyl ether (103), 3-trifluoromethylbenzyl xanthorrhizyl ether (104), (2-tetrahydro-2H-pyranyl)methyl xanthorrhizyl ether (105) and (2-tetrahydrofuranyl)methyl xanthorrhizyl ether (106). Among the six proposed inhibitors for Hyal-1 enzyme, five derivatives (76, 102-105) were successfully synthesised and characterised using spectroscopic analyses. Meanwhile, the synthesis of derivative (106) yielded an unexpected product, characterised as propyl xanthorrhizyloxyacetate (117). The in vitro analysis against Hyal-1 enzyme showed that derivative (102) was the most active against Hyal-1 enzyme with IC50 value of 44.54 µg/mL compared to XNT (IC50 = 203.56 µg/mL). The MD simulation revealed that derivative (102) displayed stable interaction in the binding site of Hyal-1 enzyme throughout 100 ns simulation. Derivative (76) was identified as potential LOX-3 inhibitor as it exerts stronger binding energy (-8.6 kcal/mol) compared to XNT (-7.7 kcal/mol). The design of XNT derivatives through in silico FBDD approach generated four XNT derivatives as LOX-3 inhibitors, however, their synthesis were unsuccessful. Derivative (76) was synthesised, and tested via in vitro LOX-3 inhibitory assay, where it displayed weaker activity (IC50 = 619.92 µg/mL) compared to XNT (IC50 = 142.34 µg/mL). These findings suggest that XNT derivatives, particularly (2-pyridinyl)methyl xanthorrhizyl ether (102) showed promising anti-inflammatory activities through inhibition of Hyal-1 enzyme.