Table 1_7-O-sulfonation redirects luteolin’s target preference from furin to NS2B/NS3 and enhances anti-dengue activity.docx

Luteolin, a natural flavonoid, exhibits anti-dengue activity but suffers from poor solubility and moderate potency. The 7-O position is the primary site of metabolic glucuronidation, whereas the 5-O position is sequestered by intramolecular hydrogen bonding, offering structurally distinct positions for derivatization.</p>Methods<p>Five site-selective derivatives of luteolin (three 5-O- and two 7-O-substituted) were synthesized.

Their anti-DENV activity was evaluated in Vero cells by qRT-PCR (measuring viral RNA abundance) and plaque assays, and cytotoxicity was determined using the CCK8 method. For mechanistic exploration, enzymatic inhibition kinetics against NS2B/NS3 and furin were measured, and target engagement was further assessed via molecular docking and cellular thermal shift assays (CETSA).</p>Results<p>Among the derivatives, compound 5, bearing a 4-fluorobenzenesulfonyl group at the 7-O position, exhibited the most potent anti-DENV activity (EC₅₀ = 0.46 μM) and the highest selectivity index (SI = 28.5), representing an 18-fold increase in antiviral potency over luteolin.