Discovery of therapeutic promising natural products to target Kv1.3 channel, a transmembrane protein regulating immune disorders, through multidimensional virtual screening, molecular dynamics simulations and biological validation.

Kv1.3 voltage-gated potassium channel, is a transmembrane protein that facilitates K movement through cell membranes via its intrinsic pores, regulating the cell signaling cascades, especially in immune disorders. In this paper, we employed multidimensional virtual screening to identify 24 potential Kv1.3 inhibitors from a library of 27,637 compounds, with electrophysiological assays confirming 8 active inhibitors (33.33 % hit rate). Structure-activity relationship (SAR) analysis demonstrated that 4-methylpentyl group in side chain and furan ring in Furanocoumarins skeleton are crucial to the bioactivity of target compounds. Orthogonal projection to latent structures model reveals that increasing the QPlogPo/w of the compound can increase activity. Molecular dynamics simulations revealed key roles of residues (VAL469 and ILE472) as active binding sites of Kv1.3 for binding of specific compound. Notopterol (Z4), the most potent Kv1.3 inhibitor (IC = 311.90 ± 1.24 nM), significantly suppressed IFN-γ release from CD4 T cells, whereas, Kv1.3 inactive compound Z20 at 5 μM showed no significant difference in IFN-γ release from CD4 T cells. In atopic dermatitis rat model, Notopterol reduced epidermal thickening, IgE, Kv1.3, IL-1β production, and infiltration of CD4 T cells and mast cells. These findings establish Notopterol as a promising Kv1.3 inhibitor for therapeutic applications in immune disorders.