Robustaflavone as a novel scaffold for inhibitors of native and auto-proteolysed human neutrophil elastase

Human neutrophil elastase (HNE) plays a key role in initiating inflammation in the cardiopulmonary and systemic contexts. Pathological auto-proteolysed two-chain (tc) HNE exhibits reduced binding affinity with inhibitors. Using AutoDock Vina v1.2.0, 66 flavonoid inhibitors, sivelestat and alvelestat were docked with single-chain (sc) HNE and tcHNE. Schrodinger PHASE v13.4.132 was used to generate a 3D-QSAR model. Molecular dynamics (MD) simulations were conducted with AMBER v18. The 3D-QSAR model for flavonoids with scHNE showed r² = 0.95 and q² = 0.91. High-activity compounds had hydrophobic A/A2 and C/C2 rings in the S1 subsite, with hydrogen bond donors at C5 and C7 positions of the A/A2 ring, and the C4' position of the B/B1 ring. All flavonoids except robustaflavone occupied the S1'-S2' subsites of tcHNE with decreased AutoDock binding affinities. During MD simulations, robustaflavone remained highly stable with both HNE forms. Principal Component Analysis suggested that robustaflavone binding induced structural stability in both HNE forms. Cluster analysis and free energy landscape plots showed that robustaflavone remained within the sc and tcHNE binding site throughout the 100 ns MD simulation. The robustaflavone scaffold likely inhibits both tcHNE and scHNE. It is potentially superior to sivelestat and alvelestat and can aid in developing therapeutics targeting both forms of HNE.