Thioguanine-based DENV-2 NS2B/NS3 protease inhibitors: Virtual screening, synthesis, biological evaluation and molecular modelling

Abstract

Dengue virus Type 2 (DENV-2) is predominant serotype causing major dengue epidemics. There are a number of studies carried out to find its effective antiviral, however to date, there is still no molecule either from peptide or small molecules released as a drug. The present study aims to identify small molecules inhibitor from National Cancer Institute database through virtual screening. One of the hits, D0713 (IC50 = 62 μM) bearing thioguanine scaffold was derivatised into 21 compounds and evaluated for DENV-2 NS2B/NS3 protease inhibitory activity. Compounds 18 and 21 demonstrated the most potent activity with IC50 of 0.38 μM and 16 μM, respectively. Molecular dynamics and MM/PBSA free energy of binding calculation were conducted to study the interaction mechanism of these compounds with the protease. The free energy of binding of 18 calculated by MM/PBSA is -16.10 kcal/mol compared to the known inhibitor, panduratin A (-11.27 kcal/mol), which corroborates well with the experimental observation. Results from molecular dynamics simulations also showed that both 18 and 21 bind in the active site and stabilised by the formation of hydrogen bonds with Asn174.

Fig 1. Docked conformation of top 24 NCI compounds in the active pocket of DENV-2 NS2B/NS3pro.

The NS2B and NS3 domains are presented as surface form (blue area = NS2B, red area = NS3pro). Insets are the two ligands that bound to the NS2B, instead of NS3pro.

Fig 2. In vitro DENV-2 NS2B/NS3pro inhibition assay of Panduratin A and selected NCI compounds; NCI code D0265, D0685, D0227, D0152, D0126, D1804, D1855, D1498, D0713 and D1853 with D in the code stands for Diversity.

The assays contained 0 μM NCI compounds were taken as 0% protease inhibition.

Fig 3. Plot of % DENV-2 NS2B/NS3pro inhibition vs log concentration of the four NCI compounds.

Panduratin A was used as a control in this experiment.

Fig 4

Docked poses of (a) D1855, (b) D1498, (c) D0713 and (d) D1853. The NS2B and NS3pro domains are presented as surface form (blue area = NS2B, red area = NS3pro). The H-bond interactions are assigned as yellow dots inside the white circles.

Fig 5. The structure of thioguanine scaffold.

Fig 6. The log dose dependent curve of Compound 18 and 21 against DENV-2 NS2B/NS3pro.

Fig 7

Binding orientation and interaction mode of compound with compound 18 (A) and compound 21 (B) in molecular docking simulation.

Fig 8

(A) Time evolution of RMSD of NS2B/NS3pro backbone CA unbounded (Apo) (black), bounded with panduratin A (red), 18 (green) and 21 (blue). (B) Time evolution of radius of gyration of NS2B/NS3pro backbone CA unbounded (Apo) (black), bounded with panduratin A (red), 18 (green) and 21 (blue).

Fig 9

Binding orientation and interaction mode of compound with 18 (A), 21 (B) and panduratin A (C) in molecular dynamics simulation.