Darunavir inhibits dengue virus replication by targeting the hydrophobic pocket of the envelope protein.

Dengue viruses (DENV) pose significant health threats, with no approved antiviral drugs currently available, creating an urgent need for new therapies. This study screened FDA-approved drugs for their antiviral ability against DENV and identified three promising candidates: darunavir (DRV), domperidone, and tetracycline. DRV demonstrated the highest efficacy against three DENV serotypes, with half-maximal effective concentrations (EC50) below 1 µM, surpassing the performance of tetracycline and domperidone. It effectively blocked DENV envelope (E) protein attachment to two type cells with EC50 values less than 0.2 μM. Domperidone reduced DENV-2 attachment to TE671 cells (EC50 = 3.08 μM) but was less effective in BHK-21 cells, while tetracycline inhibited NS3 protease (IC50 = 1.12 μM). Among DRV's structurally related drugs, fosamprenavir (FPV) significantly reduced DENV infectivity and virus yield, with EC50 values below 0.5 µM. In vivo, DRV at 1, 2, and 5 mg/kg achieved 100 % survival in suckling mice, compared to 83.5 % with FPV. Real-time RT-PCR showed DRV more effectively reduced DENV-2 RNA in mouse brains than FPV. Molecular docking showed DRV and FPV bind tightly to the DENV-2 E protein's N-octyl-β-D-glucoside (βOG) hydrophobic pocket, with DRV forming stronger interactions than FPV. Chimeric DENV-2 single-round infectious particle tests confirmed DRV's effective targeting of this pocket, though mutations at K128, L198, Q200, I270, and T280 reduced its efficacy. These findings highlight DRV as a potent antiviral agent against DENV, targeting the E protein's βOG hydrophobic pocket, with the potential for rapid deployment in treating and preventing infections.