Development of a NS2B/NS3 protease inhibition assay using AlphaScreen beads for screening of anti-dengue activities.

Background: Dengue infection is an endemic infectious disease and it can lead to dengue fever, dengue hemorrhagic fever, and/or dengue shock syndromes. Dengue NS2B/NS3 protease complex is essential for viral replication and is a primary target for anti-dengue drug development. In this study, a NS2B/NS3 protease inhibition assay was developed using AlphaScreen beads and was used to screen compounds for their protease inhibition activities.

Methods: The assay system utilized a known NS2B/NS3 peptide substrate, a recombinant of NS2B/NS3 protease with proprietary StrepTactin donor and nickel chelate acceptor beads in 384-well format.

Results: The optimized assay to screen for NS2B/NS3 protease inhibitors was demonstrated to be potentially useful with reasonable z' factor, coefficient variance and signal to background ratio. However, screening of synthesized thioguanine derivatives using the optimized AlphaScreen assay revealed weak NS2B/NS3 inhibition activities.

Conclusion: The AlphaScreen assay to screen for NS2B/NS3 protease inhibitors is potentially applicable for high throughput screening.