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 zʹ 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.