A novel cell-based high-throughput screen for inhibitors of HIV-1 gene expression and budding identifies the cardiac glycosides

Objectives: Highly active antiretroviral therapy (HAART) is the mainstay of treatment for HIV-1 infection. While current HAART regimens have been extremely effective, issues of associated toxicity, cost and resistance remain and there is a need for novel antiretroviral compounds to complement the existing therapy. We sought to develop a novel high-throughput method for identifying compounds that block later steps in the life cycle not targeted by current therapy. Methods: We designed a high-throughput screen to identify inhibitors of post-integration steps in the HIV-1 life cycle. The screening method was applied to a library of compounds that included numerous FDA-approved small molecules. Results: Among the small molecules that inhibited late stages in HIV-1 replication were members of the cardiac glycoside family. We demonstrate that cardiac glycosides potently inhibit HIV-1 gene expression, thereby reducing the production of infectious HIV-1. We demonstrate that this inhibition is dependent upon the human Na. ⁺/K. ⁺-ATPase, but independent of cardiac glycoside-induced increases in intracellular Ca. ²⁺. Conclusions: We have validated a novel high-throughput screen to identify small molecule inhibitors of HIV-1 gene expression, virion assembly and budding. Using this screen, we have demonstrated that a number of FDA-approved compounds developed for other purposes potently inhibit HIV-1 replication, including the cardiac glycosides. Our work indicates that the entire cardiac glycoside family of drugs shows potential for antiretroviral drug development.