A fluorescence-based high-throughput screening assay to identify HIV-1 inhibitors

Highly active antiretroviral therapy (HAART) dramatically increases the long-term survival rates of human immunodeficiency virus type 1 (HIV-1) infected patients. Yet, poor adherence to therapy, adverse effects and the occurrence of resistant viruses can compromise the efficacy of HAART regiments. Therefore, there remains a clear unmet medical need for novel drugs and treatment options. In this chapter, we describe an HIV-1 antiviral high-throughput screening assay based on an HIV-1 permissive T lymphoblastoid MT4 cell line, stably transfected with a construct carrying an HIV-1 long terminal repeat promoter driving the expression of a reporter gene (enhanced green fluorescent protein). This assay runs in a 384-well format and enables the identification of HIV-1 inhibitors during a high-throughput screening campaign. In parallel, a cytotoxicity assay is performed to evaluate the compound-related in vitro toxicity.