High throughput drug screening for human immunodeficiency virus type 1 reactivating compounds

Area of Science:

• Virology
• Immunology
• Drug Discovery

Background:

• Latent human immunodeficiency virus type 1 (HIV-1) in memory T cells is a major barrier to curative therapies.
• Latently infected cells are indistinguishable from uninfected cells, complicating reservoir elimination.
• Reactivation of latent HIV-1 followed by viral-induced cell death is a strategy to clear the viral reservoir.

Purpose of the Study:

• To develop a high-throughput screening (HTS) system for identifying compounds that reactivate latent HIV-1.
• To create reporter cell lines for quantitative assessment of HIV-1 reactivation and compound toxicity.

Main Methods:

• Development of latently HIV-1-infected reporter cell lines using enhanced green fluorescence protein (eGFP) for HIV-1 expression.
• Engineering cells to express a second red fluorescent protein for simultaneous cell viability assessment.
• Implementation of a 384-well plate-based HTS assay for automated drug screening.

Main Results:

• The reporter system enables direct and quantitative measurement of HIV-1 expression (eGFP) and cell viability.
• The assay demonstrated high dynamic range and reproducibility with a Z'-factor of 0.89, indicating a robust HTS system.
• A proof-of-concept screen of 1,600 compounds was performed, evaluating the system's utility.

Conclusions:

• The developed HTS system is robust and suitable for automated screening to identify HIV-1 latency-reversing agents.
• This reporter cell line platform facilitates the prioritization of compounds with a favorable therapeutic window for HIV-1 cure strategies.
• The system offers advantages and insights into limitations for future drug discovery efforts targeting the HIV-1 reservoir.