The eradication of HIV-1 from infected individuals is prevented by the persistence of the virus in a stable reservoir of latently infected CD4+ T cells. Latently infected cells can be found in all HIV-1 infected individuals at a very low frequency and allow the virus to persist despite antiretroviral therapy for the lifetime of an infected patient. Current efforts are focused on identifying small molecules or immune strategies to eliminate these latently infected cells. To assess the efficacy of these elimination strategies in HIV-1 infected patients, we must be able to measure the size of the remaining latent reservoir. While a previous assay can measure the size of this latent reservoir, it is too laborious and costly to be utilized in large-scale HIV-1 eradication trials. A new paper in PLoS Pathogens describes a rapid assay to measure the size of the HIV-1 latent reservoir more amenable to eradication trials.
Rapid Quantification of the Latent Reservoir for HIV-1 Using a Viral Outgrowth Assay. (2013) PLoS Pathog 9(5): e1003398. doi:10.1371/journal.ppat.1003398
HIV-1 persists in infected individuals in a stable pool of resting CD4+ T cells as a latent but replication-competent provirus. This latent reservoir is the major barrier to the eradication of HIV-1. Clinical trials are currently underway investigating the effects of latency-disrupting compounds on the persistence of the latent reservoir in infected individuals. To accurately assess the effects of such compounds, accurate assays to measure the frequency of latently infected cells are essential. The development of a simpler assay for the latent reservoir has been identified as a major AIDS research priority. We report here the development and validation of a rapid viral outgrowth assay that quantifies the frequency of cells that can release replication-competent virus following cellular activation. This new assay utilizes bead and column-based purification of resting CD4+ T cells from the peripheral blood of HIV-1 infected patients rather than cell sorting to obtain comparable resting CD4+ T cell purity. This new assay also utilizes the MOLT-4/CCR5 cell line for viral expansion, producing statistically comparable measurements of the frequency of latent HIV-1 infection. Finally, this new assay employs a novel quantitative RT-PCR specific for polyadenylated HIV-1 RNA for virus detection, which we demonstrate is a more sensitive and cost-effective method to detect HIV-1 replication than expensive commercial ELISA detection methods. The reductions in both labor and cost make this assay suitable for quantifying the frequency of latently infected cells in clinical trials of HIV-1 eradication strategies.