Latently infected CD4-positive memory T cells are capable of producing replication competent virus and represent a barrier to cure. These cells were presumably infected while responding to specific antigen. We hypothesized that such cells would respond again to that antigen and in particular to HIV-1 antigens. By measuring latency reversal and cellular activation in cells from virally suppressed patients ex vivo using specific antigens, we aimed to show that latency is preferentially established in cells with certain antigen-specificities.
We used peptide pools of HIV-1 Gag, CMV, C. albicans, and a mixture of peptides from CMV, EBV, influenza, and tetanus toxoid (CEFT) to stimulate primary CD8-depleted PBMC from HIV-1 infected patients who were virally suppressed by cART for at least 1 year. Cells were stimulated for 3 days with each agent and then stained for expression of the early activation marker CD69. To assess latency-reversal, we measured virion-associated RNA released by the cells 3 days after activation; we also added MOLT4/R5 cells to the cultures and measured RNA after 7 days of viral outgrowth. Real-time RT-PCR of gag RNA was used as the readout.
Preliminary results indicate that cellular activation was greatest in cells stimulated with beads coated with α-CD3 and α-CD28, followed by the peptide pools of CMV, CEFT, HIV-1 Gag, and C. albicans. The latency-reversal activity of the peptide pools was less than that of the antibody-coated beads, and the different antigens were not uniformly active among the different patients. In some patients the HIV-1 Gag peptide pool was the most active at latency reversal, even though it was less active at inducing CD69 expression than other antigens such as CMV.
Our results suggest the potential for latency to be differentially established in cells with specific antigen specificities; in some patients latency seems to be preferentially established in cells that respond to Gag peptides. By defining the specific antigens that can reactivate viral gene expression in latently infected cells, we aim to inform models of latency and in the best case derive an antigenic mixture capable of reversing latency in the majority of cells.