CROI 2017 Abstract e-Book

Abstract eBook

Poster and Themed Discussion Abstracts

310

LATENCY REVERSAL USING SPECIFIC ANTIGENS INCLUDING THOSE OF HIV-1 Thomas Vollbrecht 1 , Aaron Angerstein 2 , John Guatelli 2 1 Veterans Med Rsr Fndn, San Diego, CA, USA, 2 Univ of California San Diego, La Jolla, CA, USA

Background: 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. Methods: 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. Results: 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. Conclusion: 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 informmodels of latency and in the best case derive an antigenic mixture capable of reversing latency in the majority of cells. 311 IDENTIFICATION OF LRAs ACTIVE IN DIVERSE PRIMARY T-CELL MODELS OF HIV LATENCY Richard Barnard 1 , Jenny L. Anderson 2 , Daria Hazuda 1 , Sharon R. Lewin 2 , Laura J. Martins 3 , Mauricio Montano 4 , Vicente Planelles 3 , Celsa A. Spina 5 , for the CARE Collaboratory 1 Merck & Co, Inc, West Point, PA, USA, 2 The Univ of Melbourne, Melbourne, Australia, 3 Univ of Utah, Salt Lake City, UT, USA, 4 Univ of California San Francisco, San Francisco, CA, USA, 5 VA San Diego Hlthcare System, San Diego, CA, USA Background: It has been challenging to identify latency reversing agents (LRAs) broadly active across multiple primary cell latency models and patient cells ex vivo. A Jurkat CD4+ T cell model of HIV latency was used to screen the Merck library of 2.9 million compounds. Unlike prior efforts, this screen was conducted with and without a suboptimal concentration of SAHA, a histone deacetylase inhibitor (HDACi), with the goal of sensitizing the latency model to allow the discovery of novel and potentially synergistic LRAs. While 34% of compounds active in the screen acted via known mechanisms of action (HDACis or Farnesyl Transferase (FT) inhibitors) the remaining 66% acted via unknown mechanisms of action (uMOA). Based on the activity of uMOA compounds in different Jurkat T- cell models of HIV latency, 144 compounds were chosen for additional detailed analysis in primary CD4 T-cell models of HIV latency. Methods: The 144 compounds were analyzed in seven different primary CD4 T-cell models of HIV latency using up to a six-point dose titration, in the presence of suboptimal concentration of panobinostat or vorinostat, tailored to the HDACi responsiveness of each primary cell system. Latency reversal was quantified by changes in reporter expression (eGFP or luciferase), HIV Nef or Tat gene expression, or HIV mRNA, depending model used. Results: As expected based on previous studies of LRAs in these primary cell models (Spina, PLoS Path 2013) among the 144 selected uMOA compounds, no one tested was active in all models. However, 2 compounds displayed statistically significant activity in six of the seven cell models and 16 compounds exhibited activity in three or more systems. Of note, none of the compounds were overtly toxic at concentrations that exhibited LRA activity. Additionally, no compounds induced CD69 expression in naïve T-cells isolated from healthy volunteers, suggesting their LRA activity was not due to generalized T cell activation. Conclusion: This novel screen was able to identify LRAs that lack generalized T cell activating properties and appear non-toxic, but display activity across multiple cell line and primary cell models of latency. The compounds appear to have mechanism(s) of action distinct from known LRAs and work in concert with HDACis, suggesting the opportunity for combination LRA therapy. Validation of these compounds in patient cells is now underway. Screening for LRAs in the presence of a sensitizing agent may be a broadly useful approach. 312 STIMULATING THE RIG-I PATHWAY TO KILL LATENT HIV-INFECTED CELLS USING ACITRETIN Peilin Li 1 , Philipp Kaiser 1 , Harry Lampiris 1 , Peggy Kim 2 , Steven A. Yukl 1 , Diane V Havlir 3 , Warner C. Greene 4 , Joseph K. Wong 1 1 San Francisco VA Med Cntr/Univ of California San Francisco, San Francisco, CA, USA, 2 San Francisco VA Med Cntr, San Francisco, CA, USA, 3 San Francisco General Hosp, Univ of California San Francisco, San Francisco, CA, USA, 4 Gladstone Inst of Virology and Immunol, San Francisco, CA, USA Background: The persistence of latent HIV proviruses in long-lived CD4+ T cells despite ART represents a major obstacle to viral eradication. Because current candidate latency- reversing agents (LRAs) induce HIV transcription but appear to fail to clear these cellular reservoirs, new approaches for killing reactivated latent HIV reservoir cells should be

Poster and Themed Discussion Abstracts

CROI 2017 122