P20. Optimization of HIV-1 latency-reversing agents (LRAs) combinations to improve HIV latency reactivation

Wang Zhang1, 2, Anders Sönnerborg1, Aman Russom2, Ujjwal Neogi1
Affiliates: Karolinska Institute1, Royal Institute of Technology2.

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Human immunodeficiency virus 1 (HIV) latency is a central challenge to cure HIV infection. One potential therapeutic cure approach is to reverse the latent cellular reservoir by activating latent HIV with latency-reversing agents (LRAs). However, because multiple mechanisms are involved in maintaining the latency of HIV-1, combinations of efficient and available LRAs targeting different pathways of latency may be necessary to achieve high levels of latency reversal. In this study, we tested and optimized different LRAs combinations and tried to find an optimal combination which is capable to achieve highest HIV latency reactivation with lowest cytotoxicity.

Three J-Lat cell culture models of HIV latency (J.Lat 6.3, J.Lat 9.2, J.Lat 10.6) were treated individually or in combination with T-cell receptor engager (αCD3/αCD28-conjugated beads and PHA), protein kinase C (PKC) agonists (TNFα, prostratin, bryostatin-1 and PMA), Calcium ionophore (Ionomycin) as well as Suberoylanilide hydroxamic acid (SAHA) which is known to inhibit Histone deacetylase (HDACi). The reactivation of latent provirus and cell viability was monitored by flow cytometry analysis of GFP expression and viability dye.

Seven stimuli shown in Figure 1 were chosen on the basis of their known or proposed activities in reactivating latent HIV-1 in various systems. Among that, all J-Lat clones were completely unresponsive to αCD3/αCD28 antibody-coated beads and PHA. Except PMA, TNFα and Prostratin are highly individually effective in reactivating HIV-latency in all three J-Lat cells models with relatively low cellular toxicity. The maximal response for all three J-Lat cells models was obtained with the combination of TNFα+Prostratin+Ionomycin, but with the optimal dose combination, not the individual optimal dose.

By using three J-Lat cell-based in vitro HIV-1 latency models, we observed that single LRA ineffectively reactivates HIV latency. However, the combination of three clinically LRAs—namely, TNFα+Prostratin+Ionomycin—overcomes the limitations of single-agent approaches and can achieve maximal reactivation while keeping relatively low cytotoxicity. Although the studied compounds cannot be used in vivo, our study provides important findings that will facilitate the strategic design of future clinical trials, the goal of which will be to reactivate latently infected cells optimally and to enhance the cytotoxic immunity that recognizes and kills those cells.