Targeting the latent HIV-1 reservoir with synergistic combinations of latency reversing agents identified using genome-wide CRISPR screens

Reversing HIV-1 latency allows killing of infected cells in cure strategies, but no single latency reversing agent (LRA) has been shown to effectively reduce HIV-1 latent reservoir size in persons living with HIV-1 (PLWH). Here we describe an approach to systematically identify synergistic LRA combinations to reactivate latent HIV-1 using a novel in vitro HIV-1 latency model and genome-wide CRISPR screens. Screens on cells treated with suboptimal concentrations of an LRA can identify host genes whose knockout enhances viral gene expression. Therefore, inhibitors of these genes should synergize with the LRA. We tested this approach using AZD5582, an inhibitor of the non-canonical NF-kappa B (ncNF-kB) pathway, as an LRA and identified HDAC2, a histone deacetylation complex blocked by some HDAC inhibitors, and BRD2, part of the Bromodomain and Extra-Terminal motif (BET) protein family targeted by BET inhibitors. Using CD4+ T cells from individuals on antiretroviral therapy, we confirmed synergy between AZD5582 and several HDAC inhibitors and between AZD5582 and the BET inhibitor JQ1. Remarkably, a reciprocal screen using suboptimal concentrations of an HDAC inhibitor as an LRA identified ncNF-kB regulators, especially BIRC2, as druggable synergistic candidates for use in combination with HDAC inhibition, confirming the validity of this approach. Our studies provide novel insights into the roles of host factors in HIV-1 reactivation and validate a system for finding drug combinations for HIV-1 latency reversal.