The Intact Noninducible Latent HIV-1 Reservoir Is Established in an In Vitro Primary T CM Cell Model of Latency

Abstract

The establishment of HIV-1 latency has hindered an HIV-1 cure. “Shock and kill” strategies to target this reservoir aim to induce the latent provirus with latency-reversing agents (LRAs). However, recent studies have shown that the majority of the intact HIV-1 viral reservoir found in antiretroviral therapy (ART)-suppressed HIV-infected individuals is not inducible. We sought to understand whether this noninducible reservoir is established, and thus able to be studied, in an in vitro primary T_CM cell model of latency. Furthermore, we wanted to expand this model system to include R5-tropic and non-B-subtype viruses. To that end, we generated our T_CM cell model of latency with an R5 subtype B virus, AD8, and an R5 subtype C virus, MJ4. Our results demonstrate that both intact and defective proviruses are generated in this model. Less than 50% of intact proviruses are inducible regardless of viral strain in the context of maximal stimulation through the T cell receptor (TCR) or with different clinically relevant LRAs, including the histone deacetylase (HDAC) inhibitors suberoylanilide hydroxamic acid (SAHA) and MS-275, the protein kinase C (PKC) agonist ingenol 3,20-dibenzoate, or the second mitochondria-derived activator of caspase (SMAC) mimetic AZD-5582. Our findings suggest that current LRA strategies are insufficient to effectively reactivate intact latent HIV-1 proviruses in primary CD4 T_CM cells and that the mechanisms involved in the generation of the noninducible HIV-1 reservoir can be studied using this primary in vitro model. IMPORTANCE HIV-1 establishes a latent reservoir that persists under antiretroviral therapy. Antiretroviral therapy is able to stop the spread of the virus and the progression of the disease but does not target this latent reservoir. If antiretroviral therapy is stopped, the virus is able to resume replication and the disease progresses. Recently, it has been demonstrated that most of the latent reservoir capable of generating replication-competent virus cannot be induced in the laboratory setting. However, the mechanisms that influence the generation of this intact and noninducible latent reservoir are still under investigation. Here, we demonstrate the generation of defective, intact, and intact noninducible latent HIV-1 in a T_CM cell model of latency using different HIV-1 strains. Thus, the mechanisms which control inducibility can be studied using this primary cell model of latency, which may accelerate our understanding of the latent reservoir and the development of curative strategies.