Trained Immunity as a Strategy for HIV Latency Reactivation

National Institute Of Allergy And Infectious Diseases

Investigators

Abstract

In our previous Office of AIDS Research (OAR)-funded projects on the application of trained immunity (TI), we had identified small molecules and metabolites with the ability to train macrophages. Some of the metabolites had previously been shown to modulate metabolism in macrophage cells and to reduce the H3K9Me3 and H3K27Me3 histone markers of transcriptional repression. They had also been shown to reduce inflammation in certain immune related diseases, and had been implicated in the regulation of pathogenesis of inflammatory disease, especially rheumatoid arthritis. The initial goal of this project was to test the reactivation of HIV from monocytic and T-cell reservoirs with the application of both established and novel TI stimuli. In line with this, we tested the reactivation of latently infected HIV in trained monocytic cell lines. As hypothesized, the HIV genome was reactivated with training molecules such as muramyl dipeptide (MDP), β-glucan or SYKi IV. Our preliminary studies also showed that trained immunity on myeloid cells like macrophages can facilitate reactivation of HIV from the T-cell reservoirs. We performed a co-culture analysis and found that secreted factors from the trained macrophages are responsible for the reactivation of HIV from the T-cell reservoirs. Our preliminary studies in PBMCs from PLWH showed that, after training, there was a significant reduction in HIV proviral DNA, alluding to the possibility that TI may, not only induce reactivation, but also potentially activate the immune cells such as NK cells or cytotoxic T-lymphocytes to eliminate the HIV-infected cells. In FY25, our studies further identify trained immunity as a novel and viable strategy to reverse HIV latency across both myeloid and lymphoid compartments, with potential to enhance immune-mediated clearance in vivo. Using multiple models—including THP-1–derived monocytic latency cells, primary macrophages from people living with HIV (PLWH) on ART, and SIV-infected microglia from ART-suppressed macaques—we demonstrate that trained immunity stimuli such as β-glucan (BG), muramyl dipeptide (MDP), and SYKi IV induce robust HIV/SIV RNA reactivation. Notably, MDP-driven reactivation in primary human macrophages and macaque microglia suggests this approach may be applicable to traditionally inaccessible or understudied reservoirs, including those in the central nervous system. We further show that trained innate cells influence neighboring CD4⁺ T cells, reactivating latent HIV through both soluble and membrane-bound signaling pathways. In particular, MHC class I–mediated interactions, ICAM1, and macrophage migration inhibitory factor (MIF) emerged as key drivers of latency reversal. Importantly, this reactivation occurred in patient-derived peripheral blood mononuclear cells (PBMCs), indicating that trained immunity can operate within complex, autologous immune environments. Trained immunity not only reactivates proviral transcription but also enhances antiviral immune function. Transcriptomic profiling of trained PBMCs revealed upregulation of interferon-stimulated genes, antigen presentation machinery, and effector molecules in monocytes, dendritic cells, T cells, and natural killer (NK) cells. These findings suggest that trained immunity can prime both innate and adaptive responses, potentially supporting reservoir recognition and clearance following latency reversal. Importantly, training with MDP or BG led to significant reductions in intact and defective proviral DNA in PBMCs from PLWH with higher reservoir burdens, as measured by the intact proviral DNA assay (IPDA). These results suggest that trained immunity may not only induce viral reactivation but also promote immune-mediated clearance of infected cells—critical features for any successful "shock and kill"–based HIV cure strategy. The clinical relevance of these findings is further underscored by the translational accessibility of our interventions. MDP and BG are components of licensed vaccines and immunotherapies already approved for human use, and SYKi IV targets pathways under active investigation in oncology and inflammatory diseases. This raises the possibility of repurposing or modifying these agents for clinical HIV cure trials. Several limitations warrant consideration. HIV reactivation was not observed in all patient-derived samples, and the determinants of responsiveness remain unclear. Additionally, although trained immunity induced broad immune activation, potential risks—including bystander activation or exacerbation of inflammation—must be carefully evaluated. Our future work will prioritize patient stratification, dosing strategies, and combination approaches with immune effectors or therapeutic vaccines. In conclusion, our findings establish trained immunity as a promising, mechanistically distinct latency-reversing strategy capable of targeting both myeloid and lymphoid HIV reservoirs. By simultaneously enhancing antigen presentation, intercellular signaling, and antiviral effector function, trained immunity may offer a dual-pronged approach to latency reversal and reservoir clearance—laying the groundwork for new interventions in the pursuit of an HIV cure.