CROI 2025 Abstract eBook

Abstract eBook

Poster Abstracts

HIV from myeloid HIV reservoirs and possibly facilitate reactivation from CD4 T cell reservoirs. Methods: We used the established innate training stimuli MDP and β-glucan, along with Syk kinase inhibitors, to train the THP89GFP cell line (an experimental monocytic model for latent HIV-1), whole PBMCs of virally suppressed PLWH under ART, and macrophages derived from these PBMCs. We used qPCR and viral outgrowth assays to detect the presence of viral gene expression and viral production. We also performed single cell RNA sequencing and single cell ATAC seq on trained PBMCs of ART suppressed PLWH to study the cross talk between the myeloid and lymphoid cells and its effect on the reactivation of the CD4 T cell HIV genome. Results: All 3 trained immunity stimuli activated transcription of HIV-1 from THP89GFP cells. Consistent with the increased transcription of viral genes, we observed that multiple regions of the HIV-1 LTR had increased deposition of H3K27Ac in the trained THP89GFP macrophages. A viral outgrowth assay identified four out of 17 PBMC samples with detectable counts of HIV upon training with Syk inhibitors, MDP and β-glucan. Training of whole PBMCs of ART suppressed PLWH enhanced HIV specific viral gene expression. scRNA sequencing showed PBMC training-induced expression of HIV genes from CD4 T cells and hyper activation of T cells in response to PMA activation. Conclusions: Our studies show that induction of trained immunity is a viable and novel approach to the reactivation of latently infected HIV-1 from both T cell and macrophage reservoirs. Our data also shows beneficial cross talk between trained myeloid cells and T-lymphocytes in PBMC mixtures, resulting in the reactivation of HIV from T cell reservoirs. Collectively our data shows that the trained immunity strategy could be developed as a novel therapeutic approach for the reactivation of latently infected HIV.

post-infection and the scarcely detectable infected cells were observed by 22h after infection of untreated samples (p=0.0032). 2)No differences were observed in early and late RT in MDMs pretreated or not with dasatinib after infection with JR_FL_Renilla. However, 2-LTR circles and proviral integration were 3.9- (p=0.0259) and 1.6-fold (p=0.0467) lower in dasatinib-treated MDMs 48h post-infection, respectively. Similar results were obtained with ponatinib. 3)MDMs treated with dasatinib showed higher expression of CPSF6 in both nuclear (p<0.0001) and cytoplasmic regions (p<0.0001) than untreated MDMs, while ponatinib-treated MDMs also showed higher expression in both regions (p<0.0001) than untreated MDMs, and higher nuclear expression than dasatinib-treated MDMs (p=0.0110). 4)Ponatinib-treated MDMs had 5.8- and 5.4-fold lower expression of SC35 than dasatinib and untreated MDMs (p<0.0001). Conclusions: Dasatinib and ponatinib modified the expression and subcellular localization of CPSF6 and SC35, which are essential for HIV-1 proviral integration and transcription in MDMs. This is a new potential mechanism of action for TKIs to interfere with HIV-1 infection. Thymosin α1 Enhances IL-15 Pathway Between MoDCs and CD8+T Cells and Restrains HIV Latency In Vitro Chaoyu Chen 1 , Jingna Xun 2 , Jiangrong Wang 1 , Renfang Zhang 2 , Tangkai Qi 1 , Li Liu 1 , Xinyu Zhang 1 , Zichen Song 1 , Yinzhong Shen 1 , Hongzhou Lu 3 , Jun Chen 2 1 Fudan University, Shanghai, China, 2 Shanghai Public Health Clinical Center, Shanghai, China, 3 Shenzhen Third People's Hospital, Shenzhen, China Background: Viral reservoir presents a significant challenge in HIV-1 cure. Studies indicate that dendritic cells may limit HIV-1 reservoir by bolstering CD8+ T cell function via the IL-15 pathway and facilitating the expansion of virtual memory CD8+ T cells (TVM). We previously observed that Thymosin α1 (Tα1) may restrict the reservoir through the IL-15 pathway, however, the precise mechanism remains to be fully elucidated. Methods: Peripheral blood mononuclear cells (PBMCs) were obtained from individuals living with HIV-1 (PLWH). In vitro, THP-1 cells were differentiated into mature monocyte-derived dendritic cells (MoDCs) and co-cultured with PBMCs under various conditions in a two-layer transwell model. To assess the levels of reservoir reactivation and CD8+ T cell functionality, flow cytometry was employed to measure changes in HIV-1 p24 protein levels in CD4+ T cells, cytokine/chemokine levels in CD8+ T cells, TVM subpopulation proportions, and IL-2Rβ receptor expression on TVM cells. Fluorescence quantitative PCR measured changes in HIV-1 integrated DNA levels to assess reservoir size. Results: In vitro, Tα1 stimulation of MoDCs resulted in significant secretion of IL-15/RA complexes (p<0.001). This interaction with IL-2Rβ/γ receptors on T cells enhanced the intracellular secretion of CCL3/5, IFNγ, and TNFα in CD8+ T cells (p<0.05), as well as proliferation of TVM subpopulation (p=0.013), inhibited p24 levels in CD4+ T cells (p=0.004), and reduced HIV-1 integration DNA levels (p=0.012). The decreased degree of p24 level was negatively correlated with the secretion of intracellular cytokines in CD8+ T cells and the increase of TVM proportions (p<0.05). However, these effects were observed in PBMCs from immunological responders rather than nonresponders (INR). Neither Tα1 nor rhIL-15 stimulation altered the frequency of TVM cells in INR (p=0.126) or the expression of surface IL-2Rβ receptors (p>0.05), indicating impaired responsiveness of these subsets to IL-15 signaling. Conclusions: This study demonstrate that Tα1 enhances CD8+ T cell function, promotes TVM proliferation, and suppresses reservoir size and reactivation via IL-15 pathway activation in dendritic cells. Impaired IL-15 signaling responsiveness in INRs may contribute to poor reservoir control, highlighting a potential therapeutic target. 581.5 Cyclic Adenosine 3',5'-Monophosphate Agonist Prevents Establishment and Reversal of HIV Latency Prerna Dabral, Li Du, Mohamed Bouzidi, Satish Pillai Vitalant Research Institute, San Francisco, CA, USA Background: Cyclic adenosine 3',5'-monophosphate (cAMP) is a secondary messenger involved in the regulation of diverse cellular, metabolic, and immunologic processes. Ex vivo and in vitro studies have shown that cAMP levels are elevated in the setting of HIV infection, with T cells from PLWH harboring twice the cAMP level as uninfected individuals. In our previous studies, we identified the adenosine signaling pathway which leads to production of cAMP as a host determinant of HIV latency. Here, we used forskolin, a cell-permeable diterpene that activates adenylyl cyclase, and SCH- 581

Poster Abstracts

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Interference of HIV Integration via Changes in CPSF6 and SC35 Expression Due to TKIs in Macrophages Mario Manzanares 1 , Juliana Manosalva Perez 1 , Elena Mateos 1 , Alicia Simón Rueda 1 , Diego Megías 1 , Montserrat Torres 1 , Vicente Planelles 2 , Adam M. Spivak 2 , Mayte Coiras 1 , Clara Sánchez Menéndez 1 1 Instituto de Salud Carlos III, Madrid, Spain, 2 University of Utah School of Medicine, Salt Lake City, UT, USA Background: Despite the efficacy of ART, HIV-1 persistence in reservoirs, mostly formed by CD4 and macrophages hinders eradication. Nuclear import of pre-integration complex is assisted by proteins from mRNA processing pathways like CPSF6 (3´RNA-cleavage and polyadenylation factor 6) and SRSF2/SC35 (Serine/Arginine-rich splicing factor 2). Dasatinib and ponatinib are tyrosine kinase inhibitors (TKIs) that interfere with HIV-1 proviral integration in both CD4 and macrophages by preserving the antiviral activity of SAMHD1. Our objective was to determine if dasatinib and ponatinib may also interfere with proviral integration in MDMs via CPSF6 and/or SC35. Methods: Blood samples were obtained from healthy donors (n=15). CD14+ cells were isolated from PBMCs and differentiated into monocyte-derived macrophages (MDMs). In vitro infection of MDMs with JR_FL_Renilla and DHIV3-GFP was monitored in presence/absence of dasatinib and ponatinib by time-lapse microscopy. Influence of TKIs on HIV-1 early and late reverse transcription (RT), formation of 2-LTR circles, and proviral integration was analyzed by ddPCR. Subcellular localization and expression levels of CPSF6 and SC35 were analyzed by confocal microscopy. Results: 1)Time-lapse of dasatinib- and ponatinib-treated MDMs infected with DHIV-3-GFP revealed that GFP was nearly undetectable for 50 hours

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