CROI 2025 Abstract eBook

Abstract eBook

Poster Abstracts

515

Macrophage-Tropic TF SHIV D-Infected NHP Model of Reservoir Persistence and Decay on ART Suvadip Mallick 1 , Ryan Krause 1 , Alexander McFarland 1 , Hannah Schrader 1 , Richard W. Camp 1 , Gregory D. Whitehill 1 , Francesco E. Marino 1 , Rachel Podgorski 2 , Himanshu Garg 1 , Emily Lewis 1 , George M. Shaw 1 , Tricia Burdo 3 , Katharine Bar 1 1 University of Pennsylvania, Philadelphia, PA, USA, 2 Temple University, Philadelphia, PA, USA, 3 Rutgers Robert Wood Johnson Medical School, Piscataway, NJ, USA Background: Characterization of HIV-1 persistence in both CD4+ T cell and myeloid reservoirs within systemic tissues and the brain are goals of the cure field. Clade D HIV-1 infects both CD4+ T cells and macrophages. Thus, we have employed SHIV encoding a clade D transmitted/founder HIV-1 Env to study reservoirs in ART-suppressed rhesus macaques (RM). Methods: Ten RM were infected with barcoded SHIV.D.191859, started on ART 10 weeks post-infection, and continued through necropsy at 9 (n=5) or 20 months (n=5) of ART. Blood and tissue samples were tested for viral load, barcode clonotype distribution, total and intact proviral HIV DNA, and RNA/ DNAscope. Results: Plasma viremia peaked at week 2 post-infection (median 2.5 x 10 6 copies/mL) with barcode sequencing indicating a multiplicity of infection between 45-400 unique clonotypes. At week 4, viral loads ranged from 10 5 -10 7 copies/mL in plasma and 10 2 -10 3 copies/mL in CSF. ART was initiated during setpoint viremia (median 2 x 10 4 copies/mL) at week 10 and durably suppressed in plasma and CSF. Reservoir quantification at necropsy showed persistence of SHIV.D within CD4+ T cells and myeloid cells systemically (blood, 5 lymph nodes, spleen). In animals necropsied after 9 months on ART, total and intact proviral DNA in was ~10X higher in CD4+ T cells (median 3.5 x 10 3 total copies, 1.3 x 10 3 intact copies/10 6 cells) than in myeloid cells (median 1.2 x 10 2 total copies, 45 intact copies/10 6 cells) across compartments. Animals necropsied at 20 months showed similar ratios, with ~10X lower absolute measures. Within the brain, all necropsied animals had evidence of SHIV RNA and DNA by immunohistochemistry; 6/10 RM had detectable SHIV proviral DNA within purified brain myeloid cells. Sequencing of barcode clonotypes from plasma and CSF during viremia, and within the brain and systemic tissues at necropsy, showed substantial overlap between viruses infecting CD4+ T cells and myeloid cells systemically, and between the brain and systemic tissues. Conclusions: The dual CD4+ T cell- and macrophage-tropic SHIV.D persists on ART in myeloid and CD4+ T cells within the brain and systemic tissues. With 10 weeks of virus replication prior to ART, there was no compartmentalization of viruses between the brain and systemic tissues, and none evolved on suppressive ART. Between 9 and 20 months of ART, brain and systemic reservoirs decrease in size but remain detectable. Together, these results inform the use of SHIV.D infected RM for persistence and cure studies. Anatomical Sites in FIND 2.0 Decedents Harbor Compartmentalized, Transcriptionally Intact HIV-1 Env Melanie Moodie 1 , Maria A. Papathanasopoulos 1 , Adriaan E. Basson 1 , Nadia Sabet 1 , Ebrahim Variava 2 , Neil Martinson 3 , Tanvier Omar 1 , Monique Nijhuis 4 , Annemarie M. J. Wensing 4 1 University of the Witwatersrand, Johannesburg, South Africa, 2 Klerksdorp Tshepong Hospital Complex, Jouberton, South Africa, 3 Perinatal HIV Research Unit, Soweto, South Africa, 4 University Medical Center Utrecht, Utrecht, Netherlands Background: HIV-1 persistence poses a significant barrier to HIV eradication. Findings from the largest post-mortem study (FIND 2.0), which utilized minimally invasive tissue sampling in 38 virally suppressed individuals on antiretroviral therapy (ART) (VS; <400 copies/mL) and 22 unsuppressed (VU) untreated individuals showed the presence of proviral DNA in all 11 anatomically distinct compartments. We report detailed genotypic characterization of full length HIV-1 envelope glycoprotein ( env ) from this cohort. Methods: Full length HIV-1 env was PCR amplified from extracted DNA from bone marrow aspirate (BM) and trephine (BMT), brain, bronchoalveolar lavage (BAL), cervicovaginal lavage, kidney, lymph nodes, liver, lung, spleen and whole blood (WB) from 60 FIND 2.0 decedents. Amplicons were sequenced (population-based) on Illumina MiSeq and analysed for subtype, minority variants, premature stop codons, hypermutations, intact rev response element (RRE), and coreceptor tropism. Viral compartmentalization was assessed by comparing sequence identity, coreceptor tropism prediction, V3 loop variants and phylogenetic clustering of each env per compartment/decedent.

after an average of 56 weeks after the initial bNAb dose at a time when HIV-1 was undetectable. Results: Individuals in the 2 arms showed similar numbers of intact and defective proviruses. Consistent with early therapy, 5 out of 27 and 7 out of 21 individuals in bNAb and placebo arms respectively, had no measurable reservoir at baseline by ddPCR. The geometric mean number of intact proviruses/106 CD4+T cells was 8.4 by ddPCR and 0.28 by Q4PCR. There was no significant correlation between reservoir size and time to initial rebound in the placebo arm. 80% of the CD4+T cells carrying intact proviruses were members of expanded clones. 267 representative proviral Env genes from reservoir and rebound were produced as pseudoviruses, and subjected to sensitivity testing in TZM-bl assays. All individuals in the bNAb arm, and all but 1 in the placebo arm showed reservoir sensitivity to at least 1 of the 2 bNAbs. Despite low to non measurable autologous antibody neutralizing activity to reservoir viruses in the placebo group, rebound sequences only corresponded to reservoir sequences in 2 of 9 individuals. Therefore, autologous antibody neutralizing activity cannot account for the disparity between intact proviruses found in the reservoir and rebound in these individuals. Reservoir half-life and log fold change after bNAb dose during ATI (bNAb arm) or at ART restart (placebo arm) were not significantly different. When all measurements for both arms are considered jointly (n=32) the half-life of intact and defective reservoirs were 33 and 424 weeks respectively. Conclusions: Antibody therapy in the RIO trial was associated with an average intact reservoir half-life decay which is 7-8 fold faster than prior reports. CD4+ T Cells Harboring Inducible HIV Genomes Display a Senescent-Like Phenotype Helene Roux 1 , Amélie Pagliuzza 1 , Remi Fromentin 1 , Jean-Pierre Routy 2 , Nicolas Chomont 3 1 Centre de Recherche du CHUM, Montreal, Canada, 2 McGill University Health Centre Research Institute, Montreal, Canada, 3 Université de Montréal, Montreal, Canada Background: Latently HIV-infected cells persist during antiretroviral therapy (ART) and these viral reservoirs inevitably trigger a viral rebound when treatment is interrupted. Identifying the mechanisms contributing to the persistence of these reservoirs is essential to the development of strategies aimed at eradicating them. In this study, we used a single-cell approach to measure multiple parameters that could contribute to the persistence of HIV infected cells after prolonged ART. Methods: Using a modified version of the HIV-Flow assay, we phenotypically analyzed the cells harboring inducible proviruses (p24+ cells) in blood samples from 10 participants who maintained undetectable plasma viral loads for 3-24 years. CD4+ T cells were stimulated for 24h with anti-CD3 and anti-CD28 antibodies, and cellular markers associated with T cell quiescence, senescence, cytotoxicity and immune functions were measured. Results: The phenotype of 395 p24+ cells was determined from a total pool of 390 million CD4+ T cells. p24+ cells more frequently expressed the anti-apoptotic protein Bcl-2 (p=0.006, fold increase 1.1), the immunotoxins granzymes A and K (p<0.0001 for both, fold increase 7 and 3, respectively) and the immune checkpoint molecules PD-1 and TIGIT (p<0.0001, fold increase 1.8 and p<0.0001, fold increase 3.4). We also found that p24+ cells more frequently expressed the cytokines TNF-a, IFN-g, IL-2 and IL-17 (p<0.02 for all, fold increases 5, 9, 14 and 55 respectively). Intriguingly, simultaneous expression of multiple cytokines, a signature of senescent T cells, was much more frequent in p24+ cells than in p24- cells (p<0.0001, fold increase 10). Finally, p24+ cells co-expressed the senescence markers KLRG-1 and CD57 more frequently than p24- cells (p=0.0009, fold increase 2.5). Conclusions: Our data indicate that immunosenescence is a signature of persistently infected cells after prolonged ART, suggesting that senolytic agents may be used to reduce the HIV reservoir.

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Poster Abstracts

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CROI 2025 132