CROI 2025 Abstract eBook

Abstract eBook

Poster Abstracts

Results: Of 521 DNA samples, 242 (46%) env amplified yielding 218 quality sequences: 127 from 36 VS, 91 from 19 VU. Phylogenetic clustering confirmed all were infected with HIV-1 subtype C, with sequences clustering by patient. Higher proportions of env sequences from VS were hypermutated (19%) as compared to VU (7%), with WB, BM, and BMT harboring most defective env . RRE was intact in 64% of VS and 52% of VU env sequences with most intact RREs detected in BAL. Premature stop codons were present in 24% of sequences, classifying them as defective. Minority variants were detected in 81% of env , with greater diversity in VU decedents. CCR5-tropic viruses predominated in both groups (92% VS; 80% VU), while CXCR4-tropic variants were more common in VU decedents, particularly in brain, liver and WB. Minor variant V3 loop sequence analysis showed mixtures of CCR5/CXCR4-utilizing viruses in some compartments, while single tropism sequences were found in others. Compartmentalization was observed in 40% VS and 63% VU decedents. Conclusions: These findings offer new insights into the dynamic and complex proviral HIV-1 populations across distinct anatomical compartments, largely independent of ART status. We highlight the persistence of diverse, transcriptionally intact HIV-1 env in these anatomical sites with evidence of compartmentalization. Biomarkers of HIV Latency in Tissues From People With HIV Xinlian Zhang 1 , Andrew Qazi 2 , Roni Lobato Ventura 2 , Antoine Chaillon 1 , Davey Smith 1 , Nadejda Beliakova-Bethell 3 , Sara Gianella Weibel 1 1 University of California San Diego, La Jolla, CA, USA, 2 Veterans Medical Research Foundation, San Diego, CA, USA, 3 Veterans Affairs San Diego Healthcare System, San Diego, CA, USA Background: Identifying biomarkers of latently infected cells is essential for eliminating the HIV reservoir. Since HIV proviruses exhibit activity even during latency, single-cell (sc) RNA-seq can detect gene expression patterns in these cells. Our prior in vitro study (PMID 38156317) revealed that transcriptional profiles of latent HIV+ cells vary depending on the environment, though certain genes were consistently expressed across conditions. We now aim to validate these biomarkers in CD4+ T cells from people with HIV (PWH) and investigate their tissue specificity. Methods: We collected lymph nodes, spleen, and liver via rapid research autopsy (<6h postmortem) from four PWH enrolled in the Last Gift cohort, all on ART with undetectable HIV RNA at death. CD4+ T cells were isolated by negative selection from dissociated tissues and subjected to scRNA-seq (10X Genomics). Reads were mapped to a combined human and autologous HIV genomic reference using CellRanger, and data were analyzed with Seurat v5 in Bioconductor R. Genes with a Bonferroni-adjusted p <0.05 were considered differentially expressed between HIV+ and HIV- cells. Gene set enrichment analysis was performed using the Database for Annotation, Visualization and Integrated Discovery v2024q2. Results: Participants included three men and one woman, with a median CD4+ T cell count of 317 before death. Across tissues, a total of 78 CD4+ T cells with low-level HIV RNA were detected. Differential gene expression analysis between HIV+ and HIV- CD4+ T cells revealed 84 genes in blood, 217 in lymph nodes, 141 in spleen, and 129 in liver. Two of the markers identified in vitro , LGALS1 and S100A11 , were differentially expressed in blood and all tissues ( Figure ). Proteins encoded by these genes, along with proteins encoded by CD2 and ITGAL , function in cell-cell adhesion. Tissue dependencies were also observed ( Figure ). For example, the previously reported marker CD2 and another transcript, ITGAL , were validated in all tissues but liver, and cytotoxic genes KLRD1 , GZMA and HCST were validated in all tissues but lymph nodes. Conclusions: This study validated biomarkers of HIV persistence in PWH using scRNA-seq, highlighting the influence of tissue microenvironments on latently infected CD4+ T cells. While several in vitro markers were confirmed, tissue dependent variations underscore the complexity of HIV latency. These findings indicate that tissue-specific biomarkers must be considered when designing strategies to target and eliminate the HIV reservoir.

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HIV Can Persist in Multiple Subsets of SARS-CoV-2-Specific CD4+ T Cells in Tissues During ART Xiaoyu Luo 1 , Julie Frouard 1 , Antoine Chaillon 2 , Sushama Telwatte 3 , Jason Neidleman 1 , Pavitra Roychoudhury 4 , Cheryl Dullano 2 , Stephanie Solso 2 , Robert Deiss 5 , Steven Yukl 6 , Nancie Archin 7 , Davey Smith 2 , Nadia R. Roan 1 , Sara Gianella Weibel 2 1 Gladstone Institutes, San Francisco, CA, USA, 2 University of California San Diego, La Jolla, CA, USA, 3 Peter Doherty Institute, Melbourne, Australia, 4 Washington DC VA Medical Center, Washington, DC, USA, 5 University of California San Diego Medical Center, La Jolla, CA, USA, 6 University of California San Francisco, San Francisco, CA, USA, 7 University of North Carolina at Chapel Hill, Chapel Hill, NC, USA Background: Transcriptionally-active cells (TAC) that produce HIV RNA persist during ART contribute to chronic inflammation and co-morbidities, and can lead to viral rebound upon ART interruption. However, our understanding of these cells – particularly within tissues – is limited. We implemented HIV-seq, a technique we established to efficiently identify HIV+ cells by scRNAseq, to characterize the transcriptional profiles and antigen specificities of TAC from tissues. Methods: Our case study focused on participant LG34 enrolled in the Last Gift rapid research autopsy program. LG34 initiated ART during chronic HIV in 2016 and remained on ART with inconsistent intake until his death in 2022. Between 2021 and 2022, he experienced multiple low-level HIV blips (<75cp/ ml), received four COVID-19 vaccinations, and had a breakthrough SARS-CoV-2 infection. We hypothesized that blipping during active SARS-CoV-2-specific immune responses (from vaccination and breakthrough infection) could seed an HIV reservoir within SARS-CoV-2-specific CD4+ T cells. To investigate this, we processed LG34’s spleen and lymph nodes post-mortem, and performed multiomic HIV-seq to characterize HIV RNA+ cells through scRNAseq and VDJ-seq to evaluate SARS-CoV-2 specificities. Results: We identified 49 and 37 TAC from 5162 and 5849 memory CD4+ T cells from spleen and lymph nodes, respectively. TAC were mainly found in Tregs with proliferative and survival transcriptional signatures, and terminally-differentiated Temra cells. Compared to uninfected Tregs, infected Tregs expressed higher levels of MHC-II and Foxp3. By contrast, compared to uninfected Temras, infected Temras expressed higher MHC-I and cytolytic effectors (granzymes B/H, perforin). VDJ-seq revealed greater clonal expansion in TAC than in uninfected cells, with some clones shared across tissues. Of all TCR matches identified by VDJMatch among HIV RNA+ cells, 83% were predicted to recognize SARS-CoV-2 antigens, and these cells resided primarily among Tregs. Conclusions: HIV-seq enabled detection of high numbers of TAC from tissues of LG34 under ART. These TAC were predominantly Tregs and Temras. Infected Tregs upregulated Foxp3 and MHC-II which may increase immunosuppressive function and facilitate recognition by CD4+ T cells. Infected Temras expressed cytolytic molecules, and their increased MHC-I expression may facilitate recognition by CD8+ T cells. TAC exhibit shared clonotypes across tissues, and include Tregs with specificities for SARS-CoV-2 antigens. Women With HIV Have a Lower Frequency of Monocyte Reservoir Reactivation Than Men With HIV Celina Abreu 1 , Jordin Dixson 1 , Lily Pohlenz 1 , Ruoyu Wang 1 , Erin Shirk 1 , Hayley Romero 1 , Janice Clements 1 , Leah H. Rubin 2 , Rebecca Veenhuis 1 1 The Johns Hopkins University School of Medicine, Baltimore, MD, USA, 2 The Johns Hopkins Bloomberg School of Public Health, Baltimore, MD, USA Background: There are known sex differences in HIV infection, latency, and immune responses. Women are overrepresented in some cohorts of immune controllers of HIV, raising questions about whether sex differences in immune

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CROI 2025 133