CROI 2025 Abstract eBook

Abstract eBook

Poster Abstracts

470

Females With HIV Favor Interferon Responses Over Inflammation Upon TLR7 Activation Alisa Huber 1 , Nadira Vadaq 1 , Albert L. Groenendijk 2 , Victoria Rios Vazquez 1 , Joost A. H. Martens 3 , Vasiliki Matzaraki 1 , Maartje Jacobs-Cleophas 1 , Jan van Lunzen 1 , Mihai G. Netea 1 , Leo Joosten 1 , Andre van der Ven 1 , Jéssica dos Santos 1 , for the 2000HIV Human Functional Genomics Partnership Program 1 Radboud University Medical Center, Nijmegen, Netherlands, 2 Erasmus University Medical Center, Rotterdam, Netherlands, 3 Radboud University, Nijmegen, Netherlands Background: Females have lower viral setpoints and are more likely to control HIV infection without ART than males, though the mechanisms remain unclear. The X-chromosome-encoded ssRNA receptor TLR7 may contribute to this difference as evasion of X-inactivation in females can increase TLR7 expression. TLR7 activation induces type-I Interferon (IFN) release and production of pro-inflammatory cytokines, but its role in HIV infection across sexes is not fully understood. We investigated cytokine production and gene expression in ART-treated females and males with HIV upon ex-vivo PBMC stimulation with the TLR7 agonist Imiquimod (IMQ). Methods: The 2000HIV study (NCT03994835) was divided into a Discovery cohort (201 females, 1161 males) and a Validation cohort (46 females, 252 males). PBMCs were stimulated with 5 µg/mL IMQ for 24 h, and IL-1β, IL-1RA, IL-6, IL-8, MCP1, and MIP1α levels were measured by ELISA in supernatants. Cytokine production between sexes was compared using a rank-based regression model, adjusting for age, ethnicity, lymphocyte/monocyte ratio, COVID-19 vaccination, and time of inclusion. Bulk RNA sequencing was conducted on PBMCs from the 2000HIV-TRAINED cohort (27 females, 43 males) (NCT04968717), assessed differential gene expression (DEG), adjusting for age, ethnicity, and COVID-19 vaccination. Gene set enrichment analysis (GSEA) was performed using the HALLMARK gene sets. Results: IMQ stimulation led to lower IL-1β production in PBMCs from females compared to males (p=0.002) and a trend towards lower IL-8 levels (p=0.054). DEGs included 43 up- and 40 downregulated genes in females compared to males. Downregulated genes in females included CRISP3 , WNT5A , ARG1 , and MMP8 (p=1.864x10 -5 , p=0.041, p=0.002, p=0.002, respectively), while upregulated genes included interferon-stimulated genes like IRF7 , IFI6 , IFIT3 , ISG15 (p=0.004, p=0.004, p=0.018, p=0.015, respectively), and the anti viral factor APOBEC3A (p= 0.042) . GSEA revealed enriched "Interferon alpha response" and "Interferon-gamma response" pathways in females (p=1.195x10 -19 , p=1.194x10 -19 ) compared to males. Conclusions: Females with HIV show enhanced IFN pathway gene expression but lower pro-inflammatory IL-1β production upon TLR7 activation compared to males with HIV. This suggests TLR7 activation drives a stronger antiviral response in females, favoring IFN responses over pro-inflammatory pathways. Sex-biased immune response differences in people with HIV should be considered in the development of TLR7-agonist therapies. CITE-Seq and Single Cell αβ TCRseq Unravels HIV-1 Specific GZMB Subset in Post-Treatment Controllers Giacomo Schmidt Frattari 1 , Miriam Rosás-Umbert 2 , Mariane Høgsbjerg Schleimann 1 , Martin Tolstrup 1 , Jesper D. Gunst 3 , Ole Søgaard 1 1 Aarhus University, Aarhus, Denmark, 2 Centre de Recherche du CHUM, Montreal, Canada, 3 Aarhus University Hospital, Aarhus, Denmark Background: Earlier studies have described a subset of clonally expanded, granzyme B (GZMB) Th1 memory CD4 T cells in people living with HIV. A deeper characterization of these cells in people that display ART-free virological control may contribute to our understanding of their role in anti-HIV-1 T cell responses. Methods: We conducted single cell transcriptome, surface protein and T cell receptor (TCR) sequencing on longitudinally collected samples from 3 post treatment controllers (PTCs). For participant ID107 (6 years off ART) we analyzed 5 time points: pre-ART, on ART and 3 times off ART. For each of ID142 and ID314 (both 2.5 years off ART) we analyzed 3 time points: on ART and 2 times off ART. In parallel, we sequenced 66,318 resting CD4 T cells and 39,429 HIV-1 specific T cells sorted using the activation induced marker (AIM) assay. We used paired TCR α- and β-chain sequences to identify clonotypes. Resting CD4 T cells that shared clonotype with AIM-sorted cells were identified as HIV-1 specific. We applied weighted gene co-expression network analysis (WGCNA) to investigate modules of correlated genes. The figure, table, or graphic for this abstract has been removed.

Results: In resting CD4 T cells, GZMB cells presented a terminally differentiated effector transcriptome and surface protein profile. Relatively few, expanded clones dominated the GZMB cluster (median clonality by Gini index 0.54–0.73) and persisted through time (median overlap across time points by Morisita-Horn index 0.82–0.96). Using the AIM dataset, we tracked 744 unique clonotypes back to 4,493 resting CD4 T cells. The median frequency of HIV-1 specific cells was 4% to 7%. Most of these HIV-1 specific cells belonged to the largest clones within the GZMB cluster. Therefore, most GZMB CD4 T cells were HIV-1 specific (median 55–65%) and HIV-1 specific GZMB CD4 T cells TCR repertoire showed high clonality (median Gini index 0.52–0.6) and overlap across visits (median Morisita-Horn index 0.84–0.96). By WGCNA, we identified a module of 66 correlated, highly expressed cytotoxic and effector genes (among these CX3CR1, FCRL6, GNLY, PRF1). Within the GZMB cluster, HIV-1 specific cells showed higher scores for this module than non-HIV-1 specific cells (average log2-fold increase 0.74, adjusted P value 3.5e-48, Wilcoxon test). Conclusions: Among these PTCs, expanded clones of terminally differentiated, highly cytotoxic GZMB CD4 T cells, that were mostly HIV-1 specific, persisted unchanged over years. This subset might play a role in ART-free virological control. Diminished Immune Imprinting at the Upper Respiratory Tract Following SARS-CoV-2 Breakthrough Infection Jian Zou, Jinyuan Liu, Mengping Chen, Chunyang Bai, Linlin Zhou, Jiajing Jiang, Xuan He West China Hospital of Sichuan University, Chengdu, China Background: Immune imprinting, also known as original antigenic sin, refers to the effects prior exposures have on subsequent immunity against antigenically related viruses. The immune imprinting of humoral immunity at anatomic sites of the respiratory tract following SARS-CoV-2 breakthrough infection is not well understood. Methods: Ninety-six adult donors with COVID-19 vaccination and breakthrough infection (BTI) during the BA.5 wave were recruited for this study. Blood samples and paired swabs, including nasal swabs from the midpoint of the inferior turbinate (MT) and nasopharyngeal (NP) swabs were collected. Each sample was monitored via a pseudovirus-based virus neutralization assay to determine neutralizing antibody (NAb) titers against wild-type (WT), BA.5 and XBB variants. Blood (n=57), MT (n=26) and NP (n=33) samples with detectable NAb titers were included for further analyses on immune imprinting level. Antigenic cartography was generated based on the NAb data. B cell immunophenotyping was assessed by multiparameter flow cytometry. Results: These vaccinated subjects with BTI showed significantly higher levels of circulating NAbs against WT (4.2-fold) and BA.5 (6.7-fold) than against XBB (P<0.0001 for both), while exhibiting roughly similar NAb titers against WT and BA.5 in circulation (<2-fold difference). Mucosal NAb responses from MT and NP samples were highly correlated (P=0.003), and displayed distinct profiles compared to circulating antibodies. Both MT and NP samples showed significantly higher mucosal NAb titers against BA.5 than that against WT (2.5 fold and 2.4-fold, respectively; P<0.001 for both), and against XBB (2.6-fold and 2.7-fold, respectively; P<0.0001 for both). Additionally, the antigenic distances from WT to BA.5 or to XBB were discernibly reduced in respiratory mucosal profile compared to that in circulation (P<0.05 for all). Germinal center B cells targeting BA.5 were detectable in the majority of NP samples but were barely observed in MT samples. There was also a trend of higher BA.5-specific memory B cells in NP samples, predominantly displaying a resting memory phenotype. Conclusions: The distinct antibody profiles between the upper respiratory tract and the circulating system reveal diminished immune imprinting at the upper respiratory tract, evidenced by the enhanced BA.5-specific mucosal NAb response following BTI during the BA.5 wave. These findings have important implications for our understanding of respiratory mucosal immunity against evolving SARS-CoV-2.

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Poster Abstracts

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CROI 2025 117