Background
Anti-PD-1 immunotherapy (API) could reverse T-cell exhaustion, strengthen HIV-specific T-cell responses, and reduce the HIV reservoir in people living with HIV (PLHIV). However, API responsiveness does not directly correlate with PD-1 expression on T-cells, suggesting that receptor heterogeneity may influence therapeutic outcomes. This study aimed to assess: 1) the variation of PD-1 expression among T-cell subsets, 2) host factors in PLHIV contributing to PD-1 heterogeneity, and 3) the correlation of PD-1 expression with cell functionality.
Methods
We analyzed high-dimensional flow cytometric PD-1 measurements on peripheral blood mononuclear cells (PBMCs), serum proteome, ex-vivo PBMC stimulations, and single cell RNA-sequencing of PBMCs from the 2000HIV cohort (NCT03994835), consisting of viral suppressed PLHIV on ART. We evaluated PD-1 expression on 21 T-cell subsets of 1372 PLHIV. Participants were categorized into high, intermediate, and low PD-1 expression based on the tertiles of PD-1 distribution. We explored the relationship between PD-1 expression, clinical factors, and PBMC cytokine production, serum proteome, and single cell RNA-sequencing using Spearman’s correlation and pathway analyses.
Results
Lower PD-1 expression in exhausted CD4+ T-cells correlated with female sex, high CD4 nadir, COVID-19 vaccination, and spontaneous HIV control. Higher expression was linked to older age, immunological non-response, higher CMV titers, and larger HIV reservoirs. Exhausted CD8+ naïve Tc were associated with older age and higher CMV titers. Higher PD-1 expression on CD4+ T-cells negatively correlated with IFN-γ, IL-17, IL-22, and IL-10 secretion upon microbial stimulation of PBMCs. Differences in plasma proteins (mostly related to cytotoxic and proliferation pathways) were determined by PD-1 expression on CD4+-lymphocytes, and not by CD8+ or NK-cells. This was also reflected by single cell RNA-sequencing (Figure), showing 224 differentially expressed genes in CD4+-lymphocytes, being linked to oxidative phosphorylation and type I and II interferons in the pathway analysis.
Conclusions
PD-1 expression in PLHIV is heterogeneous across T-cell subsets and associates negatively with cytokine production, reflecting immune exhaustion. Especially PD-1 expression on CD4+-lymphocytes determines plasma proteome and transcriptional differences. PLHIV with higher CMV reactivity, larger HIV reservoirs, and immunological non-responders may benefit from API.
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