Background
HIV-1 cure strategies require a comprehensive understanding of cellular phenotype of HIV tissue reservoirs for effective and selective targeting. While recent data have demonstrated the highly heterogeneous nature of peripheral blood CD4+ T cells subsets comprising the HIV reservoir of antiretroviral therapy (ART) treated people with HIV (PWH), CD4+ T follicular helper (Tfh) cells are generally considered as the predominant cellular reservoir of virus within lymphoid tissue. However, there are limited data supporting this during ART. Here, we re-examined this notion in lymph nodes of viremic and ART-treated PWH using unbiased multiomic analysis.
Methods
To define the cellular signature of infected cells in the lymphoid tissue compartment, we profiled memory CD4+ T-cells from lymph node samples of PWH during viremia (n = 9) and ART treatment (n = 10) using single-cell DOGMAseq (scDOGMAseq) for the simultaneous assessment of epigenetics, transcriptomics, and surface antigen levels.
Results
Using the presence of transposed HIV-1 DNA during the epigenetic profiling and/or HIV RNA during transcriptomic profiling, we found HIV-1 infected cells (n = 1388 during viremia; n = 398 during ART) and determined their cellular phenotype. HIV-1 infected cells were found to be distributed between many CD4+ T-cell subsets, including central, effector, regulatory and resident memory subsets. Importantly, the reservoir during ART or viremia was not predominantly associated with T follicular helper (Tfh) cell phenotypes during viremia (16.9% of HIV DNA+ cells; 13% of HIV RNA+ cells) or during ART treatment (8.1% of HIV DNA+ cells; 5.3% of HIV RNA+ cells). HIV DNA+ cells were instead found in two different memory populations: a non-activated and non-resident central memory T subset (34.7% and 15.6% of HIV+ DNA+ cells during viremia and ART, respectively) and a phenotypically resting but recently activated NR4A1/NR4A2 high subset (16.8% of HIV+ DNA+ cells; 18.1% of RNA+ cells) during ART.
Conclusions
Our scDOGMAseq atlas of transcriptionally inactive and active HIV-1 infected CD4+ T cells from the lymphoid tissue highlights the predominance of non-Tfh cell subsets in the HIV tissue reservoir during ART. We find the HIV+ cells to be more concentrated in resting/recently activated cells that have not been previously described. These data altogether demonstrate that HIV cure strategies primarily targeting the B cell follicles during ART may miss out on a large portion of the HIV reservoir in lymphoid tissues.