Background
While CD8 T cells and autologous neutralizing antibodies have been suggested to be critical for ART-free HIV-1 suppression in post-intervention controllers (PICs), the role of CD4 T cells remains less defined.
Methods
We studied 5 PICs (4 males, 1 female) with prolonged viral suppression (median 6 years, IQR 3.6-6.4) after analytical treatment interruption (ATI) following broadly neutralizing antibody therapy. Pre-ATI CD4 profiles were compared by spectral flow in PICs and 4 non-controllers (NCs; 3 males, 1 female). CITE-seq with paired αβ TCR sequencing was performed on 143,550 unstimulated CD4 T cells from PICs and NCs pre- and during ATI. Single-cell data from 51,469 activation-induced marker (AIM)-sorted, HIV-1 peptide-stimulated T cells were used to profile HIV-1-specific T cell responses and infer specificity of unstimulated CD4 T cells. TCR specificity was further assessed by proliferation assay with bulk TCR sequencing. Genes with log2 fold change (log2FC) > 0.5 and adjusted P < 0.05 were considered differentially expressed. Medians/IQRs refer to the listed genes.
Results
Pre-ATI frequencies of CD4 subsets defined by spectral flow were comparable between PICs and NCs. Similarly, CITE-seq cluster proportions did not differ between groups pre- or during ATI. Within a cluster of antiviral Th1 cells, NCs had higher pre-ATI expression of cytotoxic (GNLY, FGFBP2, PRF1) and activation (HLA-DR) transcripts compared to PICs (median log2FC 0.97, IQR 0.62-1.34). This signature persisted during ATI, accompanied by CTLA4 upregulation (log2FC 1.44). In contrast, upon HIV-1 peptide stimulation, AIM-sorted CD4 T cells from PICs at pre-ATI showed stronger induction of activation (JUN, FOS, HLA-DQB1), and effector/antiviral (TBX21, IFNG, APOBEC3G) transcripts (median log2FC 2.47, IQR 2.34-3.08). This activation-poised antiviral signature persisted during ATI (median log2FC 1.86, IQR 1.51-2.91). TCR backtracing from AIM-sorted T cells inferred HIV-1 specificity for 1.35% (IQR 0.44%-1.41%) of unstimulated CD4 T cells, 84% (IQR 55%-86%) of which mapped to Th1 clusters. Bulk TCR sequencing after the proliferation assay identified additional clones that also preferentially mapped to Th1 clusters (52%-80%).
Conclusions
CD4 T cell responses in PICs are distinguished by their capacity to rapidly activate and mount superior antiviral transcriptional programs upon HIV-1 challenge rather than stronger cytotoxic signatures in the unstimulated state or higher frequencies of circulating HIV-1-specific cells.