Figure 7. DP CD4⁺ Th TILs recognize HPV-associated antigens.

(A) In vitro–expanded CD4⁺ TIL subsets (DN, SP, and DP) were cocultured with autologous B cells pulsed with DMSO or the indicated peptide pools. T cell reactivity was assessed by OX40 upregulation after 24 hours of culturing. Data for 1 representative patient with HPV⁺ HNSCC are shown. (B) Summary of the reactivity of CD4⁺ Th subsets to the indicated peptide pools for 6 patients with HPV⁺ HNSCC. (C) Summary of the reactivity of CD8⁺ T cells to HPV16 E6, HPV16 E7, and CEFX peptide pools for the same 6 patients with HPV⁺ HNSCC. CD8⁺ T cell activation was assessed by 4-1BB upregulation. DP CD4⁺ (D) and DP CD8⁺ (E) TILs isolated from 1 patient with HPV⁺ HNSCC were cultured with autologous B cells pulsed with DMSO, the HPV16 E6 peptide pool, or the individual overlapping peptides contained in the HPV16 E6 peptide pool (n = 37). Reactivity was measured by IFN-γ ELISPOT assay. SEB or anti-CD3 antibodies were used as positive controls for CD4⁺ and CD8⁺ T cells, respectively. (F) Summary of the reactivity of DP CD4⁺ and DP CD8⁺ TILs to HPV16 E6 individual peptides for 3 different patients with HPV⁺ HNSCC, as measured by IFN-γ ELISPOT assay. The colors in the heatmap legend represent the number of detected spots/10⁵ cells.