Maha Ayyoub

HERV-derived epitopes represent new targets for T-cell-based immunotherapies in ovarian cancer

Background Ovarian cancer represents the most lethal gynecological cancer with poor response to checkpoint inhibitors. Human endogenous retroviruses (HERVs) are aberrantly expressed by tumor cells and may represent a source of shared T-cell epitopes for cancer immunotherapy regardless of the tumor mutational burden. Methods A transcriptomic analysis based on RNA sequencing was developed to quantify the expression of HERV-K sequences containing the selected epitopes. The presence of HERV-K/HML-2 Gag antigen was then assessed by immunohistochemistry (IHC) on tumor microarrays from ovarian cancer samples and normal ovarian tissues. A specific immunopeptidomics approach was developed to detect epitopes on human leukocyte antigens (HLA) molecules. Epitope-specific CD8+ T cells were quantified by multimer staining. HERV-specific T cells were obtained after in vitro stimulation of T cells from HLA-A2-positive healthy donors or patients with ovarian cancer, and in vitro target cell killing was evaluated using real-time analysis. In vivo antitumor efficacy of HERV-specific T cells was assessed in an avian embryo model. Results Epitope-containing HERV transcripts were significantly higher in ovarian cancers compared with normal tissues. The presence of the HERV-K/HML-2 Gag antigen was confirmed by IHC in 20/40 (50%) ovarian cancers while no Gag expression was found in normal ovarian tissue samples. Immunopeptidomics analysis revealed the presence of epitopes on HLA molecules on the surface of ovarian tumor cell lines but not on normal primary cells from critical tissues. Low percentages of HERV-specific T cells were detected among tumor-infiltrating lymphocytes from ovarian cancers. Furthermore, in vitro stimulation of patient T cells induced functional epitope-specific T cells, confirming the immunogenicity of these epitopes in patients with ovarian cancer. In vitro, HERV-specific T cells specifically killed ovarian cancer cells in an HLA class I-restricted manner while sparing normal HLA-A2-positive primary cells derived from critical tissues. Epitope-specific CD8+ T cells exhibited a strong antitumoral activity in vivo, inducing a highly significant decrease in tumor volume in comparison with control groups. Conclusion These results provide the preclinical rationale for developing T-cell-based approaches against HERV-K-derived epitopes in ovarian cancer.

PD-1 blockade restores helper activity of tumor-infiltrating, exhausted PD-1hiCD39+ CD4 T cells

Tumor antigen-specific CD4 T cells accumulate at tumor sites, evoking their involvement in antitumor effector functions in situ. Contrary to CD8 cytotoxic T lymphocyte exhaustion, that of CD4 T cells remains poorly appreciated. Here, using phenotypic, transcriptomic, and functional approaches, we characterized CD4 T cell exhaustion in patients with head and neck, cervical, and ovarian cancer. We identified a CD4 tumor-infiltrating lymphocyte (TIL) population, defined by high PD-1 and CD39 expression, which contained high proportions of cytokine-producing cells, although the quantity of cytokines produced by these cells was low, evoking an exhausted state. Terminal exhaustion of CD4 TILs was instated regardless of TIM-3 expression, suggesting divergence with CD8 T cell exhaustion. scRNA-Seq and further phenotypic analyses uncovered similarities with the CD8 T cell exhaustion program. In particular, PD-1hiCD39+ CD4 TILs expressed the exhaustion transcription factor TOX and the chemokine CXCL13 and were tumor antigen specific. In vitro, PD-1 blockade enhanced CD4 TIL activation, as evidenced by increased CD154 expression and cytokine secretion, leading to improved dendritic cell maturation and consequently higher tumor-specific CD8 T cell proliferation. Our data identify exhausted CD4 TILs as players in responsiveness to immune checkpoint blockade.