Application of individual HLA-binding prediction thresholds increases the detection of target cell surface-presented HPV16 E6- and E7-derived epitopes as a basis for therapeutic HPV vaccine design

Abstract

Abstract For the development of a therapeutic vaccine against human papillomavirus (HPV), viral epitopes bona fide presented on the surface of HPV-transformed cells need to be identified. The constitutively expressed HPV oncoproteins E6 and E7 represent ideal targets for immunotherapeutic vaccination approaches. However, viral epitopes are low abundant and human leukocyte antigen (HLA)-restricted, posing a challenge for developing peptide-based vaccines. For the identification of novel epitopes, in silico HLA-binding prediction was used to preselect HPV16 E6- and E7-derived peptides for 7 major HLA-types representing 5 HLA supertypes. Based on performance evaluation of predictors, new individual binding affinity thresholds were applied. HLA-binding of synthesized predicted binders was validated in vitro in competitive cellular binding assays. Interferon-γ ELISpot assays performed with blood samples from HLA-characterized healthy donors identified memory T cell responses against HLA-binding HPV peptides. T cells of responding donors were further characterized for epitope-specific mediation of specific target cell lysis by CD8+ CTLs using a flow cytometry based cytotoxicity assay. Complementary, an immunopeptidomics approach was used to determine the surface presentation of epitopes on HPV16-transformed cancer cells. Targeted liquid chromatography-mass spectrometry was capable of detecting epitopes predicted within the new individual affinity thresholds but missed by common thresholds. Completing the identification of bona fide target cell surface-presented E6- and E7-derived T cell epitopes will provide a map of targets for the development of a therapeutic vaccine to treat HPV16-induced malignancies.