Novel strategy to identify MHC Class II-promiscuous CD4+ peptides from tumor antigens for utilization in vaccination

DATTA J; XU S; TERHUNE JH; ROSEMBLIT C; BERK E; FITZPATRICK E; CZERNIECKI BJ

National Harbor, Maryland

Congreso; Society for Immunotherapy of Cancer 29th Annual Meeting; 2014

Society for Immunotherapy of Cancer

Background: Although cytotoxic CD8+ T lymphocytes (CTL) were historically considered primary effectors of antitumor immunity, solely boosting CTL responses with CD8+ vaccines in various tumor types has yielded unpredictable clinical results, possibly because CTLs function suboptimally without adequate CD4+ T-lymphocyte help. CD4 T-helper type 1 (Th1) cells secrete INF-γ/TNF-α, inducing tumor senescence and apoptosis. As such, successful incorporation of CD4+ epitopes into cancer vaccine construction and generation of durable antigen-specific CD4+ immunity remains a challenge. Using the extracellular domain (ECD) of HER3 as a candidate ?oncodriver? tumor antigen, we sought to identify immunogenic HER3 CD4+ peptides that demonstrate Class II promiscuity and generate anti-HER3 CD4+ immunity for inclusion in a vaccine construct.   Methods: A library comprising 123 overlapping 15 amino acid-long peptide fragments was generated from the HER3 ECD. Autologous monocyte-derived DCs from donors were matured to a type 1-polarized (DC1; IL-12 secreting) phenotype, and pulsed with HER3 ECD. Harvested DC1s were co-cultured with purified CD4+ T-cells. After 10 days, sensitized CD4+ T-cells were restimulated against immature DCs (iDC) pulsed with HER3 library peptide clusters or irrelevant CD4+ peptide control. Th1 responses, measured by IFN-γ ELISA, were considered antigen-specific if IFN-γ production was at least twice that of irrelevant control.      Results: Th1 sensitization was initially performed in 5 breast cancer patients with known anti-HER3 reactivity in order to identify single immunogenic HER3 CD4+ epitopes. To achieve this, HER3 ECD-specific CD4+ Th1 were sequentially restimulated against 10-peptide clusters, narrowed to 3-peptide clusters, and ultimately to single immunogenic HER3 peptides. A representative peptide screen is depicted in Figure 1. Four immunogenic peptides ? HER351-75, HER3402-417, HER3417-432, HER3451-465 ? were reproducibly identified and promiscuous across HLA-DR, DP, and DQ subtypes. When Th1 cells from 4 non-HER3 reactive donors were sensitized using DC1s pulsed with the four identified HER3 peptides, and subsequently challenged to recognize HER3 ECD-pulsed iDCs, all donors demonstrated successful sensitization not only to individual immunogenic HER3 peptides, but also recognized native HER3 ECD.   Conclusions: DC1 pulsed with an overlapping tumor antigen-derived peptide library can identify promiscuous class II peptides for CD4+ T-cell vaccine development. In this study, immunogenic HER3 CD4+ peptides effectively overcome immune tolerance to self-tumor antigens. Utilization of these HER3 CD4+ peptides in vaccine construction warrants investigation in patients harboring HER3-overexpressing cancers. Additionally, these results represent a novel strategy to rapidly and reproducibly identify class II-promiscuous immunogenic CD4+ epitopes from any tumor antigen for cancer immunotherapy using a DC1-CD4+ Th1 platform.