Abstract 3154: Identification of CD4+ T cell epitopes specific for the breast cancer associated antigen NY-BR-1

Abstract

Abstract Questions: Whether CD4+ T cell epitopes specific for the breast cancer associated antigen NY-BR-1 can be identified using HLA-transgenic (HLAtg) mice. Introduction: Adoptive transfer of TCR-transduced autologous tumor antigen-specific T cells provides an innovative strategy for cancer immunotherapy. The differentiation antigen NY-BR-1 is over-expressed in approx. 60% of all invasive mammary carcinomas, thus representing a potential target for cancer immunotherapy. We thus screened NY-BR-1 for the presence of CD4+ T cell epitopes. Methods: Splenocytes from DNA immunized HLA-DRB1*0301 and HLA-DRB1*0401 tg mice were screened ex vivo by IFNγ ELISpot assay against a NY-BR-1-derived peptide library. In silico predicted candidate epitopes present among recognized library peptides were used to establish CD4+ T cell lines as a read tool to prove HLA-DR-restriction and natural processing of these epitopes by human cells. NY-BR-1-specific CD4+ T cell reactivity among PBMC of breast cancer patients was tested after long term in vitro restimulation with synthetic 15mers by intracellular cytokine staining. Results: Three HLA-DRB1*0301-restricted and four HLA-DRB1*0401-restricted NY-BR-1-specific CD4+ T cell epitopes were identified in HLAtg mice. HLA-DR-restriction was confirmed upon specific recognition of peptide-loaded T2/DR3 and T2/DR4 target cells by epitope-specific CD4+ T cell lines established from HLAtg mice. These T cell lines specifically recognized human dendritic cells loaded with cell lysates from Ad5-NY-BR-1 infected melanoma cells, showing natural processing of the epitopes. Moreover, upon deep sequencing of TCRs from these murine CD4+ T cell lines two NY-BR-1-specific TCRs were identified which could be considered for the generation of autologous TCR-transduced T cells lines. Finally, CD4+ T cells reactive against the NY-BR-1 specific epitopes were also detected among PBMCs of HLA-DR matched breast cancer patients. Conclusion: NY-BR-1-specific, HLA-DRB1*0301 and HLA-DRB1*0401 restricted CD4+ T cell epitopes could be identified using HLAtg mice and CD4+ T cells reactive against these epitopes are present among PBMCs of breast cancer patients. Finally, murine HLA-restricted CD4+ T cell lines might serve as source of NY-BR-1-specific TCRs for adoptive immunotherapy approaches. Citation Format: Adriane Gardyan, Wolfram Osen, Maria Agawal, Inka Zörnig, Eliana Ruggiero, Manfred Schmidt, Andreas Schneeweiss, Dirk Jäger, Stefan B. Eichmüller. Identification of CD4+ T cell epitopes specific for the breast cancer associated antigen NY-BR-1. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3154. doi:10.1158/1538-7445.AM2015-3154