PF444 IDENTIFICATION OF T‐CELL RECEPTORS FROM THE ALLOGENEIC‐HLA REPERTOIRE SPECIFIC FOR MELANOMA‐ASSOCIATED ANTIGENS

Abstract

Background:Melanoma‐associated antigens (MAGE) are highly expressed in multiple myelomas as well as solid tumors, but silent in normal cells except for male germ line cells. This tumor‐restricted expression makes it an excellent target for adoptive T cell therapy. However, due to thymic selection high‐affinity T cells against self‐antigens presented in the context of self‐HLA are deleted from the T‐cell repertoire of a patient. To overcome this problem, we identify T‐cell receptors (TCRs) from the allogeneic‐HLA repertoire instead of the autologous‐HLA repertoire.Aims:To identify high‐affinity T‐cell receptors from the allogeneic‐HLA repertoire specific for melanoma‐associated antigens.Methods:In this study we selected MAGE peptides by peptide elution experiments from HLA class I and subsequent mass spectrometric analysis of tumor cells. Only peptides binding in the most frequently occurring HLA‐alleles were selected. From the identified peptides, HLA‐tetramers were formed to isolate peptide specific CD8+ T cells from healthy allogeneic donors. Avidity of the selected T‐cell clones was determined by stimulation assays including a broad range of MAGE expressing tumor cell lines. To examine off‐target recognition, T‐cell clones were stimulated with MAGE negative cell lines and EBV‐transformed B lymphoblastic cells expressing all prevalent HLA molecules.Results:MAGE peptides were eluted from multiple myelomas, EBV‐transformed lymphoblastic cells, acute myeloid leukemia and ovarium carcinomas. For MAGE‐A1, T‐cell clones specific for four different epitopes presented in the context of HLA‐A∗01:01, HLA‐A∗03:01, HLA‐B∗07:02 and HLA‐C∗07:02 were selected. For MAGE‐A3, T‐cell clones specific for one epitope presented in the context of HLA‐A∗01:01 and HLA‐B∗35:01 were identified. The selected T‐cell clones demonstrated efficient recognition of MAGE‐A1 or MAGE‐A3 positive multiple myeloma and solid tumor cell lines without detectable cross‐reactivity.Summary/Conclusion:MAGE‐A1 and MAGE‐A3 specific T‐cell clones were isolated from the allogeneic‐HLA repertoire of healthy individuals. The identified clones were activated upon exposure to MAGE‐A1 or MAGE‐A3 positive tumor cell lines and showed limited off‐target recognition. These newly identified TCRs could potentially be used for TCR‐gene therapy strategies targeting MAGE‐A1 or MAGE‐A3 positive tumors.