Characterization of human MHC II-restricted T cell receptors with reactivity against B cells and tumor cells for therapeutic application in the context of adoptive T cell transfer of transgenic CD4 T cells.

The goal of this doctoral thesis was the isolation of tumor reactive CD4+ T cells for adoptive T cell therapy in the treatment of hematologic malignancies. We isolated two T cell clones with reactivity against B cells from the autologous system and identified the corresponding T cell receptors (TCR) which were transferred in different effector populations for further characterization. T cells transduced with both TCR showed multifunctionality in response to distinct target cells in a defined human leukocyte antigen (HLA-DR) context. Moreover, TCR-transduced T cells recognized HLA-DR matched tumor cells. Of note, target cells treated with Toll-like receptor ligands induced distinct functional modulation of T cells transduced with both TCR. Taken together, the isolation and characterization of two novel TCR demonstrating tumor reactivity and differential regulation by innate signals may have a significant impact for the development of novel immunotherapies in cancer

Isolation of human MHC class II-restricted T cell receptors from the autologous T-cell repertoire with potent anti-leukaemic reactivity.

Adoptive transfer of T cells genetically modified with tumour-specific T-cell receptors (TCR) is a promising novel approach in the treatment of cancer. We have previously isolated an allorestricted MHC class I-restricted TCR with specificity for Formin-like protein 1 (FMNL1) with potent activity against chronic lymphocytic leukaemia cells. CD4(+) T cells have been described to be highly important for tumour elimination although TCR derived from CD4(+) T cells with anti-tumour reactivity have been only rarely described. In this study we aimed to isolate MHC class-II-restricted CD4(+) T cells and TCR with specificity for leukaemia antigens. We used professional antigen-presenting cells pulsed with the leukaemia-associated and tumour-associated antigen FMNL1 for stimulation of autologous T cells in vitro. We isolated two CD4(+) HLA-DR-restricted T-cell clones and T-cell-derived TCR with so far unknown specificity but high reactivity against lymphoma cells and native malignant cells derived from HLA-matched patients with diverse leukaemias. Moreover, characterization of the TCR after TCR gene transfer revealed that specific characteristics of isolated TCR as reactivity in response to Toll-like receptors were transferable on effector cells. Our results have a major impact on the development of novel immunotherapies. They demonstrate that TCR with potent HLA-DR-restricted anti-leukaemic reactivity against so far undefined self-restricted antigens can be isolated from the healthy autorestricted CD4(+) T-cell repertoire and these TCR are highly interesting candidate tools for novel immunotherapies

A single TCRα-chain with dominant peptide recognition in the allorestricted HER2/neu-specific T cell repertoire.

T cells can recognize tumor cells specifically by their TCR and the transfer of TCR-engineered T cells is a promising novel tool in anticancer therapies. We isolated and characterized four allorestricted TCRs with specificity for the HER2/neu-derived peptide 369 (HER2(369)) demonstrating high peptide specificity. PBMCs transduced with especially one TCR, HER2-1, mediated specific tumor reactivity after TCR optimization suggesting that this TCR represents a potential candidate for targeting HER2 by TCR-transduced effector cells. Another TCR showed high-peptide specificity without tumor reactivity. However, the TCR alpha-chain of this TCR specifically recognized HER2(369) not only in combination with the original beta-chain but also with four other beta-chains of the same variable family deriving from TCRs with diverse specificities. Pairing with one beta-chain derived from another HER2(369)-specific TCR potentiated the chimeric TCRs in regard to functional avidity, CD8 independency, and tumor reactivity. Although the frequency of such TCR single chains with dominant peptide recognition is currently unknown, they may represent interesting tools for TCR optimization resulting in enhanced functionality when paired to novel partner chains. However, undirected mispairing with novel partner chains may also result in enhanced cross-reactivity and self-reactivity. These results may have an important impact on the further design of strategies for adoptive transfer using TCR-transduced T cells