It's all in the mix : γδTCR meets αβT-cell for therapy

Cancer – the “Emperor of all Maladies” is one of the most threatening diseases of our times. In one way or the other nearly everyone has been affected by this disease and the public awareness of cancer is long perceptible. Science is now requested to reveal comprehensive insights about the complex mechanisms behind cancer biology and to find applicable treatment options for patients. Cell-based cancer immunotherapy is one of the initiatives to treat a range of malignancies by activating cells of the immune system to detect and destroy cancer. T-cells play an important role in cell-mediated immunity and are critically involved in tumor immunosurveillance. Adoptive transfer of autologous tumor infiltrating lymphocytes (TILs) demonstrated the ability of T-lymphocytes to mediate cancer regression and metastatic melanoma was successfully treated with TIL-therapy. To broaden the spectrum of malignancies to be treated with adoptive cell therapy the development of a new strategy using genetically modified T-cells arouse. Here, T-cells are redirected against tumor cells by introducing a new receptor that specifically recognizes tumor-related antigens. This thesis describes one initiative to advance this concept by redirecting conventional αβT-cells against various tumor types by introducing a defined γ9δ9T-cell receptor (TCR). The γ9δ2TCR is the main activating receptor expressed on γδT-cells which represents a small subset of peripheral T-cells with an innate-like character. The γ9δ2TCR mediates recognition of various tumor cells and is therefore a potent receptor to be used in TCR-gene therapy strategies. However, the recognition mechanism by which the γ9δ2TCR recognizes a tumor cell is still puzzling. Unlike conventional αβT-cells which recognize peptide epitopes presented by MHC-molecules, the γ9δ2TCR mediates responsiveness towards small non-peptidic structures named phosphoantigens such as isopentenyl pyrophosphate. IPP is believed to accumulate in cancer cells or other ‘stressed’ cells and elevated IPP levels induce the activation of γ9δ2T-cells via their TCR. However, γδT-cells also express other activating receptors such as natural-killer cell activating receptor (NKG2D) that also contributes to γδT-cell activation. This thesis describes the generation of a highly tumor-reactive γ9δ2TCR and its way towards clinical application as a tool in TCR-gene therapy. Starting from a molecular view point selected individual γ9δ2TCRs are described that mediate different functional avidity against tumor cells, a phenomenon which is caused by small structural differences in their TCR chains. Based on these results a method named combinatorial-γδTCR-chain-exchange (CTE) was establish to design a highly tumor reactive γ9δ2TCR. Furthermore, NKG2D, which is frequently expressed on γδT-cells, was identified as a factual co-stimulatory receptor to the γ9δ2TCR. The strength of γ9δ2TCR activation further dictates whether NKG2D is recruited to the immunological synapse and also the potency of NKG2D co-stimulation. Finally, a highly tumor-reactive γ9δ2TCR designed by CTE was introduced into αβT-cells using a clinical grade retroviral vector. To reach highest purity of γ9δ2TCR expressing T-cells a GMP-grade anti-αβTCR antibody was used to deplete αβT-cells that do not express the introduced γ9δ2TCR. Accordingly, a T-cell product with high anti-tumor reactivity was obtained and its success in pre-clinical models points towards a potent tool for cancer immunotherapy