Background The T cell receptor (TCR) is uniformly expressed in monoclonal T cell lymphoma (TCL) populations and would thus represent an ideal target antigen for a CAR-based immunotherapy. However, the TCR is present on all T cells including malignant, healthy and CAR-T cells. Strategies to increase tumor specificity of TCR-specific CAR-T cells are therefore required. To address this issue, CARs targeting one of the two TCR-β constant chains and CARs targeting the CDR3 region of a malignant clone have been designed. [2,3] However, in the first case the off-tumor activity would affect about 50% of T cells, which could lead to impaired T cell immunity in the patient. The latter approach is highly specific to the malignant T cell clone but would require creation of individual CDR3-specific CARs for every patient which is currently impracticable. The TCR V-segment chains can be grouped into families and targeting TCR malignancies via V-family-specific CARs would spare the majority of healthy T cells. [4–6] Targeting the TCR V-segments would unite high tumor specificity with the limited effort of creating a panel of CAR molecules that could be used for all patients.
Materials and Methods The TCR sequence of a malignant clone from a patient with γδ-TCL was obtained and cloned into a lentiviral expression plasmid. This γδ-TCR was expressed in Jurkat E.6.1 TCL cells together with a panel of other γδ-τCRs to create target cells with different variable chain usage. A Vδ1-specific CAR molecule was designed and the CAR was expressed in a previously described Jurkat E.6.1 based T cell reporter cell line, that allows to measure T cell activation by flow cytometry.
Results A panel of γδ-TCR expressing target cells with different variable chain usage was created. This panel included a lymphoma-patient derived Vγ2Vδ1 TCR and several other TCRs consisting of Vδ1 chains with different CDR3 regions paired to various Vγ chains. Vγ9Vδ2- and αβ-TCR expressing target cells served as control cells. The Vδ1-specific CAR was highly expressed in Jurkat E.6.1 reporter cells. Co-culture experiments revealed strong activation of reporter cells by all Vδ1 chain expressing target cells whereas no reactivity towards Vδ1 negative TCRs was detected.
Conclusion We show highly specific activation of Vδ1-specific CAR T cells in a reporter cell based in-vitro model of γδ-TCL. Provided that a panel of V-segment specific CARs would be created, these CAR-T cells would be able to specifically kill malignant T cells with very limited collateral damage to the non-malignant T cell repertoire. Unlike CDR3 regions the V-segments are largely unaffected by V(D)J-recombination and the panel of CARs could be reused for multiple patients as part of a personalized, precision immunotherapeutic treatment approach. We therefore propose the TCR V-segments as ideal targets for immunotherapies against TCLs.
Disclosure Information M.A. Funk: None. P.M. Brunner: None. C. Jonak: None. M. Deseke: None. I. Prinz: None. J. Leitner: None. J. Stöckl: None. P. Steinberger: None.
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