Lysosome-associated membrane glycoprotein 1 predicts fratricide amongst T cell receptor transgenic CD8+ T cells directed against tumor-associated antigens
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Andreas Kirschner1, Melanie Thiede1, Franziska Blaeschke1,2, Günther H.S. Richter1, Julia S. Gerke3, Michaela C. Baldauf3, Thomas G.P. Grünewald3,4,5, Dirk H. Busch6, Stefan Burdach1,*, Uwe Thiel1,*
1Laboratory for Functional Genomics and Transplantation Biology, Departments of Pediatrics and Children’s Cancer Research Center, Klinikum rechts der Isar, Technische Universität München, Munich, Germany
2Laboratory for Immunotherapy, Dr. von Hauner Children’s Hospital, Medical center of the LMU Munich, Munich, Germany
3Laboratory for Pediatric Sarcoma Biology, Institute of Pathology of the LMU Munich, Munich, Germany
4German Cancer Consortium (DKTK), Heidelberg, Germany
5German Cancer Research Center (DKFZ), Heidelberg, Germany
6Institute for Medical Microbiology, Immunology and Hygiene, Technische Universität München, Munich, Germany
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Andreas Kirschner, email: [email protected]
Keywords: fratricide, CD107a, TCR transgenic T cells, cross reactivity, amino-acid exchange scan
Received: April 18, 2016 Accepted: May 30, 2016 Published: July 18, 2016
Aim: Autologous as well as allogeneic CD8+ T cells transduced with tumor antigen specific T cell receptors (TCR) may cause significant tumor lysis upon adoptive transfer. Besides unpredictable life-threatening off-target effects, these TCRs may unexpectedly commit fratricide. We hypothesized lysosome-associated membrane glycoprotein 1 (LAMP1, CD107a) to be a marker for fratricide in TCR transgenic CD8+ T cells.
Methods: We identified HLA-A*02:01/peptide-restricted T cells directed against ADRB3295. After TCR identification, we generated HLA-A*02:01/peptide restricted TCR transgenic T cells by retroviral transduction and tested T cell expansion rates as well as A*02:01/peptide recognition and ES killing in ELISpot and xCELLigence assays. Expansion arrest was analyzed via Annexin and CD107a staining. Results were compared to CHM1319-TCR transgenic T cells.
Results: Beta-3-adrenergic receptor (ADRB3) as well as chondromodulin-1 (CHM1) are over-expressed in Ewing Sarcoma (ES) but not on T cells. TCR transgenic T cells demonstrated HLA-A*02:01/ADRB3295 mediated ES recognition and killing in ELISpot and xCELLigence assays. 24h after TCR transduction, CD107a expression correlated with low expansion rates due to apoptosis of ADRB3 specific T cells in contrast to CHM1 specific transgenic T cells. Amino-acid exchange scans clearly indicated the cross-reactive potential of HLA-A*02:01/ADRB3295- and HLA-A*02:01/CHM1319-TCR transgenic T cells. Comparison of peptide motive binding affinities revealed extended fratricide among ADRB3295 specific TCR transgenic T cells in contrast to CHM1319.
Conclusion: Amino-acid exchange scans alone predict TCR cross-reactivity with little specificity and thus require additional assessment of potentially cross-reactive HLA-A*02:01 binding candidates. CD107a positivity is a marker for fratricide of CD8+ TCR transgenic T cells.
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