Generation of CD20-specific TCRs for TCR gene therapy of CD20low B-cell malignancies insusceptible to CD20-targeting antibodies

// Lorenz Jahn 1 , Dirk M. van der Steen 1 , Renate S. Hagedoorn 1 , Pleun Hombrink 1, 2 , Michel G.D. Kester 1 , Marjolein P. Schoonakker 1 , Danielle de Ridder 1 , Peter A. van Veelen 3, 4 , J.H. Frederik Falkenburg 1 , Mirjam H.M. Heemskerk 1 1 Department of Hematology, Leiden University Medical Center, 2300 RC Leiden, The Netherlands 2 Department of Hematopoiesis, Sanquin Research, 1006 AD Amsterdam, The Netherlands 3 Department of Immunohematology and Blood Transfusion, Leiden University Medical Center, 2300 RC Leiden, The Netherlands 4 Center for Proteomics and Metabolomics, Leiden University Medical Center, 2300 RC Leiden, The Netherlands Correspondence to: Lorenz Jahn, email: l.jahn@lumc.nl Mirjam H.M. Heemskerk, email: m.h.m.heemskerk@lumc.nl Keywords: CD20, TCR gene transfer, monoclonal antibodies, immunotherapy, B-cell leukemia and lymphoma Received: August 20, 2016 Accepted: October 13, 2016 Published: October 20, 2016 ABSTRACT Immunotherapy of B-cell leukemia and lymphoma with CD20-targeting monoclonal antibodies (mAbs) has demonstrated clinical efficacy. However, the emergence of unresponsive disease due to low or absent cell surface CD20 urges the need to develop additional strategies. In contrast to mAbs, T-cells via their T-cell receptor (TCR) can recognize not only extracellular but also intracellular antigens in the context of HLA molecules. We hypothesized that T-cells equipped with high affinity CD20-targeting TCRs would be able to recognize B-cell malignancies even in the absence of extracellular CD20. We isolated CD8 + T-cell clones binding to peptide-MHC-tetramers composed of HLA-A*02:01 and CD20-derived peptide SLFLGILSV (CD20 SLF ) from HLA-A*02:01 neg healthy individuals to overcome tolerance towards self-antigens such as CD20. High avidity T-cell clones were identified that readily recognized and lysed primary HLA-A2 pos B-cell leukemia and lymphoma in the absence of reactivity against CD20-negative but HLA-A2 pos healthy hematopoietic and nonhematopoietic cells. The T-cell clone with highest avidity efficiently lysed malignant cell-lines that had insufficient extracellular CD20 to be targeted by CD20 mAbs. Transfer of this TCR installed potent CD20-specificity onto recipient T-cells and led to lysis of CD20 low malignant cell-lines. Moreover, our approach facilitates the generation of an off-the-shelf TCR library with broad applicability by targeting various HLA alleles. Using the same methodology, we isolated a T-cell clone that efficiently lysed primary HLA-B*07:02 pos B-cell malignancies by targeting another CD20-derived peptide. TCR gene transfer of high affinity CD20-specific TCRs can be a valuable addition to current treatment options for patients suffering from CD20 low B-cell malignancies.