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

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-MHCtetramers composed of HLA-A*02:01 and CD20-derived peptide SLFLGILSV (CD20SLF) from HLA-A*02:01neg 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-A2pos B-cell leukemia and lymphoma in the absence of reactivity against CD20-negative but HLA-A2pos 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 CD20low malignant cell-lines. Moreover, our approach facilitates the generation of an off-theshelf TCR library with broad applicability by targeting various HLA alleles. Using the same methodology, we isolated a T-cell clone that efficiently lysed primary HLAB* 07:02pos 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 CD20low B-cell malignancies.