CD19-specific triplebody SPM-1 engages NK and γδ T cells for rapid and efficient lysis of malignant B-lymphoid cells
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Christian B. Schiller1,*, Todd A. Braciak2,*, Nadja C. Fenn1,*, Ursula J. E. Seidel3,*, Claudia C. Roskopf2,*, Sarah Wildenhain1,*, Annemarie Honegger*, Ingo A. Schubert5, Alexandra Schele1, Kerstin Lämmermann2, Georg H. Fey6, Uwe Jacob6, Peter Lang3, Karl-Peter Hopfner1,#, Fuat S. Oduncu2,#
1Department of Biochemistry and Gene Center, Ludwig-Maximilians-University, Munich, Germany
2Division of Hematology and Oncology, Medizinische Klinik und Poliklinik IV, Klinikum der Universität München, Munich, Germany
3Department of General Paediatrics, Oncology/Haematology, University Children’s Hospital Tübingen, Tübingen, Germany
4Department of Biochemistry, University of Zurich, Zurich, Switzerland
5Department of Biology, University of Erlangen-Nuremberg, Erlangen, Germany
6Westend-Innovation, Munich, Germany
*CBS, TAB, NCF, UJES, CCR and SW are co-first authors in this study
#KPH and FSO are co-senior authors in this study
Claudia C. Roskopf, email: Claudia.Roskopf@med.uni-muenchen.de
Keywords: single chain triplebody, antibody-dependent cellular cytotoxicity, gamma delta T cell, leukemia, immunotherapy
Received: June 30, 2016 Accepted: October 03, 2016 Published: November 04, 2016
Triplebodies are antibody-derived recombinant proteins carrying 3 antigen-binding domains in a single polypeptide chain. Triplebody SPM-1 was designed for lysis of CD19-bearing malignant B-lymphoid cells through the engagement of CD16-expressing cytolytic effectors, including NK and γδ T cells.
SPM-1 is an optimized version of triplebody ds(19-16-19) and includes humanization, disulfide stabilization and the removal of potentially immunogenic sequences. A three-step chromatographic procedure yielded 1.7 - 5.5 mg of purified, monomeric protein per liter of culture medium. In cytolysis assays with NK cell effectors, SPM-1 mediated potent lysis of cancer-derived B cell lines and primary cells from patients with various B-lymphoid malignancies, which surpassed the ADCC activity of the therapeutic antibody Rituximab. EC50-values ranged from 3 to 86 pM. Finally, in an impedance-based assay, SPM-1 mediated a particularly rapid lysis of CD19-bearing target cells by engaging and activating both primary and expanded human γδ T cells from healthy donors as effectors.
These data establish SPM-1 as a useful tool for a kinetic analysis of the cytolytic reactions mediated by γδ T and NK cells and as an agent deserving further development towards clinical use for the treatment of B-lymphoid malignancies.
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