CD19-specific triplebody SPM-1 engages NK and γδ T cells for rapid and efficient lysis of malignant B-lymphoid cells.

// Christian B. Schiller 1, * , Todd A. Braciak 2, * , Nadja C. Fenn 1, * , Ursula J. E. Seidel 3, * , Claudia C. Roskopf 2, * , Sarah Wildenhain 1, * , Annemarie Honegger * , Ingo A. Schubert 5 , Alexandra Schele 1 , Kerstin Lammermann 2 , Georg H. Fey 6 , Uwe Jacob 6 , Peter Lang 3 , Karl-Peter Hopfner 1, # , Fuat S. Oduncu 2, # 1 Department of Biochemistry and Gene Center, Ludwig-Maximilians-University, Munich, Germany 2 Division of Hematology and Oncology, Medizinische Klinik und Poliklinik IV, Klinikum der Universitat Munchen, Munich, Germany 3 Department of General Paediatrics, Oncology/Haematology, University Children’s Hospital Tubingen, Tubingen, Germany 4 Department of Biochemistry, University of Zurich, Zurich, Switzerland 5 Department of Biology, University of Erlangen-Nuremberg, Erlangen, Germany 6 Westend-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 Correspondence to: 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 ABSTRACT 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. EC 50 -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.