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. 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.
Highlights
CD19 is a type I transmembrane glycoprotein and a signaling receptor of the immunoglobulin superfamily
SPM-1 mediated efficient lysis of primary cells from patients with various B lineage malignancies and of cell lines derived from various B cell neoplasms by NK cells from healthy donors
The humanized and optimized triplebody SPM1 was potent in ADCC as the parental agent, and its production and purification by industry-standard procedures routinely used for commercial production of therapeutic antibodies was found to be possible
Summary
CD19 is a type I transmembrane glycoprotein and a signaling receptor of the immunoglobulin superfamily. It is a component of the B cell receptor (BCR) cocomplex, and is expressed from early to late stages of B cell development. The antigen is promising for targeted immunotherapy of B-lymphoid malignancies, because of its relatively high and lineage-specific expression on B cells of different maturation stages and its presence on the surface of cancer progenitor cells [5,6,7]. CD19 clearly is a useful target for antigen-specific immunotherapy, provided an appropriate molecular format of the therapeutic agent is chosen
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