A recombinant trispecific single‐chain Fv derivative directed against CD123 and CD33 mediates effective elimination of acute myeloid leukaemia cells by dual targeting

• Published in: British journal of haematology Vol. 150; no. 5; pp. 574 - 586
• Main Authors: Kügler, Markus, Stein, Christoph, Kellner, Christian, Mentz, Kristin, Saul, Domenica, Schwenkert, Michael, Schubert, Ingo, Singer, Heiko, Oduncu, Fuat, Stockmeyer, Bernhard, Mackensen, Andreas, Fey, Georg H.
• Format: Journal Article
• Language: English
• Published: Oxford, UK Blackwell Publishing Ltd 01-09-2010; Blackwell; Wiley
• Subjects: Antibodies; Antibody-Dependent Cell Cytotoxicity - immunology; Antibody-dependent cell-mediated cytotoxicity; Antigens, CD - immunology; Antigens, Differentiation, Myelomonocytic - immunology; Biological and medical sciences; Bone marrow; CD123; CD123 antigen; CD16 antigen; CD33; dual targeting; Effector cells; GPI-Linked Proteins; Hematologic and hematopoietic diseases; Humans; Immunoglobulin Fragments - immunology; Interleukin 3; Interleukin-3 Receptor alpha Subunit - immunology; Killer Cells, Natural - immunology; leukaemia stem cell; Leukemia, Myeloid, Acute - immunology; Leukemia, Myeloid, Acute - pathology; Leukemias. Malignant lymphomas. Malignant reticulosis. Myelofibrosis; Medical sciences; Peripheral blood; Receptors, IgG - immunology; Recombinant Proteins - immunology; Sialic Acid Binding Ig-like Lectin 3; Single-Chain Antibodies - immunology; single‐chain Fv triplebody; Tumor Cells, Cultured; Acute myelogenous leukemia; Targeting; Hematology; Stem cell; single-chain Fv triplebody; Malignant hemopathy; dual targeting; Efficiency; Recombinant protein; leukaemia stem cell; CD123; CD33; Cancer; Medicine
• ISSN: 0007-1048; 1365-2141
• DOI: 10.1111/j.1365-2141.2010.08300.x

Summary
Two trivalent constructs consisting of single‐chain Fv antibody fragments (scFvs) specific for the interleukin‐3 receptor α chain (CD123), CD33 and the Fcγ‐receptor III (CD16) were designed and characterized for the elimination of acute myeloid leukaemia (AML) cells. The dual targeting single‐chain Fv triplebody (sctb) [123 × ds16 × 33] and the mono targeting sctb [123 × ds16 × 123] both specifically bound their respective target antigens and were stable in human serum at 37°C for at least 5 d. Both constructs induced potent antibody‐dependent cellular cytotoxicity (ADCC) of two different AML‐derived CD33‐ and CD123 double‐positive cell lines in the low picomolar range using isolated mononuclear cells (MNCs) as effector cells. In these experiments the dual targeting molecule produced significantly stronger lysis than the mono targeting agent. In addition, the sctbs showed a high potency in mediating ADCC of primary leukaemia cells isolated from peripheral blood or bone marrow of seven AML patients. Hence, these novel molecules displayed potent anti‐leukaemic effects against AML cells in vitro and represent attractive candidates for further preclinical development.