ABSTRACT
Despite new treatment modalities, the clinical outcome in a substantial number of patients with multiple myeloma (MM) has yet to be improved. Antibody-based targeted therapies for myeloma patients could make use of the HM1.24 antigen (CD317), a surface molecule overexpressed on malignant plasma cells and efficiently internalized. Here, a novel immunotoxin, HM1.24-ETA', is described. HM1.24-ETA' was generated by genetic fusion of a CD317-specific single-chain Fv (scFv) antibody and a truncated variant of Pseudomonas aeruginosa exotoxin A (ETA'). HM1.24-ETA' inhibited growth of interleukin 6 (IL-6)-dependent and -independent myeloma cell lines. Half-maximal growth inhibition was observed at concentrations as low as 0.3 nM. Target cell killing occurred via induction of apoptosis and was unaffected in co-culture experiments with bone marrow stromal cells. HM1.24-ETA' efficiently triggered apoptosis of freshly isolated/cryopreserved cells of patients with plasma cell leukemia and MM and was active in a preclinical severe combined immunodeficiency (SCID) mouse xenograft model. Importantly, HM1.24-ETA' was not cytotoxic against CD317-positive cells from healthy tissue (monocytes, human umbilical vein endothelial cells). These results indicate that CD317 may represent a promising target structure for specific and efficient immunotoxin therapy for patients with plasma cell tumors.
Subject(s)
ADP Ribose Transferases/pharmacology , Bacterial Toxins/pharmacology , Exotoxins/pharmacology , Immunotoxins/pharmacology , Multiple Myeloma/drug therapy , Virulence Factors/pharmacology , ADP Ribose Transferases/chemistry , Animals , Antibodies, Monoclonal, Humanized/chemistry , Antibodies, Monoclonal, Humanized/immunology , Antibodies, Monoclonal, Humanized/pharmacology , Antigens, CD/immunology , Apoptosis/drug effects , Apoptosis/immunology , Bacterial Toxins/chemistry , Epitopes , Exotoxins/chemistry , Female , GPI-Linked Proteins/immunology , Human Umbilical Vein Endothelial Cells/drug effects , Humans , Immunotoxins/chemistry , Immunotoxins/immunology , Jurkat Cells , Mice , Mice, SCID , Multiple Myeloma/immunology , Multiple Myeloma/pathology , Virulence Factors/chemistry , Xenograft Model Antitumor Assays , Pseudomonas aeruginosa Exotoxin AABSTRACT
Antibody-drug conjugates (ADC) represent promising agents for targeted cancer therapy. To allow rational selection of human antibodies with favorable characteristics for ADC development a screening tool was designed obviating the need of preparing individual covalently linked conjugates. Therefore, α-kappa-ETA' was designed as a fusion protein consisting of a human kappa light chain binding antibody fragment and a truncated version of Pseudomonas exotoxin A. α-kappa-ETA' specifically bound to human kappa light chains of human or human-mouse chimeric antibodies and Fab fragments. Antibody-redirected α-kappa-ETA' specifically inhibited proliferation of antigen-expressing cell lines at low toxin and antibody concentrations. Selected antibodies that efficiently delivered α-kappa-ETA' in the novel assay system were used to generate scFv-based covalently linked immunotoxins. These molecules efficiently triggered apoptosis of target cells, indicating that antibodies identified in our assay system can be converted to functional immunoconjugates. Finally, a panel of human epidermal growth factor receptor (EGFR) antibodies was screened--demonstrating favorable characteristics with antibody 2F8. These data suggest that antibodies with potential for Pseudomonas exotoxin A-based ADC development can be identified using the novel α-kappa-ETA' conjugate.
