title: Simultaneous inhibition of epidermal growth factor receptor (EGFR) signaling and enhanced activation of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptor-mediated apoptosis induction by an scFv:sTRAIL fusion protein with specificity for human EGFR.
abstract: Epidermal growth factor receptor (EGFR) signaling inhibition by monoclonal antibodies and EGFR-specific tyrosine kinase inhibitors has shown clinical efficacy in cancer by restoring susceptibility of tumor cells to therapeutic apoptosis induction. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a promising anti-cancer agent with tumor-selective apoptotic activity. Here we present a novel approach that combines EGFR-signaling inhibition with target cell-restricted apoptosis induction using a TRAIL fusion protein with engineered specificity for EGFR. This fusion protein, scFv425:sTRAIL, comprises the EGFR-blocking antibody fragment scFv425 genetically fused to soluble TRAIL (sTRAIL). Treatment with scFv425:sTRAIL resulted in the specific accretion to the cell surface of EGFR-positive cells only. EGFR-specific binding rapidly induced a dephosphorylation of EGFR and down-stream mitogenic signaling, which was accompanied by cFLIP(L) down-regulation and Bad dephosphorylation. EGFR-specific binding converted soluble scFv425:sTRAIL into a membrane-bound form of TRAIL that cross-linked agonistic TRAIL receptors in a paracrine manner, resulting in potent apoptosis induction in a series of EGFR-positive tumor cell lines. Co-treatment of EGFR-positive tumor cells with the EGFR-tyrosine kinase inhibitor Iressa resulted in a potent synergistic pro-apoptotic effect, caused by the specific down-regulation of c-FLIP. Furthermore, in mixed culture experiments binding (L)of scFv425:sTRAIL to EGFR-positive target cells conveyed a potent apoptotic effect toward EGFR-negative bystander tumor cells. The favorable characteristics of scFv425:sTRAIL, alone and in combination with Iressa, as well as its potent anti-tumor bystander activity indicate its potential value for treatment of EGFR-expressing cancers.
authors.affiliation: Groningen University Institute for Drug Exploration, University Medical Center Groningen Department of Pathology and Laboratory Medicine, Section Medial Biology, Laboratory for Tumor Immunology, 9713 GZ Groningen, The Netherlands.
authors.forename: Edwin
authors.initials: E
authors.lastname: Bremer
authors.forename: Douwe F
authors.initials: DF
authors.lastname: Samplonius
authors.forename: Linda
authors.initials: L
authors.lastname: van Genne
authors.forename: Marike H
authors.initials: MH
authors.lastname: Dijkstra
authors.forename: Bart Jan
authors.initials: BJ
authors.lastname: Kroesen
authors.forename: Lou F M H
authors.initials: LF
authors.lastname: de Leij
authors.forename: Wijnand
authors.initials: W
authors.lastname: Helfrich
mesh_terms: D000911:Antibodies, Monoclonal / Q000737:chemistry; D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis; D053181:Caspase 8; D020169:Caspases / Q000378:metabolism; D002460:Cell Line; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism; D002470:Cell Survival; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D066246:ErbB Receptors / Q000037:antagonists & inhibitors*; D005434:Flow Cytometry; D000077156:Gefitinib; D006801:Humans; D007135:Immunoglobulin Variable Region / Q000737:chemistry; D019169:Jurkat Cells; D008564:Membrane Potentials; D008954:Models, Biological; D010766:Phosphorylation; D011485:Protein Binding; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011799:Quinazolines / Q000494:pharmacology; D053220:Receptors, TNF-Related Apoptosis-Inducing Ligand; D018124:Receptors, Tumor Necrosis Factor / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015398:Signal Transduction; D057127:Single-Chain Antibodies; D013997:Time Factors
chemical_list: D000911:Antibodies, Monoclonal; D000970:Antineoplastic Agents; D004791:Enzyme Inhibitors; D007135:Immunoglobulin Variable Region; D047428:Protein Kinase Inhibitors; D011799:Quinazolines; D053220:Receptors, TNF-Related Apoptosis-Inducing Ligand; D018124:Receptors, Tumor Necrosis Factor; D011993:Recombinant Fusion Proteins; D057127:Single-Chain Antibodies; C105190:TNFRSF10A protein, human; C114035:scFv 425; D066246:ErbB Receptors; C505530:CASP8 protein, human; D053181:Caspase 8; D020169:Caspases; D000077156:Gefitinib