title: Human monomeric antibody fragments to TRAIL-R1 and TRAIL-R2 that display potent in vitro agonism.
abstract: Apoptosis through the TRAIL receptor pathway can be induced via agonistic IgG to either TRAIL-R1 or TRAIL-R2. Here we describe the use of phage display to isolate a substantive panel of fully human anti-TRAIL receptor single chain Fv fragments (scFvs); 234 and 269 different scFvs specific for TRAIL-R1 and TRAIL-R2 respectively. In addition, 134 different scFvs that were cross-reactive for both receptors were isolated. To facilitate screening of all 637 scFvs for potential agonistic activity in vitro, a novel high-throughput surrogate apoptosis assay was developed. Ten TRAIL-R1 specific scFv and 6 TRAIL-R2 specific scFv were shown to inhibit growth of tumor cells in vitro in the absence of any cross-linking agents. These scFv were all highly specific for either TRAIL-R1 or TRAIL-R2, potently inhibited tumor cell proliferation, and were antagonists of TRAIL binding. Moreover, further characterization of TRAIL-R1 agonistic scFv demonstrated significant anti-tumor activity when expressed and purified as a monomeric Fab fragment. Thus, scFv and Fab fragments, in addition to whole IgG, can be agonistic and induce tumor cell death through specific binding to either TRAIL-R1 or TRAIL-R2. These potent agonistic scFv were all isolated directly from the starting phage antibody library and demonstrated significant tumor cell killing properties without any requirement for affinity maturation. Some of these selected scFv have been converted to IgG format and are being studied extensively in clinical trials to investigate their potential utility as human monoclonal antibody therapeutics for the treatment of human cancer.
authors.affiliation: MedImmune, Cambridge, Cambridgeshire, UK. dobsonc@medimmune.com
authors.forename: Claire L
authors.initials: CL
authors.lastname: Dobson
authors.forename: Sarah
authors.initials: S
authors.lastname: Main
authors.forename: Philip
authors.initials: P
authors.lastname: Newton
authors.forename: Matthieu
authors.initials: M
authors.lastname: Chodorge
authors.forename: Karen
authors.initials: K
authors.lastname: Cadwallader
authors.forename: Robin
authors.initials: R
authors.lastname: Humphreys
authors.forename: Vivian
authors.initials: V
authors.lastname: Albert
authors.forename: Tristan J
authors.initials: TJ
authors.lastname: Vaughan
authors.forename: Ralph R
authors.initials: RR
authors.lastname: Minter
authors.forename: Bryan M
authors.initials: BM
authors.lastname: Edwards
mesh_terms: D000920:Antibody-Dependent Cell Cytotoxicity / Q000235:genetics / Q000276:immunology; D017209:Apoptosis / Q000187:drug effects / Q000276:immunology; D001667:Binding, Competitive; D053148:Caspase 3 / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D005818:Genetic Engineering; D006367:HeLa Cells; D057166:High-Throughput Screening Assays; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology*; D007167:Immunotherapy; D009369:Neoplasms / Q000276:immunology / Q000473:pathology / Q000628:therapy*; D053220:Receptors, TNF-Related Apoptosis-Inducing Ligand / Q000819:agonists*; D057127:Single-Chain Antibodies / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification / Q000494:pharmacology*
chemical_list: D007140:Immunoglobulin Fab Fragments; D053220:Receptors, TNF-Related Apoptosis-Inducing Ligand; D057127:Single-Chain Antibodies; D053148:Caspase 3