Dual display: phage selection driven by co-engagement of two targets by two different antibody fragments

Article Article scientifique Article Dual display: phage selection driven by co-engagement of two targets by two different antibody fragments eng 2017 author Fagete Séverine 649164 author Botas Perez Ledicia 785572 author Rossitto-Borlat Irène 23207 author Adea Kenneth 631795 author Gueneau Franck author Ravn Ulla author Rousseau François author Kosco-Vilbois Marie author Fischer Nicolas author Hartley Oliver 0000-0002-2283-8379 507767 575-582 10.1093/protein/gzx021 28444391 1741-0126 Antibody phage display technology has supported the emergence of numerous therapeutic antibodies. The development of bispecific antibodies, a promising new frontier in antibody therapy, could be facilitated by new phage display approaches that enable pairs of antibodies to be co-selected based on co-engagement of their respective targets. We describe such an approach, making use of two complementary leucine zipper domains that heterodimerize with high affinity. Phagemids encoding a first antibody fragment (scFv) fused to phage coat protein via the first leucine zipper are rescued in bacteria expressing a second scFv fused to the second leucine zipper as a soluble periplasmic protein, so that it is acquired by phage during assembly. Using a soluble scFv specific for a human CD3-derived peptide, we show that its acquisition by phage displaying an irrelevant antibody is sufficiently robust to drive selection of rare phage (1 in 10(5)) over three rounds of panning. We then set up a model selection experiment using a cell line expressing the chemokine receptor CCR5 fused to the CD3 peptide together with a panel of phage clones capable displaying either an anti-CCR5 scFv or an irrelevant antibody, with or without the capacity to acquire the soluble anti-CD3 scFv. In this experiment we showed that rare phage (1 in 10(5)) capable of displaying the two different scFvs can be specifically enriched over four rounds of panning. This approach has the potential to be applied to the identification of pairs of ligands capable of co-engaging two different user-defined targets, which would facilitate the discovery of novel bispecific antibodies. Amino Acid Sequence Animals Antibodies Bispecific/biosynthesis/genetics Antibodies Monoclonal/biosynthesis/genetics Antibody Specificity Bacteriophages/genetics/immunology CD3 Complex/genetics/immunology CHO Cells Capsid Proteins/genetics/immunology Cricetulus Escherichia coli/genetics/metabolism/virology Gene Expression Humans Leucine Zippers Peptide Library Peptides/genetics/immunology Receptors CCR5/genetics/immunology Recombinant Fusion Proteins/genetics/immunology Sequence Analysis DNA Single-Chain Antibodies/biosynthesis/genetics Dewey 616 Dewey 616.07 54 Département de microbiologie et médecine moléculaire 21B 137 Département de pathologie et immunologie 21F false 497 Protein and peptide engineering for medicine 835 Protein Engineering, Design and Selection 30 9 0ce53c4f-8848-4b7b-875d-eec22720d3af Article (Published version) Restricted gzx021.pdf 642775 application/pdf restrictedAccess Faculté de médecine / Section de médecine fondamentale / Département de microbiologie et médecine moléculaire Faculté de médecine / Section de médecine fondamentale / Département de pathologie et immunologie

Title	Published in	Year	Views	Downloads

Dual display: phage selection driven by co-engagement of two targets by two different antibody fragments	Protein Engineering, Design and Selection	2017	483	1
Pan chemokine inhibitors : from viral natural solutions to engineered antibodies		2011	948	0

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Fagete, Séverine