title: Discovery of hapten-specific scFv from a phage display library and applications for HER2-positive tumor imaging.
abstract: In this study, an anti-hapten antibody (single chain Fv, scFv) against a hapten probe was developed as a unique reporter system for molecular imaging or therapy. The hapten peptide (histamine-succinyl-GSYK, Him) was synthesized for phage displayed scFv affinity selection and for conjugation with cypate (Cy-Him) for in vivo near-infrared (NIR) optical imaging. Hapten-specific scFvs were affinity selected from the human single fold phage display scFv libraries (Tomlinson I + J) with high specificity and affinity. Utilizing HER2 targeting as a model system, the highest affinity scFv (clone J42) was recombinantly fused to an anti-HER2 affibody (scFv-L-Aff) with no loss of affinity of either protein. The functionality of the hapten-scFv reporter system was tested in vitro with a HER2-positive human breast cancer cell line, SK-BR3, and in vivo with SK-BR3 xenografts. ScFv-L-Aff mediated the binding of the hapten to HER2 on SK-BR3 cells and from tissue from the SK-BR3 xenograft; however, scFv-L-Aff did not mediate uptake of the hapten in the SK-BR3 xenografted tumors, presumably due to rapid internalization of the HER2/scFv-L-Aff complex. Our results suggest that this hapten-peptide and anti-hapten scFv can be a universal reporter system in a wide range of imaging and therapeutic applications.
authors.affiliation: Molecular Imaging Laboratory, Department of Radiology, University of Pittsburgh , 100 Technology Drive, Pittsburgh, Pennsylvania 15219, United States.
authors.forename: Hye-Yeong
authors.initials: HY
authors.lastname: Kim
authors.forename: Xiaolei
authors.initials: X
authors.lastname: Wang
authors.forename: Brendon
authors.initials: B
authors.lastname: Wahlberg
authors.forename: W Barry
authors.initials: WB
authors.lastname: Edwards
mesh_terms: D000818:Animals; D015153:Blotting, Western; D001943:Breast Neoplasms / Q000276:immunology / Q000378:metabolism / Q000473:pathology*; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D005456:Fluorescent Dyes; D006241:Haptens / Q000276:immunology* / Q000378:metabolism; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D057054:Molecular Imaging / Q000379:methods*; D010446:Peptide Fragments / Q000276:immunology* / Q000378:metabolism; D019151:Peptide Library; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000276:immunology* / Q000378:metabolism; D057127:Single-Chain Antibodies / Q000276:immunology* / Q000378:metabolism; D019265:Spectroscopy, Near-Infrared; D014407:Tumor Cells, Cultured; D023041:Xenograft Model Antitumor Assays
chemical_list: D005456:Fluorescent Dyes; D006241:Haptens; D010446:Peptide Fragments; D019151:Peptide Library; D018719:Erb-b2 Receptor Tyrosine Kinases; D057127:Single-Chain Antibodies