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Generation of activation-specific human anti-αMβ2 single-chain antibodies as potential diagnostic tools and therapeutic agents

Eisenhardt, Steffen U and Schwarz, Meike and Schallner, Nils and Soosairajah, Juliana and Bassler, Nicole and Huang, Dexing and Bode, Christoph and Peter, Karlheinz (2006) Generation of activation-specific human anti-αMβ2 single-chain antibodies as potential diagnostic tools and therapeutic agents. Blood, 109 (8). pp. 3521-3528. ISSN 0006-4971 (print) 1528-0020 (online)

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Abstract

The leukocyte integrin Mac-1 (αMβ2) plays a pivotal role in inflammation and host defense. Upon leukocyte activation, Mac-1 undergoes a conformational change exposing interaction sites for multiple ligands. We aimed to generate single-chain antibodies (scFv's) directed against activation-specific Mac-1 ligand-binding sites. Using human scFv phage libraries, we developed subtractive strategies with depletion of phages binding to nonactivated Mac-1 and selection of phages binding to activated Mac-1, using monocytes as well as CHO cells transfected with native or mutated, activated Mac-1. Three scFv clones demonstrated exclusive binding to activated Mac-1. Mac-1 binding of the ligands fibrinogen, heparin, and ICAM-1, but not C3bi, was inhibited. Using alanine substitutions, the paratope was identified within the heavy chain HCDR3s of the scFv's. The epitope was localized to Lys245-Arg261 of the αM I-domain. In a pilot study with septicemic patients, we provide initial support for the use of these scFv's as markers of monocyte activation and as potential diagnostic tools. Potential therapeutic use was tested in adhesion assays under static and flow conditions demonstrating the selective blockade of activated monocytes only. Furthermore, scFv HCDR3–derived peptides retain selectivity for the activated integrin, providing a unique template for the potential development of inhibitors that are specific for the activated Mac-1.

Item Type: Article
Uncontrolled Keywords: antibodies, leukocyte activation, monocyte activation, diagnostic tools, inflammation, therapeutic strategy
Subjects: FOR Classification > 0304 Medicinal and Bimolecular Chemistry
Faculty/School/Research Centre/Department > College of Health and Biomedicine
Depositing User: VUIR
Date Deposited: 04 Dec 2013 00:07
Last Modified: 04 Dec 2013 00:07
URI: http://vuir.vu.edu.au/id/eprint/22435
DOI: 10.1182/blood-2006-03-007179
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Citations in Scopus: 29 - View on Scopus

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