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A general chemical synthesis platform for crosslinking multivalent single chain variable fragments.
MedLine Citation:
PMID:  22132412     Owner:  NLM     Status:  Publisher    
Abstract/OtherAbstract:
Multivalent single chain variable fragments (scFv) show increased affinity to tumor-associated antigens compared to monovalent scFv and intact monoclonal antibodies (mAb). Multivalent constructs can be derived from self-associating or covalent scFv with covalent constructs offering improved in vivo and in vitro stability. Covalent attachment of scFv can be achieved using genetically engineered expression vectors that afford scFv with site specific cysteine functionality. Expression vectors for di-scFv-C wherein the cysteine is located in the center of two scFv have also been developed for attaching chemically reactive linkers. In the example illustrated here, the di-scFv-C is derived from a mAb directed against the MUC1 epitope, which is presented on cancer cells. To achieve multivalency, a chemical crosslinking strategy utilizing various azide and multi-alkyne functionalized polyethylene glycol (PEG) linkers was implemented. Conjugation was achieved by attachment of these linkers to the scFv thiol functionality. Chemoselective ligation was employed to covalently link different protein conjugates via copper(i) catalyzed azide alkyne 1,3-dipolar cycloaddition reaction (CuAAC) chemistry. Ligations were achieved in >70% yield using a specific set of linkers as determined by SDS-PAGE and densitometry. ELISA showed increased tumor binding of a tetravalent scFv providing a versatile chemical crosslinking strategy for construction of multivalent and bi-specific immunoconjugates that retain biological activity and have potential application in pre-targeted radioimmunotherapy and imaging.
Authors:
Joan G Schellinger; Avinash Kudupudi; Arutselvan Natarajan; Wenjun Du; Sally J Denardo; Jacquelyn Gervay-Hague
Publication Detail:
Type:  JOURNAL ARTICLE     Date:  2011-12-1
Journal Detail:
Title:  Organic & biomolecular chemistry     Volume:  -     ISSN:  1477-0539     ISO Abbreviation:  -     Publication Date:  2011 Dec 
Date Detail:
Created Date:  2011-12-1     Completed Date:  -     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  101154995     Medline TA:  Org Biomol Chem     Country:  -    
Other Details:
Languages:  ENG     Pagination:  -     Citation Subset:  -    
Affiliation:
Chemistry Department, University of California Davis, One Shields Avenue, Davis, CA95616, USA. Gervay@chem.ucdavis.edu.
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