Abstract
We have designed, synthesized, and characterized peptides containing four repeats of the sequences VAALEKE (peptide E) or VAALKEK (peptide K). While the peptides alone adopt in aqueous solutions a random coil conformation, their equimolar mixture forms heterodimeric coiled coils as confirmed by CD spectroscopy. 5-Azidopentanoic acid was connected to the N-terminus of peptide E via a short poly(ethylene glycol) spacer. The terminal azide group enabled conjugation of the peptide with a synthetic drug carrier based on the N-(2-hydroxypropyl)methacrylamide copolymer containing propargyl groups using "click" chemistry. When incorporated into the polymer drug carrier, peptide E formed a stable noncovalent complex with peptide K belonging to a recombinant single-chain fragment (scFv) of the M75 antibody. The complex thereby mediates a noncovalent linkage between the polymer drug carrier and the protein. The recombinant scFv antibody fragment was selected as a targeting ligand against carbonic anhydrase IX-a marker overexpressed by tumor cells of various human carcinomas. The antigen binding affinity of the polymer-scFv complex was confirmed by ELISA. This approach offers a well-defined, specific, and nondestructive universal method for the preparation of protein (antibody)-targeted polymer drug and gene carriers designed for cell-specific delivery.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Acrylamides / chemistry*
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Antibodies, Monoclonal / chemistry*
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Antibodies, Monoclonal / genetics
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Antibodies, Monoclonal / immunology
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Antigens, Neoplasm / immunology
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Antigens, Neoplasm / metabolism
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Biomarkers, Tumor / immunology
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Biomarkers, Tumor / metabolism
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Carbonic Anhydrase IX
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Carbonic Anhydrases / immunology
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Carbonic Anhydrases / metabolism
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Carcinoma / drug therapy
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Carcinoma / enzymology
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Carcinoma / immunology
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Carcinoma / pathology
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Circular Dichroism
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Click Chemistry / methods*
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Cloning, Molecular
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Dimerization
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Drug Carriers / chemical synthesis*
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Drug Carriers / pharmacology
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Enzyme-Linked Immunosorbent Assay
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Escherichia coli
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Humans
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Immunoconjugates / chemistry*
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Immunoconjugates / immunology
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Immunoconjugates / pharmacology
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Molecular Conformation
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Oligopeptides / chemical synthesis*
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Oligopeptides / immunology
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Oligopeptides / pharmacology
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Plasmids
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Polyethylene Glycols / chemistry
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Recombinant Proteins / chemistry*
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Recombinant Proteins / genetics
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Recombinant Proteins / immunology
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Transformation, Bacterial
Substances
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Acrylamides
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Antibodies, Monoclonal
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Antigens, Neoplasm
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Biomarkers, Tumor
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Drug Carriers
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Immunoconjugates
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M75 monoclonal antibody
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Oligopeptides
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Recombinant Proteins
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Polyethylene Glycols
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CA9 protein, human
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Carbonic Anhydrase IX
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Carbonic Anhydrases
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N-(2-hydroxypropyl)methacrylamide