Identification of CD21-binding peptides with phage display and investigation of binding properties of HPMA copolymer-peptide conjugates.
Cancer targeting with peptides has become promising with the emergence of combinatorial peptide techniques such as phage display. Using phage display under stringent screening conditions, we selected five distinct peptides that specifically recognized the CD21 receptor, a cell surface marker of malignant B cell lymphoma. Two highly hydrophobic sequences were excluded (RLAYWCFSGLFLLVC and PVAAVSFVPYLVKTY). The binding affinity toward CD21 of the other three selected peptides (RMWPSSTVNLSAGRR, PNLDFSPTCSFRFGC, and GRVPSMFGGHFFFSR) was analyzed with fluorescence quenching. Their dissociation constants were determined to be within the micromolar range. On the basis of the results of phage ELISA, competitive phage ELISA, and fluorescence quenching, the binding sites of the three selected peptides were found to reside within the first four short consensus repeats of CD21 (SCR1-4). The peptide RMWPSSTVNLSAGRR (P1) was bound to the N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer, a potential drug carrier for chemotherapeutic agents, and the surface binding properties of HPMA copolymer-P1 conjugates were investigated. Specific interactions were observed between HPMA copolymer-P1 conjugates and surface-bound receptor. Binding of HPMA copolymer-P1 conjugates was directly related to the amount of surface (MaxiSorp plate) bound receptor, and the binding of the conjugates could be inhibited by the application of a 3-4 orders-of-magnitude excess of free peptide over the peptide concentration in conjugates. The enhanced binding of polymer-bound peptide was ascribed to multivalent interactions between the HPMA copolymer-P1 conjugate and the surface-bound CD21 receptor.
Department of Pharmaceutics and Pharmaceutical Chemistry, University of Utah, Salt Lake City, Utah 84112, USA.
Hui
H
Ding
Wolfgang M
WM
Prodinger
Jindrich
J
Kopecek
D000595:Amino Acid Sequence; D004797:Enzyme-Linked Immunosorbent Assay; D008024:Ligands; D008689:Methacrylates / Q000737:chemistry* / Q000378:metabolism; D008969:Molecular Sequence Data; D015394:Molecular Structure; D019151:Peptide Library; D010455:Peptides / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D017464:Receptors, Complement 3d / Q000235:genetics / Q000378:metabolism*
D008024:Ligands; D008689:Methacrylates; D019151:Peptide Library; D010455:Peptides; D017464:Receptors, Complement 3d; C032802:hydroxypropyl methacrylate