Synthesis and in vitro evaluation of cyclic NGR peptide targeted thermally sensitive liposome.
The Asn-Gly-Arg (NGR) motif in both cyclic and linear form has previously been shown to specifically bind to CD13/aminopeptidase N that is selectively overexpressed in tumor vasculature and some tumor cells. However, previous versions of cyclic NGR used a liable disulfide bridge between cysteine residues that may be problematic for liposome targeting due to disulfide bond formation between adjacent peptides on the liposomal surface. In this study, we report the design, synthesis, and characterization of a novel cyclic NGR-containing peptide, cKNGRE, which does not contain a disulfide bridge. cKNGRE was synthesized in good yield and purity and attached to the fluorescent reporter Oregon Green (cKNGRE-OG) and lysolipid-containing temperature sensitive liposomes (LTSLs). The identity of cKNGRE was verified with NMR and mass spectral techniques. In vitro fluorescence microscopy evaluation of cKNGRE-OG demonstrated binding and active uptake by CD13(+) cancer cells and minimal binding to CD13(-) cancer cells. The cKNGRE-OG ligand displayed 3.6-fold greater affinity for CD13(+) cancer cells than a linear NGR-containing peptide. Affinity for CD13(+) cancer cells was similarly improved 10-fold for both the cyclic and linear NGR when presented in a multivalent fashion on the surface of an LTSL. cKNGRE-targeted LTSLs rapidly released (>75% in <4s) doxorubicin at 41.3 degrees C with minimal release at 37 degrees C. These results demonstrate the ability to synthesize a cKNGRE-targeted temperature sensitive liposome that lacks a disulfide bridge and has sufficient binding affinity for biological applications.
Negussie
Ayele H
AH
Radiology and Imaging Sciences, Clinical Center, National Institutes of Health, Bethesda, MD 20892, USA.
Miller
Jenna L
JL
Reddy
Goutham
G
Drake
Steven K
SK
Wood
Bradford J
BJ
Dreher
Matthew R
MR
D000903:Antibiotics, Antineoplastic / Q000008:administration & dosage; D018826:CD13 Antigens / Q000378:metabolism*; D045744:Cell Line, Tumor; D004317:Doxorubicin / Q000008:administration & dosage; D006801:Humans; D008081:Liposomes / Q000138:chemical synthesis* / Q000737:chemistry / Q000378:metabolism*; D009842:Oligopeptides / Q000138:chemical synthesis* / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D013696:Temperature
D000903:Antibiotics, Antineoplastic; D008081:Liposomes; C526428:NGR peptide; D009842:Oligopeptides; D004317:Doxorubicin; D018826:CD13 Antigens