(64)Cu-labeled tetrameric and octameric RGD peptides for small-animal PET of tumor alpha(v)beta(3) integrin expression.
Integrin alpha(v)beta(3) plays a critical role in tumor angiogenesis and metastasis. Suitably radiolabeled cyclic arginine-glycine-aspartic (RGD) peptides can be used for noninvasive imaging of alpha(v)beta(3) expression and targeted radionuclide therapy. In this study, we developed (64)Cu-labeled multimeric RGD peptides, E{E[c(RGDyK)](2)}(2) (RGD tetramer) and E(E{E[c(RGDyK)](2)}(2))(2) (RGD octamer), for PET imaging of tumor integrin alpha(v)beta(3) expression.


METHODS
Both RGD tetramer and RGD octamer were synthesized with glutamate as the linker. After conjugation with 1,4,7,10-tetra-azacyclododecane-N,N',N'',N'''-tetraacetic acid (DOTA), the peptides were labeled with (64)Cu for biodistribution and small-animal PET imaging studies (U87MG human glioblastoma xenograft model and c-neu oncomouse model). A cell adhesion assay, a cell-binding assay, receptor blocking experiments, and immunohistochemistry were also performed to evaluate the alpha(v)beta(3)-binding affinity/specificity of the RGD peptide-based conjugates in vitro and in vivo.


RESULTS
RGD octamer had significantly higher integrin alpha(v)beta(3)-binding affinity and specificity than RGD tetramer analog (inhibitory concentration of 50% was 10 nM for octamer vs. 35 nM for tetramer). (64)Cu-DOTA-RGD octamer had higher tumor uptake and longer tumor retention than (64)Cu-DOTA-RGD tetramer in both tumor models tested. The integrin alpha(v)beta(3) specificity of both tracers was confirmed by successful receptor-blocking experiments. The high uptake and slow clearance of (64)Cu-DOTA-RGD octamer in the kidneys was attributed mainly to the integrin positivity of the kidneys, significantly higher integrin alpha(v)beta(3)-binding affinity, and the larger molecular size of the octamer, as compared with the other RGD analogs.


CONCLUSIONS
Polyvalency has a profound effect on the receptor-binding affinity and in vivo kinetics of radiolabeled RGD multimers. The information obtained here may guide the future development of RGD peptide-based imaging and internal radiotherapeutic agents targeting integrin alpha(v)beta(3).
Li
Zi-Bo
ZB
Molecular Imaging Program at Stanford, Department of Radiology and Bio-X Program, Stanford University School of Medicine, Stanford, California 94305, USA.
Cai
Weibo
W
Cao
Qizhen
Q
Chen
Kai
K
Wu
Zhanhong
Z
He
Lina
L
Chen
Xiaoyuan
X
D000818:Animals; D002448:Cell Adhesion; D045744:Cell Line, Tumor; D003301:Copper Radioisotopes / Q000493:pharmacokinetics*; D005260:Female; D006573:Heterocyclic Compounds, 1-Ring / Q000737:chemistry; D006801:Humans; D039302:Integrin alphaVbeta3 / Q000378:metabolism*; D051379:Mice; D008819:Mice, Nude; D009368:Neoplasm Transplantation; D009842:Oligopeptides / Q000138:chemical synthesis / Q000493:pharmacokinetics*; D049268:Positron-Emission Tomography; D019275:Radiopharmaceuticals / Q000138:chemical synthesis / Q000493:pharmacokinetics*; D014018:Tissue Distribution; D014183:Transplantation, Heterologous
D003301:Copper Radioisotopes; D006573:Heterocyclic Compounds, 1-Ring; D039302:Integrin alphaVbeta3; D009842:Oligopeptides; D019275:Radiopharmaceuticals; C071349:1,4,7,10-tetraazacyclododecane- 1,4,7,10-tetraacetic acid; C047981:arginyl-glycyl-aspartic acid