Hybrid PET/optical imaging of integrin α(V)β(3) receptor expression using a (64)Cu-labeled streptavidin/biotin-based dimeric RGD peptide

BACKGROUND: Hybrid PET/optical imaging provides quantitative and complementary information for diagnosis of tumors. Herein, we developed a (64)Cu-labeled AlexaFluor 680-streptavidin ((AF)SAv)/biotin-based dimeric cyclic RGD peptide (RGD(2)) for hybrid PET/optical imaging of integrin α(V)β(3) expression. METHODS: (64)Cu-1,4,7,10-tetraazacyclododecane-N,N′,N′′,N′′′-tetraacetic acid (DOTA)-(AF)SAv/biotin-PEG-RGD(2) was prepared by formation of a complex comprising DOTA-(AF)SAv and biotin-PEG-RGD(2), followed by radiolabeling with (64)Cu. Receptor binding studies of DOTA-(AF)SAv/biotin-PEG-RGD(2) were performed using U87MG cells and (125)I-RGDyK as the radioligand, and cellular uptake studies of (64)Cu-DOTA-(AF)SAv/biotin-PEG-RGD(2) were also performed. MicroPET imaging followed by optical imaging of U87MG tumor-bearing mice was acquired after injection of the hybrid probe, and region of interest (ROI) analysis of tumors was performed. Ex vivo PET/optical imaging and biodistribution studies of the major tissues were performed after the in vivo imaging, and immunofluorescence staining of the tumor tissue sections was carried out. RESULTS: (64)Cu-DOTA-(AF)SAv/biotin-PEG-RGD(2) was prepared in 52.1 ± 5.4 % radiochemical yield and with specific activity of 1.0 ± 0.1 GBq/mg. Receptor binding studies showed that DOTA-(AF)SAv/biotin-PEG-RGD(2) had higher binding affinity for integrin α(V)β(3) than RGD(2), reflecting a possible polyvalency effect. Moreover, the hybrid probe revealed time-dependent uptake by U87MG cells. In a microPET/optical imaging study, the hybrid probe demonstrated high accumulation in tumors; ROI analysis revealed 2.7 ± 0.2 % ID/g at 1 h and 4.7 ± 0.2 % ID/g at 21 h after injection, and subsequently acquired optical images showed tumors with strong fluorescence intensity. Ex vivo PET/optical images of the major tissues confirmed the in vivo imaging data, and biodistribution studies demonstrated high and specific uptake in tumors (4.8 ± 0.1 % ID/g). Immunofluorescence staining showed the formation of new blood vessels in tumor tissues, suggesting that the tumor uptake was due to specific binding of the hybrid probe to integrin α(V)β(3) expressed on tumor cells. CONCLUSIONS: These results indicate that a (64)Cu-DOTA-(AF)SAv/biotin-PEG-RGD(2) is able to provide quantitative information on hybrid PET/optical imaging of integrin α(V)β(3) expression.