64Cu-labeled CB-TE2A and Sar conjugated RGD peptide analogs for targeting angiogenesis: Comparison of their biological activity

233 Objectives: The αvβ3 integrin is a cell adhesion molecule known to be involved in stages of angiogenesis and metastasis. In this study, the chelators CB-TE2A and Sarcophagine (Sar) were conjugated to RGD peptide analogs cyclic RGDyK and RGDfD and the biological properties of 64Cu labeled peptides were compared. Methods: CB-TE2A-c(RGDyK) and Sar-c(RGDfD) were labeled with 64Cu in 0.1 M NH4OAc (pH = 8) at 95 °C and 25 °C, respectively, for 1 h. PET and biodistribution studies were carried out on M21 (αvβ3-positive) and M21L (αv-negative) melanoma-bearing mice at 1, 2, 4 and 24h post injection (p.i.). Binding affinity of the Cu-chelator-RGD peptides to αvβ3 integrins was determined by a competitive binding affinity assay. Results: Biological studies showed higher tumor uptake of the 64Cu labeled peptides in M21 tumor compared to M21L tumor (e.g. at 4h p.i., 64Cu-CB-TE2A-c(RGDyK): 1.66±0.8 (M21) vs. 0.84±0.41 %ID/g (M21L), p=0.04). Tumor uptakes of 64Cu-CB-TE2A-c(RGDyK) are higher than those of 64Cu-Sar-c(RGDfD)(e.g., at 1h p.i., 64Cu-CB-TE2A-c(RGDyK): 2.98±0.90 vs. 64Cu-Sar-c(RGDfD): 1.51±0.53 %ID/g, p=0.01). These differences are not due to differing binding affinities, since similar values were obtained for the agents from the binding assays (IC50: Cu-CB-TE2A-c(RGDyK): 6.0 nM vs. Cu-Sar-c(RGDfD): 4.8 nM vs. c(RGDyK): 3.7 nM). Compared to Cu-CB-TE2A-c(RGDyK), which has a longer lysine linker, Cu-Sar-c(RGDfD) is closer to the binding site of the peptide, potentially causing partial demetallation of the Cu complex. The slower liver clearance of 64Cu-Sar-c(RGDfD) (2.63±0.53 (2h p.i.) vs. 2.24±0.41 %ID/g (24h p.i.), p=NS) compared to 64Cu-CB-TE2A-c(RGDyK) (2.53±0.89 (2h p.i.) vs. 0.84±0.17 %ID/g (24h p.i.), p=0.05) is further evidence of demetallation. Administration of c(RGDyK) as a block significantly reduced the tumor uptake indicating a receptor-specific accumulation. Conclusions: Both 64Cu-CB-TE2A-c(RGDyK) and 64Cu-Sar-c(RGDfD) are potential candidates for imaging tumor angiogenesis. For Sar-c(RGDfD), a linker may be needed between the Cu-chelate moiety and the RGD peptide to achieve optimal in vivo tumor uptake. Research Support (if any): NCI grants R24 CA86307 and R01 CA93375