Preparation and characterization of 99mTc(CO)3–BPy–RGD complex as αvβ3 integrin receptor-targeted imaging agent

The aim of this study is to develop a novel arginine–glycine–aspartic acid (RGD) peptide-containing ligand for 99mTc labeling as αvβ3 integrin receptor-targeted imaging agent. BPy–RGD conjugate was successfully synthesized by coupling of 5-carboxylate-2,2′-bipyridine and c(RGDyK) peptide through EDC/SNHS in aqueous solution and was characterized by MADLI-TOF-MS (m/z=802.72, C38H48N11O9). 99mTc(CO)3–BPy–RGD was prepared by exchange reaction between [99mTc(H2O)3(CO)3]+ and BPy–RGD. Final product was purified by HPLC and tested for octanol/water partition coefficient. Cell-binding assays of BPy–RGD and unmodified c(RGDyK) were tested in MDA-MB-435 cells (125I-echistatin as radioligand). Preliminary biodistribution of the 99mTc(I)-labeled radiotracer in orthotopic MDA-MB-435 breast tumor xenograft model was also evaluated. The BPy–RGD conjugate had good integrin-binding affinity (50% inhibitory concentration (IC50)=92.51±22.69 nM), slightly lower than unmodified c(RGDyK) (IC50=59.07±11.03 nM). The hydrophilic radiotracer also had receptor-mediated activity accumulation in MDA-MB-435 tumor (1.45±0.25 percentage of injected dose per gram (%ID/g) at 1.5 h postinjection (p.i.)), which is known to be integrin positive. After blocking with c(RGDyK), the tumor uptake was reduced from 0.71±0.01%ID/g to 0.33±0.18%ID/g at 4 h p.i. 99mTc(I) tricarbonyl complex of cyclic RGD peptide is a promising strategy for integrin targeting. Further modification of the bipyridine-conjugated RGD peptide by using more potent RGD peptides and fine tuning of the tether group between the RGD moiety and 99mTc(CO)3+ core to improve the tumor targeting efficacy and in vivo kinetic profiles is currently in progress.