Improving tumor-targeting capability and pharmacokinetics of (99m)Tc-labeled cyclic RGD dimers with PEG(4) linkers.

This report describes the synthesis of two cyclic RGD (Arg-Gly-Asp) conjugates, HYNIC-2PEG(4)-dimer (HYNIC = 6-hydrazinonicotinyl; 2PEG(4)-dimer = E[PEG(4)-c(RGDfK)](2); and PEG(4) = 15-amino-4,7,10,13-tetraoxapentadecanoic acid) and HYNIC-3PEG(4)-dimer (3PEG(4)-dimer = PEG(4)-E[PEG(4)-c(RGDfK)](2)), and evaluation of their (99m)Tc complexes [(99m)Tc(HYNIC-2PEG(4)-dimer)(tricine)(TPPTS)] ((99m)Tc-2PEG(4)-dimer: TPPTS = trisodium triphenylphosphine-3,3',3''-trisulfonate) and [(99m)Tc(HYNIC-3PEG(4)-dimer)(tricine)(TPPTS)] ((99m)Tc-3PEG(4)-dimer) as novel radiotracers for imaging integrin alpha(v)beta(3) expression in athymic nude mice bearing U87MG glioma and MDA-MB-435 breast cancer xenografts. The integrin alpha(v)beta(3) binding affinities of RGD peptides were determined by competitive displacement of (125)I-c(RGDyK) on U87MG glioma cells. It was found that the two PEG(4) linkers between RGD motifs in HYNIC-2PEG(4)-dimer (IC(50) = 2.8 +/- 0.5 nM) and HYNIC-3PEG(4)-dimer (IC(50) = 2.4 +/- 0.7 nM) are responsible for their higher integrin alpha(v)beta(3) binding affinity than that of HYNIC-PEG(4)-dimer (PEG(4)-dimer = PEG(4)-E[c(RGDfK)](2); IC(50) = 7.5 +/- 2.3 nM). Addition of extra PEG(4) linker in HYNIC-3PEG(4)-dimer has little impact on integrin alpha(v)beta(3) binding affinity. (99m)Tc-2PEG(4)-dimer and (99m)Tc-3PEG(4)-dimer were prepared in high yield with >95% radiochemical purity and the specific activity of >10 Ci/mumol. Biodistribution studies clearly demonstrated that PEG(4) linkers are particularly useful for improving the tumor uptake and clearance kinetics of (99m)Tc-2PEG(4)-dimer and (99m)Tc-3PEG(4)-dimer from noncancerous organs. It was also found that there was a linear relationship between the tumor size and radiotracer tumor uptake expressed as %ID (percentage of the injected dose) in U87MG glioma and MDA-MB-435 breast tumor models. The blocking experiment showed that the tumor uptake of (99m)Tc-2PEG(4)-dimer is integrin alpha(v)beta(3)-mediated. In the metabolism study, (99m)Tc-2PEG(4)-dimer had high metabolic stability during its excretion from renal and hepatobiliary routes. (99m)Tc-3PEG(4)-dimer also remained intact during thee excretion from the renal route, but, had approximately 30% metabolism during the excretion from the hepatobiliary route. Planar imaging studies in U87MG glioma and MDA-MB-435 breast tumor models showed that the tumors of approximately 5 mm in diameter could be readily visualized with excellent contrast. Thus, (99m)Tc-3PEG(4)-dimer is a very promising radiotracer for the early detection of integrin alpha(v)beta(3)-positive tumors, and may have the potential for noninvasive monitoring of tumor growth or treatment efficacy.