Evaluation of 111In-Labeled Cyclic RGD Peptides: Tetrameric not Tetravalent

This report presents the synthesis and evaluation of In-111(DOTA-6G-RGD(4)) (DOTA = 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetracetic acid; 6G-RGD(4) = E{G(3)-E[G(3)-c(RGDfK)](2)}(2) and G(3) = Gly-Gly-Gly), In-111(DOTA-RGD(4)) (RGD(4) = E{E[c(RGDfK)](2)}(2)) and In-111(DOTA-3G-RGD(2)) (3G-RGD(2) = G(3)-E[G(3)-c(RGDfK)](2)) as new radiotracers for imaging integrin alpha(v)beta(3)-positive tumors. The IC50 values of DOTA-6G-RGD(4), DOTA-RGD(4), and DOTA-3G-RGD(2) were determined to be 0.4 +/- 0.1, 1.4 +/- 0.1 and 1.1 +/- 0.1 nM against I-125-c(RGDyK) bound to integrin alpha(v)beta(3)-positive U87MG human glioma cells. In-111(DOTA-6G-RGD(4)), (111)(DOTA-RGD(4)), and In-111(DOTA-3G-RGD(2)) were prepared by reacting (InCl3)-In-111 with the respective DOTA conjugate in NH4OAc buffer (100 mM, pH = 5.5). Radiolabeling could be completed by heating the reaction mixture at 100 degrees C for 15-20 min. The specific activity was similar to 1850 MBq/mu mol for In-111(DOTA-3G-RGD(2)) and similar to 1480 MBq/mu mol for In-111(DOTA-6G-RGD(4)). The athymic nude mice bearing U87MG human glioma xenografts were used to evaluate tumor uptake and excretion kinetics of In-111(DOTA-6G-RGD(4)), In-111(DOTA-RGD(4)), and In-111(DOTA-3G-RGD(2)). The results from both the integrin alpha(v)beta(3) binding assay and biodistribution studies suggest that the tetrameric cyclic ROD peptides, such as RGD(4) and 6G-RGD(4), are most likely bivalent in binding to the integrin alpha(v)beta(3). Both In-111(DOTA-6G-RGD(4)) and In-111(DOTA-RGD(4)) had significantly higher tumor uptake than In-111(DOTA-3G-RGD(2)) at 24-72 h post injection due to the extra RGD motifs in RGD(4) and 60-RGD(4). In-111(DOTA-3G-RGD(2)) had very little metabolism. while In-111(DOTA-6G-RGD(4)) had significant metabolism during its excretion via both renal and hepatobiliary routes over the 2 h period, probably due to its much larger size. The combination of high tumor uptake with long tumor retention suggests that their corresponding Y-90 and Lu-177 analogues M(DOTA-6G-RGD(4)) (M = Y-90 and Lu-177) might be useful as therapeutic radiotracers for treatment of integrin alpha(v)beta(3)-positive solid tumors.