Effects of linker variation on the in vitro and in vivo characteristics of an 111In-labeled RGD peptide

Introduction: Due to the selective expression of the αvβ3 integrin in tumors, radiolabeled arginine-glycine-aspartic acid (RGD) peptides are attractive candidates for tumor targeting. Minor modifications of these peptides could have a major impact on in vivo characteristics. In this study, we systematically investigated the effects of linker modification between two cyclic RGD sequences and DOTA (1,4,7,10-tetraazadodecane-N,N',N',N'''-tetraacetic acid) on the in vitro and in vivo characteristics of the tracer. Methods: A dimeric RGD peptide was synthesized and conjugated either directly with DOTA or via different linkers: PEG4 (polyethylene glycol), glutamic acid or lysine. The RGD peptides were radiolabeled with 111In, and their in vitro and in vivo αvβ3-binding characteristics were determined. Results: LogP values varied between -2.82±0.06 and -3.95±0.33. The IC50 values for DOTA-E-[c(RGDfK)]2, DOTA-PEG4-E-[c(RGDfK)]2, DOTA-E-E-[c(RGDfK)]2 and DOTA-K-E-[c(RGDfK)]2 were comparable. Two hours after injection, the tumor uptakes of the 111In-labeled compounds were not significantly different. The kidney accumulation of [111In]-DOTA-K-E-[c(RGDfK)]2 [4.05±0.20% of the injected dose per gram (ID/g)] was significantly higher as compared with that of [111In]-DOTA-E-[c(RGDfK)]2 (2.63±0.19% ID/g; P<.05) as well as that of [111In]-DOTA-E-E-[c(RGDfK)]2 (2.16±0.21% ID/g; P<.01). The liver uptake of [111In]-DOTA-E-E-[c(RGDfK)]2 (2.12±0.09% ID/g) was significantly higher as compared with that of [111In]-DOTA-E-[c(RGDfK)]2 (1.64±0.1% ID/g; P<.05) as well as that of [111In]-DOTA-K-E-[c(RGDfK)]2 (1.52±0.04% ID/g; P<.01). Conclusions: Linker variation did not affect affinity for αvβ3 and tumor uptake. Insertion of lysine caused enhanced kidney retention; that of glutamic acid also resulted in enhanced retention in the kidneys. PEG4 appeared to be the most suitable linker as compared with glutamic acid and lysine because it has the highest tumor-to-blood ratio and the lowest uptake in the kidney and liver