Tetraamine-modified octreotide and octreotate:: labeling with 99mTc and preclinical comparison in AR4-2J cells and AR4-2J tumor-bearing mice

Two somatostatin analogues, [Tc-99m]Demotide and [Tc-99m]Demotate 4, were compared with [99mTc]Demotate 1, a previously reported somatostatin receptor subtype 2 (sst(2)) targeting tracer. Conjugates were prepared by coupling an open-chain tetraamine chelator to D-Phe(1) of [Tyr(3)]-octreotide or [Tyr(3)]-octreotate, respectively, via a p-benzylaminodiglycolic acid spacer adopting solid-phase peptide synthesis techniques. Peptide conjugates were collected in a highly pure form after chromatographic purification. Eventually, [Tc-99m]Demotide and [Tc-99m]Demotate 4 were obtained in similar to 1 Ci/mu mol specific activity and > 96% purity after labeling under alkaline conditions. Demotide and Demotate 4 exhibited similar high binding affinities for the sst(2) expressed in AR4-2J cells with IC50 values 0.16 and 0.10 nM, respectively. The (radio)metallated analogues [Tc-99m]Demotide and [Tc-99m]Demotate 4 showed equally high affinities to the sst(2) during saturation binding assays in AR4-2J cell membranes (K(d)s 0.08 and 0.07 nM, respectively). During incubation at 37 degrees C with AR4-2J cells, the radiopeptides internalized effectively via a receptor-mediated process, with [Tc-99m]Demotate 4 exhibiting a faster internalization rate than [Tc-99m]Demotide. After injection in athymic mice bearing sst(2)-expressing AR4-2J tumors, the rachotraccrs showed high and specific uptake in the tumor (> 25%ID/g at 1 h) and in the sst(2)-positive organs. However, both [Tc-99m]Demotide and [Tc-99m]Demotate 4 showed unfavorably higher background activity, especially in the abdomen, in comparison to [99mTc]Demotate I and are, therefore, less suited than [Tc-99m]Demotate 1 for sst(2)-targeted tumor imaging in man. Copyright (c) 2005 European Peptide Society and John Wiley & Sons, Ltd.