Spacer Effects on in vivo Properties of DOTA‐Conjugated Dimeric [Tyr3]Octreotate Peptides Synthesized by a “CuI‐Click” and “Sulfo‐Click” Ligation Method

We report on the SSTR2‐binding properties of a series of four dimeric [Tyr3]octreotate analogues with different spacer lengths (nine, 19, 41, and 57 atoms) between the peptides. Two analogues (9 and 57 atoms) were selected as precursors for the design, synthesis, and biological evaluation of DOTA‐conjugated dimeric [Tyr3]octreotate analogues for tumor targeting. These compounds were synthesized by using a two‐stage click ligation procedure: a CuI‐catalyzed 1,3‐dipolar cycloaddition (“copper‐click” reaction) and a thio acid/sulfonyl azide amidation (“sulfo‐click” reaction). The IC50 values of these DOTA‐conjugated [Tyr3]octreotate analogues were comparable, and internalization studies showed that the nine‐atom 111In‐DOTA‐labeled [Tyr3]octreotate dimer had rapid and high receptor binding. Biodistribution studies with BALB/c nude mice bearing subcutaneous AR42J tumors showed that the 111In‐labeled [Tyr3]octreotate dimer (nine atoms) had a high tumor uptake at 1 h p.i. (38.8±8.3 % ID g−1), and excellent tumor retention at 4 h p.i. (40.9±2.5 % ID g−1). However, the introduction of the extended hydrophilic 57 atoms spacer led to rapid clearance from the circulation; this limited tumor accumulation of the radiotracer (21.4±4.9 % ID g−1 at 1 h p.i.). These findings provide important insight on dimerization and spacer effects on the in vivo properties of DOTA‐conjugated [Tyr3]octreotate dimers.