Evaluation of Re/^99mTc-labeled somatostatin receptor-targeting peptide complexes synthesized via direct metal cyclization

Abstract With interest in the development of somatostatin receptor (SSTR) targeting agents for potential application in diagnostic SPECT imaging ( 99m Tc) or Peptide Radionuclide Receptor Therapy (PRRT, 186 Re or 188 Re) of neuroendocrine tumors, we present herein 99m Tc/Re (radio)complexes synthesized by the integrated (radio)labeling approach of peptide cyclization via metal complexation. In particular, we utilized the potent SSTR2 peptide antagonist sequence DOTA-4-NO 2 -Phe-c( D Cys-Tyr- D Trp-Lys-Thr-Cys)- D Tyr-NH 2 (DOTA-sst 2 -ANT) and report the syntheses and in vitro evaluations of its respective [ 99m Tc]Tc/Re-cyclized peptides ([ 99m Tc]Tc/Re-cyc-DOTA-sst 2 -ANT). The Re-cyc-DOTA-sst 2 -ANT complex was synthesized via an on-resin Re(V)-cyclization reaction using the ReOCl 3 (PPh 3 ) 2 precursor and consisted of three isomers characterized by LC–ESI-MS. The [ 99m Tc]Tc-cyclized analogue was prepared via a ligand exchange reaction of the [ 99m Tc][TcO] 3+ core through a [ 99m Tc]Tc-glucoheptonate intermediate with linear DOTA-sst 2 -ANT and was characterized by comparative HPLC studies against Re-cyc-DOTA-sst 2 -ANT. Good in vitro binding affinity was demonstrated in SSTR-expressing cells (AR42J) by the Re-cyc-DOTA-sst 2 -ANT major isomer, similar to the potent binder Lu-DOTA-sst 2 -ANT, in which the Lu metal was complexed by the bifunctional chelator DOTA versus via peptide cyclization. [ 99m Tc]Tc-cyc-DOTA-sst 2 -ANT was obtained in high radiochemical yield, also with an elution pattern of three isomers observed by HPLC analysis, which were comparable yet not identical to those of Re-cyc-DOTA-sst 2 -ANT. The [ 99m Tc]Tc-tracer complex was shown to be hydrophilic, and stability studies at 4 h demonstrated that it remained intact in both PBS and in rat serum, with low non-specific rat serum protein binding, while exhibiting more moderate stability in 1 mM cysteine. These findings demonstrate that direct Re/[ 99m Tc]Tc-cyclization of DOTA-sst 2 -ANT is feasible and may be used as an alternative approach to the bifunctional chelate labeling strategy. However, given that the non-radioactive (Re) and radiotracer ( 99m Tc) analogues are not identical and both form isomeric products in equilibrium, additional design modifications will be necessary prior to in vivo application of [ 99m Tc]Tc/Re-cyc-DOTA-sst 2 -ANT.