Structure-activity relationship of radiocopper-labeled DMPTACN-Bombesin conjugates

Aim:

Radiometal‐labeled peptide derivatives of bombesin are very interesting targeting vectors for certain types of cancer. Bombesin derivatives have shown very high selectivity and affinity to
G‐protein‐coupled gastrin‐releasing‐peptide‐receptor (GRPR), which is over‐expressed in a variety of tumors including breast‐, prostate‐ and pancreatic‐tumors. Consequently, the
application of radiolabeled bombesin‐analogs for both the diagnosis and therapy of such tumors is being intensively investigated. However, the development of chemically and radiolytically
stable compounds which can be easily radiolabeled presents significant challenges. We recently showed that bis(2‐pyridylmethyl)triazacyclocyclononane (DMPTACN) was a promising candidate
for radiocopper‐labeling. In this study we examine structure‐activity relationships for new [64Cu]DMPTACN bombesin derivatives including their biodistribution and pharmacokinetics in
prostate cancer (PC3) xenografted tumor mice.

Methods:

Stabilized bombesin derivatives β‐Ala‐β‐Ala‐[Cha13,Nle14]BBN(7‐14) and β‐homo‐Glu‐β‐Ala‐β‐Ala‐[Cha13, Nle14]BBN(7‐14) were conjugated to the N‐terminus with DMPTACN ligands containing either a carboxylate or phenylisothiocyanate pendant arm via amide coupling and thiourea‐bridging, respectively. Radiolabeling of DMPTACN‐BBN derivatives with 64Cu was performed in aqueous ammonium acetate solution (pH=6) at 50°C using [64Cu]CuCl2. The affinity of DMPTACN‐BBN derivatives for the GRPR was determined using a competitive displacement/binding assay in human prostate (PC3) cancer cells. Internalization data for the [64Cu]Cu‐DMPTACN bombesin derivatives were obtained in the same cell line. Partition coefficients of the radiocopper‐labeled complexes of the DMPTACN‐BBN derivatives were determined in a 1‐octanol/buffer system. Biodistribution studies were performed on Wistar rats and NMRI nu/nu mice bearing the human prostate tumor PC‐3. Tumor accumulation was evaluated with small animal PET.

Results:

Radiolabeling of DMPTACNBBN‐bioconjugates was achieved in 30 min, yielding >99% radiochemical purity and specific activity up to 30 GBq/μmol after HPLC. DMPTACN‐NCS derivatives could be rapidly labeled with 64Cu under mild conditions in almost quantitative yield. The DMPTACN‐BBN conjugates showed high affinity to the GRPR and high uptake in PC‐3 cells. PET studies on tumor‐bearing PC‐3 mice revealed an accumulation in the GRPR‐positive tissue. Clear visualization of the tumor tissue and noticeable delineation from healthy tissue was achieved.

Conclusion:

DMPTACN ligands are attractive chelates for the development of radiocopper pharmaceuticals featuring very high chemical and radiolytical stability. They can be effectively coupled to target‐oriented peptides, such as bombesin. However, many issues need to be resolved, including the metabolic stabilization of the peptides and the direct fixation of radiometalated conjugates in the tumor tissue. DMPTACN‐isothiocyanate was found to be rapidly and efficiently labeled with 64Cu. These features make it a promising candidate as a pre‐labeling building block for antibody and synthetic polymers.