Trifunctional Somatostatin-Based Derivatives Designed for Targeted Radiotherapy Using Auger Electron Emitters

Auger electron-emitting radionuclides have potential for the therapy of small-size cancers because of their high level of cytotoxicity, low-energy, high linear energy transfer, and short-range biologic effectiveness. Biologic effects are critically dependent on the subcellular (and even subnuclear) localization of these radionuclides. Our goals were the design, synthesis, and in vitro preclinical assessment of new trifunctional conjugates of somatostatin that should aim at the nucleus and, therefore, ensure a longer retention time in the cell, a close approximation to the DNA, and the success of Auger electron emitters in targeted radionuclide therapy as well as also improve other targeted therapy strategies. Methods: Three trifunctional derivatives of [(1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid) 0 .Tyr 3 ]octreotide (DOTA-TOC) bearing the nuclear localization signal (NLS) (of simian virus 40 large-T antigen) PKKKRKV in 3 different positions relative to the somatostatin analog sequence were synthesized using solid and solution phase peptide synthesis. These compounds together with DOTA-TOC and DOTA-NLS derivatives were labeled with 1 1 1 In and tested for binding affinity, internalization, externalization, and nuclei localization on AR4-2J cells and on human embryonic cells stably transfected with sst2A. Results: The two N-terminal derivatives preserved the sstr2A binding affinity. Their rate of internalization in all tested sstr-expressing cell lines was always superior for the trifunctional derivatives in comparison with the parent compound. A 6-fold increase in cellular retention from the total internalized activity and a 45-fold higher accumulation in the cell nuclei were found for one of the N-terminally modified compounds compared with [ 1 1 1 In]-DOTA-TOC. The C-terminal conjugate was inferior in all tests compared with the parent compound. Conclusion: These encouraging results support our hypothesis that an additional NLS sequence to the DOTA-TOC could not only provide a better carrier for Auger electron-emitting radionuclides but also ensure a longer radioactivity retention time in the tumor cell.