J Pharm Pharmaceut Sci (www. cspsCanada.org): 10(2): 286s-297s, 2007 286s Synthesis and evaluation of intercalating somatostatin receptor binding peptide conjugates for endoradiotherapy Keith Graham a,b , Qin Wang a,c , Regine Garcia Boy a , Michael Eisenhut d , Uwe Haberkorn a and Walter Mier a a Universitätsklinikum Heidelberg, Department of Nuclear Medicine, 69120 Heidelberg, Germany b present address: Schering AG Berlin, Germany c present address: Vion Pharmaceuticals, New Haven, USA d German Cancer Research Centre, Division of Radiochemistry and Radiopharmacology, 69120 Heidelberg, Germany Keywords: Peptides, nuclear localization, endoradiotherapy, intercalators, somatostatin. Dedicated to the memory of Prof. Antoine (Tony) A. Noujaim, in recognition of his outstanding contributions to radiopharmacy, diagnostic oncology and the immunotherapy of cancer. Received December 17, 2006; Revision received March 14, 2007; Accepted March 14, 2007; Published June 18, 2007 ABSTRACT – Purpose: Intercalators, planar aromatic compounds, are able to interact with DNA by sandwiching themselves between the stacked bases at right angles to the long axis of the helix. Under certain circumstances, Auger-electron- emitting radionuclides can be extremely radiotoxic and produce extensive DNA damage. Auger electron-emitting radioisotopes, are known to be highly cytotoxic when localized in cell nuclei due to highly localized energy deposition by low energy Auger electrons. In addition binding to the DNA might increase the retention in the receptor expressing tissues. Methods: In order to exploit the cytotoxic potential of intercalator-Auger-emitter conjugates, bis-benzimidazole dyes, Hoechst 33258 and 33342, were linked to a somatostatin receptor affine carrier peptide. For this purpose a bis- benzimidazole intercalating moiety was prepared using variations on the literature methods. The intercalating moieties were coupled under normal SPPS conditions to the carrier peptide, Tyr 3 - octreotate. To attach the chelating agent (DOTA) to the intercalating moiety, a free amine derivative was prepared and coupled in solution to DOTA tris- t-butyl ester. The resulting chelator-intercalator conjugate was then coupled to a Tyr 3 -octreotate carrying resin using SPPS. Results: The peptide conjugates were obtained in good yields after HPLC chromatography. The cellular uptake of the novel conjugates was determined using fluorescence microscopy. All intercalator-peptide conjugates revealed somatostatin receptor binding affinities in the nanomolar range. Conclusions: The novel chelator-intercalator derivatives of the somatostatin receptor binding Tyr 3 -octreotate introduce a new scope to the range of tracers for therapeutic purposes. INTRODUCTION Small radiolabeled peptides have been proven as important tools in tumor diagnostic and endoradiotherapy for more than one decade. By specifically binding to receptors or other structures expressed on the surface of tumor cells, peptides are able to shuttle radionuclides to these tissues. Somatostatin receptor binding peptides labeled with a variety of gamma-, positron- and beta-emitters have been the prototypes for those constructs, as these receptors are frequently overexpressed in certain tumor types, such as neuroendocrine tumors and their metastases. When conjugated to ‘therapeutic’ nuclides such as 90 Y, somatostatin receptor binding peptides have been shown to be successful for the therapy of receptor expressing tumors. The most successful radiotherapeutic is 90 Y-DOTATOC. However, the therapeutic value of this treatment is limited by the side effects in non- target tissues such as the kidneys. The effectiveness of the treatment modality might be further increased with conjugates which enable the nuclear targeting of the low energy emitting radioactive isotopes. Intercalators, planar aromatic compounds, are able to interact with DNA by sandwiching themselves between the stacked bases at right angles to the long axis of the helix. The bis-benzimidazole dyes, Hoechst 33258 and 33342, have been shown to bind to the minor groove of DNA in A-T rich regions. The insertion of an intercalator changes the DNA conformation (1). Intercalators have thus thoroughly been examined with respect to their DNA-cleaving activity for anticancer therapy (2). Corresponding Author: PD Dr. Walter Mier, Universitätsklinikum Heidelberg, Department of Nuclear Medicine, 69120 Heidelberg, Germany. Email: walter.mier@med.uni-heidelberg.de