[CANCER RESEARCH 58. 437-441. February I. 1998] Comparison of 1!1In-labeled Somatostatin Analogues for Tumor Scintigraphy and Radionuclide Therapy1 Marion de Jong,2 Wout A. P. Breeman, Willem H. Bakker, Peter P. M. Kooij, Bert F. Bernard, Leo J. Hofland, Theo J. Visser, Ananth Srinivasan, Michelle A. Schmidt, Jack L. Erion, Joseph E. Bugaj, Helmut R. Macke, and Eric P. Krenning Departments of Nuclear Medicine {M. d. J.. W. A. P. B.. W. H. B.. P. P. M. K., B. F. B.. E. P. K.¡ and Internal Medicine HI ¡LJ. H.. T. J. V.. E. P. K.¡. University Hospital Dijkziat, 3015 CD Rotterdam, the Netherlands: Mallinckrndt Medical, St. Louis. Missouri 63134 ¡A.S.. M. A. S.. J. L. E., J. E. B.¡; und Department of Nuclear Medicine. Kunlanspital Basel. CH-4D3I Basel. Switzerland ¡H.R. M.I ABSTRACT We evaluated the following '"In-labeled somatostatin (SS) analogues (diethylenetriaminepentaacetic acid, Oil'A: tetraazacyclododecanetet- raacetic acid, DOTA): [DTPA°loctreotide, |DTPA",Tyr'loctreotide, [DTPA°,D-Tyr'loctreotide. |DTPA°,Tyr']octreotate (Thr(ol) in octreotide replaced with Thr|, and [DOTA°,Tyr'loctreotide, in vitro and in vivo. In vitro, all compounds .showed high and specific binding to SS recep tors in mouse pituitary AtT20 tumor cell membranes, and K '.,,,s were in the nanomolar range. Furthermore, all compounds showed specific inter- nalization in rat pancreatic tumor cells; uptake of |'"ln- DTPA",Tyr*]octreotate was the highest of the compounds tested, and that of |l"ln-DTPA0,l)-Tyr'loctreotide was the lowest. Biodistribution exper iments in rats showed that, 4, 24, and 48 h after injection of |'"ln- DTPA°,Tyr']octreotide, |mIn-DTPA0.Tyr3loctreotate, and ('"In- DOTA°,Tyr'loctreotide, radioactivity in the octreotide-binding, receptor- expressing tissues and tumor-to-blood ratios were significantly higher than those after injection of ["'In-DTPA"]octreotide. Uptake of l'"ln- DTPA°,Tyr']octreotate in the target organs was also, in vivo, the highest of the radiolabeled peptides tested, whereas that of ['"in-DTPA",!)- Tyr'loctreotide was the lowest. Uptake of |"'ln-DTPA",Tyr*loctreotide, ['"ln-DTPA",TyrJloctreotate, and ['"ln-DOTA0,TyrJloctreotide in tar get tissues was blocked by >90% by 0.5 mg of unlabeled octreotide, indicating specific binding to the octreotide receptors. Blockade of ['"In- DTPA°,D-Tyr'loctreotide was >70%. In conclusion, radiolabeled [DTPA°,Tyr<loctreotide and, especially, (DTPA",Tyr'|octreotate and their DOTA-coupled counterparts are most promising for scintigraphy and radionuclide therapy of SS receptor-positive tumors in humans. INTRODUCTION Radiolabeled tumor receptor-binding peptides can be used for in vivo scintigraphic imaging. An example is SS,3 which binds to its receptors on tumors of neuroendocrine origin ( 1). The native peptide. however, is susceptible to very rapid enzymatic degradation (2) and is. therefore, not useful for in vivo application. Therefore, more stable synthetic SS analogues have been developed; e.g., the octapeptide octreotide (Fig. 1: Ref. 3). Because octreotide cannot be radiolabeled easily with a -y-emitting radionuclide. Tyr'-octreotide was developed, allowing radioiodination of the molecule (Fig. 1). This compound, radiolabeled with 125Ior I23I, was the first used in in vitro SS receptor studies (4). tumor scintigraphy in animals (4). and in humans (1, 5). ["'ln-DTPA"]octreotide. consisting of the octapeptide octreotide and the chelator DTPA (Fig. 1), enabling radiolabeling with a radiometal Received 8/6/97; accepted 12/3/97. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely lo indicate this fact. 1This work was supported by Swiss National Science Foundation Gram 31-42516/94 and the "Regionale Krebsliga beider Basel" (both to H. R. M.). - To whom requests for reprints should be addressed, at Department of Nuclear Medicine. V220. University Hospital Rotterdam. 3015 GD Rotterdam, the Netherlands. Phone: 31 104635781; Fax: 31 104635997; E-mail: dejongC»'nuge.azr.nl. 3 The abbreviations used are: SS, somatostatin; DTPA. diethylenetriaminepentaacetic acid: DOTA, tetraa/.acyclododecanetetraacelic acid; AUC, area under the curve; 7rlD, percentage injected dose. like '"In, was the next SS analogue to be synthesized for scintigraphy of SS receptor-positive lesions in vivo. We have described its advan tages over radioiodinated Tyr3-octreotide and its use for scintigraphic imaging of SS receptor-positive lesions (6. 7). A new and fascinating application is the use of radiolabeled oct reotide for radionuclide therapy. Promising results with regard to tumor growth inhibition have been reported in humans using ['"in- DTPA"]octreotide (8). A ß particle emitter, such as ""Y, may, in certain cases, appear more suitable for this purpose than the Auger electron emitter '"In. However. '"'Y-DTPA is unstable, resulting in hematopoietic toxicity in vivii; therefore, Tyr'-octreotide has been derivatized with the DOTA chelator (Fig. 1). enabling stable radiolabeling with **'Â¥ and "'in. (Pre)clinical studies with [DOTA".Tyr']oetreotide showed favorable biodistribution and tumor uptake characteristics (9-11). The success of the therapeutic strategy relies upon the amount of radioligand. which can be concentrated within tumor cells, and this will, among other things, be determined by the rates of internalization. degradation, and recycling of both ligand and receptor. We have evaluated and compared the different mentioned "'in-chelator-pep- tide constructs, and we have also studied some new. recently synthe sized SS analogues, with regard to binding to octreotide receptors on mouse pituitary tumor cell membranes and internalization in rat pancreatic tumor cells. Furthermore, biodistribution in tumor-bearing rats was investigated in vivo. The newly synthesized analogues tested were [DTPA°,D-Tyr']octreotide and [DTPA°.Tyr;l]octreotate (struc tures shown in Fig. 1). MATERIALS AND METHODS Labeling of Octreotide Derivatives. |DTPA"|octreotide and ' "inCI, were provided by Mallinckrodt Medical (Petten. the Netherlands), and octreotide was supplied by Sando/. (Basel. Switzerland). |DOTA".Tyr'|octreolide was synthesized by H. R. M., and [DTPA",D-Tyr']octreotide. [DTPA".Tyr'l- octreotide. and [DTPA".Tyr']octreotale were synthesized by A. S. '"In label ing of the DTPA-analogues was as described for |DTPA"|octreotide (12), and '"In-labeling of |DOTA".Tyr']octreotide (9) and '"l-labeling of [Tyrl|octreotide (4) were performed as described. In Vitro Receptor Binding Studies. Receptor binding assays were carried out using [I2'l-Tyr']octreotide (2200 Ci/mmol) as radioligand using mouse AtT2() pituitary tumor cell membranes (13). Internalization. AR42J cells were grown in RPMI 1640 (Life Technolo gies. Inc., Grand Island, NY), CA20948 cells were grown in DMEM (Life Technologies, Inc.). and ARO cells were grown in DMEM/FI2 (Lite Tech nologies, Inc.): tor all cell lines, medium was supplemented with 2 mM glutamine and l()'/r FCS. Before the experiment, subcontinent cell cultures were transferred to six-well plates. The binding of the radiolabeled peptides to tumor cells and subsequent internalization were studied essentially as described (14). In short, before the experiments, cells were washed, and incubation was started by addition of 1 ml of internalization medium/well (culture medium without PCS but with 1% BSA) with 80 kBq of peptide (O.I nM concentration). Cells were incubated at 37°Cfor indicated periods of time. To determine nonspecific internalization. cells were incubated with an excess unlabeled octreotide (0.1 /¿M). Cellular 437 Research. on December 9, 2021. © 1998 American Association for Cancer cancerres.aacrjournals.org Downloaded from