Preparation of 66 Ga- and 68 Ga-labeled Ga(III)-deferoxamine-folate as potential folate-receptor-targeted PET radiopharmaceuticals Carla J. Mathias a , Michael R. Lewis e , David E. Reichert e , Richard Laforest e , Terry L. Sharp e , Jason S. Lewis e , Zhen-Fan Yang b , David J. Waters c , Paul W. Snyder d , Philip S. Low b , Michael J. Welch e , Mark A. Green a, * a Department of Industrial and Physical Pharmacy, Purdue University, West Lafayette, IN, USA b Department of Chemistry, Purdue University, West Lafayette, IN, USA c Department of Veterinary Clinical Sciences, Purdue University, West Lafayette, IN, USA d Department of Veterinary Pathobiology, Purdue University, West Lafayette, IN, USA e The Edward Mallinckrodt Institute of Radiology, Washington University School of Medicine, Saint Louis, MO, USA Abstract A folate-receptor-targeting radiopharmaceutical, Ga(III)-deferoxamine-folate (Ga-DF-Folate), was radiolabeled with two positron- emitting isotopes of gallium, cyclotron-produced 66 Ga (9.5 hour half-life) and generator-produced 68 Ga (68 minute half-life). The [ 66 Ga]Ga-DF-Folate was administered to athymic mice with folate-receptor-positive human KB cell tumor xenografts to demonstrate that microPET mouse tumor imaging is feasible with 66 Ga, despite the relatively high positron energy of this radionuclide. Using the athymic mouse KB tumor xenograft model, dual-isotope autoradiography was also performed following i.v. co-administration of [ 18 F]-FDG, a marker of regional metabolic activity, and folate-receptor-targeted [ 111 In]In-DTPA-Folate. The autoradiographic images of 1 mm tumor sections demonstrate the gross heterogeneity of the KB cell tumor xenograft, as well as subtle disparity in the regional accumulation of the two radiotracers. © 2003 Elsevier Inc. All rights reserved. 1. Introduction The folate receptor (or folate-binding protein, FBP) is a cell membrane protein involved in cellular accumulation of folic acid by endocytosis. The folate receptor is highly expressed in a variety of epithelial cancers [1,2,3,19,20,23,24,27,28,29,30], most notably in about 90% of ovarian and endometrial epithe- lial carcinomas, but occurs in only a limited number of normal tissues [5,20,28,29]. When folic acid is conjugated to either macromolecules or low-molecular-weight chelates via its al- pha or gamma carboxylate, the folate fragment is no longer recognized by the facilitated transport system that is partially responsible for folate uptake in all mammalian cells; however, the folate-conjugate is still recognized by the folate receptor [4,6,7,8,9,10,13,14,15,17,18,25,26]. Thus, these folate-conju- gates concentrate selectively in folate-receptor-expressing cells. Radiolabeled chelate-conjugates, such as [ 111 In]In-DTPA- Folate and [ 67 Ga]Ga-Deferoxamine-Folate ([ 67 Ga]Ga-DF-Fo- late), have been shown to concentrate selectively in cells and tissues that express the membrane folate receptor [15,17,18,22,25,26], including folate-receptor-positive tu- mors and kidneys, where the folate receptor is found in the proximal tubules [17,21,29,31]. We report here the radiola- beling of Ga(III)-DF-Folate with positron-emitting 66 Ga (9.5 hour half-life; 57%  + decay) and [ 68 ]Ga (68 minute half-life; 89%  + decay). Gallium-66 is a particularly at- tractive for tumor imaging with positron emission tomog- raphy (PET), since its relatively long half-life is suitable for PET imaging over an extended time period following ra- diopharmaceutical administration, which may result in im- proved tumor/non-target contrast using tracers with rela- tively slow clearance kinetics. 2. Materials and methods 2.1. General. The deferoxamine-folate conjugate (DF-Folate) was pre- pared as described previously [15,25]. Folate-deficient ro- * Corresponding author. Tel.: +1-765-496-1445. E-mail address: magreen@purdue.edu (M. A. Green). Nuclear Medicine and Biology 30 (2003) 725–731 www.elsevier.com/locate/nucmedbio 0969-8051/03/$ – see front matter © 2003 Elsevier Inc. All rights reserved. doi:10.1016/S0969-8051(03)00080-5