Characterization of the four isomers of 123 I-CMICE-013: A potential SPECT myocardial perfusion imaging agent Lihui Wei a,b,c,⇑,  , Corinne Bensimon a,  , Xuxu Yan a , Julia Lockwood b,c , Wei Gan b,c , R. Glenn Wells b,c , Yin Duan a,c , Pasan Fernando a,b,c,d , Bram Gottlieb a , Wayne Mullett a , Terrence D. Ruddy b,c a Nordion Inc., 447 March Road, Ottawa, ON K2K 1X8, Canada b Division of Cardiology, University of Ottawa Heart Institute, 40 Ruskin Street, Ottawa, ON K1Y 4W7, Canada c Canadian Molecular Imaging Center of Excellence (CMICE), Nordion Lab/University of Ottawa Heart Institute, 40 Ruskin Street, Ottawa, ON K1Y 4W7, Canada d Department of Cellular and Molecular Medicine, Faculty of Medicine, University of Ottawa, Ottawa, ON K1H 8M5, Canada article info Article history: Received 12 January 2014 Revised 12 February 2014 Accepted 24 February 2014 Available online 4 March 2014 Keywords: 123 I-CMICE-013 127 I-CMICE-013 Myocardial perfusion imaging SPECT abstract Myocardial perfusion imaging (MPI) with single photon emission computed tomography (SPECT) is widely used in the assessment of coronary artery disease (CAD). We have developed 123 I-CMICE-013 based on rotenone, a mitochondrial complex I (MC-1) inhibitor, as a promising new MPI agent. Our syn- thesis results in a mixture of four species of 123 I-CMICE-013 A, B, C, D. In this study, we separated the four species and evaluated their biodistribution and imaging properties. The cold analogs 127 I-CMICE-013 A, B, C, D were isolated and characterized and their chemical structures proposed. Methods: 123 I-CMICE-013 was synthesized by radiolabeling rotenone with Na 123 I in trifluoroacetic acid (TFA) with iodogen as the oxidizing agent at 60 °C for 45 min, and the four species were separated by RP-HPLC. The cold analogs 127 I-CMICE-013 A, B, C and D were isolated with a similar procedure and characterized by NMR and mass spectrometry. Biodistribution and microSPECT imaging studies were carried out on normal rats. Results: We propose the mechanism of the rotenone iodination and the structures of the four species. First, I + forms an intermediate three-membered ring with 6 0 and 7 0 carbons. Second, the lone electron pair of the water molecule attacks the 6 0 or 7 0 -carbon, following by the formation of 6 0 -OH, and 7 0 -I bonds as in major products C and D, or 6 0 -I and 7 0 -OH bonds as in minor products A and B. The weaker 6 0 -I bond in the intermediate prompts the nucleophilic attachment of water at the favorable 6 0 -carbon to generate C and D. MicroSPECT images of 123 I-CMICE-013 A, B, C, D in rats showed clear visualization of myocar- dium and little interference from lung and liver. The imaging time activity curves and biodistribution data showed complex profiles for the four isomers, which is not expected from the structure activity rela- tionship theory. Conclusion: 123/127 I-CMICE-013 A and B are constitutional isomers with C and D, while A and C are diastereomers of B and D, respectively. Overall, the biological characteristics of the four species are not correlated perfectly with their molecular structures. Ó 2014 Elsevier Ltd. All rights reserved. 1. Introduction Stress myocardial perfusion imaging (MPI) has been increasingly utilized in the assessment of coronary artery disease (CAD). 1–3 The most commonly used MPI modality is single photon emission com- puted tomography (SPECT), 4 in conjunction with one of three SPECT radiotracers, 99m Tc-sestamibi, 99m Tc-tetrofosmin, and 201 Tl. 5–7 Important information has been obtained with these agents for evaluation of myocardial perfusion and viability in patients. However, these radiotracers have certain suboptimal pharmacoki- netic characteristics. 201 Tl SPECT MPI has poor image quality in obese patients, redistribution to non-target tissues over time, and fair radiodosimetry. 2,5 A shortcoming with both 99m Tc MPI agents and 201 Tl is the disproportional low myocardial uptake under stress conditions with myocardial hyperaemia (the ‘roll-off’ phenomenon). 2,8–10 201 Tl is a potassium analog that accumulates in myocardial cells by uptake mediated by Na + /K + adenosine triphosphatase. 5,11 99m Tc- sestamibi and 99m Tc-tetrofosmin, lipophilic and cationic com- plexes, accumulate in cardiac cells based on the mitochondrial membrane potential. 12–15 Most recently, a new class of radiotracers that specifically target complex I of the mitochondrial electron http://dx.doi.org/10.1016/j.bmc.2014.02.052 0968-0896/Ó 2014 Elsevier Ltd. All rights reserved. ⇑ Corresponding author. Tel.: +1 613 798 5555x16500; fax: +1 613 595 4599. E-mail address: lihui.wei@nordion.com (L. Wei).   Lihui Wei and Corinne Bensimon contributed equally to this study. Bioorganic & Medicinal Chemistry 22 (2014) 2033–2044 Contents lists available at ScienceDirect Bioorganic & Medicinal Chemistry journal homepage: www.elsevier.com/locate/bmc