Molecular imaging is the visualization, characterization and measurement of biological processes at the molecular and cellular levels in humans and other living systems. Molecular imaging agents are probes used to visualize, characterize and measure biological processes in living systems. These two definitions were put forth by the Sociey of Nuclear Medicine (SNM) in 2007 as a way to capture the interdisciplinary nature of this relatively new field. The emergence of molecular imaging as a scientific discipline is a result of advances in chemistry, biology, physics and engineering, and the application of imaging probes and technologies has reshaped the philosophy of drug discovery in the pharmaceutical sciences by providing more cost effective ways to evaluate the efficacy of a drug candidate and allowing pharmaceutical companies to reduce the time it takes to introduce new therapeutics to the marketplace. Finally the impact of molecular imaging on clinical medicine has been extensive since it allows a physician to diagnose a patient’s illness, prescribe treatment and monitor the efficacy of that treatment non-invasively.

Single Photon Emission Computed Tomography (SPECT) and Positron Emission Tomography (PET) were the first molecular imaging modalities used clinically. SPECT requires the use of a contrast agent labeled with a gamma emitting radionuclide, which should have an ideal gamma energy of 100-250 keV. These gamma rays are recorded by the detectors of a dedicated gamma camera or SPECT instrument and after signal processing can be converted into an image indentifying the localization of the radiotracer. PET requires the injected radiopharmaceutical to be labeled with a positron emitting radionuclide. As the radionuclide decays it ejects a positron from its nucleus which travels a short distance before being annihilated with an electron to release two 511 keV gamma rays 180° apart that are detected by the PET scanner (Figure 1). After sufficient acquisition time the data are reconstructed using computer based algorithms to yield images of the radiotracer’s location within the organism. When compared to SPECT, PET has greater advantages with respect to sensitivity and resolution and has been gaining in clinical popularity, with the number of PET-based studies expected to reach 3.2 million by 2010.1 While SPECT and PET technology has been around for decades, its use remained limited because of the limited availability of relevant isotopes which had to be produced in nuclear reactors or particle accelerators. However, the introduction of the small biomedical cyclotron, the self-contained radionuclide generator and the dedicated small animal or clinical SPECT and PET scanners to hospitals and research facilities has increased the demand for SPECT and PET isotopes.



Figure 1

Cartoon depicting the fundamental principle of Positron Emission Tomography (PET). As the targeting group interacts with the cell surface receptor, the positron emitting radio-metal decays by ejecting β+ particles from its nucleus. After traveling ...



Traditional PET isotopes such as 18F, 15O, 13N and 11C have been developed for incorporation into small molecules, but due to their often lengthy radio-syntheses, short half-lives and rapid clearance, only early time points were available for imaging, leaving the investigation of biological processes, which occur over the duration of hours or days, difficult to explore. With the continuing development of biological targeting agents such as proteins, peptides, antibodies and nanoparticles, which demonstrate a range of biological half-lives, a need arose to produce new radionuclides with half-lives complementary with their biological properties. As a result, the production and radiochemistry of radiometals such as Zr, Y, In, Ga and Cu have been investigated as radionuclide labels for biomolecules since they have the potential to combine their favorable decay characteristics with the biological characteristics of the targeting molecule to become a useful radiopharmaceutical (Tables ​(Tables11 and ​and22).2



Table 1

Gamma- and Beta-Emitting Radiometals





Table 2

Positron-Emitting Radiometals



The number of papers published describing the production or use of these radiometals continues to expand rapidly, and in recognition of this fact, the authors have attempted to present a comprehensive review of this literature as it relates to the production, ligand development and radiopharmaceutical applications of radiometals (excluding 99mTc) since 1999. While numerous reviews have appeared describing certain aspects of the production, coordination chemistry or application of these radiometals,2-18 very few exhaustive reviews have been published.10,12 Additionally, this review has been written to be used as an individual resource or as a companion resource to the review written by Anderson and Welch in 1999.12 Together, they provide a literature survey spanning 50 years of scientific discovery. To accomplish this goal, this review has been organized into three sections: the first section discusses the coordination chemistry of the metal ions Zr, Y, In, Ga and Cu and their chelators in the context of radiopharmaceutical development; the second section describes the methods used to produce Zr, Y, In, Ga and Cu radioisotopes; and the final section describes the application of these radiometals in diagnostic imaging and radiotherapy.

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Coordinating Radiometals of Copper, Gallium, Indium, Yttrium and Zirconium for PET and SPECT Imaging of Disease

Radiometals comprise many useful radioactive isotopes of various metallic elements When properly harnessed, these have valuable emission properties that can be used for diagnostic imaging techniques, such as single photon emission computed tomography (SPECT, eg67Ga, 99mTc, 111In, 177Lu) and positron emission tomography (PET, eg68Ga, 64Cu, 44Sc, 86Y, 89Zr), as well as therapeutic applications (eg47Sc, 114mIn, 177Lu, 90Y, 212/213Bi, 212Pb, 225Ac, 186/188Re) A fundamental critical component of a radiometal-based radiopharmaceutical is the chelator, the ligand system that binds the radiometal ion in a tight stable coordination complex so that it can be properly directed to a desirable molecular target in vivo This article is a guide for selecting the optimal match between chelator and radiometal for use in these systems The article briefly introduces a selection of relevant and high impact radiometals, and their potential utility to the fields of radiochemistry, nuclear medicine, and molecular imaging A description of radiometal-based radiopharmaceuticals is provided, and several key design considerations are discussed The experimental methods by which chelators are assessed for their suitability with a variety of radiometal ions is explained, and a large selection of the most common and most promising chelators are evaluated and discussed for their potential use with a variety of radiometals Comprehensive tables have been assembled to provide a convenient and accessible overview of the field of radiometal chelating agents

Matching chelators to radiometals for radiopharmaceuticals.

Bifunctional coupling agents for radiolabeling of biomolecules and target-specific delivery of metallic radionuclides.

Summation Copper-64 (T1/2 = 12.7 hours; β+, 0.653 MeV [17.8 %]; β−, 0.579 MeV [38.4 %]) has decay characteristics that allow for positron emission tomography (PET) imaging and targeted radiotherapy of cancer. The well-established coordination chemistry of copper allows for its reaction with a wide variety of chelator systems that can potentially be linked to peptides and other biologically relevant small molecules, antibodies, proteins, and nanoparticles. The 12.7-hours half-life of 64Cu provides the flexibility to image both smaller molecules and larger, slower clearing proteins and nanoparticles. In a practical sense, the radionuclide or the 64Cu-radiopharmaceuticals can be easily shipped for PET imaging studies at sites remote to the production facility. Due to the versatility of 64Cu, there has been an abundance of novel research in this area over the past 20 years, primarily in the area of PET imaging, but also for the targeted radiotherapy of cancer. The biologic activity of the hypoxia imaging agen...

Copper-64 Radiopharmaceuticals for PET Imaging of Cancer: Advances in Preclinical and Clinical Research

Molecular imaging has evolved over the past several years into an important tool for diagnosing, understanding, and monitoring disease. Molecular imaging has distinguished itself as an interdisciplinary field, with contributions from chemistry, biology, physics, and medicine. The cross-disciplinary impetus has led to significant achievements, such as the development of more sensitive imaging instruments and robust, safer radiopharmaceuticals, thereby providing more choices to fit personalized medical needs. Molecular imaging is making steadfast progress in the field of cancer research among others. Cancer is a challenging disease, characterized by heterogeneity, uncontrolled cell division, and the ability of cancer cells to invade other tissues. Researchers are addressing these challenges by aggressively identifying and studying key cancer-specific biomarkers such as growth factor receptors, protein kinases, cell adhesion molecules, and proteases, as well as cancer-related biological processes such as hyp...

Molecular imaging of cancer with copper-64 radiopharmaceuticals and positron emission tomography (PET).

The kinetic inertness of copper(II) complexes of several carboxymethyl-armed cyclams and cyclens in 5 M HCl have been determined confirming that the complex derived from cross-bridged cyclam (Cu-CB-TE2A) is by far the most resistant to acid decomplexation. FT-IR studies in D2O solution revealed its unique resistance to full carboxylate protonation and its retention of coordination by both pendant arms even in 1 M DCl. The X-ray structure of its monoprotonated form, [Cu-CB-TE2AH]+, also established full coordination by both COO– and COOH pendant arms in the solid state. Cyclic voltammograms of four carboxymethyl pendant-armed cyclam and cyclen complexes in aqueous solution were obtained with only Cu-CB-TE2A displaying a quasi-reversible CuII/CuI reduction wave. These indicators correlate with the superior in vivo behavior of this complex and its bifunctional conjugate. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2005)

Kinetic Inertness and Electrochemical Behavior of Copper(II) Tetraazamacrocyclic Complexes: Possible Implications for in Vivo Stability

Copper-64, a positron emitter suitable for positron emission tomography (PET), demonstrates improved in vivo clearance when chelated by the cross-bridged tetraazamacrocycle CB-TE2A compared to TETA. Good in vivo clearance was also observed for 64Cu-CB-TE2A conjugated to a peptide, which converts one coordinating carboxylate pendant arm to an amide. To better understand the in vivo stability of peptide- conjugated CB-TE2A, cross-bridged monoamides were synthesized. Crystal structures of natCu(II)-CB-TEAMA and natCu(II)-CB-PhTEAMA revealed hexadentate, distorted octahedral coordination geometry. In vivo biodistribution showed clearance of all 64Cu-radiolabeled cross-bridged monoamides from liver and bone marrow such that uptake at 24 h was 60% of 30 min uptake from 64Cu-TETA was retained in these tissues at 24 h. Clearance of 64Cu-cross-bridged monoamides from nontarget organs suggests good in vivo stability, thus supporting the use of CB-TE2A as a bifunctional chelat...

Synthesis, characterization and in vivo studies of Cu(II)-64-labeled cross-bridged tetraazamacrocycle-amide complexes as models of peptide conjugate imaging agents.

A cross-bridged cyclam ligand bearing two N-carboxymethyl pendant arms (1) has been found to form a copper(II) complex that exhibits significantly improved biological behavior in recent research towards 64Cu-based radiopharmaceuticals. Both the kinetic inertness and resistance to reduction of Cu–1 are believed to be relevant to its enhanced performance. To explore the influence of pendant arm length on these properties, new cross-bridged cyclam and cyclen ligands with longer N-carboxyethyl pendant arms, 2 and 4, and their respective copper(II) complexes have been synthesized. Both mono- as well as di-O-protonated forms of Cu–2 have also been isolated and structurally characterized. The spectral and structural properties of Cu–2 and Cu–4, their kinetic inertness in 5 M HCl, and electrochemical behavior have been obtained and compared to those of their N-carboxymethyl-armed homologs, Cu–1 and Cu–3. Only the cyclam-based Cu–1 and Cu–2 showed unusually high kinetic inertness towards acid decomplexation. While both of these complexes also exhibited quasi-reversible Cu(II)/Cu(I) reductions, Cu–2 is easier to reduce by a substantial margin of +400 mV, bringing it within the realm of physiological reductants. Similarly, of the cyclen-based complexes, Cu–4 is also easier to reduce than Cu–3 though both reductions are irreversible. Biodistribution studies of 64Cu-labeled 2 and 4 were performed in Sprague Dawley rats. Despite comparable acid inertness to their shorter-armed congeners, both longer-armed ligand complexes have poorer bio-clearance properties. This inferior in vivo behavior may be a consequence of their higher reduction potentials.

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The long and short of it: the influence of N-carboxyethyl versusN-carboxymethyl pendant arms on in vitro and in vivo behavior of copper complexes of cross-bridged tetraamine macrocycles.

Katrina S. Woodin

Papers

Kinetic Inertness and Electrochemical Behavior of Copper(II) Tetraazamacrocyclic Complexes: Possible Implications for in Vivo Stability

Synthesis, characterization and in vivo studies of Cu(II)-64-labeled cross-bridged tetraazamacrocycle-amide complexes as models of peptide conjugate imaging agents.

The long and short of it: the influence of N-carboxyethyl versusN-carboxymethyl pendant arms on in vitro and in vivo behavior of copper complexes of cross-bridged tetraamine macrocycles.