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In vivo evaluation of [11C]N-(2-chloro-5-thiomethylphenyl)-N′- (3-methoxy-phenyl)-N′-methylguanidine ([11C]GMOM) as a potential PET radiotracer for the PCP/NMDA receptor

https://doi.org/10.1016/j.nucmedbio.2004.03.012Get rights and content

Abstract

The development of imaging methods to measure changes in NMDA ion channel activation would provide a powerful means to probe the mechanisms of drugs and device based treatments (e.g., ECT) thought to alter glutamate neurotransmission. To provide a potential NMDA/PCP receptor PET tracer, we synthesized the radioligand [11C]GMOM (ki = 5.2 ±0.3 nM; log P = 2.34) and evaluated this ligand in vivo in awake male rats and isoflurane anesthetized baboons. In rats, the regional brain uptake of [11C]GMOM ranged from 0.75±0.13% ID/g in the medulla and pons to 1.15±0.17% ID/g in the occipital cortex. MK801 (1 mg/kg i.v.) significantly reduced (24-28%) [11C]GMOM uptake in all regions. D-serine (10 mg/kg i.v.) increased [11C]GMOM %ID/g values in all regions (10-24%) reaching significance in the frontal cortex and cerebellum only. The NR2B ligand RO 25-6981 (10 mg/kg i.v.) reduced [11C]GMOM uptake significantly (24-38%) in all regions except for the cerebellum and striatum. Blood activity was 0.11±0.03 %ID/g in the controls group and did not vary significantly across groups. PET imaging in isoflurane-anesthetized baboons with high specific activity [11C]GMOM provided fairly uniform regional brain distribution volume (VT) values (12.8–17.1 ml g−1). MK801 (0.5 mg/kg, i.v., n = 1, and 1.0 mg/kg, i.v., n = 1) did not significantly alter regional VT values, indicating a lack of saturable binding. However, the potential confounding effects associated with ketamine induction of anesthesia along with isoflurane maintenance must be considered because both agents are known to reduce NMDA ion channel activation. Future and carefully designed studies, presumably utilizing an optimized NMDA/PCP site tracer, will be carried out to further explore these hypotheses. We conclude that, even though [11C]GMOM is not an optimized PCP site radiotracer, its binding is altered in vivo in awake rats as expected by modulation of NMDA ion channel activity by MK801, D-serine or RO 25-6981. The development of higher affinity NMDA/PCP site radioligands is in progress.

Introduction

The N-methyl-D-aspartate (NMDA) ion channel is a major site of action for glutamate involved in neuroprotection, neurodegeneration, long-term potentiation, memory, and cognition [1], [2], [3], [4], [5], [6], [7], [8], [9]. Alterations in normal NMDA channel composition and function have been implicated in the pathophysiology of certain neurological and neuropsychiatric disorders such as Parkinson's Disease, Huntington's Chorea, schizophrenia, alcoholism and stroke [10], [11], [12], [13], [14]. Each NMDA ion channel is formed by one subunit called NR-1, along with select combinations of the four distinct NR-2 subunits, termed NR-2A, -2B, -2C, and -2D) [2], [15], [16], [17], [18]. In addition, NR3 subunits have been recently described [19]. Several distinct functional receptors and modulatory sites exist on each channel. The best characterized of these are the receptors for the agonist L-glutamate and the co-agonists glycine and D-serine. There are also specific binding sites within the channel for Mg2+ and phencyclidine (called the PCP site); both of these mediate ion flux through the channel. Additional modulatory sites rest on the outside of the channel, including distinct binding sites for polyamines and Zn(II).

Our ultimate goal is to use positron emission tomography (PET) imaging methods to explore the possible etiological role and therapeutic potential of the NMDA ion channel in neurodegenerative and psychiatric disorders. Imaging the PCP site in particular would allow the opportunity for quantification of active NMDA channels, including assessments of the effect of drug action and the active state of the channel [20]. It is well established that the in vitro and in vivo binding of the selective PCP site ligand [3H]MK801 is affected by the state of activation of NMDA receptors [21], [22]. Agonists at NMDA associated glutamate or glycine receptors increase [3H]MK801 binding, while competitive antagonists at these sites reduce [3H]MK801 binding. Therefore, in theory the in vivo binding of a PET PCP site PET ligand could be used to assess the regional activation state of NMDA ion channels in normal subjects and in patients before and after treatment.

To date, about two dozen candidate PCP site tracers have been reported, and a recent review on NMDA/PCP site tracer development has been published [20]. Examples published tracers include [11C]ketamine [23], [24], [25], fluorine-18, iodine-123, and carbon-11 labeled MK801 derivatives [26], [27], [28], fluorine-18 and carbon-11 labeled derivatives of phencyclidine (PCP) such as [18F]fluorothienylcyclohexylpiperidine ([18F]FETCP [29], [30], [31], novel adamantane derivatives including [18F]1-amino-3-fluoromethyl-5-methyl-adamantane ([18F]AFA) [32], [33], and the benzomorphan based tracer [11C]methyl BCLIII277CL [34]. Another approach involves a group of selective non-competitive NMDA/PCP site ligands consisting of trisubstituted N-(phenyl)-N′-(phenyl)-N′-methylguanidines [35], [36], [37]. Compounds of this series comprise some of the highest affinity PCP site ligands reported to date, moderate lipophilicity, and several exhibit specificity for the PCP site versus sigma receptors and therefore should produce a variety of potential NMDA radioligands. Herein, we report the in vivo evaluation of the first novel PET tracer of this series, [11C]N-(2-chloro-3-thiomethylphenyl)-N′-(3-methoxyphenyl)-N′-methylguanidine ([11C]GMOM) [38]. A preliminary report of this work has been published [39].

Section snippets

General

N-(2-Chloro-5-thiomethylphenyl)-N′-(3-methoxyphenyl)-N′-methylguanidine (GMOM) and the corresponding phenol precursor were synthesized as previously reported [38]. All other chemicals, reagents and solvents were obtained from commercial sources and were used without further purification. [11C]Methyl iodide is prepared routinely at Columbia University Radioligand Laboratory using established methods. HPLC purification and quality control analysis of the radioligand was performed using a Waters

Results and discussion

[11C]GMOM was synthesized in good yield and high specific activity using our previously reported method [39] (Scheme 1). Radiochemical yield ranged from 5.8 to 13.0% (EOS) and high radiochemical purity (96.7±1.5%) and specific activity (1.25±0.45 Ci/μmol EOS) were obtained. The tracer was utilized for in vivo studies between 20 and 30 min after synthesis. The regional brain distribution and specific binding of [11C]GMOM was examined through distribution studies in male rats ([11C]GMOM dose: 6–8

Acknowledgements

Funding for this work was provided for by grants from the National Institutes of Health (NIAAA IP50AA-12870-01 and NIMH MH59342-01).

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