Development Of New Pet And Spect Radiotracers And New Ap

National Institute On Drug Abuse

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Abstract

Analogs of A-85380 and A-84543 have been evaluated as imaging agents for nicotinic acetylcholine receptors (nAChRs) with positron emission tomography (PET) and single photon emission computed tomography (SPECT). Radiation dosimetry and toxicological studies in animals demonstrated the safety of 5-[I-125]iodo-A-85380 (5IA). In collaboration with Yale University, the first human SPECT studies with 5IA revealed that cerebral nAChRs can be visualized in non-smokers and dosimetry data from these studies indicate that multiple studies can be performed on a single volunteer without exceeding radiation dosimetry limits. Similarly, the first human PET studies with 2-[F-18]fluoro-A-85380 (2FA) demonstrate the feasibility of quantitatively imaging nAChRs in the thalamus and visualizing these receptors in brain regions containing low to moderate receptor densities. Mice studies showed that the administration of the non-selective inhibitor of cytochrome P450, cimetidine, substantially slowed the rapid in vivo metabolism of 5IA, suggesting that the use of this or similar compounds could reduce the dose of radioactivity needed to successfully image nAChRs in human volunteers. Graphical analysis of PET data from Rhesus monkeys provided binding potential values for 2FA in the thalamus, cortex, striatum and cerebellum that were consistent with the distribution pattern of alpha4beta2 nAChRs. Methodology was developed and data were collected for estimating the density of nAChRs in the thalamus of non-human primates (NHPs).In collaboration with researchers from the University of Michigan, the loss of nAChRs in the striatum of unilaterally MPTP-lesioned NHPs with 2FA and PET was demonstrated. Kinetic studies with 2FA and PET in NHPs revealed that 2FA accumulates relatively slowly in brain, partially because its low lipophilicity slows its blood-brain-barrier penetration. Efforts were directed at developing radioligands with faster kinetics for imaging nAChRs in vivo. To that end, six novel analogs of A-85380 and A-84543 with calculated log P (Clog P) values in the range of 1.2 to 3 were synthesized, radiolabeled and evaluated in vitro and in vivo. Molecular modeling of the structures of these compounds revealed differences in their conformational profiles and the electronic properties, providing insight into structure-activity relationships with nAChRs. The molecular modeling was performed in collaboration with staff in the Medicinal Chemistry Section of the Medications Discovery Research Branch. Increasing the lipophilicity from Clog P = 0.6 (2FA) yielded ligands with faster blood brain barrier penetration and increased nonspecific binding. Although [11C]methyl-PVC, the best radioligand from the series, exhibits specific in vivo receptor binding that is half that of 2FA, [11C]methyl-PVC may be useful for studying the in vivo occupancy of nAChRs by endogenous or exogenous ligands. From in vitro and in vivo studies with these radioligands (Kd values at 37 degrees C: 9 to 600 pM, Clog P values: 0.6 to 3), an algorithm was developed for predicting binding potential (BP). BP is the one of the most important characteristic of a radioligand as it characterizes the specificity of radioligand receptor binding and reflects its suitability for qualitative and quantitative in vivo imaging. BP values for six radioligands calculated from their Kds at 37 oC, their Clog Ps, and the density of alpha4beta2 nAChRs in each brain region of interest and this algorithm were in good agreement with those observed in vivo in Rhesus monkeys, suggesting that this algorithm might be useful for screening potential compounds for a new radioligand for alpha4beta2 nAChRs with improved characteristics. As part of our effort to develop radioligands for in vivo imaging of cannabinoid receptors, we synthesized and evaluated a series of 1,5-diarylpyrazoles by modifying the substituents on positions 1,3,4 and 5 of the pyrazole core. As part of this project, we developed a practical method for preparing C-4 fluoro pyrazoles using electrophilic fluorination. One compound in the series exhibited lower lipophilicity than and comparable binding affinity to SR141716, a selective CB1 antagonist, for the CB1 cannabinoid receptor. Greater accumulation of radioactivity after administration of this compound to mice was observed in CB1-rich regions (e.g., hippocampus, striatum and cerebellum) than in receptor poor brain regions, suggesting specific binding to these receptors. Similar findings were obtained in PET studies of Rhesus monkeys with this compound.