Henrik Neu

Effects of the substituted (S)-3-phenylpiperidine (−)-OSU6162 on PET measurements in subhuman primates: Evidence for tone-dependent normalization of striatal dopaminergic activity

(−)‐OSU6162 is a substituted (S)‐3‐phenylpiperidine derivative which exhibits some affinity to the dopamine D2 receptor family. In vivo, the compound displays a unique normalizing profile on psychomotor activity by an intriguing mixture of stimulatory and inhibitory properties. In the present investigation, some of the effects of (−)‐OSU6162 on central dopaminergic function were studied by positron emission tomography (PET) and L‐[11C]DOPA in anaesthetized female rhesus monkeys. (−)‐OSU6162 displayed a dopaminergic tone‐dependent effect with a reduction in the striatal L‐[11C]DOPA influx rate in monkeys with high baseline values and an increased striatal L‐[11C]DOPA influx rate in animals with low baseline values. Infusion of (−)‐OSU6162 for a whole day resulted in a stable effect with no evidence of tolerance. (−)‐OSU6162 also stabilized dopaminergic function by attenuating the upregulation of the striatal L‐[11C]DOPA influx rate which has previously been shown to occur following 6R‐BH4 or 6R‐BH4 + L‐tyrosine infusions. This “Protean” effect of (−)‐OSU6162 on the striatal dopaminergic function corresponds to previous behavioral observations in intact animals and demonstrates a true functional correlation to the measures obtained with L‐[11C]DOPA and PET. The normalizing and stabilizing profile of (−)‐OSU6162 should be of value in treating a variety of disorders where an underlying dysregulation or disruption of dopaminergic function can be assumed. Synapse 28:280–287, 1998.

Receptor imaging technique with 11C-labeled receptor ligands in living brain slices: its application to time-resolved imaging and saturation analysis of benzodiazepine receptor using [11C]Ro15-1788

Recently we developed a novel imaging technique using positron emitter-labeled compounds as probes and a storage phosphor screen as a detector. This approach makes it possible to follow a variety of biochemical processes with spatial information in living brain slices. Further technical development is reported here in terms of time-resolved imaging and receptor characterization in a real equilibrium state. The method was validated by use of [11C]Ro15-1788, a benzodiazepine receptor antagonist. Fresh brain slices were incubated with [11C]Ro15-1788 in oxygenated Krebs-Ringer solution at 37 degrees C, in a specially designed chamber. By placing the chamber on a storage phosphor screen, we could obtain two-dimensional images of radioactivity in the slices. Time-resolved imaging was made at 5 min intervals, revealing that it took 60 min to reach equilibrium binding. The dissociation process was observed by adding an excess amount of unlabeled Ro15-1788 to the chamber, 25 min was required for the full dissociation. In the equilibrium state, i.e. in the presence of free radio-ligand, Scatchard plot analysis was performed on the cerebral cortex (Kd = 7.4 nM, Bmax = 146 fmol/mg tissue) and striatum (Kd = 7.5 nM, Bmax = 107 fmol/mg tissue), suggesting the presence of a single component of binding site in these two regions. The present method, for the first time, made it possible to study a ligand-receptor interaction in living brain slices with temporal and spatial resolutions. This technique should prove useful for studies of receptor function under physiological conditions.

Synthesis of [11C-methyl]-(−)-OSU6162, its regional brain distribution and some pharmacological effects of (−)-OSU6162 on the dopaminergic system studied in the rhesus monkey by positron emission tomography

The labelling of the presynaptic dopamine receptor antagonist (-)-OSU6162, ((S)-(-)-3-(3-(methylsulfonyl)phenyl)-1-propylpiperidine) was performed by an alkylation with [11C]methyl iodide of the thio anion (-)-OSU1281, followed by a selective oxidation to the corresponding methyl sulfone, [11C-methyl]-(-)-OSU6162. The total radiochemical yield calculated from the produced [11C]carbon dioxide to final product was about 25% and the time of synthesis was in the range of 40 min from end of bombardment. The synthesis of the precursor, (-)-OSU1281, was performed from (-)-3PPP in a three-step synthesis. The regional brain distribution of (-)-OSU6162 radiolabelled with 11C was studied in rhesus monkeys by means of positron emission tomography, PET. [11C-Methyl]-(-)-OSU6162 was rapidly and uniformly distributed to gray matters of the brain, and no decrease of radioactivity uptake in the brain was seen after pretreatment with 1 to 3 mg/kg/h of (-)-OSU6162. The effect of doses of 1 to 3 mg/kg/h of (-)-OSU6162 on the dopamine binding was studied by PET using [11C-methyl]raclopride. Radioactivity in the striatum was significantly and dose-dependently decreased by (-)-OSU6162 (r = 0.88), supporting competition with dopamine for selective binding to dopamine receptors.

Syntheses of [21-11C] and (21-13C) progesterone

[21- 11 C]Progesterone was synthesised via the cross-coupling 17β-carboxylic acid chloride-4-androsten-3-one with lithium [ 11 C]methyl(2-thienyl)cuprate (LiCN). The title compound was obtained in 30-35% decay corrected radiochemical yield, based on [ 11 C]methyl iodide, within 35 minutes after end of radionuclide production. The specific radioactivity was 14 GBq/μmole. (21- 13 C)Progesterone was synthesised using the same procedure for determination of the position of the label.

Synthesis of saturated fatty acids 11C(13C)-labelled in the ω-methyl position

A method for the preparation of saturated fatty acids 11 C ( 13 C)-labelled in the ω-methyl position is described. A highly reactive zerovalent copper complex was prepared from lithium naphtalenide reduced lithium(2-thienyl)iodocuprate. The labelling precursors were obtained by addition of tert-butyl ω-iodocarboxylates to the organocuprate and these were reacted with [ 11 C]methyl iodide to form 11 C-labelled, protected intermediates. The tert-butyl ester protecting group was rapidly removed with trifluoroacetic acid, affording fatty acids C-labelled in the ω-methyl position. A solid phase extraction method was developed and preceded final HPLC purification. In a typical run starting with 2.75 GBq of [ 11 C]methyl iodide, 375 MBq (66%) [16- 11 C]palmitic acid was obtained within 46 min from the end of radionuclide production

Lithium [11C]methyl(2-thienyl)cuprate·LiCN in 1,4-additions to α,β-unsaturated ketones. 11C and 13C labelling of the androgen mesterolone

Conjugate 1,4-addition of the [11C]methyl group to α,β-unsaturated ketones was achieved by the use of lithium [11C]methyl(2-thienyl)cuprate · LiCN in a mixture of tetrahydrofuran and diethylether. The 11C labelling procedure was applied successfully to three model compounds and the androgen mesterolone. Mesterolone labelling was also performed with 13C in order to confirm the position of the label. The labelled products were obtained in radiochemical yields of 31–50% within 35–51 minutes of the end of radionuclide production. In a typical synthesis, 1.5 GBq [1α-methyl-11C]mesterolone was synthesized starting from an 18 μA · h cyclotron irradiation.

Synthesis of [18-11C/(13C)]linoleic acid

A method for the preparation of [18-11C]linoleic acid is described. A highly reactive zerovalent copper complex was prepared from lithium (2-thienyl)iodocuprate reduced by lithium naphtalenide. This copper complex was used in a coupling reaction between 17-iodo-cis,cis-9,12-heptadecadienoic acid tert-butyl ester and [11C]methyl iodide to form [18-11C]linoleic acid tert-butyl ester as intermediate. The tert-butyl ester protecting group was rapidly removed with trifluoroacetic acid, affording [18-11C]linoleic acid in 48% radiochemical yield. In a typical run starting with 27 GBq of [11C]methyl iodide, 2.9 GBq [18-11C]linoleic acid was obtained within 44 min from the end of radionuclide production.