Norman Simpson

[11C]WAY 100635: a radioligand for imaging 5-HT1A receptors with positron emission tomography.

The potent and selective 5-HT1A antagonist WAY 100635 (N-[2-]4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-N-(2- pyridinyl)cyclohexanecarboxamide) was radiolabeled with 11C in high specific activity, and the in vivo properties of this radioligand were assessed in the brains of rats and monkeys. Following i.v. tail vein injection in rats, [11C]WAY 100635 rapidly penetrated into brain tissue and was retained over a 30-90 min time period in a manner consistent with the known distribution of 5-HT1A receptors. Pretreatment of rats with the selective 5-HT1A agonist (+/-)-8-OH-DPAT effectively blocked the retention of radioactivity in brain regions known to contain high densities of 5-HT1A receptors. The hippocampus-to-cerebellum radioactivity concentration ratio reached a maximum of 16:1 at 60 min post injection. Following i.v. injection of [11C]WAY 100635 in rhesus monkeys, the concentrations of radioactivity in brain regions were consistent with the reported distribution of 5-HT1A receptors in primates, and the frontal cortex-to-cerebellum ratio reached 5.5:1 at 80 min post injection. Pretreatment of the monkeys with (+/-)-8-OH-DPAT reduced this ratio to 1.4:1, and injection of (+/-)-8-OH-DPAT 20 min after the injection of [11C]WAY 100635 significantly displaced frontal cortex binding. The in vivo properties of [11C]WAY 100635 in rats and monkeys strongly support the future utility of this radioligand for imaging 5-HT1A receptors using positron emission tomography (PET).

Test–retest variability of serotonin 5-HT2A receptor binding measured with positron emission tomography and [18F]altanserin in the human brain

The role of serotonin in CNS function and in many neuropsychiatric diseases (e.g., schizophrenia, affective disorders, degenerative dementias) support the development of a reliable measure of serotonin receptor binding in vivo in human subjects. To this end, the regional distribution and intrasubject test–retest variability of the binding of [18F]altanserin were measured as important steps in the further development of [18F]altanserin as a radiotracer for positron emission tomography (PET) studies of the serotonin 5‐HT2A receptor. Two high specific activity [18F]altanserin PET studies were performed in normal control subjects (n = 8) on two separate days (2–16 days apart). Regional specific binding was assessed by distribution volume (DV), estimates that were derived using a conventional four compartment (4C) model, and the Logan graphical analysis method. For both analysis methods, levels of [18F]altanserin binding were highest in cortical areas, lower in the striatum and thalamus, and lowest in the cerebellum. Similar average differences of 13% or less were observed for the 4C model DV determined in regions with high receptor concentrations with greater variability in regions with low concentrations (16–20%). For all regions, the absolute value of the test–retest differences in the Logan DV values averaged 12% or less. The test–retest differences in the DV ratios (regional DV values normalized to the cerebellar DV) determined by both data analysis methods averaged less than 10%. The regional [18F]altanserin DV values using both of these methods were significantly correlated with literature‐based values of the regional concentrations of 5‐HT2A receptors determined by postmortem autoradiographic studies (r2 = 0.95, P < 0.001 for the 4C model and r2 = 0.96, P < 0.001 for the Logan method). Brain uptake studies in rats demonstrated that two different radiolabeled metabolites of [18F]altanserin (present at levels of 3–25% of the total radioactivity in human plasma 10–120 min postinjection) were able to penetrate the blood–brain barrier. However, neither of these radiolabeled metabolites bound specifically to the 5‐HT2A receptor and did not interfere with the interpretation of regional [18F]altanserin‐specific binding parameters obtained using either a conventional 4C model or the Logan graphical analysis method. In summary, these results demonstrate that the test–retest variability of [18F]altanserin‐specific binding is comparable to that of other PET radiotracers and that the regional specific binding of [18F]altanserin in human brain was correlated with the known regional distribution of 5‐HT2A receptors. These findings support the usefulness of [18F]altanserin as a radioligand for PET studies of 5‐HT2A receptors. Synapse 30:380–392, 1998.

CHAPTER 16 – Scatter Correction for Brain Receptor Quantitation in 3D PET

This chapter investigates the performance of the two correction algorithms when applied to scatter from activity outside the field of view. Two studies with the Utah phantom have been presented in the chapter: In the first, the entire water-filled phantom contains uniform activity with the exception of the short cylinder, which contains only water, and a chamber, which is empty. The fraction of scattered events in the image can then be estimated from the mean value in a region of interest placed on the short (cold) cylinder expressed as a fraction of the activity in the inner cylinder. In the second study, the annulus contains approximately four times the activity in the inner cylinder to simulate increased cortical uptake. In addition, the long cylinder contains twice the background activity to simulate uptake in subcortical structures. This chapter also explores the quantitative potential of 3D positron emission tomography (PET), and in particular the efficacy of the two scatter correction procedures, when applied to neuroreceptor ligand studies. For this, a ligand for imaging central benzodiazepine receptors is used.

PET Measurement of Endogenous Neurotransmitter Activity Using High and Low Affinity Radiotracers

Functional imaging of the in vivo competition of dopamine-D 2 radioligands with endogenous dopamine has provided indirect measures of dopamine activity. Binding parameter changes ( > 20%) have been observed for [ 11 C]raclopride positron emission tomography studies performed after d-amphetamine administration, and this sensitivity has been partially attributed to its low D 2 receptor affinity. In this work, a d-amphetamine pretreatment/bolus radiotracer injection paradigm was used to examine binding parameter changes for the high and low affinity benzamides, [ 18 F]fallypride and [ 11 C]raclopride; and a preliminary examination of [ 18 F]fallypride extrastriatal D 2 binding at baseline and after haloperidol pretreatment was performed. [ 11 C]aclopride studies were performed in baboons at baseline and after d-amphetamine pretreatment (0.6 mg/kg, n = 3 or 1.0 mg/kg, n = 2). Two baboons had additional [ 18 F]fallypride studies at baseline and after d-amphetamine, and a third baboon was studied at baseline and after haloperidol pretreatment (1 mg/kg). Kinetic analyses provided distribution volume (DV) and DV-de-rived binding potential (BP DV ) estimates. For [ 11 C]raclopride, the average striatal BP DV change was – 26 ± 2.6% for the 0.6-mg/kg amphetamine dose, with slightly greater changes of −31 and −39% after the 1.0-mg/kg dose. Lower striatal changes were observed for [ 18 F]fallypride: – 14% (0.6 mg/kg) and – 29% (1.0 mg/kg). Baseline regional BP DV values were highly correlated (r 2 > 0.9) with human D 2 receptor rank order but did not correlate after 1 mg/kg haloperidol pretreatment (r 2 0.1).Functional imaging of the in vivo competition of dopamine-D2radioligands with endogenous dopamine has provided indirect measures of dopamine activity. Binding parameter changes (> 20%) have been observed for [11C]raclopride positron emission tomography studies performed after d-amphetamine administration, and this sensitivity has been partially attributed to its low D2 receptor affinity. In this work, a d-amphetamine pretreatment/bolus radiotracer injection paradigm was used to examine binding parameter changes for the high and low affinity benzamides, [18F]fallypride and [11C]raclopride; and a preliminary examination of [18F]fallypride extrastriatal D2 binding at baseline and after haloperidol pretreatment was performed. [11C]aclopride studies were performed in baboons at baseline and after d-amphetamine pretreatment (0.6 mg/kg, n = 3 or 1.0 mg/kg, n = 2). Two baboons had additional [18F]fallypride studies at baseline and after d-amphetamine, and a third baboon was studied at baseline and after haloperidol pretreatment (1 mg/kg). Kinetic analyses provided distribution volume (DV) and DV-de-rived binding potential (BPDV) estimates. For [11C]raclopride, the average striatal BPDV change was –26 ± 2.6% for the 0.6-mg/kg amphetamine dose, with slightly greater changes of −31 and −39% after the 1.0-mg/kg dose. Lower striatal changes were observed for [18F]fallypride: –14% (0.6 mg/kg) and –29% (1.0 mg/kg). Baseline regional BPDVvalues were highly correlated (r2 > 0.9) with human D2 receptor rank order but did not correlate after 1 mg/kg haloperidol pretreatment (r2 < 0.1).

Synthesis and in vitro evaluation of [18F]FECIMBI-36: A potential agonist PET ligand for 5-HT2A/2C receptors.

Radiosynthesis and in vitro evaluation of [(18)F]-2-(4-bromo-2,5-dimethoxyphenyl)-N-(2-(2-fluoroethoxy)benzyl)ethanamine, ([(18)F]FECIMBI-36) or ([(18)F]1), a potential agonist PET imaging agent for 5-HT2A/2C receptors is described. Syntheses of reference standard 1 and the corresponding des-fluoroethyl radiolabeling precursor (2) were achieved with 75% and 65% yields, respectively. In vitro pharmacology assay of FECIMBI-36 by [(3)H]-ketanserin competition binding assay obtained from NIMH-PDSP showed high affinities to 5-HT2AR (Ki = 1nM) and 5-HT2CR (Ki=1.7 nM). Radiolabeling of FECIMBI-36 was achieved from the boc-protected precursor 2 using [(18)F]-fluoroethyltosylate in presence of Cs2CO3 in DMSO followed by removal of the protective group. [(18)F]1 was isolated using RP-HPLC in 25 ± 5% yield, purity > 95% and specific activity 1-2Ci/μmol (N = 6). In vitro autoradiography studies demonstrate that [(18)F]1 selectively label 5-HT2A and 5-HT2C receptors in slide-mounted sections of postmortem human brain using phosphor imaging. Our results indicate the potential of [(18)F]1 for imaging 5-HT2A/2C receptors in the high affinity state in vivo using PET imaging.

Stereoconservative synthesis of the enantiomerically pure precursors of [11C](+)-McN 5652 and [11C](-)-McN 5652

A stereoconservative synthetic route for the preparation of the precursors of [11C](+)-McN 5652 and [11C](−)-McN 5652, also known as [11C]McN-5652-X and [11C]McN-5652-W, is reported. The key steps involved the transformation of the (+)-trans-4-bromophenyl or (−)-trans-4-bromophenyl analogues of (+)-McN 5652 or (−)-McN 5652 to triisopropylsilyl-protected thioethers and conversion of the thioethers to benzoyl thioesters via a one-pot deprotection and esterification sequence. Subsequent formation of a tartrate salt of the benzoyl thioesters provided stable, convenient precursors for the routine radiosynthesis of [11C](+)-McN 5652 and [11C](−)-McN 5652. The radiosyntheses were accomplished by S-methylation with high specific activity [11C]methyl iodide of the phenylsulfides, which were generated in situ from the tartrate salt of the benzoyl thioesters. HPLC analyses of the final products demonstrated that all of the reactions were stereoconservative, with no detectable isomerization or racemization taking place in any of the reaction sequences. [11C](+)-McN 5652 and [11C](−)-McN 5652 were obtained with end of synthesis (EOS) yields of ∽120 mCi and EOS specific activities of ∽4000 Ci/mmol following radiosynthesis and purification procedures that required approximately 25 min. The chemical and radiochemical purities of the final products were greater than 95%.

Association of BDNF Val66Met Polymorphism and Brain BDNF Levels with Major Depression and Suicide

Abstract Background Brain-derived neurotrophic factor is implicated in the pathophysiology of major depressive disorder and suicide. Both are partly caused by early life adversity, which reduces brain-derived neurotrophic factor protein levels. This study examines the association of brain-derived neurotrophic factor Val66Met polymorphism and brain brain-derived neurotrophic factor levels with depression and suicide. We hypothesized that both major depressive disorder and early life adversity would be associated with the Met allele and lower brain brain-derived neurotrophic factor levels. Such an association would be consistent with low brain-derived neurotrophic factor mediating the effect of early life adversity on adulthood suicide and major depressive disorder. Methods Brain-derived neurotrophic factor Val66Met polymorphism was genotyped in postmortem brains of 37 suicide decedents and 53 nonsuicides. Additionally, brain-derived neurotrophic factor protein levels were determined by Western blot in dorsolateral prefrontal cortex (Brodmann area 9), anterior cingulate cortex (Brodmann area 24), caudal brainstem, and rostral brainstem. The relationships between these measures and major depressive disorder, death by suicide, and reported early life adversity were examined. Results Subjects with the Met allele had an increased risk for depression. Depressed patients also have lower brain-derived neurotrophic factor levels in anterior cingulate cortex and caudal brainstem compared with nondepressed subjects. No effect of history of suicide death or early life adversity was observed with genotype, but lower brain-derived neurotrophic factor levels in the anterior cingulate cortex were found in subjects who had been exposed to early life adversity and/or died by suicide compared with nonsuicide decedents and no reported early life adversity. Conclusions This study provides further evidence implicating low brain brain-derived neurotrophic factor and the brain-derived neurotrophic factor Met allele in major depression risk. Future studies should seek to determine how altered brain-derived neurotrophic factor expression contributes to depression and suicide.

Radiosynthesis of [18F]ATPFU: A potential PET ligand for mTOR

Mammalian target of rapamycin (mTOR) plays a pivotal role in many aspects of cellular proliferation, and recent evidence suggests that an altered mTOR signaling pathway plays a central role in the pathogenesis of aging, tumor progression, neuropsychiatric, and major depressive disorder. Availability of a mTOR-specific PET tracer will facilitate monitoring early response to treatment with mTOR inhibitors that are under clinical development. Towards this we have developed the radiosynthesis of [(18)F]1-(4-(4-(8-oxa-3-azabicyclo[3.2.1]octan-3-yl)-1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-d]pyrimidin-6-yl)phenyl)-3-(2-fluoroethyl)urea [(18)F]ATPFU ([(18)F]1) as an mTOR PET ligand. Synthesis of reference 1 and the precursor for radiolabeling, 4-(4-8-oxa-3-azabicyclo[3.2.1]-octan-3yl)-1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-d]pyrimidin-6yl)aniline (10), were achieved from beta-chloroaldehyde 3 in 4 and 5 steps, respectively, with an overall yield of 25-28%. [(18)F]Fluoroethylamine was prepared by heating N-[2-(toluene-4-sulfonyloxy)ethyl]phthalimide with [(18)F]fluoride ion in acetonitrile. [(18)F]1 was obtained by slow distillation under argon of [(18) F]FCH2CH2NH2 into amine 10 that was pre-treated with triphosgene at 0-5 °C. The total time required for the two-step radiosynthesis including semi-preparative HPLC purification was 90 min, and the overall radiochemical yield of [(18)F]1 for the process was 15 ± 5% based on [(18)F]fluoride ion (decay corrected). At the end of synthesis (EOS), the specific activity was 37-74 GBq/µmol (N = 6).

In vivo evaluation of IGF1R/IR PET ligand [18F]BMS-754807 in rodents

In vivo evaluation of [18F]BMS-754807 binding in mice and rats using microPET and biodistribution methods is described herein. The radioligand shows consistent binding characteristics, in vivo, in both species. Early time frames of the microPET images and time activity curves of brain indicate poor penetration of the tracer across the blood brain barrier (BBB) in both species. However, microPET experiments in mice and rats show high binding of the radioligand outside the brain to heart, pancreas and muscle, the organs known for higher expression of IGF1R/1R. Biodistribution analysis 2h after injection of [18F]BMS-754807 in rats show negligible [18F]defluorination as reflected by the low bone uptake and clearance from blood. Overall, the data indicate that [18F]BMS-754807 can potentially be a radiotracer for the quantification of IGF1R/IR outside the brain using PET.

Autoradiographic Evaluation of [18F]FECUMI-101, a High Affinity 5-HT1AR Ligand in Human Brain

[(18)F]FECUMI-101 ([(18)F]1) is a 5HT1AR ligand demonstrating specific binding in brain regions corresponding to the distribution of 5-HT1AR in baboons. However, we detected moderate uptake of [(18)F]1 in baboon thalamus, a brain region lacking 5-HT1AR. We sought to investigate the relative binding of [(18)F]1 to 5-HT1AR, α1R, and 5-HT7R in vitro. Using autoradiography in human brain sections, specific binding of [(18)F]1 to 5-HT1AR was confirmed. However, [(18)F]1 also showed 26% binding to α1R in PFC. The hippocampal formation exhibited 51% and 92% binding of [(18)F]1 to α1R and 5-HT1AR, respectively. Thalamus and cerebellum showed very little binding. There is no measurable specific binding of [(18)F]1 to 5-HT7R and no effect of temperature on [(18)F]1 specific binding to 5-HT1AR or α1R. These results indicate that, while [(18)F]FECUMI-101 is not a completely selective 5-HT1AR ligand for receptor quantification, it may be useful for occupancy measurements of drugs acting at 5-HT1AR in vivo.