Andri C. Tziortzi

Imaging dopamine receptors in humans with [11C]-(+)-PHNO: Dissection of D3 signal and anatomy

[(11)C]-(+)-PHNO is a D3 preferring PET radioligand which has recently opened the possibility of imaging D3 receptors in the human brain in vivo. This imaging tool allows characterisation of the distribution of D3 receptors in vivo and further investigation of their functional role. The specific [(11)C]-(+)-PHNO signal is a mixture of D3 and D2 components with the relative magnitude of each component determined by the regional receptor densities. An accurate and reproducible delineation of regions of interest (ROI) is therefore important for optimal analysis of human PET data. We present a set of anatomical guidelines for the delineation of D3 relevant ROIs including substantia nigra, hypothalamus, ventral pallidum/substantia innominata, ventral striatum, globus pallidus and thalamus. Delineation of these structures using this approach allowed for high intra- and inter-operator reproducibility. Subsequently we used a selective D3 antagonist to dissect the total [(11)C]-(+)-PHNO signal in each region into its D3 and D2 components and estimated the regional fraction of the D3 signal (f(PHNO)(D3)). In descending order of magnitude the following results for the f(PHNO)(D3) were obtained: hypothalamus=100%, substantia nigra=100%, ventral pallidum/substantia innominata=75%, globus pallidus=65%, thalamus=43%, ventral striatum=26% and precommissural-ventral putamen=6%. An automated approach for the delineation of these anatomical regions of interest was also developed and investigated in terms of its reproducibility and accuracy.

Imaging Dopamine D3 Receptors in the Human Brain with Positron Emission Tomography, [11C]PHNO, and a Selective D3 Receptor Antagonist

1. Clinical Imaging Centre, GlaxoSmithKline, London, U.K. 2. Neurosciences Centre of Excellence in Drug Discovery, GlaxoSmithKline, Verona, Italy 3. Department of Psychiatry, Columbia University, New York, USA 4. Centre for Addiction and Mental Health, Toronto, Canada 5. Department of Engineering Science, University of Oxford, U.K. 6. Division of Neuroscience and Mental Health, Imperial College, London, U.K.BACKGROUND Dopamine D(3) receptors are involved in the pathophysiology of several neuropsychiatric conditions. [(11)C]-(+)-PHNO is a radiolabeled D(2) and D(3) agonist, suitable for imaging the agonist binding sites (denoted D(2HIGH) and D(3)) of these receptors with positron emission tomography (PET). PET studies in nonhuman primates documented that, in vivo, [(11)C]-(+)-PHNO displays a relative selectivity for D(3) compared with D(2HIGH) receptor sites and that the [(11)C]-(+)-PHNO signal is enriched in D(3) contribution compared with conventional ligands such as [(11)C] raclopride. METHODS To define the D(3) contribution (f(PHNO)(D3)) to [(11)C]-(+)-PHNO binding potential (BP(ND)) in healthy humans, 52 PET scans were obtained in 19 healthy volunteers at baseline and following oral administration of various doses of the selective D(3) receptor antagonist, GSK598809. RESULTS The impact of GSK598809 on [(11)C]-(+)-PHNO was regionally selective. In dorsal regions of the striatum, GSK598809 did not significantly affect [(11)C]-(+)-PHNO BP(ND) (f(PHNO)(D3) approximately 0%). Conversely, in the substantia nigra, GSK598809 dose-dependently reduced [(11)C]-(+)-PHNO binding to nonspecific level (f(PHNO)(D3) approximately 100%). In ventral striatum (VST), globus pallidus and thalamus (THA), [(11)C]-(+)-PHNO BP(ND) was attributable to a combination of D(2HIGH) and D(3) receptor sites, with f(PHNO)(D3) of 26%, 67% and 46%, respectively. D(3) receptor binding potential (BP(ND)(D3)) was highest in globus pallidus (1.90) and substantial nigra (1.39), with lower levels in VST (.77) and THA (.18) and negligible levels in dorsal striatum. CONCLUSIONS This study elucidated the pharmacologic nature of the [(11)C]-(+)-PHNO signal in healthy subjects and provided the first quantification of D(3) receptor availability with PET in the living human brain.

Connectivity-Based Functional Analysis of Dopamine Release in the Striatum Using Diffusion-Weighted MRI and Positron Emission Tomography

The striatum acts in conjunction with the cortex to control and execute functions that are impaired by abnormal dopamine neurotransmission in disorders such as Parkinsons and schizophrenia. To date, in vivo quantification of striatal dopamine has been restricted to structure-based striatal subdivisions. Here, we present a multimodal imaging approach that quantifies the endogenous dopamine release following the administration of d-amphetamine in the functional subdivisions of the striatum of healthy humans with [(11)C]PHNO and [(11)C]Raclopride positron emission tomography ligands. Using connectivity-based (CB) parcellation, we subdivided the striatum into functional subregions based on striato-cortical anatomical connectivity information derived from diffusion magnetic resonance imaging (MRI) and probabilistic tractography. Our parcellation showed that the functional organization of the striatum was spatially coherent across individuals, congruent with primate data and previous diffusion MRI studies, with distinctive and overlapping networks. d-amphetamine induced the highest dopamine release in the limbic followed by the sensory, motor, and executive areas. The data suggest that the relative regional proportions of D2-like receptors are unlikely to be responsible for this regional dopamine release pattern. Notably, the homogeneity of dopamine release was significantly higher within the CB functional subdivisions in comparison with the structural subdivisions. These results support an association between local levels of dopamine release and cortical connectivity fingerprints.

Within-Subject Comparison of [11C]-( + )-PHNO and [11C]raclopride Sensitivity to Acute Amphetamine Challenge in Healthy Humans

[11C]PHNO is a D2/D3 agonist positron emission tomography radiotracer, with higher in vivo affinity for D3 than for D2 receptors. As [11C]-( + )-PHNO is an agonist, its in vivo binding is expected to be more affected by acute fluctuations in synaptic dopamine than that of antagonist radiotracers such as [11C]raclopride. In this study, the authors compared the effects of an oral dose of the dopamine releaser amphetamine (0.3 mg/kg) on in vivo binding of [11C]-( + )-PHNO and [11C]raclopride in healthy subjects, using a within-subjects, counterbalanced, open-label design. In the dorsal striatum, where the density of D3 receptors is negligible and both tracers predominantly bind to D2 receptors, the reduction of [11C]-( + )-PHNO binding potential (BPND) was 1.5 times larger than that of [11C]raclopride. The gain in sensitivity associated with the agonist [11C]-( + )-PHNO implies that ~65% of D2 receptors are in the high-affinity state in vivo. In extrastriatal regions, where [11C]-( + )-PHNO predominantly binds to D3 receptors, the amphetamine effect on [11C]-( + )-PHNO BPND was even larger, consistent with the higher affinity of dopamine for D3. This study indicates that [11C]- ( + )-PHNO is superior to [11C]raclopride for studying acute fluctuations in synaptic dopamine in the human striatum. [11C]-( + )-PHNO also enables measurement of synaptic dopamine in D3 regions.

Loss of phosphodiesterase 10A expression is associated with progression and severity in Parkinson’s disease

The mechanisms underlying neurodegeneration and loss of dopaminergic signalling in Parkinsons disease are still only partially understood. Phosphodiesterase 10A (PDE10A) is a basal ganglia expressed dual substrate enzyme, which regulates cAMP and cGMP signalling cascades, thus having a key role in the regulation of dopaminergic signalling in striatal pathways, and in promoting neuronal survival. This study aimed to assess in vivo the availability of PDE10A in patients with Parkinsons disease using positron emission tomography molecular imaging with (11)C-IMA107, a highly selective PDE10A radioligand. We studied 24 patients with levodopa-treated, moderate to advanced Parkinsons disease. Their positron emission tomography imaging data were compared to those from a group of 12 healthy controls. Parametric images of (11)C-IMA107 binding potential relative to non-displaceable binding (BPND) were generated from the dynamic (11)C-IMA107 scans using the simplified reference tissue model with the cerebellum as the reference tissue. Corresponding region of interest analysis showed lower mean (11)C-IMA107 BPND in the caudate (P < 0.001), putamen (P < 0.001) and globus pallidus (P = 0.025) in patients with Parkinsons disease compared to healthy controls, which was confirmed with voxel-based analysis. Longer Parkinsons duration correlated with lower (11)C-IMA107 BPND in the caudate (r = -0.65; P = 0.005), putamen (r = -0.51; P = 0.025), and globus pallidus (r = -0.47; P = 0.030). Higher Unified Parkinsons Disease Rating Scale part-III motor scores correlated with lower (11)C-IMA107 BPND in the caudate (r = -0.54; P = 0.011), putamen (r = -0.48; P = 0.022), and globus pallidus (r = -0.70; P < 0.001). Higher Unified Dyskinesia Rating Scale scores in those Parkinsons disease with levodopa-induced dyskinesias (n = 12), correlated with lower (11)C-IMA107 BPND in the caudate (r = -0.73; P = 0.031) and putamen (r = -0.74; P = 0.031). Our findings demonstrate striatal and pallidal loss of PDE10A expression, which is associated with Parkinsons duration and severity of motor symptoms and complications. PDE10A is an enzyme that could be targeted with novel pharmacotherapy, and this may help improve dopaminergic signalling and striatal output, and therefore alleviate symptoms and complications of Parkinsons disease.

Altered PDE10A expression detectable early before symptomatic onset in Huntington’s disease

There is an urgent need for early biomarkers and novel disease-modifying therapies in Huntingtons disease. Huntingtons disease pathology involves the toxic effect of mutant huntingtin primarily in striatal medium spiny neurons, which highly express phosphodiesterase 10A (PDE10A). PDE10A hydrolyses cAMP/cGMP signalling cascades, thus having a key role in the regulation of striatal output, and in promoting neuronal survival. PDE10A could be a key therapeutic target in Huntingtons disease. Here, we used combined positron emission tomography (PET) and multimodal magnetic resonance imaging to assess PDE10A expression in vivo in a unique cohort of 12 early premanifest Huntingtons disease gene carriers with a mean estimated 90% probability of 25 years before the predicted onset of clinical symptoms. We show bidirectional changes in PDE10A expression in premanifest Huntingtons disease gene carriers, which are associated with the probability of symptomatic onset. PDE10A expression in early premanifest Huntingtons disease was decreased in striatum and pallidum and increased in motor thalamic nuclei, compared to a group of matched healthy controls. Connectivity-based analysis revealed prominent PDE10A decreases confined in the sensorimotor-striatum and in striatonigral and striatopallidal projecting segments. The ratio between higher PDE10A expression in motor thalamic nuclei and lower PDE10A expression in striatopallidal projecting striatum was the strongest correlate with higher probability of symptomatic conversion in early premanifest Huntingtons disease gene carriers. Our findings demonstrate in vivo, a novel and earliest pathophysiological mechanism underlying Huntingtons disease with direct implications for the development of new pharmacological treatments, which can promote neuronal survival and improve outcome in Huntingtons disease gene carriers.

In Vivo Imaging of Cerebral Dopamine D3 Receptors in Alcoholism

Animal studies support the role of the dopamine D3 receptor (DRD3) in alcohol reinforcement or liking. Sustained voluntary alcohol drinking in rats has been associated with an upregulation of striatal DRD3 gene expression and selective blockade of DRD3 reduces ethanol preference, consumption, and cue-induced reinstatement. In vivo measurement of DRD3 in the living human brain has not been possible until recently owing to a lack of suitable tools. In this study, DRD3 status was assessed for the first time in human alcohol addiction. Brain DRD3 availability was compared between 16 male abstinent alcohol-dependent patients and 13 healthy non-dependent age-matched males using the DRD3-preferring agonist positron emission tomography (PET) radioligand [11C]PHNO with and without blockade with a selective DRD3 antagonist (GSK598809 60 mg p.o.). In striatal regions of interest, where the [11C]PHNO PET signal represents primarily DRD2 binding, no differences were seen in [11C]PHNO binding between the groups at baseline. However, baseline [11C]PHNO binding was higher in alcohol-dependent patients in hypothalamus (VT: 16.5±4 vs 13.7±2.9, p=0.040), a region in which the [11C]PHNO signal almost entirely reflects DRD3 availability. The reductions in regional receptor binding (VT) following a single oral dose of GSK598809 (60 mg) were consistent with those observed in previous studies across all regions. There were no differences in regional changes in VT following DRD3 blockade between the two groups, indicating that the regional fractions of DRD3 are similar in the two groups, and the increased [11C]PHNO binding in the hypothalamus in alcohol-dependent patients is explained by elevated DRD3 in this group. Although we found no difference between alcohol-dependent patients and controls in striatal DRD3 levels, increased DRD3 binding in the hypothalamus of alcohol-dependent patients was observed. This may be relevant to the development of future therapeutic strategies to treat alcohol abuse.

Mathematical modelling of [11C]-(+)-PHNO human competition studies

The D(2)/D(3) agonist radioligand [(11)C]-(+)-PHNO is currently the most suitable D(3) imaging agent available, despite its limited selectivity for the D(3) over the D(2). Given the collocation of D(2) and D(3) receptors, and generally higher densities of D(2), the separation of D(2) and D(3) information from [(11)C]-(+)-PHNO PET data are somewhat complex. This complexity is compounded by recent data suggesting that [(11)C]-(+)-PHNO PET scans might be routinely performed in non-tracer conditions (with respect to D(3) receptors), and that the cerebellum (used as a reference region) might manifest some displaceable binding signal. Here we present the modelling and analysis of data from two human studies which employed an adequate dose range of selective D(3) antagonists (GSK598809 and GSK618334) to interrogate the [(11)C]-(+)-PHNO PET signal. Models describing the changes observed in the PET volume of distribution (V(T)) and binding potential (BP(ND)) were used to identify and quantify a [(11)C]-(+)-PHNO mass dose effect at the D(3), and displaceable signal in the cerebellum, as well as providing refined estimates of regional D(3) fractions of [(11)C]-(+)-PHNO BP(ND). The dose of (+)-PHNO required to occupy half of the available D(3) receptors (ED(50)(PHNO,D3)) was estimated as 40ng/kg, and the cerebellum BP(ND) was estimated as 0.40. These findings confirm that [(11)C]-(+)-PHNO human PET studies are in fact routinely performed under non-tracer conditions. This suggests that (+)-PHNO injection masses should be minimised and tightly controlled in order to mitigate the mass dose effect. The specific binding detected in the cerebellum was modest but could have a significant effect, for example on estimates of D(3) potency in drug occupancy studies. A range of methods for the analysis of future [(11)C]-(+)-PHNO data, incorporating models for the effects quantified here, were developed and evaluated. The comparisons and conclusions drawn from these can inform the design and analysis of future PET studies with [(11)C]-(+)-PHNO.

Convergence optimization of parametric MLEM reconstruction for estimation of Patlak plot parameters

In dynamic positron emission tomography data many researchers have attempted to exploit kinetic models within reconstruction such that parametric images are estimated directly from measurements. This work studies a direct parametric maximum likelihood expectation maximization algorithm applied to [(18)F]DOPA data using reference-tissue input function. We use a modified version for direct reconstruction with a gradually descending scheme of subsets (i.e. 18-6-1) initialized with the FBP parametric image for faster convergence and higher accuracy. The results compared with analytic reconstructions show quantitative robustness (i.e. minimal bias) and clinical reproducibility within six human acquisitions in the region of clinical interest. Bland-Altman plots for all the studies showed sufficient quantitative agreement between the direct reconstructed parametric maps and the indirect FBP (--0.035x+0.48E--5).

Spatio-temporal pharmacokinetic model based registration of 4D PET neuroimaging data

In dynamic positron emission tomography (PET) neuroimaging studies, where scan durations often exceed 1h, registration of motion-corrupted dynamic PET images is necessary in order to maintain the integrity of the physiological, pharmacological, or biochemical information derived from the tracer kinetic analysis of the scan. In this work, we incorporate a pharmacokinetic model, which is traditionally used to analyse PET data following any registration, into the registration process itself in order to allow for a groupwise registration of the temporal time frames. The new method is shown to achieve smaller registration errors and improved kinetic parameter estimates on validation data sets when compared with image similarity based registration approaches. When applied to measured clinical data from 10 healthy subjects scanned with [(11)C]-(+)-PHNO (a dopamine D3/D2 receptor tracer), it reduces the intra-class variability on the receptor binding outcome measure, further supporting the improvements in registration accuracy. Our method incorporates a generic tracer kinetic model which makes it applicable to different PET radiotracers to remove motion artefacts and increase the integrity of dynamic PET studies.