Low convergent validity of [11C]raclopride binding in extrastriatal brain regions: A PET study of within-subject correlations with [11C]FLB 457.

Dopamine D2 receptors (D2-R) in extrastriatal brain regions are of high interest for research in a wide range of psychiatric and neurologic disorders. Pharmacological competition studies and test-retest experiments have shown high validity and reliability of the positron emission tomography (PET) radioligand [11C]FLB 457 for D2-R quantification in extrastriatal brain regions. However, this radioligand is not available at most research centers. Instead, the medium affinity radioligand [11C]raclopride, which has been extensively validated for quantification of D2-R in the high-density region striatum, has been applied also in studies on extrastriatal D2-R. Recently, the validity of this approach has been questioned by observations of low occupancy of [11C]raclopride in extrastriatal regions in a pharmacological competition study with quetiapine. Here, we utilise a data set of 16 healthy control subjects examined with both [11C]raclopride and [11C]FLB 457 to assess the correlation in binding potential (BPND) in extrastriatal brain regions. BPND was quantified using the simplified reference tissue model with cerebellum as reference region. The rank order of mean regional BPND values were similar for both radioligands, and corresponded to previously reported data, both post-mortem and using PET. Nevertheless, weak to moderate within-subject correlations were observed between [11C]raclopride and [11C]FLB 457 BPND extrastriatally (Pearson's R: 0.30-0.56), in contrast to very strong correlations between repeated [11C]FLB 457 measurements (Pearson's R: 0.82-0.98). In comparison, correlations between repeated [11C]raclopride measurements were low to moderate (Pearson's R: 0.28-0.75). These results are likely related to low signal to noise ratio of [11C]raclopride in extrastriatal brain regions, and further strengthen the recommendation that extrastriatal D2-R measures obtained with [11C]raclopride should be interpreted with caution.

Distinct inactive conformations of the dopamine D2 and D3 receptors correspond to different extents of inverse agonism.

By analyzing and simulating inactive conformations of the highly homologous dopamine D2 and D3 receptors (D2R and D3R), we find that eticlopride binds D2R in a pose very similar to that in the D3R/eticlopride structure but incompatible with the D2R/risperidone structure. In addition, risperidone occupies a sub-pocket near the Na+ binding site, whereas eticlopride does not. Based on these findings and our experimental results, we propose that the divergent receptor conformations stabilized by Na+-sensitive eticlopride and Na+-insensitive risperidone correspond to different degrees of inverse agonism. Moreover, our simulations reveal that the extracellular loops are highly dynamic, with spontaneous transitions of extracellular loop 2 from the helical conformation in the D2R/risperidone structure to an extended conformation similar to that in the D3R/eticlopride structure. Our results reveal previously unappreciated diversity and dynamics in the inactive conformations of D2R. These findings are critical for rational drug discovery, as limiting a virtual screen to a single conformation will miss relevant ligands.

Almost a third of prescribed drugs work by acting on a group of proteins known as GPCRs (short for G-protein coupled receptors), which help to transmit messages across the cell's outer barrier. The neurotransmitter dopamine, for instance, can act in the brain and body by attaching to dopamine receptors, a sub-family of GPCRs. The binding process changes the three-dimensional structure (or conformation) of the receptor from an inactive to active state, triggering a series of molecular events in the cell. However, GPCRs do not have a single 'on' or 'off' state; they can adopt different active shapes depending on the activating molecule they bind to, and this influences the type of molecular cascade that will take place in the cell. Some evidence also shows that classes of GPCRs can have different inactive structures; whether this is also the case for the dopamine D₂ and D₃ receptors remained unclear. Mapping out inactive conformations of receptors is important for drug discovery, as compounds called antagonists can bind to inactive receptors and interfere with their activation. Lane et al. proposed that different types of antagonists could prefer specific types of inactive conformations of the dopamine D₂ and D₃ receptors. Based on the structures of these two receptors, the conformations of D₂ bound with the drugs risperidone and eticlopride (two dopamine antagonists) were simulated and compared. The results show that the inactive conformations of D₂ were very different when it was bound to eticlopride as opposed to risperidone. In addition D₂ and D₃ showed a very similar conformation when attached to eticlopride. The two drugs also bound to the inactive receptors in overlapping but different locations. These computational findings, together with experimental validations, suggest that D₂ and D₃ exist in several inactive states that only allow the binding of specific drugs; these states could also reflect different degrees of inactivation. Overall, the work by Lane et al. contributes to a more refined understanding of the complex conformations of GPCRs, which could be helpful to screen and develop better drugs.

Multi-target dopamine D3 receptor modulators: Actionable knowledge for drug design from molecular dynamics and machine learning.

Local changes in the structure of G-protein coupled receptors (GPCR) binders largely affect their pharmacological profile. While the sought efficacy can be empirically obtained by introducing local modifications, the underlining structural explanation can remain elusive. Here, molecular dynamics (MD) simulations of the eticlopride-bound inactive state of the Dopamine D3 Receptor (D3DR) have been clustered using a machine learning-based approach in the attempt to rationalize the efficacy change in four congeneric modulators. Accumulating extended MD trajectories of receptor-ligand complexes, we observed how the increase in ligand flexibility progressively destabilized the crystal structure of the inactivated receptor. To prospectively validate this model, a partial agonist was rationally designed based on structural insights and computational modeling, and eventually synthesized and tested. Results turned out to be in line with the predictions. This case study suggests that the investigation of ligand flexibility in the framework of extended MD simulations can assist and inform drug design strategies, highlighting its potential role as a powerful in silico counterpart to functional assays.

Inhibition of Classical and Alternative Modes of Respiration in Candida albicans Leads to Cell Wall Remodeling and Increased Macrophage Recognition.

The human fungal pathogen Candida albicans requires respiratory function for normal growth, morphogenesis, and virulence. Mitochondria therefore represent an enticing target for the development of new antifungal strategies. This possibility is bolstered by the presence of characteristics specific to fungi. However, respiration in C. albicans, as in many fungal organisms, is facilitated by redundant electron transport mechanisms, making direct inhibition a challenge. In addition, many chemicals known to target the electron transport chain are highly toxic. Here we made use of chemicals with low toxicity to efficiently inhibit respiration in C. albicans We found that use of the nitric oxide donor sodium nitroprusside (SNP) and of the alternative oxidase inhibitor salicylhydroxamic acid (SHAM) prevents respiration and leads to a loss of viability and to cell wall rearrangements that increase the rate of uptake by macrophages in vitro and in vivo We propose that treatment with SNP plus SHAM (SNP+SHAM) leads to transcriptional changes that drive cell wall rearrangement but which also prime cells to activate the transition to hyphal growth. In line with this, we found that pretreatment of C. albicans with SNP+SHAM led to an increase in virulence. Our data reveal strong links between respiration, cell wall remodeling, and activation of virulence factors. Our findings demonstrate that respiration in C. albicans can be efficiently inhibited with chemicals that are not damaging to the mammalian host but that we need to develop a deeper understanding of the roles of mitochondria in cellular signaling if they are to be developed successfully as a target for new antifungals.IMPORTANCE Current approaches to tackling fungal infections are limited, and new targets must be identified to protect against the emergence of resistant strains. We investigated the potential of targeting mitochondria, which are organelles required for energy production, growth, and virulence, in the human fungal pathogen Candida albicans Our findings suggest that mitochondria can be targeted using drugs that can be tolerated by humans and that this treatment enhances their recognition by immune cells. However, release of C. albicans cells from respiratory inhibition appears to activate a stress response that increases the levels of traits associated with virulence. Our results make it clear that mitochondria represent a valid target for the development of antifungal strategies but that we must determine the mechanisms by which they regulate stress signaling and virulence ahead of successful therapeutic advance.

Kallikrein 5 inhibitors identified through structure based drug design in search for a treatment for Netherton Syndrome.

Netherton syndrome (NS) is a rare and debilitating severe autosomal recessive genetic skin disease with high mortality rates particularly in neonates. NS is caused by loss-of-function SPINK5 mutations leading to unregulated kallikrein 5 (KLK5) and kallikrein 7 (KLK7) activity. Furthermore, KLK5 inhibition has been proposed as a potential therapeutic treatment for NS. Identification of potent and selective KLK5 inhibitors would enable further exploration of the disease biology and could ultimately lead to a treatment for NS. This publication describes how fragmentation of known trypsin-like serine protease (TLSP) inhibitors resulted in the identification of a series of phenolic amidine-based KLK5 inhibitors 1. X-ray crystallography was used to find alternatives to the phenol interaction leading to identification of carbonyl analogues such as lactam 13 and benzimidazole 15. These reversible inhibitors, with selectivity over KLK1 (10-100 fold), provided novel starting points for the guided growth towards suitable tool molecules for the exploration of KLK5 biology.

Dopamine D3 receptor antagonist reveals a cryptic pocket in aminergic GPCRs.

The recent increase in the number of X-ray crystal structures of G-protein coupled receptors (GPCRs) has been enabling for structure-based drug design (SBDD) efforts. These structures have revealed that GPCRs are highly dynamic macromolecules whose function is dependent on their intrinsic flexibility. Unfortunately, the use of static structures to understand ligand binding can potentially be misleading, especially in systems with an inherently high degree of conformational flexibility. Here, we show that docking a set of dopamine D3 receptor compounds into the existing eticlopride-bound dopamine D3 receptor (D3R) X-ray crystal structure resulted in poses that were not consistent with results obtained from site-directed mutagenesis experiments. We overcame the limitations of static docking by using large-scale high-throughput molecular dynamics (MD) simulations and Markov state models (MSMs) to determine an alternative pose consistent with the mutation data. The new pose maintains critical interactions observed in the D3R/eticlopride X-ray crystal structure and suggests that a cryptic pocket forms due to the shift of a highly conserved residue, F6.52. Our study highlights the importance of GPCR dynamics to understand ligand binding and provides new opportunities for drug discovery.

The Effect of Treatment with Guanfacine, an Alpha2 Adrenergic Agonist, on Dopaminergic Tone in Tobacco Smokers: An [11C]FLB457 PET Study.

Guanfacine, a noradrenergic alpha2a agonist, reduced tobacco smoking in a 4-week trial and in animal models has been shown to reduce cortical dopamine release, which is critically involved in the reinforcing effect of tobacco smoking. We measured amphetamine-induced extrastriatal dopamine release before and after treatment with guanfacine with [11C]FLB457, a dopamine D2/D3 receptor radiotracer, and positron emission tomography (PET). Sixteen tobacco smokers had one set of [11C]FLB457 PET scans on the same day, one before and one at 2.5-3 h after amphetamine (0.4-0.5 mg/kg, PO). A subset (n=12) then underwent guanfacine treatment (3 mg/day for 3 weeks) and the set of scans were repeated. [11C]FLB457-binding potential (BPND) was measured pre- and post amphetamine in extrastriatal brain regions. The fractional change in BPND after vs before amphetamine (Δ BPND) is an indirect measure of DA release and was compared between the untreated and guanfacine-treated conditions. Guanfacine treatment attenuated amphetamine-induced DA release; however, the change was due to a global 8% decrease in baseline BPND from the untreated to the guanfacine-treated condition. Chronic guanfacine treatment reduced [11C]FLB457 BPND in tobacco smokers, suggesting an increase in dopaminergic tone. Guanfacine-induced normalization of dopamine signaling may be an important mesocortical mechanism contributing to its ability to aid in tobacco smoking cessation.

Effects of point spread function-based image reconstruction on neuroreceptor binding in positron emission tomography study with [(11)C]FLB 457.

The ordered subset expectation maximization with a point spread function (OSEM-PSF) was developed to improve the spatial resolution of reconstructed positron emission tomography (PET) images and has been reported to improve the contrast of hot spots in PET studies for oncology. However, in neuroreceptor imaging, the regional radioactivity concentration changes dynamically during the scan, and the effects of the PSF may differ among various radioligands or quantification methods. In this study, we investigated the effects of the PSF on quantification in PET studies with [(11)C]FLB 457 of dopamine D2 receptors, using both phantom and human data acquired by the Siemens Biograph 16 imaging platform. In the phantom studies, we evaluated the hot contrast recovery coefficient (HCRC) for variously sized hot spheres and the linearity between the measured and true radioactivities in OSEM-PSF images. Next, in the human studies with [(11)C]FLB 457, radioactivity concentrations and binding potentials for the cerebral cortex and thalamus were compared between images reconstructed with and without PSF. In the phantom studies, the OSEM-PSF images showed a better HCRC compared to images without PSF, and they showed a good linear correlation with true radioactivity. In the human studies, the radioactivity concentration increased especially in small regions with high accumulation of [(11)C]FLB 457 when the PSF was included. However, little difference in the binding potentials was observed for the target regions between both types of reconstructed images. In conclusion, PSF-based reconstruction reduced the spill-over phenomena in small hot regions; however, it caused no increase in the binding potentials in the [(11)C]FLB 457 studies.

Reference region modeling approaches for amphetamine challenge studies with [11C]FLB 457 and PET.

Detecting fluctuations in synaptic dopamine levels in extrastriatal brain regions with [(11)C]FLB 457 and positron emission tomography (PET) is a valuable tool for studying dopaminergic dysfunction in psychiatric disorders. The evaluation of reference region modeling approaches would eliminate the need to obtain arterial input function data. Our goal was to explore the use of reference region models to estimate amphetamine-induced changes in [(11)C]FLB 457 dopamine D2/D3 binding. Six healthy tobacco smokers were imaged with [(11)C]FLB 457 at baseline and at 3 hours after amphetamine (0.4 to 0.5 mg/kg, per os) administration. Simplified reference tissue models, SRTM and SRTM2, were evaluated against the 2-tissue compartmental model (2TC) to estimate [(11)C]FLB 457 binding in extrastriatal regions of interest (ROIs), using the cerebellum as a reference region. No changes in distribution volume were observed in the cerebellum between scan conditions. SRTM and SRTM2 underestimated binding, compared with 2TC, in ROIs by 26% and 9%, respectively, with consistent bias between the baseline and postamphetamine scans. Postamphetamine, [(11)C]FLB 457 binding significantly decreased across several brain regions as measured with SRTM and SRTM2; no significant change was detected with 2TC. These data support the sensitivity of [(11)C]FLB 457 for measuring amphetamine-induced dopamine release in extrastriatal regions with SRTM and SRTM2.

Reproducibility of post-amphetamine [11C]FLB 457 binding to cortical D2/3 receptors.

In a recent positron emission tomography (PET) study, we demonstrated the ability to measure amphetamine-induced dopamine (DA) release in the human cortex with the relatively high affinity dopamine D2/3 radioligand [(11)C]FLB 457. Herein we report on reproducibility and reliability of [(11)C]FLB 457 binding potential relative to non-displaceable uptake (BP(ND)) following an acute amphetamine challenge. Ten healthy human subjects were studied twice with [(11)C]FLB 457 following an acute amphetamine (oral, 0.5 mg kg(-1) dose) challenge on two-separate days approximately one week apart. D2/3 receptor binding parameters were estimated using a two-tissue compartment kinetic analysis in the cortical regions of interest and cerebellum (reference region). The test-retest variability and intraclass correlation coefficient were assessed for distribution volume (V(T)), binding potential relative to plasma concentration (BP(P)), and BP(ND) of [(11)C]FLB 457. The test-retest variability of [(11)C]FLB 457 V(T), BP(P) and BP(ND) were ≤ 17%, 22% and 11% respectively. These results, which are consistent with the published test-retest variability for this ligand measured under baseline conditions demonstrate that the post-amphetamine [(11)C]FLB 457 BP(ND) is reproducible. These data further support the use [(11)C]FLB 457 and amphetamine to characterize cortical dopamine transmission in neuropsychiatric disorders.

Striatal and extrastriatal dopamine D2 receptor occupancy by a novel antipsychotic, blonanserin: a PET study with [11C]raclopride and [11C]FLB 457 in schizophrenia.

Blonanserin is a novel antipsychotic with high affinities for dopamine D(2) and 5-HT(2A) receptors, and it was recently approved for the treatment of schizophrenia in Japan and Korea. Although double-blind clinical trials have demonstrated that blonanserin has equal efficacy to risperidone, and with a better profile especially with respect to prolactin elevation, its profile of in vivo receptor binding has not been investigated in patients with schizophrenia. Using positron emission tomography (PET), we measured striatal and extrastriatal dopamine D(2) receptor occupancy by blonanserin in 15 patients with schizophrenia treated with fixed doses of blonanserin (ie, 8, 16, and 24 mg/d) for at least 4 weeks before PET scans, and in 15 healthy volunteers. Two PET scans, 1 with [(11)C]raclopride for the striatum and 1 with [(11)C]FLB 457 for the temporal cortex and pituitary, were performed on the same day. Striatal dopamine D(2) receptor occupancy by blonanserin was 60.8% (3.0%) [mean (SD)] at 8 mg, 73.4% (4.9%) at 16 mg, and 79.7% (2.3%) at 24 mg. The brain/plasma concentration ratio calculated from D(2) receptor occupancy in the temporal cortex and pituitary was 3.38, indicating good blood-brain barrier permeability. This was the first study to show clinical daily dose amounts of blonanserin occupying dopamine D(2) receptors in patients with schizophrenia. The clinical implications obtained in this study were the optimal therapeutic dose range of 12.9 to 22.1 mg/d of blonanserin required for 70% to 80% dopamine D(2) receptor occupancy in the striatum, and the good blood-brain barrier permeability that suggested a relatively lower risk of hyperprolactinemia.

Involvement of alternative oxidase in the regulation of growth, development, and resistance to oxidative stress of Sclerotinia sclerotiorum.

Sclerotinia sclerotiorum is a cosmopolitan, filamentous, fungal pathogen that can cause serious disease in many kinds of crops. Alternative oxidase is the terminal oxidase of the alternative mitochondrial respiratory pathway in fungi and higher plants. We report the presence of this alternative pathway respiration and demonstrate its expression in two isolates of S. sclerotiorum under unstressed, normal culture conditions. Application of salicylhydroxamic acid, a specific inhibitor of alternative oxidase, severely inhibited the mycelial growth of S. sclerotiorum both on potato dextrose agar plates and in liquid culture media. Inhibition of alternative oxidase could influence the growth pattern of S. sclerotiorum, as salicylhydroxamic acid treatment induced obvious aerial mycelia growing on potato dextrose agar plates. Under the treatment with salicylhydroxamic acid, S. sclerotiorum formed sclerotia much more slowly than the control. Treatment with hydrogen peroxide in millimolar concentrations greatly decreased the growth rate of mycelia and delayed the formation of sclerotia in both tested S. sclerotiorum isolates. As well, this treatment obviously increased their alternative pathway respiration and the levels of both mRNA and protein of the alternative oxidase. These results indicate that alternative oxidase is involved in the regulation of growth, development, and resistance to oxidative stress of S. sclerotiorum.

Striatal and extrastriatal dopamine D2 receptor occupancy by the partial agonist antipsychotic drug aripiprazole in the human brain: a positron emission tomography study with [¹¹C]raclopride and [¹¹C]FLB457.

RATIONALE: Second-generation antipsychotics demonstrate clinical efficacy with fewer extrapyramidal side effects compared with first-generation antipsychotics. One of the proposed explanations is the hypothesis of preferential extrastriatal dopamine D2 receptor occupancy (limbic selectivity) by antipsychotics. In the present study, we focused on aripiprazole, which has a unique pharmacological profile with partial agonism at dopamine D2 receptors and the minimal risk of extrapyramidal side effects. Previous positron emission tomography (PET) studies using high-affinity radioligands for dopamine D2 receptors have reported inconsistent results regarding regional differences of dopamine D2 receptor occupancy by aripiprazole. OBJECTIVE: To test the hypothesis of preferential binding to extrastriatal dopamine D2 receptors by aripiprazole, we investigated its regional dopamine D2 receptor occupancies in healthy young subjects. MATERIALS AND METHODS: Using PET and two radioligands with different affinities for dopamine D2 receptors, [¹¹C]raclopride and [¹¹C]FLB457, striatal and extrastriatal dopamine D2 receptor bindings at baseline and after oral administration of 6 mg aripiprazole were measured in 11 male healthy subjects. RESULTS: Our data showed that dopamine D2 receptor occupancies in the striatum measured with [¹¹C]raclopride were 70.1% and 74.1%, with the corresponding values for the extrastriatal regions measured with [¹¹C]FLB457 ranging from 46.6% to 58.4%. CONCLUSIONS: In the present study, preferential extrastriatal dopamine D2 receptor occupancy by aripiprazole was not observed. Our data suggest partial agonism at dopamine D2 receptors is the most likely explanation for the minimal risk of extrapyramidal side effects in the treatment by aripiprazole.

Chronic quercetin feeding decreases plasma concentrations of salicylamide phase II metabolites in pigs following oral administration.

We investigated acute effects and effects after chronic intake of the orally administered flavonol quercetin on pharmacokinetics of salicylamide metabolites (SAM) after oral administration of salicylamide in pigs. Salicylamide (8 mg/kg body weight) was orally administered to seven pigs either without or with quercetin (10 mg/kg body weight). Additionally, salicylamide was administered to five pigs that had received a diet supplemented with the flavonol for 1 week. Daily quercetin intake was 10 mg/kg in these animals. Co-ingestion of quercetin with the drug did not alter area under the concentration-time curve (AUC(0→∞)), time to achieve maximum plasma concentration (t(max)), mean residence time (MRT) or half-life (t(1/2)) of SAM. However, maximum plasma concentration (c(max)) of SAM was lower when quercetin was administered concomitantly. After quercetin pre-treatment for 1 week AUC(0→∞), t(1/2) and MRT of SAM were decreased, while other parameters investigated were not affected. Co-ingestions and dietary pre-treatment with quercetin influenced SAM metabolism after oral salicylamide intake. But effects seen after acute concomitant intake are rather explained by induced salicylamide excretion from the intestinal mucosa, whereas quercetin pre-treatment seemed to induce hepatic enzymes involved in phase-II metabolism and thereby enhanced elimination of SAM.

Promysalin, a salicylate-containing Pseudomonas putida antibiotic, promotes surface colonization and selectively targets other Pseudomonas.

Under control of the Gac regulatory system, Pseudomonas putida RW10S1 produces promysalin to promote its own swarming and biofilm formation, and to selectively inhibit many other pseudomonads, including the opportunistic pathogen Pseudomonas aeruginosa. This amphipathic antibiotic is composed of salicylic acid and 2,8-dihydroxymyristamide bridged by a unique 2-pyrroline-5-carboxyl moiety. In addition to enzymes for salicylic acid synthesis and activation, the biosynthetic gene cluster encodes divergent type II fatty acid biosynthesis components, unusual fatty acid-tailoring enzymes (two Rieske-type oxygenases and an amidotransferase), an enzyme resembling a proline-loading module of nonribosomal peptide synthetases, and the first prokaryotic member of the BAHD family of plant acyltransferases. Identification of biosynthetic intermediates enabled to propose a pathway for synthesis of this bacterial colonization factor.

Positron emission tomography imaging of dopamine D2/3 receptors in the human cortex with [¹¹C]FLB 457: reproducibility studies.

In a recent PET study, we demonstrated the ability to measure amphetamine-induced DA release in the human cortex with the relatively high affinity dopamine D2/3 radioligand [¹¹C]FLB 457 (Narendran et al., [2009] Synapse 63:447-461). The aim of this study was to evaluate the reproducibility and reliability of [¹¹C]FLB 457 in the same imaging paradigm we used to measure amphetamine-induced DA transmission. Six healthy human subjects (three males/three females)were studied twice with [¹¹C]FLB 457, once at baseline and again 3 h following the end of the baseline scan. D2/3 receptor binding parameters were estimated using a two-tissue compartment kinetic analysis in the cortical regions of interest and cerebellum (reference region). The test-retest variability and intraclass correlation coefficient were assessed for distribution volume (VT), binding potential relative to plasma concentration (BP(P)), and binding potential relative to non-displaceable uptake (BP(ND)) of [¹¹C]FLB 457. The test-retest variability of [¹¹C]FLB 457 VT, BPP, and BP(ND) were ≤15%, consistent with the published test-retest variability for this ligand in other brain regions (Sudo et al., [2001] Nucl Med Commun 22:1215-1221; Vilkman et al., [2000] Eur J Nucl Med 27:1666-1673). In addition, no significant decrease in [¹¹C]FLB457 BP(ND) was observed in the second scan compared to the first one. This suggests that the contribution of carryover mass of [¹¹C]FLB 457 to the measured reduction in[¹¹C]FLB 457 BP(ND) following amphetamine was relatively low. These data support the further validation of [¹¹C]FLB 457 as a tool to measure amphetamine-induced dopamine release in the human cortex.

Positron emission tomography inter-scanner differences in dopamine D2 receptor binding measured with [¹¹C]FLB457.

OBJECTIVE: It is well known that the positron emission tomography (PET) system is subject to inter-scanner differences of regional radioactivity distribution. In the present study, the effect of inter-scanner difference of regional radioactivity on estimated binding potential (BP(ND)) of [¹¹C]FLB457 using the simplified reference tissue model (SRTM) was investigated. METHODS: Each of the 11 subjects was given two PET scans using [¹¹C]FLB457, one each with both SET-3000 GCT/X (Shimadzu) and with ECAT EXACT HR+ (Siemens/CTI). In order to assess regional differences between the two scanners, estimated BP(ND) values in six volumes of interest (VOIs) by SRTM method were compared in both individual PET space and anatomical template space after anatomical normalization. Statistical voxel-by-voxel paired t test of BP(ND) images between SET-3000 GCT/X and ECAT EXACT HR+ was also performed. RESULTS: Shapes of time-activity curves of the two PET scanners were slightly different in each VOI, with estimated BP(ND) values from ECAT EXACT HR+ appearing greater in the cerebral cortical regions and thalamus than that of SET-3000 GCT/X in both individual PET space and anatomical template space after anatomical normalization. Statistical voxel-by-voxel analysis showed similar tendency to BP(ND) value estimation, with greater BP(ND) values from ECAT EXACT HR+ than from SET-3000 GCT/X. CONCLUSIONS: We demonstrated the inter-scanner differences in dopamine D(2) receptor binding measured with [¹¹C]FLB457. In particular, statistically significant differences of BP(ND) in certain regions were observed between two PET scanners, despite the subject groups being the same. Our results suggest that we reconsider the effect of the scanner model on the measurement of receptor binding.

No effect of dopamine depletion on the binding of the high-affinity D 2/3 radiotracer [11C]FLB 457 in the human cortex.

The use of PET and SPECT endogenous competition-binding techniques has contributed to the understanding of the role of dopamine (DA) in several neuropsychiatric disorders. An important limitation of these imaging studies is the fact that measurements of changes in synaptic DA have been restricted to the striatum. The ligands previously used, such as [(11)C]raclopride and [(123)I]IBZM, do not provide sufficient signal-to-noise ratio to quantify D(2) receptors in extrastriatal areas, such as cortex, where the concentration of D(2) receptors is much lower than that in the striatum. Recently, we published a comparison study of the ability of two high-affinity DA D(2) radioligands [(11)C]FLB 457 and [(11)C]fallypride to measure amphetamine-induced changes in DA transmission in the human cortex. Our findings support the use of [(11)C]FLB 457 to measure changes in cortical synaptic DA induced by amphetamine. The goal of this study is to examine the effects of DA depletion with α-methyl-para-tyrosine (α-MPT) on [(11)C]FLB 457 binding in the cortex. Six healthy volunteers underwent two PET scans, first under control conditions and subsequently after DA depletion. The simplified reference tissue model as well as kinetic modeling with an arterial input function was used to derive the binding potential (BP(ND)) in seven cortical regions. We found no effect of DA depletion with α-MPT on [(11)C]FLB 457 binding in any of the regions examined. In contrast to the measurement of DA release, the combination of low D(2) receptor density and low basal DA levels in the cortex greatly reduce the power to detect alterations in [(11)C]FLB 457 binding secondary to DA depletion.

The value of the dopamine D2/3 receptor ligand 18F-desmethoxyfallypride for the differentiation of idiopathic and nonidiopathic parkinsonian syndromes.

UNLABELLED: We evaluated the utility of the selective dopamine D(2/3) receptor ligand (18)F-desmethoxyfallypride ((18)F-DMFP) for the differential diagnosis of patients with idiopathic parkinsonian syndrome (IPS) and nonidiopathic parkinsonian syndrome (non-IPS). On the basis of the superior sensitivity of PET, we hypothesized that (18)F-DMFP should have properties for the differential diagnosis of these syndromes superior to what has been reported for the more conventional SPECT procedures. METHODS: A series of 81 patients with parkinsonism (26 women, 55 men; mean age +/- SD, 68 +/- 11 y) were included in this retrospective analysis. A 30-min (18)F-DMFP PET recording was acquired starting 1 h after injection of the tracer (180-200 MBq, intravenously). The specific binding (SB) in divisions of the striatum was calculated relative to the occipital cortex using an observer-independent semiautomatic volume-of-interest-based technique. The optimal SB threshold was defined by means of receiver-operating-characteristic analysis, which was also used for the evaluation of the diagnostic performance of SB, ratios between striatal subregions, and absolute asymmetries in SB. RESULTS: Significant differences (P < 0.001) were found in striatal SB between IPS and non-IPS, most notably in the posterior putamen, for which the diagnostic power for discrimination of IPS and non-IPS was the highest (sensitivity, 87%; specificity, 96%; and accuracy, 91%). A further gain of diagnostic power (sensitivity, 92%; specificity, 96%; and accuracy, 94%) was obtained through discriminant analysis combining 3 parameters: SB of the posterior putamen, the posterior-to-anterior putamen ratio, and the posterior putamen-to-caudate ratio. CONCLUSION: (18)F-DMFP PET is useful for the differential diagnosis of IPS and non-IPS in patients with parkinsonism. The findings are consistent with relative sparing of D(2/3) receptors in the dopamine-denervated putamen of IPS patients, in contrast to a more substantial loss of striatal dopamine receptors in non-IPS patients. The PET procedure for this differential diagnosis was superior to the reported experience with (123)I-iodobenzamide SPECT.

PET studies of D2-receptor binding in striatal and extrastriatal brain regions: Biochemical support in vivo for separate dopaminergic systems in humans.

Most molecular imaging studies of the dopamine (DA) system performed to date have focused on the striatum, a region receiving dense dopaminergic innervation. In clinical research on the DA D2-receptor, striatal binding has often been regarded as an index of global DA function, based on the underlying assumption of common regulatory mechanisms for receptor expression across brain regions. Recent data has challenged this view, suggesting differences in genetic regulation between striatal and extrastriatal brain regions. The relationship between binding levels in brain regions has, however, not been directly examined in the same sample. In this study, we searched for interregional correlations between DA D2-receptor availability as determined with Positron Emission Tomography in 16 control subjects. The radioligands [11C]raclopride and [11C]FLB 457 were used for measurements of D2-receptor binding in striatal and extrastriatal regions, respectively. No correlation was observed between D2-receptor availability in striatum and any of the extrastriatal regions, as assessed using both region of interest- and voxel-based analyses. Instead, the pattern of correlations was consistent with the model of separate dopaminergic systems as has been originally observed in rodents. These preliminary results encourage approaches searching for individual patterns of receptor binding across the whole brain volume in clinical studies on the dopamine system.