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Purpose: Elevated dopamine synthesis capacity is part of the pathophysiology of schizophrenia thought to underlie psychosis. Drugs that reduce this phenomenon could thus be potential treatments for these disorders. In this study, we evaluated the ability of the trace amine-associated receptor 1 (TAAR1) partial agonist ralmitaront to reduce presynaptic dopamine synthesis capacity. Procedures: Ralmitaront (3 mg/kg, i.p.), a TAAR1 partial agonist, was evaluated using [18F]DOPA PET for its ability to modulate presynaptic dopamine synthesis capacity in naïve mice as well as mice in an induced hyperdopaminergic state following acute cocaine administration (20 mg/kg, i.p.). Results: Cocaine treatment on its own did not induce elevated dopamine synthesis capacity when compared to the control group. Pretreatment with ralmitaront significantly reduced dopamine synthesis capacity when given either alone (44%) or in combination with the psychostimulant cocaine (50%) when compared to the control group. Conclusions: The TAAR1 agonist ralmitaront reduces striatal dopamine synthesis capacity, indexed as KiMod, both in naïve animals and when given prior to acute cocaine. This indicates the potential of TAAR1 agonism to address disorders characterized by striatal hyperdopaminergia. [ABSTRACT FROM AUTHOR]

Positron emission tomography (PET) using the radiotracer [18F]-FDOPA provides a tool for studying brain dopamine synthesis capacity in animals and humans. We have previously standardised a micro-PET methodology in mice by intravenously administering [18F]-FDOPA via jugular vein cannulation and assessment of striatal dopamine synthesis capacity, indexed as the influx rate constant K i Mod of [18F]-FDOPA, using an extended graphical Patlak analysis with the cerebellum as a reference region. This enables a direct comparison between preclinical and clinical output values. However, chronic intravenous catheters are technically difficult to maintain for longitudinal studies. Hence, in this study, intraperitoneal administration of [18F]-FDOPA was evaluated as a less-invasive alternative that facilitates longitudinal imaging. Our experiments comprised the following assessments: (i) comparison of [18F]-FDOPA uptake between intravenous and intraperitoneal radiotracer administration and optimisation of the time window used for extended Patlak analysis, (ii) comparison of K i Mod in a within-subject design of both administration routes, (iii) test-retest evaluation of K i Mod in a within-subject design of intraperitoneal radiotracer administration, and (iv) validation of K i Mod estimates by comparing the two administration routes in a mouse model of hyperdopaminergia induced by subchronic ketamine. Our results demonstrate that intraperitoneal [18F]-FDOPA administration resulted in good brain uptake, with no significant effect of administration route on K i Mod estimates (intraperitoneal: 0.024 ± 0.0047 min − 1 , intravenous: 0.022 ± 0.0041 min − 1 , p = 0.42) and similar coefficient of variation (intraperitoneal: 19.6%; intravenous: 18.4%). The technique had a moderate test-retest validity (intraclass correlation coefficient ICC = 0.52 , N = 6) and thus supports longitudinal studies. Following subchronic ketamine administration, elevated K i Mod as compared to control condition was measured with a large effect size for both methods (intraperitoneal: Cohen's d = 1.3 ; intravenous: Cohen's d = 0.9), providing further evidence that ketamine has lasting effects on the dopamine system, which could contribute to its therapeutic actions and/or abuse liability. [ABSTRACT FROM AUTHOR]

Background: Mutations in the epidermal growth factor receptor (EGFR) kinase domain are common in non-small cell lung cancer. Conventional tyrosine kinase inhibitors target the mutation site in the ATP binding pocket, thereby inhibiting the receptor's function. However, subsequent treatment resistance mutations in the ATP binding site are common. The EGFR allosteric inhibitor, EAI045, is proposed to have an alternative mechanism of action, disrupting receptor signaling independent of the ATP-binding site. The antibody cetuximab is hypothesized to increase the number of accessible allosteric pockets for EAI045, thus increasing the potency of the inhibitor. This work aimed to gain further knowledge on pharmacokinetics, the EGFR mutation-targeting potential, and the influence of cetuximab on the uptake by radiolabeling EAI045 with carbon-11 and tritium. Results: 2-(5-fluoro-2-hydroxyphenyl)-2-((2-iodobenzyl)amino)-N-(thiazol-2-yl)acetamide and 2-(5-fluoro-2-hydroxyphenyl)-N-(5-iodothiazol-2-yl)-2-(1-oxoisoindolin-2-yl)acetamide were synthesized as precursors for the carbon-11 and tritium labeling of EAI045, respectively. [11C]EAI045 was synthesized using [11C]CO in a palladium-catalyzed ring closure in a 10 ± 1% radiochemical yield (decay corrected to end of [11C]CO2 production), > 97% radiochemical purity and 26 ± 1 GBq/µmol molar activity (determined at end of synthesis) in 51 min. [3H]EAI045 was synthesized by a tritium-halogen exchange in a 0.2% radiochemical yield, 98% radiochemical purity, and 763 kBq/nmol molar activity. The ability of [11C]EAI045 to differentiate between L858R/T790M mutated EGFR expressing H1975 xenografts and wild-type EGFR expressing A549 xenografts was evaluated in female nu/nu mice. The uptake was statistically significantly higher in H1975 xenografts compared to A549 xenografts (0.45 ± 0.07%ID/g vs. 0.31 ± 0.10%ID/g, P = 0.0166). The synergy in inhibition between EAI045 and cetuximab was evaluated in vivo and in vitro. While there was some indication that cetuximab influenced the uptake of [3H]EAI045 in vitro, this could not be confirmed in vivo when tumor-bearing mice were administered cetuximab (0.5 mg), 24 h prior to injection of [11C]EAI045. Conclusions: EAI045 was successfully labeled with tritium and carbon-11, and the in vivo results indicated [11C]EAI045 may be able to distinguish between mutated and non-mutated EGFR in non-small cell lung cancer mouse models. Cetuximab was hypothesized to increase EAI045 uptake; however, no significant effect was observed on the uptake of [11C]EAI045 in vivo or [3H]EAI045 in vitro in H1975 xenografts and cells. [ABSTRACT FROM AUTHOR]

Positron emission tomography (PET) using the radiotracer [18F]-FDOPA provides a tool for studying brain dopamine synthesis capacity in animals and humans. We have previously standardised a micro-PET methodology in mice by intravenously administering [18F]-FDOPA via jugular vein cannulation and assessment of striatal dopamine synthesis capacity, indexed as the influx rate constant K i Mod of [18F]-FDOPA, using an extended graphical Patlak analysis with the cerebellum as a reference region. This enables a direct comparison between preclinical and clinical output values. However, chronic intravenous catheters are technically difficult to maintain for longitudinal studies. Hence, in this study, intraperitoneal administration of [18F]-FDOPA was evaluated as a less-invasive alternative that facilitates longitudinal imaging. Our experiments comprised the following assessments: (i) comparison of [18F]-FDOPA uptake between intravenous and intraperitoneal radiotracer administration and optimisation of the time window used for extended Patlak analysis, (ii) comparison of K i Mod in a within-subject design of both administration routes, (iii) test-retest evaluation of K i Mod in a within-subject design of intraperitoneal radiotracer administration, and (iv) validation of K i Mod estimates by comparing the two administration routes in a mouse model of hyperdopaminergia induced by subchronic ketamine. Our results demonstrate that intraperitoneal [18F]-FDOPA administration resulted in good brain uptake, with no significant effect of administration route on K i Mod estimates (intraperitoneal: 0.024 ± 0.0047 min − 1 , intravenous: 0.022 ± 0.0041 min − 1 , p = 0.42) and similar coefficient of variation (intraperitoneal: 19.6%; intravenous: 18.4%). The technique had a moderate test-retest validity (intraclass correlation coefficient ICC = 0.52 , N = 6) and thus supports longitudinal studies. Following subchronic ketamine administration, elevated K i Mod as compared to control condition was measured with a large effect size for both methods (intraperitoneal: Cohen's d = 1.3 ; intravenous: Cohen's d = 0.9), providing further evidence that ketamine has lasting effects on the dopamine system, which could contribute to its therapeutic actions and/or abuse liability. [ABSTRACT FROM AUTHOR]

Cocaine is a recreational drug of abuse that binds to the dopamine transporter, preventing reuptake of dopamine into pre-synaptic terminals. The increased presence of synaptic dopamine results in stimulation of both pre- and post-synaptic dopamine receptors, considered an important mechanism by which cocaine elicits its reinforcing properties. However, the effects of acute cocaine administration on pre-synaptic dopamine function remain unclear. Non-invasive imaging techniques such as positron emission tomography have revealed impaired pre-synaptic dopamine function in chronic cocaine users. Similar impairments have been seen in animal studies, with microdialysis experiments indicating decreased basal dopamine release. Here we use micro positron emission tomography imaging techniques in mice to measure dopamine synthesis capacity and determine the effect of acute cocaine administration of pre-synaptic dopamine function. We show that a dose of 20 mg/kg cocaine is sufficient to elicit hyperlocomotor activity, peaking 15-20 min post treatment ( p < 0.001). However, dopamine synthesis capacity in the striatum was not significantly altered by acute cocaine treatment ( [ABSTRACT FROM AUTHOR]

The environmental day-night cycle provides the principal synchronizing signal for behavioral activity in most mammals. Light information is relayed to the master circadian pacemaker, the suprachiasmatic nucleus (SCN), via synaptic transmission from the retina directly to the SCN, where a predominately glutamate-driven cellular signaling pathway is able to reset biochemical, physiological, and behavioral activities. In the present study, we aimed to decipher the key roles played by protein kinase C (PKC) in regulating light-induced behavioral resetting under both a temporal and intensity-dependent manner; in addition, we also investigate PKC contributions to advancing and delaying re-entrainment paradigms. Our findings show that during the early night PKC acts in a temporal manner, where PKC inhibition selectively attenuates light-induced behavioral resetting in response to subsaturating and saturating light intensities. Declines in light response were also evident upon PKC inhibition during the late night, but restricted to bright light stimuli. The positive regulatory actions of PKC were further demonstrated in response to an 8-h delayed re-entrainment paradigm where inhibition of PKC resulted in slower re-entrainment. Further, analysis of both classic and novel PKC isozymes present within the SCN showed significant circadian variation in the mRNA expression of PKCα, indicating possible isozyme-specific mediators in photic signaling. Our data provide evidence of a PKC contribution to both acute light-induced clock resetting, which is intensity and time of day dependent, and a functional role in circadian photoentrainment. (Author correspondence: ) [ABSTRACT FROM AUTHOR]

This paper describes the preclinical validation of the radioligand [11C]ORM-13070 and its tritiated analog for addressing selectivity and occupancy of the selective alpha-2C adrenergic receptor (α2CR) antagonist BAY 292 in the cynomolgus brain. BAY 292 is a novel drug candidate being developed for the treatment of obstructive sleep apnea (OSA) via binding to central α2CR. In vitro autoradiography studies with sections from non-diseased post-mortem human caudate revealed an excellent specific binding window (>80%) using [3H]ORM-13070. BAY 292 bound to the same binding site as [3H]ORM-13070 and generated a good specific binding signal, with greater selectivity for α2CR. In non-human primates in vivo, [11C]ORM-13070 demonstrated a reversible behavior, with uptake at baseline highest in striatum (putamen, caudate, ventral striatum, and pallidum) and low in the cerebellar cortex, consistent with the known distribution of the α2CR. A dose dependent increase in receptor occupancy after BAY 292 administration was observed, confirming BBB penetration and target engagement. The estimated EC50 for BAY 292 is 33.39 ± 11.91 ng/mL. This study aimed to demonstrate the suitability of [11C]ORM-13070 as a PET-radioligand for the study of α2CR in the non-human primate brain, and to pave the way for future clinical PET tracer studies with BAY 292. [ABSTRACT FROM AUTHOR]

This paper describes the preclinical validation of the radioligand [ 11 C]ORM-13070 and its tritiated analog for addressing selectivity and occupancy of the selective alpha-2C adrenergic receptor (α 2C R) antagonist BAY 292 in the cynomolgus brain. BAY 292 is a novel drug candidate being developed for the treatment of obstructive sleep apnea (OSA) via binding to central α 2C R. In vitro autoradiography studies with sections from non-diseased post-mortem human caudate revealed an excellent specific binding window (>80%) using [ 3 H]ORM-13070. BAY 292 bound to the same binding site as [ 3 H]ORM-13070 and generated a good specific binding signal, with greater selectivity for α 2C R. In non-human primates in vivo , [ 11 C]ORM-13070 demonstrated a reversible behavior, with uptake at baseline highest in striatum (putamen, caudate, ventral striatum, and pallidum) and low in the cerebellar cortex, consistent with the known distribution of the α 2C R. A dose dependent increase in receptor occupancy after BAY 292 administration was observed, confirming BBB penetration and target engagement. The estimated EC 50 for BAY 292 is 33.39 ± 11.91 ng/mL. This study aimed to demonstrate the suitability of [ 11 C]ORM-13070 as a PET-radioligand for the study of α 2C R in the non-human primate brain, and to pave the way for future clinical PET tracer studies with BAY 292., Competing Interests: Declaration of conflicting interestsThe author(s) declared the following potential conflicts of interest with respect to the research, authorship, and/or publication of this article: DPB, IP, TP and WP are employees of Bayer AG and may own shares or share options. MD was employee of Bayer AG at the time of conducting the study and data analysis. The study was funded by Bayer AG.ALA, CCC, DRB, EAR, KRZ, LAW are full-time employees of Invicro LLC.

Cocaine is a recreational drug of abuse that binds to the dopamine transporter, preventing reuptake of dopamine into pre-synaptic terminals. The increased presence of synaptic dopamine results in stimulation of both pre- and post-synaptic dopamine receptors, considered an important mechanism by which cocaine elicits its reinforcing properties. However, the effects of acute cocaine administration on pre-synaptic dopamine function remain unclear. Non-invasive imaging techniques such as positron emission tomography have revealed impaired pre-synaptic dopamine function in chronic cocaine users. Similar impairments have been seen in animal studies, with microdialysis experiments indicating decreased basal dopamine release. Here we use micro positron emission tomography imaging techniques in mice to measure dopamine synthesis capacity and determine the effect of acute cocaine administration of pre-synaptic dopamine function. We show that a dose of 20 mg/kg cocaine is sufficient to elicit hyperlocomotor activity, peaking 15-20 min post treatment (p < 0.001). However, dopamine synthesis capacity in the striatum was not significantly altered by acute cocaine treatment (KiCer: 0.0097 per min vs. 0.0112 per min in vehicle controls, p > 0.05). Furthermore, expression levels of two key enzymes related to dopamine synthesis, tyrosine hydroxylase and aromatic l-amino acid decarboxylase, within the striatum of scanned mice were not significantly affected by acute cocaine pre-treatment (p > 0.05). Our findings suggest that while the regulation of dopamine synthesis and release in the striatum have been shown to change with chronic cocaine use, leading to a reduced basal tone, these adaptations to pre-synaptic dopaminergic neurons are not initiated following a single exposure to the drug., (© 2017 International Society for Neurochemistry.)