Acute Effects of Hallucinogens on Functional Connectivity: Psilocybin and Salvinorin-A.

The extent of changes in functional connectivity (FC) within functional networks as a common feature across hallucinogenic drug classes is under-explored. This work utilized fMRI to assess the dissociative hallucinogens Psilocybin, a classical serotonergic psychedelic, and Salvinorin-A, a kappa-opioid receptor (KOR) agonist, on resting-state FC in nonhuman primates. We highlight overlapping and differing influence of these substances on FC relative to the thalamus, claustrum, prefrontal cortex (PFC), default mode network (DMN), and DMN subcomponents. Analysis was conducted on a within-subject basis. Findings support the cortico-claustro-cortical network model for probing functional effects of hallucinogens regardless of serotonergic potential, with a potential key paradigm centered around the claustrum, PFC, anterior cingulate cortices (ACC), and angular gyrus relationship. Thalamo-cortical networks are implicated but appear dependent on 5-HT2AR activation. Acute desynchronization relative to the DMN for both drugs was also shown. Our findings provide a framework to understand broader mechanisms at which hallucinogens in differing classes may impact subjects regardless of the target receptor.

A Fast-Binding, Functionally Reversible, COX-2 Radiotracer for CNS PET Imaging.

Cyclooxygenase-2 (COX-2) is an enzyme that plays a pivotal role in peripheral inflammation and pain via the prostaglandin pathway. In the central nervous system (CNS), COX-2 is implicated in neurodegenerative and psychiatric disorders as a potential therapeutic target and biomarker. However, clinical studies with COX-2 have yielded inconsistent results, partly due to limited mechanistic understanding of how COX-2 activity relates to CNS pathology. Therefore, developing COX-2 positron emission tomography (PET) radiotracers for human neuroimaging is of interest. This study introduces [11C]BRD1158, which is a potent and uniquely fast-binding, selective COX-2 PET radiotracer. [11C]BRD1158 was developed by prioritizing potency at COX-2, isoform selectivity over COX-1, fast binding kinetics, and free fraction in the brain. Evaluated through in vivo PET neuroimaging in rodent models with human COX-2 overexpression, [11C]BRD1158 demonstrated high brain uptake, fast target-engagement, functional reversibility, and excellent specific binding, which is advantageous for human imaging applications. Lastly, post-mortem samples from Huntington's disease (HD) patients and preclinical HD mouse models showed that COX-2 levels were elevated specifically in disease-affected brain regions, primarily from increased expression in microglia. These findings indicate that COX-2 holds promise as a novel clinical marker of HD onset and progression, one of many potential applications of [11C]BRD1158 human PET.

Neuroimmune signatures in chronic low back pain subtypes.

We recently showed that patients with different chronic pain conditions (such as chronic low back pain, fibromyalgia, migraine and Gulf War illness) demonstrated elevated brain and/or spinal cord levels of the glial marker 18-kDa translocator protein (TSPO), which suggests that neuroinflammation might be a pervasive phenomenon observable across multiple aetiologically heterogeneous pain disorders. Interestingly, the spatial distribution of this neuroinflammatory signal appears to exhibit a degree of disease specificity (e.g. with respect to the involvement of the primary somatosensory cortex), suggesting that different pain conditions may exhibit distinct 'neuroinflammatory signatures'. To explore this hypothesis further, we tested whether neuroinflammatory signal can characterize putative aetiological subtypes of chronic low back pain patients based on clinical presentation. Specifically, we explored neuroinflammation in patients whose chronic low back pain either did or did not radiate to the leg (i.e. 'radicular' versus 'axial' back pain). Fifty-four patients with chronic low back pain, 26 with axial back pain [43.7 ± 16.6 years old (mean ± SD)] and 28 with radicular back pain (48.3 ± 13.2 years old), underwent PET/MRI with 11C-PBR28, a second-generation radioligand for TSPO. 11C-PBR28 signal was quantified using standardized uptake values ratio (validated against volume of distribution ratio; n = 23). Functional MRI data were collected simultaneously to the 11C-PBR28 data (i) to functionally localize the primary somatosensory cortex back and leg subregions; and (ii) to perform functional connectivity analyses (in order to investigate possible neurophysiological correlations of the neuroinflammatory signal). PET and functional MRI measures were compared across groups, cross-correlated with one another and with the severity of 'fibromyalgianess' (i.e. the degree of pain centralization, or 'nociplastic pain'). Furthermore, statistical mediation models were used to explore possible causal relationships between these three variables. For the primary somatosensory cortex representation of back/leg, 11C-PBR28 PET signal and functional connectivity to the thalamus were: (i) higher in radicular compared to axial back pain patients; (ii) positively correlated with each other; (iii) positively correlated with fibromyalgianess scores, across groups; and finally (iv) fibromyalgianess mediated the association between 11C-PBR28 PET signal and primary somatosensory cortex-thalamus connectivity across groups. Our findings support the existence of 'neuroinflammatory signatures' that are accompanied by neurophysiological changes and correlate with clinical presentation (in particular, with the degree of nociplastic pain) in chronic pain patients. These signatures may contribute to the subtyping of distinct pain syndromes and also provide information about interindividual variability in neuroimmune brain signals, within diagnostic groups, that could eventually serve as targets for mechanism-based precision medicine approaches.

Imaging Kappa Opioid Receptors in the Living Brain with Positron Emission Tomography.

Kappa opioid receptor (KOR) neuroimaging using positron emission tomography (PET) has been immensely successful in all phases of discovery and validation in relation to radiotracer development from preclinical imaging to human imaging. There are now several KOR-specific PET radiotracers that can be utilized for neuroimaging, including agonist and antagonist ligands, as well as C-11 and F-18 variants. These technologies will increase KOR PET utilization by imaging centers around the world and have provided a foundation for future studies. In this chapter, I review the advances in KOR radiotracer discovery, focusing on ligands that have been translated into human imaging, and highlight key attributes unique to each KOR PET radiotracer. The utilization of these radiotracers in KOR PET neuroimaging can be subdivided into three major investigational classes: the first, measurement of KOR density; the second, measurement of KOR drug occupancy; the third, detecting changes in endogenous dynorphin following activation or deactivation. Given the involvement of the KOR/dynorphin system in a number of brain disorders including, but not limited to, pain, itch, mood disorders and addiction, measuring KOR density in the living brain will offer insight into the chronic effects of these disorders on KOR tone in humans. Notably, KOR PET has been successful at measuring drug occupancy in the human brain to guide dose selection for maximal therapeutic efficacy while avoiding harmful side effects. Lastly, we discuss the potential of KOR PET to detect changes in endogenous dynorphin in the human brain, to elucidate neural mechanisms and offer critical insight into disease-modifying therapeutics. We conclude with comments on other translational neuroimaging modalities such as MRI that could be used to study KOR-dynorphin tone in the living human brain.

A New Positron Emission Tomography Probe for Orexin Receptors Neuroimaging.

The orexin receptor (OX) is critically involved in motivation and sleep-wake regulation and holds promising therapeutic potential in various mood disorders. To further investigate the role of orexin receptors (OXRs) in the living human brain and to evaluate the treatment potential of orexin-targeting therapeutics, we herein report a novel PET probe ([11C]CW24) for OXRs in the brain. CW24 has moderate binding affinity for OXRs (IC50 = 0.253 µM and 1.406 µM for OX1R and OX2R, respectively) and shows good selectivity to OXRs over 40 other central nervous system (CNS) targets. [11C]CW24 has high brain uptake in rodents and nonhuman primates, suitable metabolic stability, and appropriate distribution and pharmacokinetics for brain positron emission tomography (PET) imaging. [11C]CW24 warrants further evaluation as a PET imaging probe of OXRs in the brain.

Novel radioligands for imaging sigma-1 receptor in brain using positron emission tomography (PET).

The sigma-1 receptor (σ 1R) is a unique intracellular protein. σ 1R plays a major role in various pathological conditions in the central nervous system (CNS), implicated in several neuropsychiatric disorders. Imaging of σ 1R in the brain using positron emission tomography (PET) could serve as a noninvasively tool for enhancing the understanding of the disease's pathophysiology. Moreover, σ 1R PET tracers can be used for target validation and quantification in diagnosis. Herein, we describe the radiosynthesis, in vivo PET/CT imaging of novel σ 1R 11C-labeled radioligands based on 6-hydroxypyridazinone, [11C]HCC0923 and [11C]HCC0929. Two radioligands have high affinities to σ 1R, with good selectivity. In mice PET/CT imaging, both radioligands showed appropriate kinetics and distributions. Additionally, the specific interactions of two radioligands were reduced by compounds 13 and 15 (self-blocking). Of the two, [11C]HCC0929 was further investigated in positive ligands blocking studies, using classic σ 1R agonist SA 4503 and σ 1R antagonist PD 144418. Both σ 1R ligands could extensively decreased the uptake of [11C]HCC0929 in mice brain. Besides, the biodistribution of major brain regions and organs of mice were determined in vivo. These studies demonstrated that two radioligands, especially [11C]HCC0929, possessed ideal imaging properties and might be valuable tools for non-invasive quantification of σ 1R in brain.

Discrepancies in Kappa Opioid Agonist Binding Revealed through PET Imaging.

Kappa opioid receptor (KOR) modulation has been pursued in many conceptual frameworks for the treatment of human pain, depression, and anxiety. As such, several imaging tools have been developed to characterize the density of KORs in the human brain and its occupancy by exogenous drug-like compounds. While exploring the pharmacology of KOR tool compounds using positron emission tomography (PET), we observed discrepancies in the apparent competition binding as measured by changes in binding potential (BPND, binding potential with respect to non-displaceable uptake). This prompted us to systematically look at the relationships between baseline BPND maps for three common KOR PET radioligands, the antagonists [11C]LY2795050 and [11C]LY2459989, and the agonist [11C]GR103545. We then measured changes in BPND using kappa antagonists (naloxone, naltrexone, LY2795050, JDTic, nor-BNI), and found BPND was affected similarly between [11C]GR103545 and [11C]LY2459989. Longitudinal PET studies with nor-BNI and JDTic were also examined, and we observed a persistent decrease in [11C]GR103545 BPND up to 25 days after drug administration for both nor-BNI and JDTic. Kappa agonists were also administered, and butorphan and GR89696 (racemic GR103545) impacted binding to comparable levels between the two radiotracers. Of greatest significance, kappa agonists salvinorin A and U-50488 caused dramatic reductions in [11C]GR103545 BPND but did not change [11C]LY2459989 binding. This discrepancy was further examined in dose-response studies with each radiotracer as well as in vitro binding experiments.

Site-selective 18F fluorination of unactivated C-H bonds mediated by a manganese porphyrin.

The first direct C-H 18F fluorination reaction of unactivated aliphatic sites using no-carrier-added [18F]fluoride is reported. Under the influence of a manganese porphyrin/iodosylbenzene system, a variety of unactivated aliphatic C-H bonds can be selectively converted to C-18F bonds. The mild conditions, broad substrate scope and generally inaccessible regiochemistry make this radio-fluorination a powerful alternate to established nucleophilic substitution for the preparation of 18F labeled radio tracers.

Metal Protein-Attenuating Compound for PET Neuroimaging: Synthesis and Preclinical Evaluation of [11C]PBT2.

Dyshomeostasis or abnormal accumulation of metal ions such as copper, zinc, and iron have been linked to the pathogenesis of multiple neurodegenerative disorders including Alzheimer's disease (AD) and Huntington's disease (HD). 5,7-Dichloro-2-((dimethylamino)methyl)quinolin-8-ol, PBT2, is a second generation metal protein-attenuating compound that has recently advanced in Phase II clinical trials for the treatment of AD and HD based on promising preclinical efficacy data. Herein, we report the first radiosynthesis and preclinical positron emission tomography (PET) neuroimaging evaluation of [11C]PBT2 in rodents and nonhuman primates. Carbon-11 labeled PBT2 was synthesized in 4.8 ± 0.5% (nondecay corrected) radiochemical yield (RCY) at end-of-synthesis, based upon [11C]CH3I (n = 6), with >99% radiochemical purity and 80-90 GBq/µmol molar activity (Am) from the corresponding normethyl precursor. In the nonhuman primate brain, [11C]PBT2 uptake was extensive with peak concentration SUVpeak of 3.2-5.2 within 2.5-4.5 min postinjection in all cortical and subcortical gray matter regions (putamen > caudate > cortex ≫ white matter) followed by rapid washout from normal brain tissues. Furthermore, it is shown that [11C]PBT2 binds specifically in AD human brain tissue in vitro. The results presented here, combined with the clinical data available for PBT2, warrant the evaluation of [11C]PBT2 as an exploratory PET radiotracer in humans.

In Vivo [18F]GE-179 Brain Signal Does Not Show NMDA-Specific Modulation with Drug Challenges in Rodents and Nonhuman Primates.

As one of the major excitatory ion channels in the brain, NMDA receptors have been a leading research target for neuroscientists, physicians, medicinal chemists, and pharmaceutical companies for decades. Molecular imaging of NMDA receptors by means of positron emission tomography (PET) with [18F]GE-179 quickly progressed to clinical PET studies, but a thorough understanding of its binding specificity has been missing and has thus limited signal interpretation. Here a preclinical study with [18F]GE-179 in rodents and nonhuman primates (NHPs) is presented in an attempt to characterize [18F]GE-179 signal specificity. Rodent PET/CT was used to study drug occupancy and functional manipulation in rats by pretreating animals with NMDA targeted blocking/modulating drug doses followed by a single bolus of [18F]GE-179. Binding competition with GE-179, MK801, PCP, and ketamine, allosteric inhibition by ifenprodil, and brain activation with methamphetamine did not alter the [18F]GE-179 brain signal in rats. In addition, multimodal imaging with PET/MRI in NHPs was used to evaluate changes in radiotracer binding as a function of pharmacological challenges. Drug-induced hemodynamic changes were monitored simultaneously using functional MRI (fMRI). Comparisons of baseline and signal after drug challenge in NHPs demonstrated that the [18F]GE-179 signal cannot be manipulated in a predictable fashion in vivo. fMRI data acquired simultaneously with PET data supported this finding and provided evidence that radiotracer delivery is not altered by blood flow changes. In conclusion, the [18F]GE-179 brain signal is not readily interpretable in the context of NMDA receptor binding on the basis of the results shown in this study.

[11C]Cyanation of arylboronic acids in aqueous solutions.

A copper-mediated 11C-cyanation method employing arylboronic acids and [11C]HCN has been developed. This method was applied to the radiochemical synthesis of a wide range of aromatic 11C-nitriles in aqueous solutions. The use of readily accessible arylboronic acids as precursors makes this method complementary to the well-established 11C-cyanation methods that utilize aryl halide precursors.

Preclinical PET Neuroimaging of [11C]Bexarotene.

Activation of retinoid X receptors (RXRs) has been proposed as a therapeutic mechanism for the treatment of neurodegeneration, including Alzheimer's and Parkinson's diseases. We previously reported radiolabeling of a Food and Drug Administration-approved RXR agonist, bexarotene, by copper-mediated [(11)C]CO2 fixation and preliminary positron emission tomography (PET) neuroimaging that demonstrated brain permeability in nonhuman primate with regional binding distribution consistent with RXRs. In this study, the brain uptake and saturability of [(11)C]bexarotene were studied in rats and nonhuman primates by PET imaging under baseline and greater target occupancy conditions. [(11)C]Bexarotene displays a high proportion of nonsaturable uptake in the brain and is unsuitable for RXR occupancy measurements in the central nervous system.

(11)C[double bond, length as m-dash]O bonds made easily for positron emission tomography radiopharmaceuticals.

The positron-emitting radionuclide carbon-11 ((11)C, t1/2 = 20.3 min) possesses the unique potential for radiolabeling of any biological, naturally occurring, or synthetic organic molecule for in vivo positron emission tomography (PET) imaging. Carbon-11 is most often incorporated into small molecules by methylation of alcohol, thiol, amine or carboxylic acid precursors using [(11)C]methyl iodide or [(11)C]methyl triflate (generated from [(11)C]carbon dioxide or [(11)C]methane). Consequently, small molecules that lack an easily substituted (11)C-methyl group are often considered to have non-obvious strategies for radiolabeling and require a more customized approach. [(11)C]Carbon dioxide itself, [(11)C]carbon monoxide, [(11)C]cyanide, and [(11)C]phosgene represent alternative reactants to enable (11)C-carbonylation. Methodologies developed for preparation of (11)C-carbonyl groups have had a tremendous impact on the development of novel PET tracers and provided key tools for clinical research. (11)C-Carbonyl radiopharmaceuticals based on labeled carboxylic acids, amides, carbamates and ureas now account for a substantial number of important imaging agents that have seen translation to higher species and clinical research of previously inaccessible targets, which is a testament to the creativity, utility and practicality of the underlying radiochemistry.

PET Neurochemical Imaging Modes.

PET has deep roots in neuroscience stemming from its first application in brain tumor and brain metabolism imaging. PET emerged over the past few decades and continues to play a prominent role in the study of neurochemistry in the living human brain. Over time, neurochemical imaging with PET has been expanded to address a host of research questions related to, among many others, protein density, drug occupancy, and endogenous neurochemical release. Each of these imaging modes has distinct design and analysis considerations that are critical for enabling quantitative measurements. The number of considerations required for a neurochemical PET study can make it unapproachable. This article aims to orient those interested in neurochemical PET imaging to three of the common imaging modes and to provide some perspective on needs that exist for expansion of neurochemical PET imaging.

Development of a Fluorinated Class-I HDAC Radiotracer Reveals Key Chemical Determinants of Brain Penetrance.

Despite major efforts, our knowledge about many brain diseases remains remarkably limited. Epigenetic dysregulation has been one of the few leads toward identifying the causes and potential treatments of psychiatric disease over the past decade. A new positron emission tomography radiotracer, [(11)C]Martinostat, has enabled the study of histone deacetylase in living human subjects. A unique property of [(11)C]Martinostat is its profound brain penetrance, a feature that is challenging to engineer intentionally. In order to understand determining factors for the high brain-uptake of Martinostat, a series of compounds was evaluated in rodents and nonhuman primates. The study revealed the major structural contributors to brain uptake, as well as a more clinically relevant fluorinated HDAC radiotracer with comparable behavior to Martinostat, yet longer half-life.

A Novel Radiotracer for Imaging Monoacylglycerol Lipase in the Brain Using Positron Emission Tomography.

Monoacylglycerol lipase (MAGL) is a serine hydrolase that hydrolyzes monoacylglycerols to glycerol and fatty acid and plays an important role in neuroinflammation. MAGL inhibitors are a class of molecules with therapeutic potential for human diseases of the central nervous system (CNS), in areas such as pain and inflammation, immunological disorders, and neurological and psychiatric conditions. Development of a noninvasive imaging probe would elucidate the distribution and functional roles of MAGL in the brain and accelerate medical research and drug discovery in this domain. Herein, we describe the synthesis and pilot rodent imaging of a novel MAGL imaging agent, [(11)C]SAR127303. Our imaging results demonstrate the high specificity, good selectivity, and appropriate kinetics and distribution of [(11)C]SAR127303, validating its utility for imaging MAGL in the brain. Our findings support the translational potential for human CNS MAGL imaging.

Immediate and Persistent Effects of Salvinorin A on the Kappa Opioid Receptor in Rodents, Monitored In Vivo with PET.

Monitoring changes in opioid receptor binding with positron emission tomography (PET) could lead to a better understanding of tolerance and addiction because altered opioid receptor dynamics following agonist exposure has been linked to tolerance mechanisms. We have studied changes in kappa opioid receptor (KOR) binding availability in vivo with PET following kappa opioid agonist administration. Male Sprague-Dawley rats (n=31) were anesthetized and treated with the (KOR) agonist salvinorin A (0.01-1.8 mg/kg, i.v.) before administration of the KOR selective radiotracer [(11)C]GR103545. When salvinorin A was administered 1 min prior to injection of the radiotracer, [(11)C]GR103545 binding potential (BPND) was decreased in a dose-dependent manner, indicating receptor binding competition. In addition, the unique pharmacokinetics of salvinorin A (half-life ~8 min in non-human primates) allowed us to study the residual impact on KOR after the drug had eliminated from the brain. Salvinorin A was administered up to 5 h prior to [(11)C]GR103545, and the changes in BPND were compared with baseline, 2.5 h, 1 h, and 1 min pretreatment times. At lower doses (0.18 mg/kg and 0.32 mg/kg) we observed no prolonged effect on KOR binding but at 0.60 mg/kg salvinorin A induced a sustained decrease in KOR binding (BPND decreased by 40-49%) which persisted up to 2.5 h post administration, long after salvinorin A had been eliminated from the brain. These data point towards an agonist-induced adaptive response by KOR, the dynamics of which have not been previously studied in vivo with PET.

Virtually instantaneous, room-temperature [(11)C]-cyanation using biaryl phosphine Pd(0) complexes.

A new radiosynthetic protocol for the preparation of [(11)C]aryl nitriles has been developed. This process is based on the direct reaction of in situ prepared L·Pd(Ar)X complexes (L = biaryl phosphine) with [(11)C]HCN. The strategy is operationally simple, exhibits a remarkably wide substrate scope with short reaction times, and demonstrates superior reactivity compared to previously reported systems. With this procedure, a variety of [(11)C]nitrile-containing pharmaceuticals were prepared with high radiochemical efficiency.

Investigation of the lactam side chain length necessary for optimal indenoisoquinoline topoisomerase I inhibition and cytotoxicity in human cancer cell cultures.

Indenoisoquinolines with lactam substituents such as ethylamino, propylamino, and butylamino have previously demonstrated potent biological activity, but an optimal length has never been established. In the present study, a series of simplified indenoisoquinoline analogues possessing a linker spacing of 0-12 carbon atoms between the lactam nitrogen and the terminal amino group have been prepared, determining that 2-4-atom lengths are optimal for topoisomerase I inhibition and cytotoxicity. Using these lengths, analogues were prepared with the amino group and portions of the linker replaced by a pyridine ring. A three-carbon spacer within the pyridine series still demonstrated potent topoisomerase I inhibition.