Magnetic resonance imaging detects white adipose tissue beiging in mice following PDE10A inhibitor treatment.

Weight gain is a common harmful side effect of atypical antipsychotics used for schizophrenia treatment. Conversely, treatment with the novel phosphodiesterase-10A (PDE10A) inhibitor MK-8189 in clinical trials led to significant weight reduction, especially in patients with obesity. This study aimed to understand and describe the mechanism underlying this observation, which is essential to guide clinical decisions. We hypothesized that PDE10A inhibition causes beiging of white adipose tissue (WAT), leading to weight loss. Magnetic resonance imaging (MRI) methods were developed, validated, and applied in a diet-induced obesity mouse model treated with a PDE10A inhibitor THPP-6 or vehicle for measurement of fat content and vascularization of adipose tissue. Treated mice showed significantly lower fat fraction in white and brown adipose tissue, and increased perfusion and vascular density in WAT versus vehicle, confirming the hypothesis, and matching the effect of CL-316,243, a compound known to cause adipose tissue beiging. The in vivo findings were validated by qPCR revealing upregulation of Ucp1 and Pcg1-α genes, known markers of WAT beiging, and angiogenesis marker VegfA in the THPP-6 group. This work provides a detailed understanding of the mechanism of action of PDE10A inhibitor treatment on adipose tissue and body weight and will be valuable to guide both the use of MK-8189 in schizophrenia and the potential application of the target for weight loss indication.

Structures of active-state orexin receptor 2 rationalize peptide and small-molecule agonist recognition and receptor activation.

Narcolepsy type 1 (NT1) is a chronic neurological disorder that impairs the brain's ability to control sleep-wake cycles. Current therapies are limited to the management of symptoms with modest effectiveness and substantial adverse effects. Agonists of the orexin receptor 2 (OX2R) have shown promise as novel therapeutics that directly target the pathophysiology of the disease. However, identification of drug-like OX2R agonists has proven difficult. Here we report cryo-electron microscopy structures of active-state OX2R bound to an endogenous peptide agonist and a small-molecule agonist. The extended carboxy-terminal segment of the peptide reaches into the core of OX2R to stabilize an active conformation, while the small-molecule agonist binds deep inside the orthosteric pocket, making similar key interactions. Comparison with antagonist-bound OX2R suggests a molecular mechanism that rationalizes both receptor activation and inhibition. Our results enable structure-based discovery of therapeutic orexin agonists for the treatment of NT1 and other hypersomnia disorders.

Discovery of highly potent and selective orexin 1 receptor antagonists (1-SORAs) suitable for in vivo interrogation of orexin 1 receptor pharmacology.

While a correlation between blockade of the orexin 2 receptor (OX2R) with either a dual orexin receptor antagonist (DORA) or a selective orexin 2 receptor antagonist (2-SORA) and a decrease of wakefulness is well established, less is known about selective blockade of the orexin 1 receptor (OX1R). Therefore, a highly selective orexin 1 antagonist (1-SORA) with suitable properties to allow in vivo interrogation of OX1R specific pharmacology in preclinical species remains an attractive target. Herein, we describe the discovery of an optimized 1-SORA series in the piperidine ether class. Notably, a 4,4-difluoropiperidine core coupled with a 2-quinoline ether linkage provides OX1R selective compounds. The combination with an azabenzimidazole or imidazopyridine amide substituent leads to analogs 47 and 51 with >625-fold functional selectivity for OX1R over OX2R in rat. Compounds 47 and 51 possess clean off-target profiles and the required pharmacokinetic and physical properties to be useful as 1-SORA tool compounds.

Quantitative electroencephalography within sleep/wake states differentiates GABAA modulators eszopiclone and zolpidem from dual orexin receptor antagonists in rats.

Dual orexin receptor antagonists (DORAs) induce sleep by blocking orexin 1 and orexin 2 receptor-mediated activities responsible for regulating wakefulness. DORAs represent a potential alternative mechanism to the current standard of care that includes the γ-aminobutyric acid (GABA)A receptor-positive allosteric modulators, eszopiclone and zolpidem. This work uses an innovative method to analyze electroencephalogram (EEG) spectral frequencies within sleep/wake states to differentiate the effects of GABAA modulators from DORA-22, an analog of the DORA MK-6096, in Sprague-Dawley rats. The effects of low, intermediate, and high doses of eszopiclone, zolpidem, and DORA-22 were examined after first defining each compound's ability to promote sleep during active-phase dosing. The EEG spectral frequency power within specific sleep stages was calculated in 1-Hz intervals from 1 to 100 Hz within each sleep/wake state for the first 4 h after the dose. Eszopiclone and zolpidem produced marked, dose-responsive disruptions in sleep stage-specific EEG spectral profiles compared with vehicle treatment. In marked contrast, DORA-22 exhibited marginal changes in the spectral profile, observed only during rapid eye movement sleep, and only at the highest dose tested. Moreover, while eszopiclone- and zolpidem-induced changes were evident in the inactive period, the EEG spectral responses to DORA-22 were absent during this phase. These results suggest that DORA-22 differs from eszopiclone and zolpidem whereby DORA-22 promotes somnolence without altering the neuronal network EEG activity observed during normal sleep.

Orexin receptor antagonists differ from standard sleep drugs by promoting sleep at doses that do not disrupt cognition.

Current treatments for insomnia, such as zolpidem (Ambien) and eszopiclone (Lunesta), are γ-aminobutyric acid type A (GABAA)-positive allosteric modulators that carry a number of side effects including the potential to disrupt cognition. In an effort to develop better tolerated medicines, we have identified dual orexin 1 and 2 receptor antagonists (DORAs), which promote sleep in preclinical animal models and humans. We compare the effects of orally administered eszopiclone, zolpidem, and diazepam to the dual orexin receptor antagonist DORA-22 on sleep and the novel object recognition test in rat, and on sleep and two cognition tests (delayed match to sample and serial choice reaction time) in the rhesus monkey. Each compound's minimal dose that promoted sleep versus the minimal dose that exerted deficits in these cognitive tests was determined, and a therapeutic margin was established. We found that DORA-22 has a wider therapeutic margin for sleep versus cognitive impairment in rat and rhesus monkey compared to the other compounds tested. These data were further supported with the demonstration of a wider therapeutic margin for DORA-22 compared to the other compounds on sleep versus the expression of hippocampal activity-regulated cytoskeletal-associated protein (Arc), an immediate-early gene product involved in synaptic plasticity. These findings suggest that DORAs might provide an effective treatment for insomnia with a greater therapeutic margin for sleep versus cognitive disturbances compared to the GABAA-positive allosteric modulators currently in use.

Design, synthesis, and evaluation of novel 3,6-diaryl-4-aminoalkoxyquinolines as selective agonists of somatostatin receptor subtype 2.

Agonists of somatostatin receptor subtype 2 (sst(2)) have been proposed as therapeutics for the treatment of proliferative diabetic retinopathy and exudative age-related macular degeneration. An HTS screen identified 2-quinolones as weak agonists of sst(2), and these were optimized to provide small molecules with sst(2) binding and functional potency comparable to peptide agonists. Agonist 21 was shown to inhibit rat growth hormone secretion following systemic administration and to inhibit ocular neovascular lesion formation after local administration.

Discovery of 3-substituted aminocyclopentanes as potent and orally bioavailable NR2B subtype-selective NMDA antagonists.

A series of 3-substituted aminocyclopentanes has been identified as highly potent and selective NR2B receptor antagonists. Incorporation of a 1,2,4-oxadiazole linker and substitution of the pendant phenyl ring led to the discovery of orally bioavailable analogues that showed efficient NR2B receptor occupancy in rats. Unlike nonselective NMDA antagonists, the NR2B-selective antagonist 22 showed no adverse affects on motor coordination in the rotarod assay at high dose. Compound 22 was efficacious following oral administration in a spinal nerve ligation model of neuropathic pain and in an acute model of Parkinson's disease in a dose dependent manner.

Allosteric modulation of group III metabotropic glutamate receptor 4: a potential approach to Parkinson's disease treatment.

Parkinson's disease (PD) is a debilitating movement disorder that afflicts >1 million people in North America. Current treatments focused on dopamine-replacement strategies ultimately fail in most patients because of loss of efficacy and severe adverse effects that worsen as the disease progresses. The recent success of surgical approaches suggests that a pharmacological intervention that bypasses the dopamine system and restores balance in the basal ganglia motor circuit may provide an effective treatment strategy. We previously identified the metabotropic glutamate receptor 4 (mGluR4) as a potential drug target and predicted that selective activation of mGluR4 could provide palliative benefit in PD. We now report that N-phenyl-7-(hydroxylimino)cyclopropa[b]chromen-1a-carboxamide (PHCCC) is a selective allosteric potentiator of mGluR4. This compound selectively potentiated agonist-induced mGluR4 activity in cultured cells expressing this receptor and did not itself act as an agonist. Furthermore, PHCCC potentiated the effect of l-(+)-2-amino-4-phosphonobutyric acid in inhibiting transmission at the striatopallidal synapse. Modulation of the striatopallidal synapse has been proposed as a potential therapeutic target for PD, in that it may restore balance in the basal ganglia motor circuit. Consistent with this, PHCCC produced a marked reversal of reserpine-induced akinesia in rats. The closely related analogue 7-(hydroxylimino)cyclopropachromen-1a-carboxamide ethyl ester, which does not potentiate mGluR4, had no effect in this model. These results are evidence for in vivo behavioral effects of an allosteric potentiator of mGluRs and suggest that potentiation of mGluR4 may be a useful therapeutic approach to the treatment of PD.

Neuropeptide FF receptor 2 (NPFF2) is localized to pain-processing regions in the primate spinal cord and the lower level of the medulla oblongata.

Studies have suggested that NPFF-like peptides and their receptors play important roles in physiological and pathological conditions. Here, we show, using multiple expression modalities, that the type 2 NPFF receptor (hNPFF2) is expressed in regions of the primate spinal cord and brainstem mediating pain sensation. In situ hybridization using an NPFF2 riboprobe, and immunohistochemistry using a novel NPFF2 antibody, demonstrated strong NPFF2 expression in the superficial layer of the dorsal horn, and in the spinal trigeminal nucleus of the brainstem of the African green monkey (AGM). In addition, autoradiography using a radiolabeled NPFF analog ([125I]1DMe) revealed dense binding signal in the superficial layer of the dorsal horn in the spinal cord. The distribution pattern of hNPFF2 in the AGM spinal cord and the lower level of the brainstem are consistent with a hypothesized potential role for NPFF peptides in modulation of sensory input, opioid analgesia and morphine tolerance through spinal and supraspinal mechanisms.