Vijay Chhajlani

Acute pharmacological modulation of mGluR8 reduces measures of anxiety

Metabotropic glutamate receptors (mGluRs), which are coupled to second messenger pathways via G proteins, modulate glutamatergic and GABAergic neurotransmission. Because of their role in modulating neurotransmission, mGluRs are attractive therapeutic targets for anxiety disorders. Previously we showed that mGluR8(-/-) male mice showed higher measures of anxiety in the open field and elevated plus maze than age-matched wild-type mice. In this study, we assessed the potential effects of acute pharmacological modulation of mGluR8 on measures of avoidable and unavoidable anxiety. In addition to wild-type mice, we also tested apolipoprotein E-deficient (Apoe(-/-)) mice, as these mice show increased levels of anxiety-like behaviors and therefore might show an altered sensitivity to mGluR8 stimulation. mGluR8 stimulation with the specific agonist DCPG, or modulation with AZ12216052, a new, positive allosteric modulator of mGluR8 reduced measures of anxiety in both wild-type mice. The effects of mGluR8 positive allosteric modulators, which only affect neurotransmission in the presence of extracellular glutamate, seem particularly promising for patients with anxiety disorders showing benzodiazepine insensitivity.

Preclinical profile of a novel metabotropic glutamate receptor 5 positive allosteric modulator

Recent reports have indicated that patients with schizophrenia have a profound hypo-functionality of glutamatergic signaling pathways. Positive allosteric modulation of mGlu(5) receptor has been postulated to augment NMDA function and thereby alleviate the glutamatergic hypo-function observed in schizophrenic patients. Here we report the in vitro and in vivo characterization of CPPZ (1-(4-(2-chloro-4-fluorophenyl)piperazin-1-yl)-2-(pyridin-4-ylmethoxy)ethanone), a structurally novel positive allosteric modulator selective for mGlu(5) receptor. In HEK293 cells stably over-expressing human mGlu(5) receptor, CPPZ potentiates the intracellular calcium response elicited by a suboptimal concentration of the endogenous agonist glutamate. CPPZ does not have any intrinsic agonist activity and behaves functionally as a positive allosteric modulator. This is further supported by binding data, which demonstrate that CPPZ is able to displace the negative allosteric modulator MPEP but does not compete with the orthosteric ligand quisqualic acid. Instead, CPPZ enhances the binding of the orthosteric ligand. In native preparations, CPPZ potentiates calcium flux in rat cortical neurons stimulated with the group I agonist dihydroxyphenylglycine (DHPG). In addition, CPPZ modulates long-term potentiation in rat hippocampal slices, a process known to be NMDA dependent. In vivo, CPPZ reverses hyper locomotion triggered by the NMDA open channel blocker MK801 in CD1 mice. CPPZ was also able to reduce rat conditioned avoidance responding to electric shock. Both in vitro and in vivo data demonstrate that this novel compound acts as an mGlu(5) receptor positive allosteric modulator, which modulates NMDA dependent responses and suggests that the enhancement of mGlu(5) receptor activity may prove useful in the treatment of schizophrenia.

4-Aryl piperazine and piperidine amides as novel mGluR5 positive allosteric modulators

Positive allosteric modulation of metabotropic glutamate receptor 5 (mGluR5) is regarded as a potential novel treatment for schizophrenic patients. Herein we report the synthesis and SAR of 4-aryl piperazine and piperidine amides as potent mGluR5 positive allosteric modulators (PAMs). Several analogs have excellent activity and desired drug-like properties. Compound 2b was further characterized as a PAM using several in vitro experiments, and produced robust activity in several preclinical animal models.

Synthesis and Biological activity of kappa opioid receptor agonists. Part 2: Preparation of 3-aryl-2-pyridone analogues generated by solution- and solid-phase parallel synthesis methods

New analogues of the previously described 3-aryl pyridone KOR agonists have been synthesised by parallel synthetic methods, both in solution- and with solid-phase chemistry, making use of the well known and versatile Mitsunobu, Suzuki and Buchwald reactions. Opioid receptor binding data for the compounds produced is reported.

3-Aryl pyridone derivatives. Potent and selective kappa opioid receptor agonists

A new series of 3-aryl pyridone based kappa opioid receptor agonists was designed and synthesised, based on an understanding of the classical kappa opioid receptor pharmacophore. The most potent of the new compounds were comparable to U-69,593 in receptor affinity, selectivity and functional agonist effect at the cloned human kappa opioid receptor.

[3H]Chiba-1001(methyl-SSR180711) has low in vitro binding affinity and poor in vivo selectivity to nicotinic alpha-7 receptor in rodent brain

Neuronal nicotinic acetylcholine receptor (nAChR) agonists active at the alpha‐7 (α‐7) receptor subtype are potential therapeutics for cognitive deficits in schizophrenia, Alzheimers disease, and other mental disorders. SSR180711, an α‐7 selective partial agonist, has been shown to improve preclinical cognition. A novel positron emission tomography (PET) radioligand, 11C‐Chiba1001, is a close analog of SSR180711. We labeled Chiba‐1001 with tritium in order to evaluate its utility as a preclinical radioligand tool. In vitro, the binding affinity of [3H]Chiba‐1001 at the α‐7 receptor was low (Kd = 120–180 nM) in both HEK239 cell membranes expressing human α‐7 receptor and in native rat hippocampus membranes. The α‐7 selective ligands AZD0328, ARR17779, and MLA did not inhibit [3H]Chiba‐1001 binding (Ki > 10,000 nM). In rat hippocampal membranes, Chiba‐1001 and SSR180711 inhibited [3H]Chiba‐1001 binding (Ki = 220 and 230 nM, respectively), consistent with the literature reports. The in vivo binding profile of the radioligand was examined in normal rat, wild type mouse, and α‐7 knockout mouse brain. We found that [3H]Chiba‐1001 lacks adequate and specific brain regional uptake in rat and mouse brain. No significant inhibition of the radioligand binding was obtained following pretreatment of the animal with AZ11637326, AZD0328, or MLA. Our results indicate that [3H]Chiba‐1001 has low affinity for α‐7 nAChRs in vitro and poor α‐7 regional and pharmacological selectivity in the rodent brain. Synapse, 2012.

Absence of direct effects on the dopamine D2 receptor by mGluR2/3-selective receptor agonists LY 354,740 and LY 379,268.

We previously reported the absence of high‐affinity binding of the group II metabotropic glutamate receptor agonists LY 354,740 and LY 379,268 to the D2L dopamine receptor. A rebuttal to our findings has since been reported (see Introduction section); this study represents our response. Analysis by LCMS of LY 354,740 and LY 379,268 used in this study revealed the correct molecular mass for these compounds. Both LY 354,740 and LY 379,268 exhibited potent agonist activity for mGluR2 in the 35S‐GTPγS assay. Functionally, neither compound displayed antagonist activity in the GTPγS assay with recombinant D2. At concentrations up to 10 μM, both compounds failed to displace [3H]‐raclopride, [3H]‐PHNO, or [3H]‐domperidone in filter‐binding assays under isotonic (120 mM NaCl or N‐methyl glucamine) or low‐ionic strength (no NaCl or N‐methyl glucamine) conditions. Some displacement of [3H]‐domperidone (20–40%) was observed at 30 μM of LY 354,740 under low‐ionic strength and under isotonic conditions in the absence of NaCl. No displacement of [3H]‐domperidone was detected in a two site model at lower (<100 nM) concentrations of either compound. Moreover, no D2 activity was observed for LY 354,740 or LY 379,268 in the CellKey™ (cellular dielectric spectroscopy) assay. In this communication, we discuss the possible reasons for differences in our study and the previously published work and implications of these studies for mechanisms of antipsychotic action. Synapse 65:64–68, 2011.

In vitro binding of a radio-labeled positive allosteric modulator for metabotropic glutamate receptor subtype 5

The positive allosteric modulator (PAM) binding site for metabotropic glutamate receptor subtype 5 (mGlu5) lacks a readily available radio‐labeled tracer fordetailed structure‐activity studies. This communication describes a selective mGlu5 compound, 7‐methyl‐2‐(4‐(pyridin‐2‐yloxy)benzyl)‐5‐(pyridin‐3‐yl)isoindolin‐1‐one (PBPyl) that binds with high affinity to human mGlu5 and exhibits functional PAM activity. Analysis of PBPyl by FLIPR revealed an EC50 of 87 nM with an 89% effect in transfected HEK293 cells and an EC50 of 81 nM with a 42% effect in rat primary neurons. PBPyl exhibited 5‐fold higher functional selectivity for mGlu5 in a full mGlu receptor panel. Unlabeled PBPyl was tested for specific binding using a liquid chromatography mass spectrometry (LC/MS/MS)‐based filtration binding assay and exhibited 40% specific binding in recombinant membranes, a value higher than any candidate compound tested. In competition binding studies with [3H]MPEP, the mGlu5 receptor negative allosteric modulator (NAM), PBPyl exhibited a k i value of 34 nM. PBPyl also displaced [3H]ABP688, a mGluR5 receptor NAM, in tissue sections from mouse and rat brain using autoradiography. Areas of specific binding included the frontal cortex, striatum and nucleus accumbens. PBPyl was radiolabeled to a specific activity of 15 Ci/mmol and tested for specific binding in a filter plate format. In recombinant mGlu5b membranes, [3H] PBPyl exhibited saturable binding with a Kd value of 18.6 nM. In competition binding experiments, [3H] PBPyl was displaced by high affinity mGlu5 positive and negative modulators. Further tests showed that PBPyl displays less than optimal characteristics as an in vivo tool, including a high volume of distribution and ClogP, making it more suitable as an in vitro compound. However, as a first report of direct binding of an mGlu5 receptor PAM, this study offers value toward the development of novel PET imaging agents for this important therapeutic target. Synapse, 2013.