Evidence suggests that 30-50% of patients suffering from major depressive disorder (MDD) are classified as suffering from treatment resistant depression (TRD) as they have an inadequate response to standard antidepressants. A key feature of this patient population is the increased incidence of co-morbid symptoms like anxiety and pain. Recognizing that current standards of care are largely focused on monoaminergic mechanisms of action (MOAs), innovative approaches to drug discovery for TRD are targeting glutamate hyperfunction. Here we describe the in vitro and in vivo profile of GRN-529, a novel negative allosteric modulator (NAM) of metabotropic glutamate receptor 5 (mGluR5). In cell based pharmacology assays, GRN-529 is a high affinity (Ki 5.4 nM), potent (IC50 3.1 nM) and selective (>1000-fold selective vs mGluR1) mGluR5 NAM. Acute administration of GRN-529 (0.1-30 mg/kg p.o.) had dose-dependent efficacy across a therapeutically relevant battery of animal models, comprising depression (decreased immobility time in tail suspension and forced swim tests) and 2 of the co-morbid symptoms overrepresented in TRD, namely anxiety (attenuation of stress-induced hyperthermia, and increased punished crossings in the four plate test) and pain (reversal of hyperalgesia due to sciatic nerve ligation or inflammation). The potential side effect liability of GRN-529 was also assessed using preclinical models: GRN-529 had no effect on rat sexual behavior or motor co-ordination (rotarod), however it impaired cognition in mice (social odor recognition). Efficacy and side effects of GRN-529 were compared to standard of care agents (antidepressant, anxiolytic or analgesics) and the tool mGluR5 NAM, MTEP. To assess the relationship between target occupancy and efficacy, ex vivo receptor occupancy was measured in parallel with efficacy testing. This revealed a strong correlation between target engagement, exposure and efficacy across behavioral endpoints, which supports the potential translational value of PET imaging to dose selection in patients. Collectively this broad spectrum profile of efficacy of GRN-529 supports our hypothesis that negative allosteric modulation of mGluR5 could represent an innovative therapeutic approach to the treatment of TRD. This article is part of a Special Issue entitled 'Metabotropic Glutamate Receptors'.

Positive allosteric modulators (PAMs) of metabotropic glutamate receptor subtype 5 (mGlu5) enhance N-methyl-d-aspartate receptor function and may represent a novel approach for the treatment of schizophrenia. ADX47273 [S-(4-fluoro-phenyl)-{3-[3-(4-fluoro-phenyl)-[1,2,4]oxadiazol-5-yl]-piperidin-1-yl}-methanone], a recently identified potent and selective mGlu5 PAM, increased (9-fold) the response to threshold concentration of glutamate (50 nM) in fluorometric Ca2+ assays (EC50 = 170 nM) in human embryonic kidney 293 cells expressing rat mGlu5. In the same system, ADX47273 dose-dependently shifted mGlu5 receptor glutamate response curve to the left (9-fold at 1 μM) and competed for binding of [3H]2-methyl-6-(phenylethynyl)pyridine (Ki = 4.3 μM), but not [3H]quisqualate. In vivo, ADX47273 increased extracellular signal-regulated kinase and cAMP-responsive element-binding protein phosphorylation in hippocampus and prefrontal cortex, both of which are critical for glutamate-mediated signal transduction mechanisms. In models sensitive to antipsychotic drug treatment, ADX47273 reduced rat-conditioned avoidance responding [minimal effective dose (MED) = 30 mg/kg i.p.] and decreased mouse apomorphine-induced climbing (MED = 100 mg/kg i.p.), with little effect on stereotypy or catalepsy. Furthermore, ADX47273 blocked phencyclidine, apomorphine, and amphetamine-induced locomotor activities (MED = 100 mg/kg i.p.) in mice and decreased extracellular levels of dopamine in the nucleus accumbens, but not in the striatum, in rats. In cognition models, ADX47273 increased novel object recognition (MED = 1 mg/kg i.p.) and reduced impulsivity in the five-choice serial reaction time test (MED = 10 mg/kg i.p.) in rats. Taken together, these effects are consistent with the hypothesis that allosteric potentiation of mGlu5 may provide a novel approach for development of antipsychotic and procognitive agents.

The nuclear receptors LXRα and LXRβ are ligand-activated transcription factors that induce genes controlling cholesterol homeostasis and lipogenesis. The ability of non-selective LXR agonists to reduce brain Aβ levels, improve cognition and to inhibit inflammatory gene expression in APP transgenic mice has generated widespread interest in these nuclear hormone receptors as potential targets for Alzheimer's disease therapy. However, these non-selective compounds also have the undesirable side effect of inducing liver steatosis and hypertriglyceridemia by activating hepatic LXRα. In order to avoid this LXRα-based toxicity many pharmaceutical companies have begun to develop selective LXRβ agonists as potential therapeutics. However, the question of whether the selective activation LXRβ in the CNS will be sufficient to reduce Aβ levels has not been addressed. We therefore studied the effect of a non-selective LXR agonist, TO901317, in LXRα and LXRβ knockout mice to assess the relative contribution of each LXR isoform on the regulation of hippocampal Aβ levels. We treated wild type, LXRα-/- and LXRβ-/- animals with vehicle or 30 mg/kg T0901317 for 7 days. This dose of TO901317 has previously been shown to maximally activate LXR in the CNS. In wild type mice, TO901317 induced the LXR target genes, ABCA1 and apoE by 400 % and 132 % respectively, leading to a corresponding 28 % reduction in hippocampal Aβ42 levels. In contrast, mice deficient in either LXRα or LXRβ, showed a blunted induction of hippocampal ABCA1 and apoE following TO901317 treatment, indicating that both isoforms significantly contribute to the overall CNS efficacy of TO901317. Indeed, when measuring the relative contribution of each LXR isoform on hippocampal ABCA1 and apoE induction, we see a consistent ratio of 40% of efficacy due to LXRα expression and 60% of efficacy due to LXRβ. Importantly, the loss of either LXR isoform completely blocked the ability of TO901317 to reduce hippocampal Aβ42 levels. Taken together these data suggest that activation of both LXR isoforms in the CNS is required to effectively lower Aβ levels and that LXRβ selective compounds currently being developed for atherosclerosis will have limited utility as an Aβ-lowering therapy for Alzheimer's disease.

Recent studies show that intracellular cholesterol levels can modulate the processing of amyloid precursor protein to Aβ peptide. Moreover, cholesterol-rich apoE-containing lipoproteins may also promote Aβ clearance. Agonists of the liver X receptor (LXR) transcriptionally induce genes involved in intracellular lipid efflux and transport, including apoE. Thus, LXR agonists have the potential to both inhibit APP processing and promote Aβ clearance. Here we show that LXR agonist, TO901317, increased hippocampal ABCA1 and apoE and decreased Aβ42 levels in APP transgenic mice. TO901317 had no significant effects on levels of Aβ40, full length APP, or the APP processing products. Next, we examined the effects of TO901317 in the contextual fear conditioning paradigm; TO901317 completely reversed the contextual memory deficit in these mice. These data demonstrate that LXR agonists do not directly inhibit APP processing but rather facilitate the clearance of Aβ42 and may represent a novel therapeutic approach to Alzheimer's disease.

Brain derived neurotrophic factor (BDNF) exhibits a sequence of actions on neurons ranging from acute enhancement of transmission to long-term promotion of neurite outgrowth and synaptogenesis associated with learning and memory. The manifold effects of BDNF on neuronal modifications may be mediated by genomic alterations. We previously found that BDNF treatment acutely increases transcription of the synaptic vesicle protein Rab3A, required for trophin-induced synaptic plasticity, as well as the peptide VGF, which increases during learning. To elucidate comprehensive transcriptional programs associated with short- and long-term BDNF exposure, we now examine mRNA abundance and complexity using Affymetrix GeneChips in cultured hippocampal neurons. Consistent with the modulation of synaptic plasticity, BDNF treatment (3-6 h) induced mRNAs encoding the synapse-associated proteins synaptojanin 2, neuronal pentraxin 1, septin 9, and ryanodine receptor 2. BDNF also induced expression of mRNAs encoding neuropeptides (6-12 h), including prepronociceptin, neuropeptide Y, and secretogranin. To determine whether these neuropeptides induced by BDNF mediate neuronal development, we examined their effects on hippocampal neurons. The four mature peptides derived from post-translational processing of the ppNociceptin propeptide induced the expression of several immediate early genes in hippocampal cultures, indicating neuronal activation. To examine the significance of activation, the effects of nociceptin (orphanin FQ) and nocistatin on neurite outgrowth were examined. Quantitative morphometric analysis revealed that nociceptin significantly increased both average neurite length and average number of neurites per neuron, while nocistatin had no effect on these parameters. These results reveal a novel role for nociceptin and suggest that these neuropeptide systems may contribute to the regulation of neuronal function by BDNF.