Fragile X mental retardation protein (FMRP) and metabotropic glutamate receptor subtype 5 (mGlu5) control stress granule formation in astrocytes

Fragile X syndrome (FXS) is a common form of intellectual disability and autism caused by the lack of Fragile X Mental Retardation Protein (FMRP), an RNA-binding protein involved in RNA transport and protein synthesis. Upon cellular stress, global protein synthesis is blocked and mRNAs are recruited into stress granules (SGs), together with RNA-binding proteins including FMRP. Activation of group-I metabotropic glutamate (mGlu) receptors stimulates FMRP-mediated mRNA transport and protein synthesis, but their role in SGs formation is unexplored. To this aim, we pre-treated wild type (WT) and Fmr1 knockout (KO) cultured astrocytes with the group-I-mGlu receptor agonist (S)-3,5-Dihydroxyphenylglycine (DHPG) and exposed them to sodium arsenite (NaAsO2), a widely used inducer of SGs formation. In WT cultures the activation of group-I mGlu receptors reduced SGs formation and recruitment of FMRP into SGs, and also attenuated phosphorylation of eIF2α, a key event crucially involved in SGs formation and inhibition of protein synthesis. In contrast, Fmr1 KO astrocytes, which exhibited a lower number of SGs than WT astrocytes, did not respond to agonist stimulation. Interestingly, the mGlu5 receptor negative allosteric modulator (NAM) 2-methyl-6-(phenylethynyl)pyridine (MPEP) antagonized DHPG-mediated SGs reduction in WT and reversed SGs formation in Fmr1 KO cultures. Our findings reveal a novel function of mGlu5 receptor as modulator of SGs formation and open new perspectives for understanding cellular response to stress in FXS pathophysiology

Effect of mofezolac-galactose distance in conjugates targeting cyclooxygenase (COX)-1 and CNS GLUT-1 carrier

Neuroinflammation is the earliest stage of several neurological and neurodegenerative diseases. In the case of neurodegenerative disorders, it takes place about 15â 20 years before the appearance of specific neurodegenerative clinical symptoms. Constitutive microglial COX-1 is one of the pro-inflammatory players of the neuroinflammation. Novel compounds 3, 14 and 15 (Galmof0, Galmof5and Galmof11, respectively) were projected, and their synthetic methodologies developed, by linking by an ester bond, directly or through a C5 or C11 unit linker the highly selective COX-1 inhibitor mofezolac (COXs selectivity index > 6000) to galactose in order to obtain substances capable to cross blood-brain barrier (BBB) and control the CNS inflammatory response. 3, 14 and 15 (Galmofs) were prepared in good to fair yields. Galmof0(3) was found to be a selective COX-1 inhibitor (COX-1 IC50= 0.27 μM and COX-2 IC50= 3.1 μM, selectivity index = 11.5), chemically and metabolically stable, and capable to cross Caco-2 cell monolayer, resembling BBB, probing that its transport is GLUT-1-mediated. Furthermore, Galmof0(3) powerfully inhibits PGE2release higher than mofezolac (1) in LPS-stimulated mouse BV2 microglial cell line, a worldwide recognized neuroinflammation model. In addition, Fingerprints for Ligands and Proteins (FLAP) was used to explain the different binding interactions of Galmofs with the COX-1 active site

Fluorochrome Selection for Imaging Intraoperative Ovarian Cancer Probes

The identification and removal of all gross and microscopic tumor to render the patient disease free represents a huge challenge in ovarian cancer treatment. The presence of residual disease is an independent negative prognostic factor. Herein, we describe the synthesis and the “in vitro” evaluation of compounds as cyclooxygenase (COX)-1 inhibitors, the COX-1 isoform being an ovarian cancer biomarker, each bearing fluorochromes with different fluorescence features. Two of these compounds N-[4-(9-dimethylimino-9H-benzo[a]phenoxazin-5-ylamino) butyl]-2-(3,4-bis(4-methoxyphenyl)isoxazol-5-yl)acetamide chloride (RR11) and 3-(6-(4-(2-(3,4-bis(4-methoxyphenyl)isoxazole-5-yl)acetamido)butyl)amino-6-oxohexyl)-2-[7-(1,3-dihydro-1,1-dimethyl-3-ethyl 2H-benz[e]indolin-2-yl-idene)-1,3,5-heptatrienyl]-1,1-dimethyl-3-(6-carboxilato-hexyl)-1H-benz[e]indolium chloride, 23 (MSA14) were found to be potent and selective inhibitors of cyclooxygenase (COX)-1 “in vitro”, and thus were further investigated “in vivo”. The IC50 values were 0.032 and 0.087 µM for RR11 and 23 (MSA 14), respectively, whereas the COX-2 IC50 for RR11 is 2.4 µM while 23 (MSA14) did not inhibit COX-2 even at a 50 µM concentration. Together, this represented selectivity index = 75 and 874, respectively. Structure-based virtual screening (SBVS) performed with the Fingerprints for Ligands and Proteins (FLAP) software allowed both to differentiate highly active compounds from less active and inactive structures and to define their interactions inside the substrate-binding cavity of hCOX1. Fluorescent probes RR11 and 23 (MSA14), were used for preliminary near-infrared (NIR) fluorescent imaging (FLI) in human ovarian cancer (OVCAR-3 and SKOV-3) xenograft models. Surprisingly, a tumor-specific signal was observed for both tested fluorescent probes, even though this signal is not linked to the presence of COX-1.publishedVersio

Group I metabotropic glutamate receptors: A role in neurodevelopmental disorders?

Group I metabotropic glutamate receptors (mGlu1 and mGlu5) are coupled to polyphosphoinositide hydrolysis and are involved in activity-dependent forms of synaptic plasticity, both during development and in the adult life. Group I mGlu receptors can also regulate proliferation, differentiation, and survival of neural stem/progenitor cells, which further support their role in brain development. An exaggerated response to activation of mGlu5 receptors may underlie synaptic dysfunction in Fragile X syndrome, the most common inherited form of mental retardation. In addition, group I mGlu receptors are overexpressed in dysplastic neurons of focal cortical dysplasia and hemimegaloencephaly, which are disorders of cortical development associated with chronic epilepsy. Drugs that block the activity of group I mGlu receptors (in particular, mGlu5 receptors) are potentially helpful for the treatment of Fragile X syndrome and perhaps other neurodevelopmental disorders. © Humana Press Inc. 2007

Group I metabotropic glutamate receptors: a role in neurodevelopmental disorders?

Group I metabotropic glutamate receptors (mGlu1 and mGlu5) are coupled to polyphosphoinositide hydrolysis and are involved in activity-dependent forms of synaptic plasticity, both during development and in the adult life. Group I mGlu receptors can also regulate proliferation, differentiation, and survival of neural stem/progenitor cells, which further support their role in brain development. An exaggerated response to activation of mGlu5 receptors may underlie synaptic dysfunction in Fragile X syndrome, the most common inherited form of mental retardation. In addition, group I mGlu receptors are overexpressed in dysplastic neurons of focal cortical dysplasia and hemimegaloencephaly, which are disorders of cortical development associated with chronic epilepsy. Drugs that block the activity of group I mGlu receptors (in particular, mGlu5 receptors) are potentially helpful for the treatment of Fragile X syndrome and perhaps other neurodevelopmental disorder

Metabotropic glutamate receptors in glial cells

Glutamate is the major excitatory neurotransmitter in the mammalian central nervous system (CNS) and exerts its actions via a number of ionotropic glutamate receptors/channels and metabotropic glutamate (mGlu) receptors. In addition to being expressed in neurons, glutamate receptors are expressed in different types of glial cells including astrocytes, oligodendrocytes, and microglia. Astrocytes are now recognized as dynamic signaling elements actively integrating neuronal inputs. Synaptic activity can evoke calcium signals in astrocytes, resulting in the release of gliotransmitters, such as glutamate, ATP, and D-serine, which in turn modulate neuronal excitability and synaptic transmission. In addition, astrocytes, and microglia may play an important role in pathology such as brain trauma and neurodegeneration, limiting or amplifying the pathologic process leading to neuronal death. The present review will focus on recent advances on the role of mGlu receptors expressed in glial cells under physiologic and pathologic condition

A metabolomic and systems biology perspective on the brain of the Fragile X syndrome mouse model

Fragile X syndrome (FXS) is the first cause of inherited intellectual disability, due to the silencing of the X-linked Fragile X Mental Retardation 1 gene encoding the RNA-binding protein FMRP. While extensive studies have focused on the cellular and molecular basis of FXS, neither human Fragile X patients nor the mouse model of FXS—the Fmr1-null mouse—have been profiled systematically at the metabolic and neurochemical level to provide a complementary perspective on the current, yet scattered, knowledge of FXS. Using proton high-resolution magic angle spinning nuclear magnetic resonance (1H HR-MAS NMR)-based metabolic profiling, we have identified a metabolic signature and biomarkers associated with FXS in various brain regions of Fmr1-deficient mice. Our study highlights for the first time that Fmr1 gene inactivation has profound, albeit coordinated consequences in brain metabolism leading to alterations in: (1) neurotransmitter levels, (2) osmoregulation, (3) energy metabolism, and (4) oxidative stress response. To functionally connect Fmr1-deficiency to its metabolic biomarkers, we derived a functional interaction network based on the existing knowledge (literature and databases) and show that the FXS metabolic response is initiated by distinct mRNA targets and proteins interacting with FMRP, and then relayed by numerous regulatory proteins. This novel “integrated metabolome and interactome mapping” (iMIM) approach advantageously unifies novel metabolic findings with previously unrelated knowledge and highlights the contribution of novel cellular pathways to the pathophysiology of FXS. These metabolomic and integrative systems biology strategies will contribute to the development of potential drug targets and novel therapeutic interventions, which will eventually benefit FXS patients

Synthesis and Experimental Validation of New Designed Heterocyclic Compounds with Antiproliferative Activity versus Breast Cancer Cell Lines MCF-7 and MDA-MB-231

Recent drug discovery efforts are highly focused towards identification, design, and synthesis of small molecules as anticancer agents. With this aim, we recently designed and synthesized novel compounds with high efficacy and specificity for the treatment of breast tumors. Based on the obtained results, we constructed a Volsurf+ (VS+) model using a dataset of 59 compounds able to predict the in vitro antitumor activity against MCF-7 cancer cell line for new derivatives. In the present paper, in order to further verify the robustness of this model, we report the results of the projection of more than 150 known molecules and 9 newly synthesized compounds. We predict their activity versus MCF-7 cell line and experimentally verify the in silico results for some promising chosen molecules in two human breast cell lines, MCF-7 and MDA-MB-231