title: In vitro and in vivo pharmacological characterization of SSD114, a novel GABAB positive allosteric modulator.
abstract: Positive allosteric modulators (PAMs) of the GABABreceptor have emerged as a novel approach to the pharmacological manipulation of the GABABreceptor, enhancing the effects of receptor agonists with few side effects. Here, we identified N-cyclohexyl-4-methoxy-6-(4-(trifluoromethyl)phenyl)pyrimidin-2-amine (SSD114) as a new compound with activity as a GABABPAM in in vitro and in vivo assays. SSD114 potentiated GABA-stimulated [35S]GTPγS binding to native GABABreceptors, whereas it had no effect when used alone. Its effect on GTPγS stimulation was suppressed when GABA-induced activation was blocked with CGP54626, a competitive antagonist of the GABABreceptor. SSD114 failed to potentiate WIN55,212,2-, morphine- and quinpirole-induced [35S]GTPγS binding to cortical and striatal membranes, respectively, indicating that it is a selective GABABPAM. Increasing SSD114 fixed concentrations induced a leftward shift of the GABA concentration-response curve, enhancing the potency of GABA rather than its efficacy. SSD114 concentration-response curves in the presence of fixed concentrations of GABA (1, 10, and 20μM) revealed a potentiating effect on GABA-stimulated binding of [35S]GTPγS to rat cortical membranes, with EC50values in the low micromolar range. Bioluminescence resonance energy transfer (BRET) experiments in Chinese Hamster Ovary (CHO)-cells expressing GABABreceptors showed that SSD114 potentiates the GABA inhibition of adenylyl-cyclase mediated by GABABreceptors. Our compound is also effective in vivo potentiating baclofen-induced sedation/hypnosis in mice, with no effect when tested alone. These findings indicate that SSD114, a molecule with a different chemical structure compared to known GABABPAMs, is a novel GABABPAM with potential usefulness in the GABAB-receptor research field.
authors: Department of Biomedical Sciences, Division of Neuroscience and Clinical Pharmacology, University of Cagliari, Cagliari, Italy.
authors: Alessandra
authors: A
authors: Porcu
authors: Neuroscience Institute, National Research Council of Italy, Section of Cagliari, Cagliari, Italy.
authors: Carla
authors: C
authors: Lobina
authors: Biomolecular Chemistry Institute, National Research Council of Italy, Section of Sassari, Sassari, Italy.
authors: Daniela
authors: D
authors: Giunta
authors: Biomolecular Chemistry Institute, National Research Council of Italy, Section of Sassari, Sassari, Italy.
authors: Maurizio
authors: M
authors: Solinas
authors: Department of Biotechnology Chemistry and Pharmacy, University of Siena, Siena, Italy.
authors: Claudia
authors: C
authors: Mugnaini
authors: Department of Biomedical Sciences, Division of Neuroscience and Clinical Pharmacology, University of Cagliari, Cagliari, Italy. Electronic address: castelli@unica.it.
authors: M Paola
authors: MP
authors: Castelli
mesh_terms: D000494:Allosteric Regulation / Q000187:drug effects; D000818:Animals; D001418:Baclofen / Q000494:pharmacology; D016466:CHO Cells; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002540:Cerebral Cortex / Q000166:cytology; D006224:Cricetinae; D003412:Cricetulus; D003514:Cyclohexylamines / Q000378:metabolism / Q000494:pharmacology*; D008297:Male; D051379:Mice; D011743:Pyrimidines / Q000378:metabolism / Q000494:pharmacology*; D051381:Rats; D018080:Receptors, GABA-B / Q000737:chemistry / Q000378:metabolism*; D057897:Reflex, Righting / Q000187:drug effects; D005680:gamma-Aminobutyric Acid / Q000378:metabolism
chemical_list: D003514:Cyclohexylamines; C000616257:N-cyclohexyl-4-methoxy-6-(4-(trifluoromethyl)phenyl)pyrimidin-2-amine; D011743:Pyrimidines; D018080:Receptors, GABA-B; D005680:gamma-Aminobutyric Acid; D001418:Baclofen
keywords: Baclofen-induced sedation/hypnosis; GABA(B) receptors; GTPγS binding; Positive allosteric modulator