Synthesis and Biological Properties of New Constrained CCK-B Antagonists: Discrimination of Two Affinity States of the CCK-B Receptor on Transfected CHO Cells

International audienceTo improve our knowledge of the bioactive conformation of CCK-B antagonists, we have developed a new series of constrained dipeptoids whose synthesis and biochemical properties are reported here. These compounds, of general structure N alpha-[(2-adamantyloxy)carbonyl]-alpha-methyltryptophanyl-(4 -X)-proline, were designed by introducing a cyclization in the structure of the previously described CCK-B/peptoid antagonist RB 210, N-[N-[(2-adamantyloxy)carbonyl]-DL-alpha-methyltryptophanyl] -N-(2-phenylethyl)glycine (Blommaert et al. J. Med. Chem. 1993, 36, 2868-2877), by means of a five-membered ring. Structure-affinity relationship studies showed that an R configuration of Trp-C alpha and a cis configuration of the pyrrolidine substituents were favorable for receptor recognition. The most potent compounds of this new series had similar affinities for the CCK-B receptor as RB 210 and proved to be far more efficient in inhibiting inositol phosphate production in CHO cells stably transfected with rat brain CCK-B receptor, with IC50 values approaching those of the commonly used antagonists L-365,260 and PD-134,308. Moreover, binding studies performed using transfected CHO cells showed that two affinity states of the CCK-B receptor can be discriminated by some of these compounds which also have different biological profiles and are therefore highly interesting tools for the biochemical and pharmacological characterization of CCK-B receptor heterogeneity

Conformational pathway provides unique sensitivity to a synaptic mGluR

International audienceMetabotropic glutamate receptors (mGluRs) are dimeric G-protein-coupled receptors that operate at synapses. Macroscopic and single molecule FRET to monitor structural rearrangements in the ligand binding domain (LBD) of the mGluR7/7 homodimer revealed it to have an apparent affinity~4000-fold lower than other mGluRs and a maximal activation of onlỹ 10%, seemingly too low for activation at synapses. However, mGluR7 heterodimerizes, and we find it to associate with mGluR2 in the hippocampus. Strikingly, the mGluR2/7 hetero-dimer has high affinity and efficacy. mGluR2/7 shows cooperativity in which an unliganded subunit greatly enhances activation by agonist bound to its heteromeric partner, and a unique conformational pathway to activation, in which mGluR2/7 partially activates in the Apo state, even when its LBDs are held open by antagonist. High sensitivity and an unusually broad dynamic range should enable mGluR2/7 to respond to both glutamate transients from nearby release and spillover from distant synapses

Conformational pathway provides unique sensitivity to a synaptic mGluR.

Metabotropic glutamate receptors (mGluRs) are dimeric G-protein-coupled receptors that operate at synapses. Macroscopic and single molecule FRET to monitor structural rearrangements in the ligand binding domain (LBD) of the mGluR7/7 homodimer revealed it to have an apparent affinity ~4000-fold lower than other mGluRs and a maximal activation of only ~10%, seemingly too low for activation at synapses. However, mGluR7 heterodimerizes, and we find it to associate with mGluR2 in the hippocampus. Strikingly, the mGluR2/7 heterodimer has high affinity and efficacy. mGluR2/7 shows cooperativity in which an unliganded subunit greatly enhances activation by agonist bound to its heteromeric partner, and a unique conformational pathway to activation, in which mGluR2/7 partially activates in the Apo state, even when its LBDs are held open by antagonist. High sensitivity and an unusually broad dynamic range should enable mGluR2/7 to respond to both glutamate transients from nearby release and spillover from distant synapses

The mGlu7 receptor provides protective effects against epileptogenesis and epileptic seizures

International audienceFinding new targets to control or reduce seizure activity is essential to improve the management of epileptic patients. We hypothesized that activation of the pre-synaptic and inhibitory metabotropic glutamate receptor type 7 (mGlu7) reduces spontaneous seizures. We tested LSP2-9166, a recently developed mGlu7/4 agonist with unprecedented potency on mGlu7 receptors, in two paradigms of epileptogenesis. In a model of chemically induced epileptogenesis (pentylenetetrazole systemic injection), LSP2-9166 induces an anti-epileptogenic effect rarely observed in preclinical studies. In particular, we found a bidirectional modulation of seizure progression by mGlu4 and mGlu7 receptors, the latter preventing kindling. In the intra-hippocampal injection of kainic acid mouse model that mimics the human mesial temporal lobe epilepsy, we found that LSP2-9166 reduces seizure frequency and hippocampal sclerosis. LSP2-9166 also acts as an anti-seizure drug on established seizures in both models tested. Specific modulation of the mGlu7 receptor could represent a novel approach to reduce pathological network remodeling

Amino Acids Bearing Aromatic or Heteroaromatic Substituents as a New Class of Ligands for the Lysosomal Sialic Acid Transporter Sialin

International audienceSialin, encoded by the SLC17A5 gene, is a lysosomal sialic acid transporter defective in Salla disease, a rare inherited leukodystrophy. It also enables metabolic incorporation of exogenous sialic acids, leading to autoanti-bodies against N-glycolylneuraminic acid in humans. Here, we identified a novel class of human sialin ligands by virtual screening and structure−activity relationship studies. The ligand scaffold is characterized by an amino acid backbone with a free carboxylate, an N-linked aromatic or heteroaromatic substituent, and a hydrophobic side chain. The most potent compound, 45 (LSP12-3129), inhibited N-acetylneuraminic acid 1 (Neu5Ac) transport in a non-competitive manner with IC 50 ≈ 2.5 μM, a value 400-fold lower than the K M for Neu5Ac. In vitro and molecular docking studies attributed the non-competitive character to selective inhibitor binding to the Neu5Ac site in a cytosol-facing conformation. Moreover, compound 45 rescued the trafficking defect of the pathogenic mutant (R39C) causing Salla disease. This new class of cell-permeant inhibitors provides tools to investigate the physiological roles of sialin and help develop pharmacological chaperones for Salla disease

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A group III metabotropic glutamate (mGlu) receptor agonist (PCEP) was identified by virtual HTS. This orthosteric ligand is composed by an l-AP4-derived fragment that mimics glutamate and a chain that binds into a neighboring pocket, offering possibilities to improve affinity and selectivity. Herein we describe a series of derivatives where the distal chain is replaced by an aromatic or heteroaromatic group. Potent agonists were identified, including some with a mGlu₄ subtype preference, e.g., <b>17m</b> (LSP1-2111) and <b>16g</b> (LSP4-2022). Molecular modeling suggests that aromatic functional groups may bind at either one of the two chloride regulatory sites. These agonists may thus be considered as particular bitopic/dualsteric ligands. <b>17m</b> was shown to reduce GABAergic synaptic transmission at striatopallidal synapses. We now demonstrate its inhibitory effect at glutamatergic parallel fiber–Purkinje cell synapses in the cerebellar cortex. Although these ligands have physicochemical properties that are markedly different from typical CNS drugs, they hold significant therapeutic potential