In vitro activity: CTEP inhibits quisqualate-induced Ca2+ mobilization with an IC50 of 11.4 nM and [3H]IP accumulation with an IC50 of 6.4 nM in HEK293 cells stably expressing human mGlu5. CTEP inhibits the constitutive activity of human mGlu5 by approximately 50% with an IC50 of 40.1 nM in HEK293 cells stably expressing human mGlu5.

Kinase Assay: For all filtration radioligand binding assays, membrane preparations expressing the target receptors or receptor combinations are resuspended in radioligand binding buffer (15 mM Tris-HCl, 120 mM NaCl, 5 mM KCl, 1.25 mM CaCl2, and 1.25 mM MgCl2, pH 7.4), and the membrane suspension is mixed with the appropriate concentrations of radioligand and nonlabeled drugs in 96-well plates in a total volume of 200 μL and incubated for 60 min at the appropriate temperature. At the end of the incubation, membranes are filtered onto Whatman Unifilter preincubated with 0.1% polyethyleneimine in wash buffer (50 mM Tris-HCl, pH 7.4) with a Filtermate 196 harvester and washed three times with ice-cold wash buffer. Radioactivity captured on the filter is quantified on a Topcount microplate scintillation counter with quenching correction after the addition of 45 μL of MicroScint 40 per well and shaking for 20 min. The concentration of membranes and incubation time is determined for each assay in pilot experiments.

Cell Assay: In HEK293 cells expressing mGlu5, CTEP inhibits quisqualate-induced Ca2+ mobilization and inositol phosphate accumulation with IC50 value of 11.4nM and 6.4nM, respectively. In addition, it shows an IC50 value of 40.1nM in the IP accumulation assay, demonstrating its inverse agonist activity. CTEP is proved to be a highly selective inhibitor of mGlu5. It shows no significant activity against mGlu1, mGlu2, mGlu3, mGlu4, mGlu6, mGlu7 or mGlu8 at concentration up to 10μM.

J Pharmacol Exp Ther. 2011 Nov;339(2):474-86; Neuron. 2012 Apr 12;74(1):49-56.