18 research outputs found
Label-Free Phenotypic Profiling Identified D-Luciferin as a GPR35 Agonist
Fluorescent and luminescent probes are essential to both in vitro molecular assays and in vivo imaging techniques, and have been extensively used to measure biological function. However, little is known about the biological activity, thus potential interferences with the assay results, of these probe molecules. Here we show that D-luciferin, one of the most widely used bioluminescence substrates, is a partial agonist for G protein-coupled receptor-35 (GPR35). Label-free phenotypic profiling using dynamic mass redistribution (DMR) assays showed that D-luciferin led to a DMR signal in native HT-29 cells, whose characteristics are similar to those induced by known GPR35 agonists including zaprinast and pamoic acid. DMR assays further showed that D-luciferin is a partial agonist competitive to several known GPR35 agonists and antagonists. D-luciferin was found to cause the phosphorylation of ERK that was suppressed by known GPR35 antagonists, and also result in β-arrestin translocation signal but with low efficacy. These results not only suggest that D-luciferin is a partial agonist of GPR35, but also will evoke careful interpretation of biological data obtained using molecular and in vivo imaging assays when these probe molecules are used
Development of Chromen-4-one Derivatives as (Ant)agonists for the Lipid-Activated G Protein-Coupled Receptor GPR55 with Tunable Efficacy
Identification of Crucial Amino Acid Residues Involved in Agonist Signaling at the GPR55 Receptor
A Cell-Based High-Content Screening Assay Reveals Activators and Inhibitors of Cancer Cell Invasion
ERN1 and ALPK1 inhibit differentiation of bi-potential tumor-initiating cells in human breast cancer
Targeting of the Orphan Receptor GPR35 by Pamoic Acid: A Potent Activator of Extracellular Signal-Regulated Kinase and β-Arrestin2 with Antinociceptive ActivityS⃞
Known agonists of the orphan receptor GPR35 are kynurenic acid, zaprinast,
5-nitro-2-(3-phenylproplyamino) benzoic acid, and lysophosphatidic acids. Their
relatively low affinities for GPR35 and prominent off-target effects at other
pathways, however, diminish their utility for understanding GPR35 signaling and for
identifying potential therapeutic uses of GPR35. In a screen of the Prestwick Library
of drugs and drug-like compounds, we have found that pamoic acid is a potent GPR35
agonist. Pamoic acid is considered by the Food and Drug Administration as an inactive
compound that enables long-acting formulations of numerous drugs, such as the
antihelminthics oxantel pamoate and pyrantel pamoate; the psychoactive compounds
hydroxyzine pamoate (Vistaril) and imipramine pamoate (Tofranil-PM); and the peptide
hormones triptorelin pamoate (Trelstar) and octreotide pamoate (OncoLar). We have
found that pamoic acid induces a Gi/o-linked, GPR35-mediated increase in
the phosphorylation of extracellular signal-regulated kinase 1/2, recruitment of
β-arrestin2 to GPR35, and internalization of GPR35. In mice, it
attenuates visceral pain perception, indicating an antinociceptive effect, possibly
through GPR35 receptors. We have also identified in collaboration with the
Sanford-Burnham Institute Molecular Libraries Probe Production Center new classes of
GPR35 antagonist compounds, including the nanomolar potency antagonist
methyl-5-[(tert-butylcarbamothioylhydrazinylidene)methyl]-1-(2,4-difluorophenyl)pyrazole-4-carboxylate
(CID2745687). Pamoic acid and potent antagonists such as CID2745687 present novel
opportunities for expanding the chemical space of GPR35, elucidating GPR35
pharmacology, and stimulating GPR35-associated drug development. Our results indicate
that the unexpected biological functions of pamoic acid may yield potential new uses
for a common drug constituent
Identification of the GPR55 Antagonist Binding Site Using a Novel Set of High-Potency GPR55 Selective Ligands
GPR55 is a class A G protein-coupled
receptor (GPCR) that has been
implicated in inflammatory pain, neuropathic pain, metabolic disorder,
bone development, and cancer. Initially deorphanized as a cannabinoid
receptor, GPR55 has been shown to be activated by non-cannabinoid
ligands such as l-α-lysophosphatidylinositol (LPI).
While there is a growing body of evidence of physiological and pathophysiological
roles for GPR55, the paucity of specific antagonists has limited its
study. In collaboration with the Molecular Libraries Probe Production
Centers Network initiative, we identified a series of GPR55 antagonists
using a β-arrestin, high-throughput, high-content screen of
∼300000 compounds. This screen yielded novel, GPR55 antagonist
chemotypes with IC<sub>50</sub> values in the range of 0.16–2.72
μM [Heynen-Genel, S., et al. (2010) Screening for Selective
Ligands for GPR55: Antagonists (ML191, ML192, ML193) (Bookshelf ID
NBK66153; PMID entry 22091481)]. Importantly, many of the GPR55 antagonists
were completely selective, with no agonism or antagonism against GPR35,
CB1, or CB2 up to 20 μM. Using a model of the GPR55 inactive
state, we studied the binding of an antagonist series that emerged
from this screen. These studies suggest that GPR55 antagonists possess
a head region that occupies a horizontal binding pocket extending
into the extracellular loop region, a central ligand portion that
fits vertically in the receptor binding pocket and terminates with
a pendant aromatic or heterocyclic ring that juts out. Both the region
that extends extracellularly and the pendant ring are features associated
with antagonism. Taken together, our results provide a set of design
rules for the development of second-generation GPR55 selective antagonists