Antagonism of the prostaglandin E2 EP1 receptor in MDCK cells increases growth through activation of Akt and the epidermal growth factor receptor.

The actions of prostaglandin E2 (PGE2) in the kidney are mediated by G protein-coupled E-prostanoid (EP) receptors, which affect renal growth and function. This report examines the role of EP receptors in mediating the effects of PGE2 on Madin-Darby canine kidney (MDCK) cell growth. The results indicate that activation of Gs-coupled EP2 and EP4 by PGE2 results in increased growth, while EP1 activation is growth inhibitory. Indeed, two EP1 antagonists (ONO-8711 and SC51089) stimulate, rather than inhibit, MDCK cell growth, an effect that is lost following an EP1 knockdown. Similar observations were made with M1 collecting duct and rabbit kidney proximal tubule cells. ONO-8711 even stimulates growth in the absence of exogenous PGE2, an effect that is prevented by ibuprofen (indicating a dependence upon endogenous PGE2). The involvement of Akt was indicated by the observation that 1) ONO-8711 and SC51089 increase Akt phosphorylation, and 2) MK2206, an Akt inhibitor, prevents the increased growth caused by ONO-8711. The involvement of the EGF receptor (EGFR) was indicated by 1) the increased phosphorylation of the EGFR caused by SC51089 and 2) the loss of the growth-stimulatory effect of ONO-8711 and SC51089 caused by the EGFR kinase inhibitor AG1478. The growth-stimulatory effect of ONO-8711 was lost following an EGFR knockdown, and transduction of MDCK cells with a dominant negative EGFR. These results support the hypothesis that 1) signaling via the EP1 receptor involves Akt as well as the EGFR, and 2), EP1 receptor pharmacology may be employed to prevent the aberrant growth associated with a number of renal diseases.

Targeting the EP1 receptor reduces Fas ligand expression and increases the antitumor immune response in an in vivo model of colon cancer.

Despite studies demonstrating that inhibition of cyclooxygenase-2 (COX-2)-derived prostaglandin E2 (PGE2 ) has significant chemotherapeutic benefits in vitro and in vivo, inhibition of COX enzymes is associated with serious gastrointestinal and cardiovascular side effects, limiting the clinical utility of these drugs. PGE2 signals through four different receptors (EP1-EP4) and targeting individual receptor(s) may avoid these side effects, while retaining significant anticancer benefits. Here, we show that targeted inhibition of the EP1 receptor in the tumor cells and the tumor microenvironment resulted in the significant inhibition of tumor growth in vivo. Both dietary administration and direct injection of the EP1 receptor-specific antagonist, ONO-8713, effectively reduced the growth of established CT26 tumors in BALB/c mice, with suppression of the EP1 receptor in the tumor cells alone less effective in reducing tumor growth. This antitumor effect was associated with reduced Fas ligand expression and attenuated tumor-induced immune suppression. In particular, tumor infiltration by CD4(+) CD25(+) Foxp3(+) regulatory T cells was decreased, whereas the cytotoxic activity of isolated splenocytes against CT26 cells was increased. F4/80(+) macrophage infiltration was also decreased; however, there was no change in macrophage phenotype. These findings suggest that the EP1 receptor represents a potential target for the treatment of colon cancer.

Novel prostaglandin receptor modulators--part II: EP receptor modulators; a patent review (2002 - 2012).

INTRODUCTION: Prostaglandins and their G-protein-coupled receptors play numerous physiological and pathophysiological roles, especially in inflammation and its resolution. The variety of effects mediated by prostanoids makes prostanoid receptors valuable drug targets and the research on prostaglandin receptor modulators is intensive. The physiological and pathophysiological effects of prostaglandin E(2) are especially complex and numerous. The four subtypes of EP receptor have gained a lot of industrial and academic interest, in particular EP(2) and EP(4) for various indications. AREAS COVERED: Evaluation of the patent activity over the last decade (2002 - 2012) illustrates several potent compounds targeting the distinct prostaglandin E(2) receptors. Many novel methods for the use of EP receptor modulators have been developed, in addition to the classical indications for agents modulating the arachidonic acid cascade such as pain and inflammation. EXPERT OPINION: Several EP targeting agents with good potency and selectivity have been developed but their pharmacological use and utility has not yet been satisfactorily investigated. More research is necessary, and clinical use of these agents might therefore take some more time.

Antagonism of neuronal prostaglandin E(2) receptor subtype 1 mitigates amyloid ß neurotoxicity in vitro.

Multiple lines of evidence indicate that regional brain eicosanoid signaling is important in initiation and progression of neurodegenerative conditions that have neuroinflammatory pathologic component, such as AD. We hypothesized that PGE(2) receptor subtype 1 (EP1) signaling (linked to intracellular Ca(2+) release) regulates Aß peptide neurotoxicity and tested this in two complementary in vitro models: a human neuroblastoma cell line (MC65) producing Aß(1-40) through conditional expression of the APP C-terminal portion, and murine primary cortical neuron cultures exposed to Aß(1-42). In MC65 cells, EP1 receptor antagonist SC-51089 reduced Aß neurotoxicity ~50 % without altering high molecular weight Aß immunoreactive species formation. Inositol-3-phosphate receptor antagonist 2-aminoethoxy-diphenyl borate offered similar protection. SC-51089 largely protected the neuron cultures from synthetic Aß(1-42) neurotoxicity. Nimodipine, a Ca(2+) channel blocker, was completely neuroprotective in both models. Based on these data, we conclude that suppressing neuronal EP1 signaling may represent a promising therapeutic approach to ameliorate Aß peptide neurotoxicity.

Prostanoid EP1 receptor as the target of (-)-epigallocatechin-3-gallate in suppressing hepatocellular carcinoma cells in vitro.

AIM: To investigate the effects of (-)-epigallocatechin-3-gallate (EGCG), an active compound in green tea, on prostaglandin E(2) (PGE(2))-induced proliferation and migration, and the expression of prostanoid EP(1) receptors in hepatocellular carcinoma (HCC) cells. METHODS: HCC cell line HepG2, human hepatoma cell lines MHCC-97L, MHCC-97H and human hepatocyte cell line L02 were used. Cell viability was analyzed using MTT assay. PGE(2) production was determined with immunoassay. Wound healing assay and transwell filter assay were employed to assess the extent of HCC cell migration. The expression of EP(1) receptor and Gq protein were examined using Western blot assay. RESULTS: PGE(2) (4-40000 nmol/L) or the EP(1) receptor agonist ONO-DI-004 (400-4000 nmol/L) increased the viability and migration of HepG2 cells in concentration-dependent manners. EGCG (100 µg/mL) significantly inhibited the viability and migration of HepG2 cells induced by PGE(2) or ONO-DI-004. HepG2 cells secreted an abundant amount of PGE(2) into the medium, and EGCG (100 µg/mL) significantly inhibited the PGE(2)production and EP(1) receptor expression in HepG2 cells. EGCG (100 µg/mL) also inhibited the viability of MHCC-97L cells, but not that of MHCC-97H cells. Both EGCG (100 µg/mL) and EP(1) receptor antagonist ONO-8711 inhibited PGE(2) 4 µmol/L and ONO-DI-004 400 nmol/L-induced growth and migration of HepG2 cells. Both EGCG (100 µg/mL) and ONO-8711 210 nmol/L inhibited PGE(2)- and ONO-DI-004-induced EP(1) expression. EGCG and ONO-8711 had synergistic effects in inhibiting EP(1) receptor expression. PGE(2), ONO-DI-004, ONO-8711, and EGCG had no effects on Gq expression in HepG2 cells, respectively. CONCLUSION: These findings suggest that the anti-HCC effects of EGCG might be mediated, at least partially, through the suppressing EP(1) receptor expression and PGE(2) production.

Capsaicin-induced vasodilatation in human nasal vasculature is mediated by modulation of cyclooxygenase-2 activity and abrogated by sulprostone.

Extensively based on evidence gained from experimental animal models, the transient receptor potential vanilloid receptor type 1 (TRPV1)-activator capsaicin is regarded as a valuable tool in the research on neurogenic inflammation. Although capsaicin-related drugs gained renewed interest as a therapeutic tool, there is also controversy as whether neurogenic inflammation actually takes place in humans. In this study, we verified the involvement of capsaicin in vascular responses that are regarded to be implicated in the cascade of neurogenic inflammatory mechanisms. By means of ex vivo functional experiments on human nasal mucosal vascular beds, the effect and mechanism of action of capsaicin was assessed in the absence and presence of various agents that interfere with potentially related transduction pathways. Ten micromolars of capsaicin induced vasodilatations that were reduced by the selective EP(1) prostanoid receptor antagonist SC19220 (10 µM) and almost abolished by the selective COX-2 inhibitor NS398 (1 µM) and the EP(1/3) receptor agonist sulprostone (0.1-10 nM), but not affected by the TRPV1-antagonists capsazepine (5 µM), the neurokinin NK(1) receptor antagonist GR20517A (1 µM), and the calcitonin-gene-related peptide (CGRP) receptor antagonist CGRP8-37 (100 nM). Spontaneously released PGE(2) and PGD(2) levels were significantly reduced in the presence of capsaicin. In conclusion, capsaicin-at concentrations clinically applied or under investigation for diverse disease backgrounds-induces a vasodilatory response in human nasal mucosa via a mechanism involving TRPV1-independent reduction of PGE(2) production by modulation of COX-2 enzymatic activity. These vasodilatations can be suppressed by the EP(1/3) receptor agonist sulprostone at subnanomolar concentrations.