Identification and pharmacological characterization of the prostaglandin FP receptor and FP receptor variant complexes

Abstract

Background and purpose: A prostamide analogue, bimatoprost, has been shown to be effective in reducing intraocular pressure, but its precise mechanism of action remains unclear. Hence, to elucidate the molecular mechanisms of this effect of bimatoprost, we focused on pharmacologically characterizing prostaglandin FP receptor (FP) and FP receptor variant (altFP) complexes. Experimental approach: FP receptor mRNA variants were identified by reverse transcription-polymerase chain reaction. The FP-altFP4 heterodimers were established in HEK293/EBNA cells co-expressing FP and altFP4 receptor variants. A fluorometric imaging plate reader was used to study Ca 2 þ mobilization. Upregulation of cysteine-rich angiogenic protein 61 (Cyr61) mRNA was measured by Northern blot analysis, and phosphorylation of myosin light chain (MLC) by western analysis. Key results: Six splicing variants of FP receptor mRNA were identified in human ocular tissues. Immunoprecipitation confirmed that the FP receptor is dimerized with altFP4 receptors in HEK293/EBNA cells co-expressing FP and altFP4 receptors. In the studies of the kinetic profile for Ca 2 þ mobilization, prostaglandin F 2a (PGF 2a ) elicited a rapid increase in intracellular Ca 2 þ followed by a steady state phase. In contrast, bimatoprost elicited an immediate increase in intracellular Ca 2 þ followed by a second phase. The prostamide antagonist, AGN211335, selectively and dose-dependently inhibited the bimatoprost-initiated second phase of Ca 2 þ mobilization, Cyr61 mRNA upregulation and MLC phosphorylation, but did not block the action of PGF 2a . Conclusion and implications: Bimatoprost lacks effects on the FP receptor but may interact with the FP-altFP receptor heterodimer to induce alterations in second messenger signalling. Hence, FP-altFP complexes may represent the underlying basis of bimatoprost pharmacology.

Anandamide-derived Prostamide F2α Negatively Regulates Adipogenesis

“…Although PGF 2␣ does not activate such heterodimers, PGF 2␣ EA is nearly inactive at the wild-type FP receptor homodimers (24 -28). PGF 2␣ EA pharmacology has been mostly investigated through the use of its synthetic structural analog bimatoprost (37), which has been approved for the treatment of glaucoma and eyelash hypotrichosis (56,57), and of antagonists specific for the FP/FP splice variant heterodimer versus FP homodimers, such as AGN211335 (28).…”

“…Interestingly, however, bimatoprost (AGN 192024), a stable, synthetic, and potent analog of PGF 2␣ EA approved for the treatment of ocular hypertension and also capable of activating the FP/FP splice variant heterodimers (28,37), decreases periorbital fat deposits in glaucoma patients through an unknown mechanism (38 -41). This observation may suggest that bimatoprost and, by extension, PGF 2␣ EA inhibit adipogenesis, perhaps through the stimulation of the aforementioned heterodimers.…”

“…PGF 2␣ EA and bimatoprost signal through prostamide receptors, which are splice variants of the FP receptor encoded by the PTGFR gene responsible for PGF 2␣ signaling (28). To date, there has been no report of the expression of these variants in adipose tissue.…”

“…Prostamide Receptor Antagonism Reverses Prostamide-mediated Inhibition of Adipogenesis-We next tested the ability of bimatoprost to inhibit adipogenesis in 3T3-L1 preadipoctyes in the absence or presence of the prostamide receptor antagonist AGN211335, which specifically blocks bimatoprost/prostamide activity at 1 M in HEK293 cells and not PGF 2␣ activity (28) (Fig. 4A).…”

Prostaglandin E2-Glyceryl Ester: In Vivo Evidence for a Distinct Pharmacological Identity from Intraocular Pressure Studies

“…The pharmacology of these resultant electrochemically neutral prostanoids is gradually being elucidated. Greater attention has been applied to the PG-ethanolamides (prostamides), where the pharmacology has been characterized by the prostamide F 2a analog bimatoprost, selective antagonists, and structural elucidation of the receptor (Woodward et al, 2003(Woodward et al, , 2007Liang et al, 2008). PG-glyceryl ester pharmacology, by comparison, is in its early infancy.…”