Improved homology modeling of the human & rat EP4 prostanoid receptors

Background: The EP₄ prostanoid receptor is one of four GPCRs that mediate the diverse actions of prostaglandin E₂ (PGE₂). Novel selective EP₄ receptor agonists would assist to further elucidate receptor sub-type function and promote development of therapeutics for bone healing, heart failure, and other receptor associated conditions. The rat EP₄ (rEP₄) receptor has been used as a surrogate for the human EP₄ (hEP₄) receptor in multiple SAR studies. To better understand the validity of this traditional approach, homology models were generated by threading for both receptors using the RaptorX server. These models were fit to an implicit membrane using the PPM server and OPM database with refinement of intra and extracellular loops by Prime (Schrödinger). To understand the interaction between the receptors and known agonists, induced-fit docking experiments were performed using Glide and Prime (Schrödinger), with both endogenous agonists and receptor sub-type selective, small-molecule agonists. The docking scores and observed interactions were compared with radioligand displacement experiments and receptor (rat & human) activation assays monitoring cAMP.

Results: Rank-ordering of in silico compound docking scores aligned well with in vitro activity assay EC₅₀ and radioligand binding K_i. We observed variations between rat and human EP₄ binding pockets that have implications in future small-molecule receptor-modulator design and SAR, specifically a S103G mutation within the rEP4 receptor. Additionally, these models helped identify key interactions between the EP₄ receptor and ligands including PGE₂ and several known sub-type selective agonists while serving as a marked improvement over the previously reported models.

Conclusions: This work has generated a set of novel homology models of the rEP₄ and hEP₄ receptors. The homology models provide an improvement upon the previously reported model, largely due to improved solvation. The hEP₄ docking scores correlates best with the cAMP activation data, where both data sets rank order Rivenprost>CAY10684 > PGE₁ ≈ PGE₂ > 11-deoxy-PGE₁ ≈ 11-dexoy-PGE₂ > 8-aza-11-deoxy-PGE₁. This rank-ordering matches closely with the rEP₄ receptor as well. Species-specific differences were noted for the weak agonists Sulprostone and Misoprostol, which appear to DOCK more readily within human receptor versus rat receptor.

Bone healing; EP4; Heart failure; Homology model; PGE2; Prostaglandin E2; Prostaglandin E2 receptor; Prostaglandin E2 receptor subtype 4; Structure-based drug design.