The Current Status of Drug Discovery for the Oxytocin Receptor

Structural basis for the activation and ligand recognition of the human oxytocin receptor

“…Forty-eight hours post-transfection the cells were labelled with 50 nM SNAP-Lumi4-Tb (Cisbio) in labelling buffer (20 mM Hepes (pH 7.5), 100 mM NaCl, 3 mM MgCl 2 , and 0.05% (w/v) bovine serum albumin (BSA)) for 1.5 hours at 37°C. The cells were washed two times with labelling buffer and two times with assay buffer (20 mM Hepes (pH 7.5), 100 mM KCl, 3 mM MgCl 2, and 0.05% (w/v) BSA)) and subsequently incubated for 1 h at room temperature with a concentration range of fluorescently labelled peptide HiLyte Fluor 488-Orn 8 oxytocin (Eurogentec) in assay buffer. Fluorescence intensities were measured on a Spark fluorescence plate reader with an excitation wavelength of 340 nm and emission wavelengths of 620 nm and 510 nm for Tb 3+ and the fluorophore HiLyte Fluor 488, respectively.…”

“…Currently, several clinical trials are evaluating the efficacy of OTR-mediated signalling through administration of OT to treat malfunctions such as autism-spectrum disorders 3 , anxiety 4 and schizophrenia 5 . While OT itself is an approved peripheral drug in obstetrics 6 , OT-based treatments of socio-behavioural deficiencies, requiring central administration of the hormone, have yet failed, potentially due to its poor drug-like properties 7 and limited penetration through the blood brain barrier 8 . Despite the high demand for such drugs, the identification and development of OTR-specific molecules with satisfactory pharmacokinetic properties and favourable biodistribution has been impeded by the lack of structural information on the OTR:OT signalling complex.…”

“…Interestingly, the amidated C-terminus of Gly 9 , known to be important for activation 13 , is located in the proximity of residues E42 1.35 and D100 2.65 (numbers in superscripts correspond to Ballesteros-Weinstein numbering) of transmembrane helices I and II, which are involved in magnesium-dependent modulation of OT binding 14 . Together with the neighbouring Leu 8 , the extracellular surface of the orthosteric pocket is lined by Pro 7 and Asn 5 , which pack against extracellular loop 3 (ECL3) and ECL2, respectively. Leu 8 is oriented towards the extracellular space, explaining why position 8 is best suited for the attachment of fluorophores in OT 15 .…”

“…Together with the neighbouring Leu 8 , the extracellular surface of the orthosteric pocket is lined by Pro 7 and Asn 5 , which pack against extracellular loop 3 (ECL3) and ECL2, respectively. Leu 8 is oriented towards the extracellular space, explaining why position 8 is best suited for the attachment of fluorophores in OT 15 . Gln 4 is the only residue pointing out perpendicularly from the ring plane and stabilizes the cyclic ring position through a hydrogen bond to Q295 6.55 .…”

“…The largest structural differences are found for positions eight and nine of the ligand in the tripeptide C-terminus. Leu 8 and Gly 9 in OT are located along the ring plane, with Gly 9 facing helix I (Figs. 1d and 2b and Supplementary Fig.…”