TY - JOUR
T1 - Pharmacological characterization of low molecular weight biased agonists at the follicle stimulating hormone receptor
AU - De Pascali, Francesco
AU - Ayoub, Mohammed Akli
AU - Benevelli, Riccardo
AU - Sposini, Silvia
AU - Lehoux, Jordan
AU - Gallay, Nathalie
AU - Raynaud, Pauline
AU - Landomiel, Flavie
AU - Jean-Alphonse, Frédéric
AU - Gauthier, Christophe
AU - Pellissier, Lucie P.
AU - Crépieux, Pascale
AU - Poupon, Anne
AU - Inoue, Asuka
AU - Joubert, Nicolas
AU - Viaud-Massuard, Marie Claude
AU - Casarini, Livio
AU - Simoni, Manuela
AU - Hanyaloglu, Aylin C.
AU - Nataraja, Selva G.
AU - Yu, Henry N.
AU - Palmer, Stephen S.
AU - Yvinec, Romain
AU - Reiter, Eric
N1 - Funding Information:
Funding: This publication was funded with support from the French National Research Agency under the program “Investissements d’avenir” Grant Agreement LabEx MabImprove: ANR-10-LABX-53; “ARD2020 Biomédicaments” and “APR-IR MODUPHAC” grants from Région Centre Val de Loire. F.D.P. was the recipient of a doctoral fellowship from INRAE and Région Centre Val de Loire. R.B. was the recipient of an Erasmus plus fellowship from the University of Modena and Reggio Emilia and the European Union. M.A.A., F.J.-A., and M.S. were LE STUDIUM RESEARCH FELLOWS, Loire Valley Institute for Advanced Studies, Orléans & Tours, France, receiving funding from the European Union’s Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie grant agreement No 665790. A.I. was funded by the PRIME 18gm5910013 and the LEAP 18gm0010004 from the Japan Agency for Medical Research and Development (AMED) and KAKENHI 17K08264 by the Japan Society for the Promotion of Science (JSPS). A.C.H. was supported by a Biotechnology and Biological Sciences Research Council (BBSRC) project grant (BB/S001565/1) and a Society for Endocrinology Early Career Grant to S.S.
Publisher Copyright:
© 2021 by the authors. Licensee MDPI, Basel, Switzerland.
PY - 2021/9
Y1 - 2021/9
N2 - Follicle-stimulating hormone receptor (FSHR) plays a key role in reproduction through the activation of multiple signaling pathways. Low molecular weight (LMW) ligands composed of biased agonist properties are highly valuable tools to decipher complex signaling mechanisms as they allow selective activation of discrete signaling cascades. However, available LMW FSHR ligands have not been fully characterized yet. In this context, we explored the pharmacological diversity of three benzamide and two thiazolidinone derivatives compared to FSH. Concentration/activity curves were generated for Gαs, Gαq, Gαi, β-arrestin 2 recruitment, and cAMP production, using BRET assays in living cells. ERK phosphorylation was analyzed by Western blotting, and CRE-dependent transcription was assessed using a luciferase reporter assay. All assays were done in either wild-type, Gαs or β-arrestin 1/2 CRISPR knockout HEK293 cells. Bias factors were calculated for each pair of read-outs by using the operational model. Our results show that each ligand presented a discrete pharmacological efficacy compared to FSH, ranging from super-agonist for β-arrestin 2 recruitment to pure Gαs bias. Interestingly, LMW ligands generated kinetic profiles distinct from FSH (i.e., faster, slower or transient, depending on the ligand) and correlated with CRE-dependent transcription. In addition, clear system biases were observed in cells depleted of either Gαs or β-arrestin genes. Such LMW properties are useful pharmacological tools to better dissect the multiple signaling pathways activated by FSHR and assess their relative contributions at the cellular and physio-pathological levels.
AB - Follicle-stimulating hormone receptor (FSHR) plays a key role in reproduction through the activation of multiple signaling pathways. Low molecular weight (LMW) ligands composed of biased agonist properties are highly valuable tools to decipher complex signaling mechanisms as they allow selective activation of discrete signaling cascades. However, available LMW FSHR ligands have not been fully characterized yet. In this context, we explored the pharmacological diversity of three benzamide and two thiazolidinone derivatives compared to FSH. Concentration/activity curves were generated for Gαs, Gαq, Gαi, β-arrestin 2 recruitment, and cAMP production, using BRET assays in living cells. ERK phosphorylation was analyzed by Western blotting, and CRE-dependent transcription was assessed using a luciferase reporter assay. All assays were done in either wild-type, Gαs or β-arrestin 1/2 CRISPR knockout HEK293 cells. Bias factors were calculated for each pair of read-outs by using the operational model. Our results show that each ligand presented a discrete pharmacological efficacy compared to FSH, ranging from super-agonist for β-arrestin 2 recruitment to pure Gαs bias. Interestingly, LMW ligands generated kinetic profiles distinct from FSH (i.e., faster, slower or transient, depending on the ligand) and correlated with CRE-dependent transcription. In addition, clear system biases were observed in cells depleted of either Gαs or β-arrestin genes. Such LMW properties are useful pharmacological tools to better dissect the multiple signaling pathways activated by FSHR and assess their relative contributions at the cellular and physio-pathological levels.
KW - Allosteric ligands
KW - Biased signaling
KW - FSHR
KW - Operational model
KW - System bias
UR - http://www.scopus.com/inward/record.url?scp=85114624850&partnerID=8YFLogxK
U2 - 10.3390/ijms22189850
DO - 10.3390/ijms22189850
M3 - Article
C2 - 34576014
AN - SCOPUS:85114624850
SN - 1661-6596
VL - 22
JO - International Journal of Molecular Sciences
JF - International Journal of Molecular Sciences
IS - 18
M1 - 9850
ER -