OR03-01 Functional Rescue Of Trafficking-Defective Mutant Follicle-Stimulating Hormone Receptors (FSHR) Leading To Premature Ovarian Failure With The Pharmacoperone CAN1405

Disclosure: A. Ulloa-Aguirre: None. T. Zariñán: None. R. Gutiérrez-Sagal: None. S. Nataraja: None. H.N. Yu: None. E. Jardón-Valadez: None. Inactivating mutations in the follicle-stimulating hormone (FSH) receptor (FSHR) cause premature ovarian failure. Approximately 30% of mutant FSHRs (mutFSHR) are misfolded proteins unable to route to the plasma membrane (PM). We herein tested the effect of a small molecule FSHR ago-allosteric modulator (CAN1405) on the functional rescue of several inactivating mutFSHR. Wild-type (WT) and mutFSHRs A189V, A191I and D224V (at the ectodomain), and four FSHRs with mutations in the serpentine region (SR) of the receptor (D408Y, A419I, I423T, and P519T) were expressed in HEK-293 cells and stimulated with recombinant FSH in the presence of vehicle or CAN1405. Changes in plasma membrane expression and function provoked by CAN1405 exposure were analyzed by immunoblotting and by activation of the cAMP-sensitive pSOMLuc reporter plasmid and ERK1/2 phosphoryation, respectively. Molecular interactions between FSHR and CAN1405, were predicted in silico by computational docking. Exposure of WT and mutFSHR-expressing cells to CAN1405 increased the ratio of mature (PM-expressed) to immature (intracellular) FSHR forms (m/i ratio) of all SR mutant FSHRs (from 0.5 to 3.0) as disclosed by immunoblotting. Similar results were observed when activation of the cAMP-sensitive pSOMLuc reporter and ERK1/2 phosphorylation were examined in FSH-stimulated cells. PM expression of FSHR WT and the activation-defective mutFSHR I423T, but not the FSHRs with mutations in the ectodomain, were also increased by CAN1405. Interesting, in the presence of CAN1405, signaling of the mutFSHR I423T was marginal in terms of activation of the pSOMLuc reporter but not of pERK1/2, suggesting that binding of CAN1405 to the mutFSHR changed the conformation of the receptor allowing functional selectivity on FSH-stimulated signaling. CAN1405 was located in a binding pocket at interhelical upper half of the FSHR. Side chains of residues Y530, A450, T449 S453, L415, H615, M585, V612, I588, and K608 were detected in close interaction with the CAN1045 and the FSHR-CAN1405 complex was likely stabilized by hydrophobic interactions. It is concluded that the small molecule CAN1405 with ago-allosteric properties is an effective pharmacological chaperone that corrects trafficking of SR misfolded mutFSHRs and also increases WT FSHR expression and function. These data demonstrate that allosteric modulators of the FSHR may serve as an effective therapeutic approach for the treatment of infertility in women bearing the WT FSHR or in heterozygous carriers of mutations at the transmembrane domain of this receptor. Presentation: Thursday, June 15, 2023