Genetic overlap in Kallmann syndrome, combined pituitary hormone deficiency, and septo-optic dysplasia

Abstract

Context: Kallmann syndrome (KS), combined pituitary hormone deficiency (CPHD), and septo-optic dysplasia (SOD) all result from development defects of the anterior midline in the human forebrain.

Objective: The objective of the study was to investigate whether KS, CPHD, and SOD have shared genetic origins.

Design and participants: A total of 103 patients with either CPHD (n = 35) or SOD (n = 68) were investigated for mutations in genes implicated in the etiology of KS (FGFR1, FGF8, PROKR2, PROK2, and KAL1). Consequences of identified FGFR1, FGF8, and PROKR2 mutations were investigated in vitro.

Results: Three patients with SOD had heterozygous mutations in FGFR1; these were either shown to alter receptor signaling (p.S450F, p.P483S) or predicted to affect splicing (c.336C>T, p.T112T). One patient had a synonymous change in FGF8 (c.216G>A, p.T72T) that was shown to affect splicing and ligand signaling activity. Four patients with CPHD/SOD were found to harbor heterozygous rare loss-of-function variants in PROKR2 (p.R85G, p.R85H, p.R268C).

Conclusions: Mutations in FGFR1/FGF8/PROKR2 contributed to 7.8% of our patients with CPHD/SOD. These data suggest a significant genetic overlap between conditions affecting the development of anterior midline in the human forebrain.

Trial registration: ClinicalTrials.gov NCT00494169.

Figures

Fig. 1.

A, Western blotting analysis showing similar overall expression and maturation levels of mutant FGFR1 compared with WT. FGFR1 was detected using an anti-myc antibody and the blot was reprobed with heat-shock protein 90 to demonstrate equal loading. UT, Untreated; PG, PNGase F treated; EH, endoglycosidase H treated; HSP90, heat-shock protein 90. B, Luciferase reporter assay using the osteocalcin FGF response elements reporter showing decreased signaling activity by FGFR1 S450F and FGFR1 P483S. ***, P < 0.001 at maximal dose. C, Quantitative PCR showing that expression of both the FGF8e and FGF8f isoforms is significantly increased compared with WT. ***, P < 0.001; *, P < 0.05). D, Representative experiment of a transcriptional reporter (AP1-luciferase) assay of FGF8 WT and T72T mutant minigene showing increased signaling activity by the mutant ligand compared with WT. *, P < 0.05 at maximal dose. E, Western analysis showing that overall expression of the PROKR2 mutant (R85G) is reduced compared with WT. F, Expression of the PROKR2 R85G mutant at the cell surface is significantly reduced compared with WT. *, P < 0.05 in a radiolabeled antibody binding assay. G, The PROKR2 R85G mutant receptor is loss of function in the aequorin reporter assay. ***, P < 0.001. H, Prokr2 is expressed in the adult mouse pituitary. Immunofluorescent staining for GFP in the pituitary of Prokr2-GFP transgenic mice shows immunoreactivity in all pituitary structures and is most pronounced in the pars nervosa. AVP, Arginine vasopressin.