Mutations In Fibroblast Growth Factor 23 Research Articles

Bone characteristics of autosomal dominant hypophosphatemic rickets patients.

Autosomal dominant hypophosphatemic rickets (ADHR) is a rare disease caused by activating mutations in fibroblast growth factor 23 (FGF23) gene. With FGF23 activation, ADHR is a good model to explore the effects of FGF23 on skeletal development and mineralization. However, the bone microarchitecture of ADHR patients is poorly investigated. This study aims to illustrate the bone properties of ADHR patients and clarify the effect of FGF23 on load bearing and non-load bearing bone. Bone microarchitectures of 11 ADHR subjects and sex- and age-matched healthy controls were analyzed by HR-pQCT. The effect of FGF23 mutations on load bearing and non-load bearing bone was explored by comparison of bone microarchitecture in distal radius and distal tibia. The BMD, bone microarchitecture and bone strength were compared between 7 ADHR patients and 7 age- and sex-matched XLH patients. Among 11 subjects with FGF23 mutations, 10 patients presented with obvious symptoms, five of which had received 1-3years of iron supplement, neutral phosphate, and calcitriol treatments. The symptomatic patients presented with low bone density and fractures in X rays, with decreased Z score of aBMD (L1-L4: -1.3±1.4, femoral neck: -2.1±1.8, total hip: -1.85±1.6). Compared with controls, HR-pQCT analysis of 5 untreated ADHR patients showed increased total area (+61.6%, p=0.03) and cortical perimeter (+17.2%, p=0.03) in distal radius. No significant differences were found in other parameters in distal radius. In distal tibia, the patients presented obvious defects in cancellous bone, with decreased trabecular vBMD (-62.9%, p=0.003), trabecular BV/TV (-48.7%, p=0.003) and trabecular number (-42.2%, p=0.001). The trabecular separation (+113.3%, p=0.007) and trabecular network inhomogeneity (+226.7%, p=0.001) were accordingly increased. In addition to another 5 treated patients, the bone microarchitecture changes revealed similar pattern, but the increase of total area and cortical perimeter in distal radius was no longer statistically significant. The non-symptomatic ADHR patient demonstrated slightly decreased total vBMD, trabecular vBMD and trabecular BV/TV in distal tibia. The changing pattern of bone geometry and microarchitecture of ADHR patients were similar to XLH patients but showed less deficit and stronger bone strength. ADHR patients presented increased total area and cortical perimeter in distal radius, and obvious defect in cancellous bone in distal tibia. FGF23 have impairment effect on trabecular bone especially in weight bearing site.

Defective O-glycosylation of novel FGF23 mutations in a Chinese family with hyperphosphatemic familial tumoral calcinosis.

Hyperphosphatemic familial tumoral calcinosis/hyperostosis-hyperphosphatemia syndrome (HFTC/HHS) is a rare disorder caused by deficiency or resistance of fibroblast growth factor 23 (FGF23). Here we reported a Chinese family with HFTC/HHS, aiming at clarifying the clinical features, bone microarchitectures and molecular mechanisms of the disease. Clinical manifestations, laboratory examinations and genetic analyses were collected from two HFTC patients. Bone microarchitectures were detected by HR-pQCT. In vitro expression and glycosylation of mutant and wild-type FGF23 proteins were analyzed by western blotting and wheat germ agglutinin affinity chromatography. Subcellular localizations of FGF23 proteins were detected by immunocytochemistry. The two brothers carried previously unreported c.413T>G, p.Leu138Arg and c.491T>A, p.Ile164Asn compound heterozygous variants in the FGF23 gene, which was "likely pathogenic" according to American College of Medical Genetics (ACMG) Standards and Guidelines. Both patients had severe hyperphosphatemia and significantly elevated C-terminal FGF23. With HHS, patient 1 presented with lower extremity pain and widespread cardiovascular calcification. HR-pQCT of his distal radius and tibia revealed decreased volume BMD and cortical thickness, which were inconsistent with hyperostosis manifestations in X-ray. He received etidronate treatment, which improved his BMD and the ectopic calcification. His brother exhibited less bone involvement but had experienced recurrent painful calcified mass from a young age and undergone several resections. In vitro experiments showed that the mutant FGF23 proteins had defective O-glycosylation and impaired secretion. However, no difference in subcellular localization was found between the wild-type and mutant FGF23 proteins. We have presented a Chinese HFTC/HHS family with novel FGF23 c.413T>G, p.Leu138Arg and c.491T>A, p.Ile164Asn variants. We clarified the bone microarchitectures of HFTC/HHS patients by HR-pQCT, and expanded the genotype-phenotype spectrum of the disease. In vivo studies suggested that O-glycosylation of FGF23 plays an important role in the pathogenesis of HFTC/HHS, providing further understanding of the disease mechanism.

Oral Iron for Prevention and Treatment of Rickets and Osteomalacia in Autosomal Dominant Hypophosphatemia.

ABSTRACT Autosomal dominant hypophosphatemia (ADH) causes rickets, osteomalacia, and taurodontism due to heterozygous mutations in FGF23, which inhibit the inactivation (cleavage) of the encoded protein, the hormone fibroblast growth factor 23 (FGF23). Iron deficiency increases FGF23 mRNA expression and recent evidence suggests that the recurrent, late-onset, or waxing-waning hypophosphatemic phenotype may be linked to synchronous variations in iron status. The fact that most adult symptomatic ADH patients are females during reproductive age supports the notion of a gene-environmental interaction. Practically all symptomatic hypophosphatemic patients described in the recent literature were also iron deficient (with/without anemia) at presentation, when measured. Given its interaction with FGF23, correcting iron deficiency should therefore also correct FGF23 excess. Following the original report of successful phenotype reversal in an iron-deficient ADH child using oral iron supplementation in 2015, more evidence has emerged that supports the use of the element iron to restore homoeostasis of the element phosphorus (in addition to its own). We put into perspective the recent evidence and add 14 years observational data on the original case that demonstrates the correlation of serum phosphorus and renal tubular phosphate reabsorption in mass per unit volume of glomerular filtrate (TmP/GFR) with serum ferritin. Presentation and relapse of ADH, 12 years apart, occurred during iron deficiency, and the onset of menstrual periods was associated with relapse. Here we propose management guidance for patients affected by ADH throughout the lifespan based on iron stores. Because ferritin correlates best with hypophosphatemia historically, and in long-term observation of the originally treated case, it should be used as the monitoring tool and kept in the normal range. Women with ADH who are of reproductive age and other risk groups require supplementation with oral iron using WHO guidelines. Treatment of this form of FGF23 excess may not require phosphate and active vitamin D, or burosumab. © 2020 American Society for Bone and Mineral Research

Development and Validation of a Simple Diagnostic Method to Detect Gain and Loss of Function Defects in Fibroblast Growth Factor-23

Background: Fibroblast growth factor-23 (FGF23) is a bone-derived hormone that regulates the homeostasis of phosphate and vitamin D. Three substitutions in the hormone are reported to cause autosomal dominant hypophosphatemic rickets and seven substitutions to cause autosomal recessive hyperphosphatemic familial tumoral calcinosis (HFTC). Both disorders are rare in the general population and occur most often in the Eastern Mediterranean region and Africa. None of the mutations could be identified using standard restriction fragment length polymorphism. The only technique currently available to confirm the clinical diagnosis is DNA sequencing. Methods: Using a tri-primer ARMS-PCR, in vitro site-directed mutagenesis and DNA sequencing, we developed, verified and validated a rapid and reliable diagnostic test for the ten mutations in FGF23. Results: We generated a test for all ten mutations and confirmed each test by DNA sequencing. We increased the specificity of the test by introducing a mismatch at position -2 in the 3′-terminus of the reverse primer of the normal and the mutant sequences. Finally, using DNA sequencing, we validated the technique for FGF23/S129F substitution by testing samples from 80 individuals from two unrelated Arab families harboring HFTC. Conclusions: This inexpensive and specific method could be adopted where DNA sequencing is not available or affordable.

Recent advance in FGF23 – clinical perspectives

Fibroblast growth factor 23 (FGF23) is a circulating factor that plays a central role in the renal reabsorption of Pi and metabolism of vitamin D. It is mainly produced by osteocytes in bone and exerts its effects on distant organs such as the kidney and parathyroid in an endocrine fashion. FGF23 increases renal Pi excretion by reducing the expression of type 2a and 2c sodium/phosphate (Na+/Pi) co-transporters in the proximal tubules. In addition, it decreases the renal production of 1,25-dihydroxyvitamin D [1,25(OH)2D] by suppressing the expression of 25-hydroxyvitamin D-1a-hydroxylase and increasing that of 25-hydroxyvitamin D-24-hydroxylase. FGF23 requires a-Klotho as well as FGF receptor to exert its effects. FGF23 consists of 251 amino acids. Loss of function mutations in FGF23 cause familial tumoral calcinosis, which is characterized by hyperphosphatemia and increased levels of 1,25(OH)2D. Autosomal dominant hereditary hypophosphatemic rickets (ADHR) is caused by mutations in the cleavage RXXR motif, which prevent inactivation of FGF23. Excess of FGF23 action results in renal phosphate wasting and an inappropriately low level of serum 1,25(OH)2D. Recent studies have implicated the iron-deficiency in the late manifestation of ADHR. Tumor-induced osteomalacia is an acquired paraneoplastic syndrome of renal phosphate wasting caused by overproduction of FGF23 by tumors. Among hereditary hypophosphatemic rickets/osteomalacia, X-linked hypophosphatemic rickets (XLH) is the most common form and is caused by deletion or inactivating mutations of the phosphate-regulating gene homologous to endopeptidase on X chromosome (PHEX). Autosomal recessive hypophosphatemic rickets type I and type II are caused by inactivating mutations of dentin matrix protein 1 (DMP1) and ectonucreotide pyrophosphatase/phosphodiesterase 1 (ENPP1), respectively. In these conditions, serum FGF23 levels are increased, which is responsible for the renal phosphate wasting and impaired vitamin D metabolism. FAM20C is a secreted kinase that phosphorylates extracellular proteins including DMP1. Recently, inactivating mutations in FAM20C have been reported in patients with increased levels of FGF23 and hypophosphatemia. PHEX, DMP1 and FAM20C as well as FGF23 are highly expressed in osteocytes, indicating the critical role of osteocytes in mineral metabolism. Hypophosphatemic rickets/osteomalacia caused by the increased bioactivity of FGF23 are classified into FGF23-mediated hypophosphatemic rickets/osteomalacia. Measurement of serum levels of FGF23 is useful for diagnosis of these conditions. Neutralization of FGF23 will be a potential treatment for XLH and other FGF23-mediated hypophosphatemic rickets/osteomalacia.

Genetic diseases of renal phosphate handling.

Renal control of systemic phosphate homeostasis is critical as evident from inborn and acquired diseases causing renal phosphate wasting. At least three transport proteins are responsible for renal phosphate reabsorption: NAPI-IIa (SLC34A1), NAPI-IIc (SLC34A3) and PIT-2 (SLC20A2). These transporters are highly regulated by various cellular mechanisms and factors including acid-base status, electrolyte balance and hormones such as dopamine, glucocorticoids, growth factors, vitamin D3, parathyroid hormone and fibroblast growth factor 23 (FGF23). Whether renal phosphate wasting is caused by inactivating mutations in the NAPI-IIa transporter is controversial. Mutations in the NAPI-IIc transporter cause hereditary hypophosphatemic rickets with hypercalciuria. Besides the primary inherited defects, there are also inherited defects in major regulators of phosphate homeostasis that lead to alterations in phosphate handling. Autosomal dominant hypophosphatemic rickets is due to FGF23 mutations leading to resistance against its own degradation. Similarly, inactivating mutations in the PHEX gene, which causes FGF23 inactivation, cause X-linked hypophosphatemia due to renal phosphate losses. In contrast, mutations in galactosamine:polypeptide N-acetyl-galactosaminyltransferase, responsible for O-glycosylation of FGF23, or in klotho, a cofactor for FGF23 signalling result in hyperphosphatemia. Acquired syndromes of renal phosphate wasting, hypophosphatemia and osteomalacia (tumour-associated osteomalacia) can be due to the excessive synthesis or release of phosphaturic factors (FGF23, FGF-7, MEPE and sFRP4) from mesenchymal tumours. bone, FGF23, kidney, phosphate, PTH.

Coupling fibroblast growth factor 23 production and cleavage: iron deficiency, rickets, and kidney disease.

High levels of fibroblast growth factor 23 (FGF23) cause the rare disorders of hypophosphatemic rickets and are a risk factor for cardiovascular disease and death in patients with chronic kidney disease (CKD). Despite major advances in understanding FGF23 biology, fundamental aspects of FGF23 regulation in health and in CKD remain mostly unknown. Autosomal dominant hypophosphatemic rickets (ADHR) is caused by gain-of-function mutations in FGF23 that prevent its proteolytic cleavage, but affected individuals experience a waxing and waning course of phosphate wasting. This led to the discovery that iron deficiency is an environmental trigger that stimulates FGF23 expression and hypophosphatemia in ADHR. Unlike osteocytes in ADHR, normal osteocytes couple increased FGF23 production with commensurately increased FGF23 cleavage to ensure that normal phosphate homeostasis is maintained in the event of iron deficiency. Simultaneous measurement of FGF23 by intact and C-terminal assays supported these breakthroughs by providing minimally invasive insight into FGF23 production and cleavage in bone. These findings also suggest a novel mechanism of FGF23 elevation in patients with CKD, who are often iron deficient and demonstrate increased FGF23 production and decreased FGF23 cleavage, consistent with an acquired state that mimics the molecular pathophysiology of ADHR. Iron deficiency stimulates FGF23 production, but normal osteocytes couple increased FGF23 production with increased cleavage to maintain normal circulating levels of biologically active hormone. These findings uncover a second level of FGF23 regulation within osteocytes, failure of which culminates in elevated levels of biologically active FGF23 in ADHR and perhaps CKD.

RAS in FGF23: Another Piece in the Puzzle

The first piece of the puzzle that is the evolving story of fibroblast growth factor-23 (FGF23) physiology was identified in 1959 by Andrea Prader (1). He postulated the existence of a phosphate-regulating hormone as the cause of acquired rickets in a child who was cured by excision of a bone-derived tumor. More pieces were added in 1976, 1989, and 1992, respectively, by the discovery of the Hyp mouse as a model of X-linked hypophosphatemic rickets (XLH) and elegant parabiosis and renal transplant studies in hyp mice that clearly demonstrated that a circulating factor was responsible for the phosphate wasting in Hyp mice (2,–4). A key piece was identified in 2000 when the autosomal dominant hypophosphatemic rickets (ADHR) consortium discovered that mutations in FGF23 were responsible for ADHR (5). In rapid succession, FGF23 was found to be elevated in patients with XLH, tumor-induced osteomalacia, and fibrous dysplasia (FD) of bone (6, 7). Another important piece was added when it was discovered that bone-derived cells were the source of FGF23, not only in the dysplastic bone cells of FD but also in normal bone (7).

Autosomal dominant hypophosphatemic rickets in an 85 year old woman: Characterization of her disease from infancy through adulthood

BackgroundAutosomal dominant hypophosphatemic rickets (ADHR) is a rare genetic disorder of phosphate homeostasis characterized, when severely expressed, by osteomalacia, suppressed levels of calcitriol, and renal phosphate wasting due to elevated levels of fibroblast growth factor 23 (FGF23). The disease is caused by heterozygous FGF23 mutations at the RXXR site that prevent cleavage of the intact hormone. ObjectivesAn FGF23 mutation was identified in the proband an 85-year-old woman with elevated FGF23 levels, and her clinical course was characterized. Medical records revealed she was treated for rickets as an infant. She was then asymptomatic until soon after her 4th pregnancy, when she suffered incapacitating bone pain and weakness, age 37. Symptoms remitted with brief treatment. ResultsThe proband and one son, but not other family members, were found to be heterozygous for the R176Q mutation in FGF23. Expression of this germ line mutation was strikingly different in both individuals in terms of skeletal health, FGF23 levels and disease activity. ConclusionsThe identified FGF23 mutation in two members of this family raises questions about molecular mechanisms that have led to intermittent increases in FGF23 synthesis and secretion, and disease expression.

The chicken or the egg: PHEX, FGF23 and SIBLINGs unscrambled.

The eggshell is an ancient innovation that helped the vertebrates' transition from the oceans and gain dominion over the land. Coincident with this conquest, several new eggshell and noncollagenous bone-matrix proteins (NCPs) emerged. The protein ovocleidin-116 is one of these proteins with an ancestry stretching back to the Triassic. Ovocleidin-116 is an avian homolog of Matrix Extracellular Phosphoglycoprotein (MEPE) and belongs to a group of proteins called Small Integrin-Binding Ligand Interacting Glycoproteins (SIBLINGs). The genes for these NCPs are all clustered on chromosome 5q in mice and chromosome 4q in humans. A unifying feature of the SIBLING proteins is an Acidic Serine Aspartate-Rich MEPE (ASARM)-associated motif. The ASARM motif and the released ASARM peptide play roles in mineralization, bone turnover, mechanotransduction, phosphate regulation and energy metabolism. ASARM peptides and motifs are physiological substrates for phosphate-regulating gene with homologies to endopeptidases on the X chromosome (PHEX), a Zn metalloendopeptidase. Defects in PHEX are responsible for X-linked hypophosphatemic rickets. PHEX interacts with another ASARM motif containing SIBLING protein, Dentin Matrix Protein-1 (DMP1). DMP1 mutations cause bone-renal defects that are identical with the defects caused by loss of PHEX function. This results in autosomal recessive hypophosphatemic rickets (ARHR). In both X-linked hypophosphatemic rickets and ARHR, increased fibroblast growth factor 23 (FGF23) expression occurs, and activating mutations in FGF23 cause autosomal dominant hypophosphatemic rickets (ADHR). ASARM peptide administration in vitro and in vivo also induces increased FGF23 expression. This review will discuss the evidence for a new integrative pathway involved in bone formation, bone-renal mineralization, renal phosphate homeostasis and energy metabolism in disease and health.

Clinical variability of familial tumoral calcinosis caused by novel GALNT3 mutations

The GALNT3 gene encodes GalNAc-T3, which prevents degradation of the phosphaturic hormone, fibroblast growth factor 23 (FGF23). Biallelic mutations in either GALNT3 or FGF23 result in hyperphosphatemic familial tumoral calcinosis or its variant, hyperostosis-hyperphosphatemia syndrome. Tumoral calcinosis is characterized by the presence of ectopic calcifications around major joints, whereas hyperostosis-hyperphosphatemia syndrome is characterized by recurrent long bone lesions with hyperostosis. Here we investigated four patients with hyperphosphatemia and clinical manifestations including tumoral calcinosis and/or hyperostosis-hyperphosphatemia syndrome to determine underlying genetic cause and delineate phenotypic heterogeneity of these disorders. Mutational analysis of FGF23 and GALNT3 in these patients revealed novel homozygous mutations in GALNT3. Although the presence of massive calcifications, cortical hyperostosis, or dental anomalies was not shared by all patients, all had persistent hyperphosphatemia. Three of the patients also had inappropriately normal 1,25-dihyroxyvitamin D [1,25(OH)(2)D] and confirmed low circulating intact FGF23 concentrations. The four novel GALNT3 mutations invariably resulted in hyperphosphatemia as a result of low intact FGF23, but other clinical manifestations were variable. Therefore, tumoral calcinosis and hyperostosis-hyperphosphatemia syndrome represent a continuous spectrum of the same disease caused by increased phosphate levels, rather than two distinct disorders.

Loss-of-Function ENPP1 Mutations Cause Both Generalized Arterial Calcification of Infancy and Autosomal-Recessive Hypophosphatemic Rickets

The analysis of rare genetic disorders affecting phosphate homeostasis led to the identification of several proteins that are essential for the renal regulation of phosphate homeostasis; for example, fibroblast growth factor 23 (FGF23), which inhibits renal phosphate reabsorption and 1,25-dihydroxyvitamin D synthesis. Here, we report presumable loss-of-function mutations in the ENPP1 gene (ectonucleotide pyrophosphatase/phosphodiesterase) in members of four families affected with hypophosphatemic rickets. We provide evidence for the conclusion that ENPP1 is the fourth gene-in addition to PHEX, FGF23, and DMP1-that, if mutated, causes hypophosphatemic rickets resulting from elevated FGF23 levels. Surprisingly, ENPP1 loss-of-function mutations have previously been described in generalized arterial calcification of infancy, suggesting an as yet elusive mechanism that balances arterial calcification with bone mineralization.

A case of familial tumoral calcinosis/hyperostosis–hyperphosphatemia syndrome due to a compound heterozygous mutation in GALNT3 demonstrating new phenotypic features

A new case of familial tumoral calcinosis (FTC)/hyperostosis-hyperphosphatemia syndrome (HHS) due to a novel compound heterozygous mutation in N-acetylgalactosaminyltransferase 3 (GALNT3) and with new phenotypic findings is presented. The response in serum phosphate and fibroblast growth factor 23 (FGF23) to medical treatment is detailed. This case expands the genotype and phenotype of FTC/HHS and gives insight into its treatment and pathophysiology. FTC and HHS are caused by mutations in FGF23, GALNT3, or KLOTHO. They are characterized by hyperphosphatemia, increased phosphate reabsorption, and elevated or inappropriately normal serum 1,25-dihydroxyvitamin D(3) (1,25-D(3)); FTC is associated with calcific masses, and HHS with diaphyseal hyperostosis. A 36-year-old woman presented with abnormal dental X-rays at age 12 and was hyperphosphatemic at 22. She underwent radiographic, biochemical and genetic testing, and medical treatment. Serum phosphorus was 7.3 mg/dL (2.5-4.8), TmP/GFR 6.99 mg/100 mL (2.97-4.45), 1,25-D(3) 35 pg/mL (22-67). Radiographs revealed tooth anomalies, thyroid cartilage calcification, calcific masses in vertebral spaces, calcification of the interstitial septa of the soft tissue in the lower extremities, and cortical thickening of the long bones. Her total hip Z score was 1.9. C-terminus serum FGF23 was 1,210 RU/mL (20-108), but intact FGF23 was 7.4 pg/mL (10-50). DNA sequencing determined she was a compound heterozygote for mutations in GALNT3. Treatment with niacinamide and acetazolamide decreased TmP/GFR and serum phosphate, which was paralleled by a decrease in serum C-terminus FGF23. This case broadens the spectrum of phenotypic and genotypic features of FTC/HHS and suggests treatments to decrease renal phosphate reabsorption in the setting of a low intact FGF23.

Tumoral Calcinosis

Abstract: Tumoral calcinosis (TC) is a disorder of enhanced renal tubular phosphate reabsorption resulting in hyperphosphatemia and ectopic calcifications. Gene analyses have confirmed that destabilizing mutations within fibroblast growth factor-23 (FGF23) and GalNAc transferase-3 (GALNT3) are responsible for heritable TC. Herein, we present a patient with phenotypic and biochemical features of TC, whose evaluation revealed no mutations in either the FGF23 or GALNT3 genes and normal circulating C-terminal and intact FGF23 concentrations. The patient is a 12-year-old African American man with ectopic, subcutaneous mineral deposits, with a diagnosis of tumoral calcinosis based on collective clinical features. Plasma FGF23 concentrations were determined using both an assay that measures full-length FGF23 and one that recognizes the C-terminal portion. Gene analysis was performed on DNA isolated from peripheral WBCs. The patient's calcium and PTH concentrations were normal. Serum phosphorus concentrations ranged 5 to 6.7 mg/dL. 1,25-Hydroxy vitamin D levels were elevated. FGF23 C-terminal fragment concentrations were 53.79 RU/mL (reference interval 55 ± 50 RU/mL) and intact FGF23 concentrations were 11 pg/mL (reference interval 28 ± 2.2 pg/mL). Gene analysis showed no mutations in either FGF23 or GALNT3 genes. Reported cases of TC due to FGF23 and GALNT3 mutations have similar clinical and laboratory features. Our patient has most of the “classic” TC phenotypic features, but has normal circulating levels of C-terminal and intact FGF23, and no detected mutations of either the FGF23 or GALNT3 genes. Therefore, our case likely represents a new, yet undetermined, cause of TC.

Emerging role of fibroblast growth factor 23 in a bone–kidney axis regulating systemic phosphate homeostasis and extracellular matrix mineralization

To describe emerging understanding of fibroblast growth factor 23 (FGF23) - a bone-derived hormone that inhibits phosphate reabsorption and calcitriol production by kidney and participates as the principle phosphaturic factor in a bone-kidney axis coordinating systemic phosphate homeostasis and bone mineralization. FGF23 (a circulating factor made by osteocytes in bone) inhibits phosphate reabsorption and 1,25(OH)2D production by kidney. Physiologically, FGF23 is a counter-regulatory phosphaturic hormone for vitamin D and coordinates systemic phosphate homeostasis with skeletal mineralization. Pathologically, high circulating FGF23 levels cause hypophosphatemia, decreased 1,25(OH)2D production, elevated parathyroid hormone and rickets/osteomalacia. FGF23 mutations impairing its degradation cause autosomal dominant hypophosphatemic rickets. Respective loss-of-function mutations of osteocyte gene products DMP1 and Phex cause autosomal recessive hypophosphatemic rickets and X-linked hypophosphatemic rickets, initiating increased FGF23 production. Low FGF23 levels lead to hyperphosphatemia, elevated 1,25(OH)2D, and soft-tissue calcifications. FGF23 is markedly increased in chronic renal disease, but its role remains undefined. FGF23 discovery has uncovered primary regulatory pathways and new systems biology governing bone mineralization, vitamin D metabolism, parathyroid gland function, and renal phosphate handling. FGF23 assessment will become important in diagnosing hypophosphatemic and hyperphosphatemic disorders, for which pharmacological regulation of FGF23 levels may provide novel treatments.

FGF23 Concentrations Vary With Disease Status in Autosomal Dominant Hypophosphatemic Rickets

We measured FGF23 concentrations in subjects with ADHR. FGF23 and serum phosphate correlated positively in controls and negatively in subjects with ADHR. Elevated FGF23 concentrations were associated with lower phosphate values. The variable phenotype in ADHR may be caused by fluctuations in FGF23. Autosomal dominant hypophosphatemic rickets (ADHR) is a rare disorder of phosphaturia, hypophosphatemia, inappropriately low/normal calcitriol, and rickets/osteomalacia. ADHR is caused by mutations in a circulating peptide, fibroblast growth factor 23 (FGF23). We present the first report of FGF23 concentrations in subjects with ADHR. The aim was to test the hypotheses that subjects with ADHR have elevated FGF23 concentrations and that FGF23 concentrations are associated with disease severity. This was an observational study at a tertiary referral center. Subjects were from three kindreds with FGF23 mutations causing ADHR (n = 42). Controls were participants from these families without mutations (n = 55). Fasting blood and urine samples were obtained. Biochemistries were determined, and FGF23 concentrations were measured using two ELISAs. Three cases are presented illustrating activity of disease and FGF23 concentrations. One case shows persistent hypophosphatemia and elevation of FGF23 concentrations, whereas another shows remission of hypophosphatemia corresponding to a decrease in previously elevated FGF23 concentrations. Overall cross-sectional group differences were nonsignificant for serum phosphate and FGF23 concentrations. C-terminal FGF23 concentration in controls was 61.0 +/- 28.6 (SD) RU/ml (median, 52.5 RU/ml), and in subjects with mutations was 148.8 +/- 374.5 RU/ml (median, 63.1 RU/ml). Mean intact FGF23 concentration in controls was 44.7 +/- 14.9 pg/ml (median, 40.4 pg/ml), and in subjects with mutations was 83.2 +/- 233.0 pg/ml (median, 39.0 pg/ml). C-terminal FGF23 concentrations were at least +2 SD in 10/42(24%), and intact FGF23 concentrations were at least +2 SD in 3/34(9%). Phosphate correlated positively with C-terminal and intact FGF23 in both controls and in subjects with mutations with phosphate >2.5 mg/dl but correlated significantly negatively with C-terminal and intact FGF23 in ADHR subjects with phosphate < or =2.5 mg/dl. Elevated FGF23 concentrations are associated with hypophosphatemia in ADHR, and remission of the phenotype is associated with lower FGF23 concentrations. FGF23 has an opposite relationship with phosphate in ADHR compared with controls. We conclude that ADHR symptoms and disease severity likely fluctuate with FGF23 concentrations.

Novel GALNT3 Mutations Causing Hyperostosis-Hyperphosphatemia Syndrome Result in Low Intact Fibroblast Growth Factor 23 Concentrations

Hyperostosis-hyperphosphatemia syndrome (HHS) is a rare metabolic disorder characterized by hyperphosphatemia and localized hyperostosis. HHS is caused by mutations in GALNT3, which encodes UDP-N-acetyl-alpha-D-galactosamine:polypeptide N- acetylgalactosaminyltransferase 3. Familial tumoral calcinosis (TC), characterized by ectopic calcifications and hyperphosphatemia, is caused by mutations in the GALNT3 or fibroblast growth factor 23 (FGF23) genes. Our objective was to identify mutations in FGF23 or GALNT3 and determine serum FGF23 levels in an HHS patient. Mutation detection in FGF23 and GALNT3 was performed by DNA sequencing, and serum FGF23 concentrations were measured by ELISA. A 5-year-old French boy with HHS and his family members participated. The patient presented with painful cortical lesions in his leg. Radiographs of the affected bone showed diaphyseal hyperostosis. The lesional tissue comprised trabeculae of immature, woven bone surrounded by fibrous tissue. Biochemistry revealed elevated phosphate, tubular maximum rate for phosphate reabsorption per deciliter of glomerular filtrate, and 1,25-dihydroxyvitamin D levels. The patient was a compound heterozygote for two novel GALNT3 mutations. His parents and brother were heterozygous for one of the mutations and had no biochemical abnormalities. Intact FGF23 level in the patient was low normal, whereas C-terminal FGF23 was elevated, a pattern similar to TC. The presence of GALNT3 mutations and elevated C-terminal, but low intact serum FGF23, levels in HHS resemble those seen in TC, suggesting that HHS and TC are different manifestations of the same disorder. The absence of biochemical abnormalities in the heterozygous individuals suggests that one normal allele is sufficient for secretion of intact FGF23.

Tumoral Calcinosis Presenting with Eyelid Calcifications due to Novel Missense Mutations in the Glycosyl Transferase Domain of theGALNT3Gene

Familial tumoral calcinosis (TC) is a rare autosomal recessive disorder characterized by metastatic calcifications, often periarticular. Biochemical findings include hyperphosphatemia, high 1,25-dihydroxyvitamin D levels, and elevated tubular maximum for phosphate reabsorption per deciliter of glomerular filtrate (TmP/GFR). TC is caused by biallelic mutations of the genes encoding either fibroblast growth factor 23 (FGF23) or uridine diphosphate-N-acetyl-alpha-D-galactosamine:polypeptide N-acetylgalactosaminyltransferase 3 (GalNAc transferase 3 or GALNT3). The objective was to identify mutations in FGF23 or GALNT3 responsible for a mild TC phenotype by DNA sequencing and to determine serum FGF23 levels by ELISA. The subject was a 25-yr-old Caucasian woman with eyelid calcifications and biochemical features of TC. Eyelid biopsy revealed superficial dermis calcifications. There was no history of metastatic calcifications, mineral homeostasis abnormalities, or renal dysfunction. Biochemistry revealed normal levels of calcium, creatinine, PTH, and 25-hydroxyvitamin D, with elevated phosphorous, TmP/GFR, and high normal 1,25-dihydroxyvitamin D levels. Intact FGF23 was undetectable (< 3 pg/ml), whereas C-terminal FGF23 was elevated (698.2 RU/ml). Mutation detection revealed compound heterozygosity for two novel mutations in the glycosyl transferase domain of the GALNT3 gene. Previously reported GALNT3 mutations in TC have been null mutations. This study shows that missense mutations affecting the glycosyl transferase domain of GalNAc transferase 3 also cause TC. Elevated C-terminal FGF23 fragments with undetectable intact FGF23 suggest that the mutant enzyme lacks the ability to glycosylate FGF23 and that glycosylation by GalNAc transferase 3 is necessary for secretion of functional full-length FGF23.

Fibroblast Growth Factor-23 Mutants Causing Familial Tumoral Calcinosis Are Differentially Processed

Familial tumoral calcinosis (TC, OMIM 211900) is a heritable disorder characterized by hyperphosphatemia, normal or elevated serum 1,25-dihydroxyvitamin D, and often severe ectopic calcifications. Two recessive mutations in fibroblast growth factor-23 (FGF23), serine 71/glycine (S71G) and serine 129/phenylalanine (S129F), were identified as causing TC. Herein, we undertook comprehensive biochemical analyses of an extended TC family carrying the S71G FGF23 mutation, which revealed that heterozygous (serine/glycine, S/G) individuals had elevated serum FGF23 C-terminal fragments compared with wild-type (serine/serine, S/S) family members (P < 0.025). To understand the differential processing of FGF23 in TC patients, we transiently expressed S71G as well as S129F FGF23. FGF23 ELISA in tandem with Western analyses revealed increased proteolytic cleavage of mutant FGF23 and a limited secretion of intact protein. Furthermore, S71G and S129F FGF23 carrying mutations that disrupt the furin-like protease RXXR motif in FGF23 rescued the secretion of the intact protein, and both TC mutant proteins harboring the R176Q mutation revealed no altered sensitivity to trypsin compared with the native (R176Q)FGF23. Finally, S71G, but not S129F mutant FGF23, is rescued by temperature. In summary, FGF23 mutations causing TC lead to increased intracellular proteolysis of FGF23, most likely by furin-like proteases, due to conformational changes of the mutant protein. The destabilizing nature of these mutations provides new insight into the pathophysiology of TC and exemplifies the physiological importance of FGF23 in phosphate and vitamin D metabolism.

FGF23 and disorders of phosphate homeostasis

It is well known that fibroblast growth factor (FGF) family members are associated with embryonic development and are critical for basic metabolic functions. This review will focus upon fibroblast growth factor-23 (FGF23) and its roles in disorders associated with phosphate handling. The discovery that mutations in FGF23 were responsible for the isolated renal phosphate wasting disorder autosomal dominant hypophosphatemic rickets (ADHR) has ascribed novel functions to the FGF family. FGF23 circulates in the bloodstream, and animal models demonstrate that FGF23 controls phosphate and Vitamin D homeostasis through the regulation of specific renal proteins. The ADHR mutations in FGF23 produce a protein species less susceptible to proteolytic processing. X-linked hypophosphatemic rickets (XLH), tumor-induced osteomalacia (TIO), and fibrous dysplasia of bone (FD) are disorders involving phosphate homeostasis that share phenotypes with ADHR, indicating that FGF23 may be a common denominator for the pathophysiology of these syndromes. Our understanding of FGF23 will help to develop novel therapies for phosphate wasting disorders, as well as for disorders of increased serum phosphate, such as tumoral calcinosis, a rare disorder, and renal failure, a common disorder.