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FGF23 (and Klotho): what’s new?

Justine Bacchetta, MD, PhD

Reference Center for Rare Renal Diseases

Long Beach, CA, 2017

Brief introduction to FGF23

1-25 vitamin D

FGF23 PTH

Phosphorus Calcium

Stimulating effect / Inhibiting effect

Calcium and phosphate metabolism

Bacchetta, EMC 2015

Introduction to FGF23

• FGF23 synthezised by osteocytes

• FGF-R: tyrosine kinase receptor– FGFR-1/2/3/4

• Klotho: cofactor– Anti-aging protein– Tissue-specific expression – Kidney and parathyroid gland

Unakawa, Nature 2006; Bonewald JBMR 2010

FGF23: phosphorylation pathways

• Klotho-dependent and Klotho-independent activation of the

phosphorylation pathways

• Erk 1/2 and Akt pathway

• Calcineurin-NFAT pathway

Faul, JCI 2011

FGF23: phosphorylation pathways

• Klotho-dependent and Klotho-independent activation of the

phosphorylation pathways

• Erk 1/2 and Akt pathway

• Calcineurin-NFAT pathway

Faul, JCI 2011

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Proximal tubule Parathyroid

PTH inhibition

1α hydroxylase

stimulation

Hypophosphatemia, PTH and 1-25 OH2 vitamin D inhibition

FGF23: 3 main ‘historical’ actions

Prie, Kidney Int 2009

Npt inhibition 2a-2c

1α hydroxylase inhibition

24 hydroxylase stimulation

FGF23

Saito, J Biol Chem 2003

Andrukhova, EMBO 2014

Kidney

Silver, PN 2010

Olauson Plos One 2013

Parathyroid

Bacchetta, JBMR 2012

Chonchol, JASN 2015 & JASN 2016

Immune system

Cartilage

Kawai, JBC 2012 Dentoalveolar complex

Chu, Anat Rec 2010

FGF23: an endocrine FGF with off-target effects

Bone

Wang JBMR 2008, Wesseling-Perry JCEM

2009, Allard CTI 2015

Faul, JCI 2011

Heart &

Cardiomyocytes

Hippocampal cells

and CNS

Hensel, J Neurochem 2016

Other targets?

New insights: FGF23 physiology in bone

FGF23 as an inhibitor of bone mineralization

• Model of genetic hyperphosphatemia (GALNT 3 mutation)

• Low intact FGF23 levels

• High C-terminal FGF23 levels

• Familial tumoral calcinosis

Masi CTI 2015

FGF23 as an inhibitor of bone mineralization

• Comparison of mineralization in vitro in preosteoblastic cells from a patient with

GALNT3 mutation (low intact FGF23 levels) and from controls

• Higher capability to form mineralization

Masi CTI 2015

Patient

LOW

FGF23

Control

NORMAL

FGF23

Biphasic effects of FGF23 on osteoclast biology

Allard, Calcified Tissue International 2015

Inhibition of osteoclast

differentiation

An expression of FGF-R1

An activation of Erk and Akt

Effects driven through FGF-R

A moderate albeit significant

stimulation of resorption

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Klotho expression in long bones regulates FGF23 production

during renal failure

Kaludjerovic FASEB 2017

• Murine model: targeted deletion of Klotho

in long bones

• Two main results

• Long bones from these mice did not

increase FGF23 expression after adenine

diet or 5/6th nephrectomy

• FGF23-treated bone cells required Klotho

to increase FGF23 expression and induce

Erk phosphorylation

New insights: FGF23 physiology in the

central nervous system

Impact of FGF23 on neuronal morphology and synaptic density

• In vitro model of primary murine hippocampal cultures

• Incubation with FGF23 ± α-Klotho

• Enhanced number of primary neurites

• And reduced arborization

=> Resulting in a less complex neuronal morphology

Hensel J Neurochemist 2016

New insights: FGF23 and adipose tissue

And the fat lady sings about phosphate and calcium…

Wagner Kidney International 2017

Towards new endocrine regulations…

Rutkowski, Kidney International 2017; editorial comment by Wagner Kidney International 2017

• High adiponectin levels

• Suppressed Klotho renal

expression

• Decreased FGF23 levels

• Caused renal loss of

calcium

New insights: FGF23 and (pediatric) CKD

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Faul et al, J Clin Invest 2011

Gutierrez et al, N Eng J Med 2008

Fliser, JASN 2007; Isakova JAMA 2011

Wolf, Kidney International 2012 Pereira, Bone 2010

The paradigm: both FGF23 and Klotho are deregulated early

in CKD

Bacchetta, J Clin Endocrinol Metab 2010

FGF23 in pediatric CKD

Cohort of 227 followed in a pediatric

nephrology unit

Normal renal function and CKD (pre-dialysis)

GFR: inulin clearance

Influence of GFR, age, solid organ

transplantation and BMI on FGF23 levels

A likely influence of corticosteroids on FGF23

loevels

An association between FGF23 and uric acid

An association between FGF23 and IGF1

Bacchetta, Pediatr Nephrol 2012

• Similar results of an open label prospective study in Pediatric Endocrinology,

Birmingham, Alabama

– 23 children with GH deficiency

– Clinical trial: C terminal FGF23 before and one year after rhGH

– Increased FGF23, TmP/GFR and phosphorus after rhGH

– The increase of FGF23 remained significant after adjustment on age and phosphorus

Gardner et al, J Pediatr Endocrinol Metab 2011

Recombinant growth hormone increases FGF23 levels

Portale, clin J Am Soc Nephrol 2014

FGF23 increases early in pediatric CKD

Data from the CKID cohort

N=464 children ages 1–16 years with predialysis CKD

Portale, clin J Am Soc Nephrol 2016

FGF23 as a risk factor of CKD progression

Data from the CKID cohort

N=419 children ages 1–16 years with predialysis CKD

New insights: FGF23 and infections in CKD

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FGF23 as an inhibitor of extra-renal 1-alpha hydroxylase in

monocytes

Bacchetta, JBMR 2012

Vehicle

IL-15

1,2

5(O

H) 2

D1

,25

(OH

) 2D

1,2

5(O

H) 2

D

25OHD

25OHD

25OHD

1,25(OH)2D

1,25(OH)2D

IL-15+

FGF23

1,25(OH)2D

*

0.00

0.20

0.40

0.60

0.80

1.00

1.20

CYP27B1 CYP24A1 VDR LL37 TNF

Fold

change in m

RN

A r

ela

tive t

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ehic

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reate

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ells

** ***

FGF23: 100 ng/mL, 24 hours

Controls = PBS

6 hours = black

24 hours = grey

Data obtained in monocytes from healthy donors, but similar data obtained in PD cells from pediatric patients undergoing

chronic PD

High FGF23 (and low 25-D) as risk factors for clinical

infections in the HEMO-study

Chonchol, JASN 2015 and further confirmed JASN 2016

New insights: FGF23 and cardiovascular

disease in CKD

FGF23 and left ventricular hypertrophy

• Increased FGF 23 levels and left

ventricular hypertrophy

– Epidemiological prospective

– CRIC cohort, 3070 CKD adults

Faul et al, J Clin Invest 2011

• Increased FGF 23 levels at baseline

and increased risk for onset of LVH

– RR= 2.4

• Rat cardiomyocytes + FGF23

– Hypertrophy, dose-dependent

– Reactivation fetal genes

– Increased markers for LVH

FGF23 and cardiomyocytes: a Klotho-independent effect

Faul et al, J Clin Invest 2011

• Klotho is not expressed in CM, but FGF-R1 and 4 are

• Signaling pathways

– Through FGF-R signaling

– Through PLCγ and NFAT

– No activation of Akt

• Intramyocardial and intravenous injection of

FGF23 induces ventricular hypertrophy in mice

– Wild type

– Klotho deficient mice: high FGF23 and LVH

FGF23 and LVH in pediatric CKD

Leifheit-Nestler, Nephrol Dial Transplant 2016

German autopsy study

17 dialysis, 17 KTx, 24 controls

67% LVH in patients

Induction of cardiac FGF23-FGFR4

expression as well as a decreased

Klotho expression is associated with

LVH

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LVH depends on FGF23 but also on Klotho levels: loss of

Klotho in CKD breaks one’s heart!

Xie JASN 2015 / Fu & Liu JASN 2015

Heterozygous Klotho deficient mice

Klotho-deficient CKD mice display aggravated

cardiac fibrosis as compared to WT-CKD mice

Transgenic expression of soluble Klotho improves

cardiomyopathy in Klotho-deficient CKD mice

Anti FGF23 antibodies: can they be

relevant in CKD?

Antibodies against FGF23 improve

hyperparathyroidism…

Shalhoub JCI 2012

Early decreased PTH levels

But increased phosphate, calcium and 1-25 D

levels

But they also increase mortality…

Shalhoub JCI 2012

FGF23 and cardiomyocytes: a potential therapeutic target

Faul et al, J Clin Invest 2011

• FGFR inhibitors in a rat model of CKD (5/6th nephrectomy)

– Decreased LVH

– No modification of blood pressure

From prevention to treatment…

FGFR blockade can improve LVH in animal models

Di Marco NDT 2014

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So… should we trash anti FGF23

antibodies?

Wolf et al, JASN 2010

FGF23 and human diseases

Hypophosphatemic ricketsThe revolution in 2014: KRN23 in patients, short-term

Carpenter JCI 2014

• Adults with XLH

• 38 patients receiving KRN23 or placebo, IV or sc

• Single dose

Hypophosphatemic ricketsThe revolution in 2015: KRN23 in patients, ‘long-term’

Imel JCEM 2015

• Adults with XLH

• 28 patients receiving KRN23: 0.05 to 0.6 mg/kg every 28 days (SC)

• Then extension study 22 patients receiving 0.1 to 1 mg/kg every 28

days (SC)

International pediatric trials are ongoing

Conclusion

Scialla, Kidney International 2013

Back in the real life of a CKD patient…

Phosphate is a/the vascular toxin/silent killer

Shroff, Pediatric Nephrology 2013

Vascular calcifications

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Phosphate and longevity

Kuro-O Mech Ageing Dev 2010

Klotho -/-

mice

Rhinoceros

Elephant

Centenarian

human

Guinea pig

Gerbil

Dialysis patient

Conversion mg/dL: 0.323 mmol/L

FGF23, Klotho and CKD in 2017

• Both low Klotho and high FGF23

levels are delerious in CKD for LVH

• Both low 25-D and high FGF23 levels

are deleterious in CKD for infections

• FGF23 has become a therapeutic

target in genetic hypophosphatemic

diseases: the ongoing revolution!

• However FGF23 antibodies have

proved to be deleterious in CKD

• A potential role for drugs modulating

FGF-R and for recombinant Klotho in

the future?

• Clinical practice: target phosphate