KLOTHO: CLINICAL ASPECTSKLOTHO: CLINICAL ASPECTS

Pr Dominique Prié,

Université Paris Descartes, Faculté de Médecine,

Institut Necker-Enfants Malades INSERMInstitut Necker-Enfants Malades INSERM

Service des Explorations Fonctionnelles

Hôpital Necker-Enfants Malades, Paris, France

THE DIFFERENT FORMS OF KLOTHO

FGF-23

Circulating Klotho

1014 amino acids, short transmembrane and cytoplasmic segment

FGF-23

FGFR

FGF-23

membrane formof Klotho

FRS2αShedding

(ADAMs 10, 17?)

KL1

KL2

PLCγFGFR (ADAMs 10, 17?)

KL1?Alternative splicing

ERK

AKT

PLCγ

PKCIP3

βcatenine

HOW KLOTHO COULD MODIFY CLINICAL PRACTICE?

1- Is Klotho important for the diagnosis?

2- Can Klotho be a biomarker?

Can Klotho predict a risk? does this lead to a change of caring or treatment?

• Variants in Klotho gene

• mRNA expression in specific tissues

• Protein expression in specific tissues

• Protein level in blood

3- Is Klotho a surrogate or an independent biomarker?

4- Can Klotho be a therapeutic target?

• Can over or down expression of Klotho improve the prognosis of a disorder or

lower adverse outcomes?

• How can we modify Klotho expression?

Kuro-o, Nature

1997, 390:45

EXPRESSION OF KLOTHO

KIDNEY: proximal + distal tubule Parathyroid gland

1997, 390:45

Ben-Dov, JCI 2007;

117, 12: 4003

Hu Faseb J 2010

The tissue-specific expression of Klotho is controlled by the methylation of Klotho promoter

(Azuma FASEB J. 2012)

IS KLOTHO EXPRESSED IN THE ARTERY WALL?

Healthy humanMICE

(Lim Circulation. 2012;125:2243-2255)

Lindberg (2013). PLoS ONE 8: e60658.

Specific disruption of Klotho gene

in smooth muscle cell of artery

wall: No phenotypeSpecificity of the antibody?

Jimbo. Intern. J. Hypertension Volume 2014

DOES KLOTHO HAVE EFFECTS INDEPENDENT OF FGF23 ?

FGF23 -/- = Klotho -/- = Double KO (Nakatani

Faseb J 2009)

Serum phosphate, tissue calcifications are

similar

Injection of Klotho in FGF23 -/- mice increases urinary

phosphate excretion and lowers serum phosphate

concentration (Hu Faseb J 2010)

Klotho modifies channels and ion transporters activity in the absence of FGF23

(enzymatic effets?)

Inhibe la voie WNT/β-catenine & IGF1 (liaison Klotho-β-catenine)

FACTORS THAT MODIFY KLOTHO EXPRESSION

Stimulating factors:

• Calcitriol.

• EPO: rat CKD model (Sugiura Am J Nephrol 2010;32:137)

• Exercice

Inhibiting factors:

• FGF23 lowers Klotho mRNA expression in the kidney

- FGF23 transgenic mice (Marsell, Nephrol. Dial. Transplant. 2008, 23: 827),

- Hyp mice (Farrow. Journal of Endocrinology. 2010, 207: 67) no change of FGF23

signalingsignaling

• Angiotensin II via AT1 (Yoon. Nephrol Dial Transplant 2011 26: 800)

• TGFβ1 (Zhou. J Am Soc Nephrol 24: 771, 2013)

• Methylation of Klotho promoter (renal insufficiency)

RENAL EXPRESSION OF KLOTHO DURING CKD

mRNA expression of the

membrane form of Klotho in

the kidney

Protein expression in

the kidney

Patients on dialysis

Koh BBRC 2001,280:1015Sakan.2014. PLoS ONE 9(1): e86301.

Akimoto et al. BMC Nephrology 2012, 13:155

PLASMA KLOTHO CONCENTRATION IN CKD

Yamazaki BBRC 2010, 513 Pavik. Nephrol Dial Transplant (2012)

Scholze. 2014. J Clin Endocrinol MetabSeiler. Kidney Intern 2012

KLOTHO AND SURVIVAL

Klotho -/- in mice increases mortality

Overexpression of Klotho in mice extends life span (Kurosu Science. 2005; 309: 1829)

Specific renal Klotho deletion does not increase mortality nor alter growth (Olauson J

Am Soc Nephrol 23:2012)

Association between survival and plasma Klotho concentration in 804 subjects > 65

KLOTHO AND SURVIVAL

Variants in Klotho gene in healthy humans have been inconstantly associated with

life expectancy.

ans follow up 6 years. independent predictor of all-cause mortality (Semba J Gerontol A Biol

Sci Med Sci 2011)

VARIANTS IN KLOTHO GENE, VITAMIN D, MORTALITY ON DIALYSIS

Treatment VitD+ Treatment VitD-

Friedman. 2009, J Bone Miner Res;24:1847

KLOTHO AND RENAL SURVIVAL

Overexpression of Klotho slows down renal function degradation in various animal

models of kidney disease.

Very few data in human

IgA nephropathy (Ko. Kidney Blood Press Res 2012;36:191)

Arking. Am. J. Hum. Genet. 72:1154, 2003

Rhee. Metabolism Clinical and Experimental 55 (2006) 1344

KLOTHO GENE VARIANTS AND CARDIOVASCULAR DISEASE

SIBS II = African-American

Lack of association of Klotho gene variants with valvular and vascular calcification in

Caucasians: a candidate gene study of the Framingham Offspring Cohort. Tangri. Nephrol

Dial Transplant (2011) 26: 3998

Semba. J Am Geriatr Soc. 59:1596, 2011

CIRCULATING KLOTHO AND CARDIOVASCULAR DISEASE IN NON-CKD

SUBJECTS

Navarro-González. Heart 2014;100:34

CIRCULATING KLOTHO AND ATHEROSCLEROSIS

Jeong AIDS RESEARCH AND HUMAN RETROVIRUSES Vol 29,

2013

69 healthy, postmenopausal women (50–76 years old)

CIRCULATING KLOTHO ARTERIAL COMPLIANCE, EXERCISE

Matsubara. Am J Physiol Heart Circ Physiol 306: H348, 2014

444 patients with CKD stages 2–4

CIRCULATING KLOTHO DEATH AND CARDIOVASCULAR EVENTS IN

CKD STAGES 2-4

Seiler. Clin J Am Soc Nephrol 9 2014

atherosclerotic events/death decompensated heart failure/death

EFFECTS OF KLOTHO ON HEART

Xie (2012). Nat Commun 3: 1238.

100 cardiology inpatients

CIRCULATING KLOTHO AND CARDIAC HYPERTROPHY

Shibata. 2013. PLoS ONE 8(9): e73184

KLOTHO AND CANCER

• Increase in KL1 mRNA expression is associated with poor survival in ovarian cancer

• Full length Klotho protein is expressed in normal breast, and its expression

decreases in breast cancer

• Klotho gene expression is decreased in various types of cancer (breast, gastric,

colorectal, lung cancers, cervical carcinoma, pancreatic adenocarcinoma, colorectal, lung cancers, cervical carcinoma, pancreatic adenocarcinoma,

hepatocellular carcinoma) due to methylation of Klotho promoter. It might

hallmark cancer “aggressiveness”

J. Usuda. Lung Cancer 74 (2011) 332

Small cell lung cancer

J. Usuda. Lung Cancer.2010

Small cell lung cancer large cell neuroendocrine

carcinoma

immunohistochemical expression of Klotho is cytoplasmic

CONCLUSIONS

1- Klotho genomic variant analysis: contradictory results, no clinical application to

date

2- Assessment of mRNA expression or Klotho promoter methylation in tumors: may

be interesting in cancers to predict prognosis. The consequences on the treatment

remain to be determinedremain to be determined

3- Klotho protein expression in tissues: requires antibodies with unquestionable

specificity. Interesting results to determine cancer prognosis

4- Measurement of circulating Klotho in blood and/or in urine:

• need an improvement of the assay sensitivity in the lower concentrations

• might be important to select patients with increased risks (death, vascular • might be important to select patients with increased risks (death, vascular

stiffness or calcification) independently of the renal function

5- Klotho as a therapeutic target:

• Many encouraging data in animals but almost no data in human

• ARA and calcitriol are already largely used as treatment in particular in CKD.

Promote physical exercise. Future: injection of recombinant Klotho