PHYSIOPATHOLOGIE

• Défaut du transporteur

lysosomal de cystine:

CYSTINOSINE

• Gène CTNS (23 kb)

• Transmission autosomique

récessive: 1/200 000

naissances vivantes

• Accumulation et cristallisation

intralysosomiale de cystine =

Maladie systémique

W Gahl, 2002 04/03/2019 2

TROIS FORMES CLINIQUES selon la sévérité et l’âge du diagnostic

1. Forme infantile (96%)

2. Forme juvénile (4%)

• Atteinte rénale et extra-rénale

moins sévère

3. Forme adulte (<1%)

• Atteinte cornéenne

• Absence d’atteinte rénale

04/03/2019 3

MODE DE PRESENTATION

• Cassure de la courbe de croissance après 6 mois • Rachitisme hypophosphatémique • Anorexie, Vomissements, Déshydratation extracellulaire

sévère • Syndrome de Fanconi

– Perte de sel, hypokaliémie, – Diabète phosphaté – Acidose métabolique, – Hypo-uricémie, – glucosurie orthoglycémique, amino-acidurie – Protéinurie tubulaire, a1 et b2 microglobinurie, RBPurie

• Alcalose hypokaliémique • Atteinte cornéenne isolée

04/03/2019 4

DIAGNOSTIC

• EXAMEN OPHTALMOLOGIQUE

• CYSTINE INTRALEUCYTAIRE

• GENETIQUE MOLECULAIRE

04/03/2019 5

TUBE CONTOURNÉ PROXIMAL syndrome de Fanconi

Sodium Pump

K+

ATP

ADP

Na+

GL

UC

OS

E

O2

GLUT2

FGF23

AQP3

AQP1

H2 O

Na-G

luco

se

Na-A

cides am

inés

Na-A

cides o

rgan

iqu

es

NPT2a

NPT2c

NHERF

+

Lysosome

PROTEINES

Cl-

OCRL1

Cystin

+

AA

ClC5

CT

NS

mégaline-

cubuline

exocytosis

endocytosis CO2

+ H2O

Car4,Car14

Car2

H++ HCO3--

CO2 + H2O

H++ HCO3--

Na-bicarbonates

HCO3-- Na+

NH

E 4

geo

rges

.des

chen

es@

rdb

.ap

hp

.fr

04/03/2019 6

TRAITEMENT DU SYNDROME DE FANCONI

• Supplémentations

– Bicarbonate de sodium

– Bicarbonate de potassium

– Chlorure de sodium

– Phosphate disodique

– 1 a calcidol

• AINS

– Indométacine 1-3 mg/Kg/jour

– ou Ibuprofen 10-30 mg/kg/jour

– Arrêt quand le DFG est < 30 ml/min/1.73m²

04/03/2019 7

PROGRESSION DE L’INSUFFISANCE RENALE

W Gahl, 2002 04/03/2019 8

CYSTINOSIS A SYSTEMIC DISEASE

• Growth failure

• Poor nutritional status

• Renal failure

• Endocrine disease

– Hypothyroidism

– Diabetes

– Delayed puberty

– Hypofertility or infertility

• Liver disease

– Portal hypertension

– Hepatomegaly

• Bone disease

– Fracture

– Deformations

– Teeth delay & enamel defects

• Dysmorphology

• Neuromuscular involvement – Myopathy, muscle weakness

– Respiratory failure

– Swallowing deficiency

– Intracranial hypertension

– Brain calcification

– Cognition disability

– Mental deterioration

• Ocular deposition – Cornea

– Retinopathy

04/03/2019 9

La myopathie

04/03/2019 10

A STANDARD FOLLOW-UP

M. CA… age 9 years, • BW 24 kg 700. Height 121,8 cm BP 97/70 mmHg. • Plasma Creatinine 56 µmol/l, blood urea 6 mmol/l, cystatin C 0,89 mg/l,

sodium 139 mmol/l, potassium 4,1 mmol/l, chloride 102 mmol/l, bicarbonates 31 mmol/l, total protides 75 g/l.

• Plasma renin 130 mU/L (11-110) • Plasma aldosterone 2153 pg/mL (N 70-700) • Glomerular filtration rate (integrated Schwartz formula 2009) : 72

ml/min/1.73 m². (N>90) • Total Calcium 2,50 mmol/l, ionized calcium 1,25 mmol/l, alcaline

phosphatases 154 UI/l, PTH 25 ng/L. • T3 : 6 pmol/l, T4 : 24,7 pmol/l, TSH 3,33 mUI/l. • Serum Glucose 5,1 mmol/l, HbA1c 5,2 %, glucosuria 1,8 mmol/l. • Total carnitin 49 µmol/L (N25-50); free carnitin 37 µmol/L (20-40) • Hemoglobin 14,2 g/dl, VGM 87 µ3, leucocytes 5 000/mm3, plaquettes 478

000/mm3, ferritine 57 µg/l, transferrin saturation 19 %. • Proteinuria 0,75 g/l, calciuria 3,74 mmol/l, ur creatinine 3,60 mmol/l. • WBC cystin 0.69 nmol/mg protein

04/03/2019 11/45

A STANDARD PRESCRIPTION

S. CO… age 6 years, BW 16 Kg Paris, April 4, 2012 • INDOCID : 15 mg x 2/day, • PHOSPHONEUROS : 1 ml x 2/jour, • BICARBONATE DE SODIUM : 10 g/day, • NaCl : 1 g x 2/day, • POTASSIUM BICARBONATE : 1 g x 2/day, • DIFFU-K 1 pill x2/day • UN ALFACALCIDOL : 0,4 µg/day, • UVESTEROL : 1 dose n°1/day, • LEVOCARNIL : 500 mg x 2/day • L-THYROXIN 75 µg/day • RP103 : 325 mg x 2/day • CYSTEAMINE eyedrops : 1 drop/eye x 4/day.

31 pills/day 04/03/2019 12

the seminal work

Thoene 1976

04/03/2019 14

the seminal work

04/03/2019 15

the seminal work

Thoene 1976 IV Cysteamine 10 mg/kgBW

Cysteamine every 6 h Dosage at 3 hours

04/03/2019 16

Mechanism of action the disulphide interchange

Wilmer 2011, adapted from Gahl, 1985

PQLC2

04/03/2019 17

Cysteamine bitartrate

• CYSTAGON® – Immediate release

(stomach) – 4 times a day

• Dose kids 1900 mg/m²/d • Dose adults 2 g/day

• PROCYSBI® – Delayed release (small

intestine) – Twice a day – Cost x 20

• Dose kids 1900 mg/m²/d • Dose adults 2 g/day

04/03/2019 18

Pharmacokinetics dosage of cysteamine

• Blood sample into ACD

• Denaturation of plasma by DTT (reduction of disulfide bonds)

• Deproteination of plasma by sulfosalicylic acid

• Addition of NEM

• Dosage using LC-MS/MS

04/03/2019 19

Pharmacodynamics dosage of intracellular cystin

The classical method • Blood sample intocitric

acidcitrate-dextrose (ACD) • Total WBC • liquid chromatography-

tandem mass spectrometry: dosage of cysteine

• Protein dosage by BCA • Released in

nmol/mg protein of hemicystine

• Target < 1

The accurate method • Blood sample intocitric

acidcitrate-dextrose (ACD) • Neutrophils (PNN) • liquid chromatography-

tandem mass spectrometry: dosage of cysteine

• Protein dosage by Lowry • Released in

nmol/mg protein of hemicystine

• Target < 2.5

04/03/2019 20

No clear correlation between PD & PK

Langman 2014

04/03/2019 21/45

Pharmacokinetics & pharmacodynamics individual variability

Plasma cysteamin PNM cystin concentration

Bouazza 2011

2 µmol/L

30 µmol/L Methods of dosage: PNN-MS-Lowry

04/03/2019 22

LA VRAIE VIE

0

500

1000

1500

2000

2500

3000

3500

4000

4500

5000

20/

01/1

999

08/07/1

999

24/05/2

000

22/11/2

000

30/05/2

001

30/05/2

002

24/09/2

003

21/06/2

004

24/02/2

005

04/10/2

005

24/05/2

006

12/02/2

007

28/08/2

007

08/07/2

008

09/12/2

008

23/06/2

009

21/01/2

010

08/07/2

010

cy

sta

go

n m

g/d

-6

-4

-2

0

2

4

6

8

10

12

14

PN

N c

ys

tin

nm

ol/m

g

04/03/2019 23

CYS

TAG

ON

(m

g/j

)

Cys

tin

e in

tra-

leu

cocy

tair

e (n

mo

l/m

g p

roté

ine)

Adaptation to body weight or to body surface ?

04/03/2019 24

CYSTAGON: Night pause 6 hours vs 9 hours

Levtchenko 2006

n = 17 n = 5

Methods: HPLC on PNM/BCA 04/03/2019 25

Residual plasma cysteamine concentration µmol/L

Bouaza 2011

Weight (kg) Observed with PNN cystin < 2

Estimated by the PK-PD model

10-17 4.8 4.3

17-25 6.1 5.4

25-40 6.4 6.5

40-70 7.0 7.9

Median in µmol/L

In my clinical experience: 2-3 µmol/L

04/03/2019 26

Effect of diet on PK

Dohil, 2012

04/03/2019 27

intestinal absorption

Dohill 2006 04/03/2019 28

Delayed release cysteamine Cystagon vs Procysbi – PK & PD

Langman 2012

Methods of dosage: WBC-MS-BCA 04/03/2019 29

Quality of life

0,0

5,0

10,0

15,0

20,0

25,0

Physical Emotional Social School Total

PEDSQL score (%) improvement after switch from cysteamine IR Q6H to cysteamine DR Q12H and

estimated trend at 24 months

% Change in PedsQL score (6 weeks)

% Change in PedsQL score at 24 months (trend)

p=0,05

NS NS

NS

NS

P<0,05

P<0,01

NS

P<0,05

NS

Langman 2014

04/03/2019 30

Prevention of Fanconi syndrome in sibling: a success in 1 case

Da silva 1985

04/03/2019 31/45

Prevention of Fanconi syndrome in sibling: a failure in 3 cases

Reznik 1991

1 2 3 N

Cysteamin dose

g/m²

1.30 1.95 1.95 -

Age at start 12 h 9 d 3 w -

FE phosphate % 45 35 38 <20

Proteinuria

g/mmol 1.3 0.5 1.6 <0.20

Aminoaciduria

µmol/kg/d

900 433 1000 <200

GFR

mL/min/1.73 m² 95 113 82 >90

Age at Fanconi syndrome in 1st Siblings F1: 2 years,

F2: 9 months F3: 6 months

04/03/2019 32

death

Brodin Sartorius 2012

treatment

started < 5 years of age

no treatment

treatment

started

< 5 years

of age

04/03/2019 33

delayed ESRD

Brodin Sartorius 2012

treatment

started

< 5 years of age

or no treatment

treatment

started

< 5 years of age

04/03/2019 34

extrarenal disease

Brodin Sartorius 2012

Treatment started < 5 years of age

Treatment started > 5 years of age

No treatment

04/03/2019 35

Adverse events halitosis

Besouw 2007

04/03/2019 36

Adverse events gastrointestinal intolerance

Dohil, J Pediatr 2003;143:224-30 04/03/2019 37

Adverse events skin & angioendotheliomatosis

Besouw 2011 & 2013

HDMVEC viability Newly formed vessels

04/03/2019 38

Adverse events cancer?

type Age (years) reference

Renal carcinoma 18 Cameron 2008

Cerebral medulloblastoma 4 Greco 2010

hemangioblastoma 13 Greco 2010

Thyroid follicular adenoma 23 Greco 2010

04/03/2019 39

Adverse events pseudo-osteolathyrism?

• Neurotoxin – β-N-oxalyle-L-alpha β-

diaminopropionic acid (βODAP)

• Bone toxin – beta-aminopropionitrile

(BAPN)

– Antagonist of lysyl oxydase

Lativus sathyris Or

Spanish lentils

Besouw 2013 04/03/2019 40

Compliance to Cystagon a major issue at adolescence

patient motivation self-reported adherence

self-reported daily doses

Ariceta 2014

04/03/2019 41/45

Compliance a major issue to delay ESRD

Nesterova 2015

Compliance score

score Level of WBC cystin nmol/mg proteins

0 ≥3

1 ≥2 to <3

2 ≥1 to <2

3 <1

Sum of score/year prior to ESRD

04/03/2019 42

No effect on mTOR down-regulation

Andrzejewska 2015

Level of phosphorylation of the ribosomal protein S6 kinase beta-1 [SISK1] in %

aminoacids Fetal Bovine Serum

04/03/2019 43

an alternative to cysteamine or a dream ?

4-month post transplant 8-month post transplant

KO Knock-Out for cystinosin gene HSC Hematopoietic Stem Cells pCCL Construct of CTNS gene in lentivirus WT Wild Type

Harrisson et al 2013

Cys

tin

co

nte

nt

leve

l (n

mo

l/m

g p

rote

in)

KO transplanted with [pCCL ctns-/- HSC] or with [WT congenic HSC]

04/03/2019 44

Omar Fakhouri

Hôpital Universitaire Saint-Georges Beyrouth

Ocular Cystinosis:

< 1 year old: Periphery of the cornea DIAGNOSTIC TOOL

3 to 12 years old: All the cornea PHOTOPHOBIA

13 to 40 years old: All the eye RETINAL BLINDNESS

Ocular Cystinosis:

Systemic Cysteamine Systemic Cysteamine

PHOTOPHOBIA

Corneal Cystine Crystal Score (CCCS or Gahl’s score)

From photophobia…to blindness without treatment!

Corneal scaring

Cysteamine (gel-formulation)

CystadropsTM

V1 / Day 1 Score = 3,8

V2/Day 30 Score = 2,6

Crystals reduction evidence in the stroma between D1 & D30

Cysteamine (gel-formulation): Decrease of IVCM score after 30 days treatment

A, Labbé, 2014

-0.10

-1.16

-1.02

-0.74

-0.52

-0.43

-0.88

The efficacy of Cystadrops® was shown in each corneal layer

Cysteamine (gel-formulation): Crystal deposits decreased over time in all layers

A, Labbé, 2014

make them grow in length HOW?

• Prevent extracellular dehydration

• Prevent metabolic acidosis

• Prevent bone deformation

• Maintain nutritional intake

• If necessary, rGH, L-thyroxin

• Early treatment by cysteamine

SODIUM DEPLETION

1- FANCONI syndrome = sodium wasting

LOOP

30%

COLLECTING

DUCT

0-3%

Proximal

Convoluted

Tubule

60%

Distal

Convoluted

Tubule

7%

geo

rges

.des

chen

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rdb

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G

Initial volume

175 liters

Final urine

<1 liter

Final urine

>3 liters

Na + H2O

urinary

output

ECF - interstitium

Na + H2O

intake

ECF -

plasma

en

do

th

eliu

m

INTRACELLULAR FLUID

K+ Na+

Lymphatics &

Thoracic duct

H2O

ECF = EXTRACELLULAR FLUID

Homeostatic for the osmotic pressure

cell mass

SODIUM DEPLETION

2- sodium wasting = sodium depletion

= ECF contraction = chronic dehydration

– Hyponatremia

– Increased plasma Protides

– High plasma renin and aldosterone

– And urine sodium>0

– Growth impairment

– Severe acute dehydration and cardiovascular shock in case of gastroenteritis, vomiting, diarrhea

SODIUM DEPLETION

3- ECF contraction = failure to thrive

• ADD SODIUM INTAKE AND PREVENT

WASTING

– sodium chloride, sodium bicarbonates

– Non Steroidal Anti Inflamatory drugs • Indometacin 2-3 mg/kgBW/day

• Ibuprofen 10-30 mg/KgBW/day – pediatric suspension 20 mg/mL

– One dose for 1 KgBW = 0.5 mL

• STOP when GFR is < 30 ml/min/1,73m²

• BIOMARKERS TO FOLLOW

– Plasma protides, serum potassium

– Plasma renin

– Height

SODIUM DEPLETION

4- TREATMENT OF SODIUM DEPLETION

INDOMETACIN IMPROVES •Sodium reabsorption •Chronic extracellular

dehydration •Growth in length

ALDOSTERONE

PROSTAGLANDINS

+

-

-

PCT 60%

LOOP 30%

DCT 7%

CO

LLECTIN

G D

UC

T 0-3%

+

INDOMETHACIN X

SODIUM DEPLETION

5- prevent wasting: indomethacin

Sodium Pump

K+

ATP

ADP

Na+

GL

UC

OS

E

O2

GLUT2

FGF23

AQP3

AQP1

H2 O

Na-G

luco

se

Na-A

cides am

inés

Na-A

cides o

rgan

iqu

es

NPT2a

NPT2c

NHERF

+

Lysosome

PROTEINES

Cl-

OCRL1

Cystin

+

AA

ClC5

CT

NS

mégaline-

cubuline

exocytosis

endocytosis CO2

+ H2O

Car4,Car14

Car2

H++ HCO3--

CO2 + H2O

H++ HCO3--

Na-bicarbonates

HCO3-- Na+

NH

E 4

geo

rges

.des

chen

es@

rdb

.ap

hp

.fr

METABOLIC ACIDOSIS

1- BICARBONATES WASTING

H+

• BLOOD ACIDOSIS

– Growth retardation

– Inceased respiratory frequency

– Worsened tubular function

• ADD BICARBONATES or CITRATE

or BOTH

– Sodium bicarbonates or Na-citrate

– Potassium bicarbonates or K-citrate

• BIOMARKERS TO FOLLOW

– Serum bicarbonates and chlorides

– Height

METABOLIC ACIDOSIS 2- EFFECT AND TREATMENT

Sodium Pump

K+

ATP

ADP

Na+

GL

UC

OS

E

O2

GLUT2

FGF23

AQP3

AQP1

H2 O

Na-G

luco

se

Na-A

cides am

inés

Na-A

cides o

rgan

iqu

es

NPT2a

NPT2c

NHERF

+

Lysosome

PROTEINES

Cl-

OCRL1

Cystin

+

AA

ClC5

CT

NS

mégaline-

cubuline

exocytosis

endocytosis CO2

+ H2O

Car4,Car14

Car2

H++ HCO3--

CO2 + H2O

H++ HCO3--

Na-bicarbonates

HCO3-- Na+

NH

E 4

geo

rges

.des

chen

es@

rdb

.ap

hp

.fr

CONTROL PHOSPHATE DEPLETION

1- PHOSPHATE WASTING & VITAMIN D DEFICIENCY

H+

• SIGNS

– Hypophosphatemia

– Fatigue and muscle weakness

– Bone deformation

– Rickets

BONE DEFORMATION

2- PHOSPHATE DEPLETION & VITAMIN D DEFICIENCY

– DiSodium Phosphate (phocytan ®) • Vials of 10 mL at 0.66%: 10 ml = 6.6 mmol eq. to 200 mg

of phosphorus or 620 mg of phosphates (Na intake = 13.2 mmol)

• Flask of 100 ml at 0.33%: 100 mL = 33 mmol eq. to 1000 mg

of phosphorus or 3100 mg of phosphates (Na intake = 66 mmol)

– Phosphoric acid (phosphoneuros®) • 10 drops = 0.43 mL = 79 mg phosphorus

– alfacalcidol® • 1 drop = 0,1 µg • or capsules at 0.25, 0.50 & 1µg • Initial dose 0.25 µg/day then adapt to biomarkers

– BIOMARKERS

• Serum calcium and phosphate • Plasma PTH and FGF23

Age

Serum Phosphorus

[mg/dL]

Plasma

Phosphates mmol/L

Alcalines

Phosphatase

[UI/L]

0-12 months 4.8 - 7.4 1.55-2.39 >350

1-5 yrs 4.5 - 6.5 1.45-2.10 100 - 350

6-12 yrs 3.6 - 5.8 1.16-1.87 60 - 450

13-20 yrs 2.3 - 4.5 0.74-1.45 40 - 180

BONE DEFORMATION

2- TREATMENT BY SUPPLEMENTS

NUTRITIONAL STATUS

1- FEEDING TROUBLES

Elenberg 1998

Consider gastrostomy

in very young patients

NUTRITIONAL STATUS

2- GASTROSTOMY

Coleman 2000

Lean mass in KgBW

Fre

e c

arn

itin

in µ

mo

l/L

NO CARNITIN DEFECT IN GRAFTED PATIENTS

LEVOCARNIL 500 to 2000 mg per day

relationship between carnitin and lean mass

NUTRITIONAL STATUS

3- L-CARNITINE

Guillen Lopez 2015

RECOMBINANT GROWTH HORMONE

Recombinant Growth Hormone : 0.20 mg/kgBW/WEEK subcutaneous

Genotonorm ® Humatropen ® Nordipen ®

For instance final height 160 cm

indocid

GH

NaCl

deux soeurs

x x

x

x x

x x x

x x

x x

x

x

x

x x

x

x

traiter le plus tôt possible

CONCLUSIONS

• The cysteamine dramatically improves the prognosis of cystinosis but does not prevent the Fanconi syndrome

• Gastro-intestinal adverse events are frequent and often discouraging

• Compliance is a major issue

• Cysteamine is not all – other treatments remain to be discovered to complete lysosomal depletion

04/03/2019 73