BeSHG Course 2016

8 January 2016

Center for Inherited Metabolic Diseases

Institut de Pathologie et de Génétique (IPG)

Dominique.roland@ipg.be

Therapeutic options in Inborn Errors of Metabolism (IEM)

Toxic compounds

Accumulation

Substrate

Deficiency

Consequences of protein/enzyme deficiency in IEM

DNA mutation

Non functional/ missing protein/enzyme

Severe phenotype absence of protein

mutant protein with no residual activity

Mild phenotype mutant protein with residual activity

treatment will try to increase the small amount of function

Prenatal symptoms (Echo, RMN)

Neonatal screening

Acute neonatal symptoms

Symptoms after a « free interval »

Specific symptoms (eyes, skin, liver, heart, kidney,..)

Chronic/progressive symptoms (failure to thrive, neurologic

deterioration, ..)

Persistent and without explanation symptoms

Diagnostic circumstances of IEM

For the same enzymatic defect

Neonatal / late onset/ asymtomatic

Groupe 1 Complex Molecules

Groupe 2 Intermediary metabolism : Intoxication type

Groupe 3

Intermediary metabolism : Energy depletion

Classification of IEM

Complex Molecules

PROTEINS POLYSACCHARIDES LIPIDS

Amino Acids Monosaccharides Glycerol

Fatty Acids

Organic Acids

Pyruvate

Acetyl-CoA Simples

Molecules

NH3

Krebs Cycle CR ATP

Energy Metabolism

H2O CO2

1. IEM of cellular organelles (trafficking, processing)

2. Disturb synthesis or catabolism of complex molecules

3. Embryo-fetal development possible interference

4. No provocative circumstances

5. Symptoms are permanent, progressive, independent of the diet

6. Storage disorders (lysosome)

7. Specific diagnostic methods (substrates, enzyme assays, molecular analyses)

8. Enzyme replacement & substrate reduction treatments

Lysosome Peroxisome

Endoplasmic Reticulum

Classification of IEM

- Lysosomal storage diseases (LSD) - Peroxisomal diseases - Congenital disorders of glycosylation (CDG)

Group 1 : Complex Molecules

(simple molecules) A B

- Aminoacidopathy (PKU, Tyrosinemia, …)

- Urea cycle defect (UCD)

- Organic aciduria (MMA, IVA, PA, …)

- Carbohydrate (galactosemia, hereditary fructose intolerance)

1. Accumulation of toxic compounds

2. No consequence on embryo-fetal development

3. Symptom free interval

4. Clinical signs of intoxication (acute or chronic)

5. Provocative circumstances (catabolism, food intake)

6. Can be late in onset and intermittent

7. Diagnosis based on plasma / urine analyses

8. Most are treatable (extra-corporeal procedures, special diets, « cleaning drugs »)

Group 2 : Intermediary metabolism – Intoxication type

Classification of IEM

- Fatty acids ß-oxidation defects

- Ketone bodies synthesis defects

- Neoglucogenesis defects

- Mitochondrial diseases (Krebs cycle defects and oxidative phosphorylation)

1. Deficiency in energy production or utilization

2. Main target organs (liver, heart, muscle, brain)

3. Embryo-fetal development possible interference

4. Provocative circumstances (catabolism, food)

5. Diagnosis is difficult (function tests, enzymatic assays on various tissues, molecular analyses)

6. Only few are amenable to treatment

Group 3 : Intermediary metabolism – Energy depletion

Classification of IEM

Day 3-5 of life

NEONATAL SCREENING in IEM Neonatal screening in IEM on DBS

Dried blood spot

(DBS)

With Tandem Mass Spectrometry (MS/MS)

> 30 Inborn Errors of Metabolism diagnosed

> 40 metabolites analyzed (amino acids, acylcarnitines, ..)

• « Consider IEM in parallel with other more common conditions

• Be aware of symptoms that persist and remain unexplained after initial

treatment.

• Don’t confuse a symptom or a syndrome with etiology.

• IEM can present at any age from fetal life to old age.

• Although most IEM are autosomal recessive disorders, majority of

cases appear sporadic.

• Take care first of the patient (emergency treatment), and then of the

family (genetic counselling).

• Initially consider IEM amenable to treatment : »

DON’T MISS A TREATABLE disorder

When to think about IEM ?

Therapeutic approaches to IEM

Pharmacologic enzyme replacement Substrate reduction

Correcting product deficiency

- Replenish depleted product

- Increasing substance supply

- Providing alternate substrate

Transplantation

- Inhibition of enzyme

within pathway :

e.g NTBC in TYR1

- Dietary restriction :

e.g. PKU, galactosemia

Decreasing metabolic toxicity

- Removing toxic metabolite

- Blocking the effect of toxic metabolites

- Co-enzyme treatment

- Enzyme enhancement therapy

Stimulation residual enzyme

A B -

+

- Hematopoietic Stem cell Transfer

- Other organ transplantation

Lysosome

- Mucopolysaccharidosis (MPS

- Oligosaccharidosis

- Sphingolipidosis

Group 1 : Complex molecules

- Lysosomal storage diseases (LSD)

• LSD 1/6.000 – 7.000 birth

• LSD are progressive and multisystemic

• > 40 LSD identified

Synthesis

Degradation

inhibition

Lysosomal

storage

Reduce substrat synthesis :

Substrate reduction therapy (SRT)

Synthesis Degradation

Storage

Principles of treatments in LSD

Increase degradation : Enzyme Replacement Therapy (ERT) Enzyme Enhancement Therapy (EET) or « Chaperones »

Deficient breakdown of glycosaminoglyvans (GAGs)

Dermatan sulfate degradation Incidence 1/100.000

Mucopolysaccharidosis (MPS)

Sometimes, the same enzyme participates

to different GAGs breakdown

Biochemical Diagnosis in MPS

• The primary diagnostic test for MPS disorders is ALWAYS urine

glycosaminoglycans (GAGs) analysis.

• 10mls urine is required.

• Quantitative analysis. Total GAGs concentration is measured using

Dimethylene Blue (DMB).

• False negative exists mainly in attenuated forms of MPS

Total Glycosaminoglycans

age mg/g creat

<6 mo 135 460

6-12 mo 134 279

1-2 y 81 265

2-4 y 68 188

4-6 y 67 127

6-8 y 50 114

8-10 y 46 103

10-15 y 30 94

15-20 y 13 59

>20 y 13 45

CS

CS : Chondroïtine sulfate

HS : Heparan sulfate

DS : Dermatan sulfate

CS

CS DS

HS

HS

• Qualitative test establishes the identity of the GAGs present

• GAGs are extracted from 2ml urine and precipitated

with Alcian Blue.

• Extract is washed and analysed by 2D electrophoresis.

DS

Biochemical Diagnosis in MPS

Biochemical Diagnosis in MPS – GAGs and enzyme deficiency

Genet. Mol. Biol. 1998, vol. 21 n.1, 1678 TLC : thin layer chromatography

Disease-specific treatments - MPS

Treatment options, no curative therapy

MPS I Hurler (MPS IH) : severe SCT (< 2 y), ERT, symptomatic

Hurler-Scheie (mPS IH/S) : less severe ERT, symptomatic

Scheie (MPS IS) : least severe ERT, symptomatic

MPS II Hunter (MPS IIA), severe ERT (SCT), symptomatic

Hunter (MPS IIAB), less severe Symptomatic, (ERT)

MPS III Sanfilippo A (MPS IIIA) Symptomatic, ERT in development

Sanfilippo B (MPS IIIB) Symptomatic

Sanfilippo C (MPS IIIC) Symptomatic, ERT in development

Sanfilippo D (MPS IIID) Symptomatic

MPS IV Morquio A (MPS IVA) Symptomatic, ERT

Morquio B (MPS IVB) Symptomatic

MPS VI Maroteaux-Lamy Symptomatic, ERT (SCT)

MPS VII Sly Symptomatic, SCT

MPS IX Hyaluronidase deficiency Symptomatic

ERT : Enzyme Replacement Therapy; SCT : Stem Cell Therapy

Age at diagnosis 0.2–7 years 0.2–36 years 2–54 years

Effect on cognition Pronounced mental delay with

loss of acquired skills

No/mild mental delay;

learning disabilities

No

impairment

Mean life expectancy

(untreated) 7 years Approximately

20 years

Adulthood

HURLER HURLER-SCHEIE SCHEIE

SEVERE FORM ATTENUATED FORM

Phenotype distribution* ~65% ~25% ~10%

*based on Moore et al. Orphanet J Rare Dis 2008;3:24 and MPS I Registry data

Treatment according to clinical phenotype in MPS I

*Prediction of disease severity based on clinical

picture, neurodevelopmental testing, genotype and

other relevant information

2 years of age

DQ < 70

MPS I Diagnosis

> 2 years of age

DQ 70 Severe or unknown

phenotype predicted

Attenuated

phenotype predicted*

DQ < 70 DQ 70

ERT (Consider

HSCT

in rare cases)

ERT ERT

Consider

ERT

Consider

HSCT

Treatment algorithm for MPS I

From Muenzer et al. Pediatrics 2009;123:19-29

DQ = Developmental quotient

HSCT = Hematopoietic stem cell transplant

Treatment is adapted to phenotype

Hematopoietic Stem Cell Transplantation (HSCT)-Principle

- Enzyme deficiency corrected by donor cells

- Better respons is some diseases.

Stem cells are self-renewing cells defined by 2 properties

Stem cell transplantation

1. Ability to proliferate to form the differentiated cell types of a

tissue in vivo

2. Ability to self-renew to form another stem cell

Origin : embryonic, fetal, cord blood, adult

Stem cell transplantation

iPSC cells: Induced-pluripotent stem cells

Stem cell

Two mechanisms by which bone marrow transplantation or gene transfer into bone

marrow may reduce the substrate accumulation in LSD

Hematopoietic Stem Cell Transplantation (HSCT)-Principle

a. Hematopoietic stem cells from bone marrow

• Limitations

– Early diagnosis (before irreversible brain damage)

– Matching Donor

– Procedure-related mortality and morbidity

– Variable results on brain and bone

– Do not cure the disease but changes the natural history

BMT - Limitations and Evolution

b. Hematopoietic stem cells from placental cord blood

- Increased tolerance of histoincompatible donor cells

- Reduced risk of graft-versus host disease

- Widely available

HSCT (since 1981) ERT (since 2003)

Patients • Hurler patients <2y who have a

DQ >70

• Due to risks, not recommended

for attenuated forms

• Hurler patients who are not

candidates for HSCT

• Hurler-Scheie and Scheie

patients

Risks • 5-30% mortality

• 60 % complication

• 10-15 % failure

• 40 % risk of mild to moderate

infusion associated reactions

during first 6 months of

treatment

• Very small risk of life-

threatening infusion reactions

Benefits • If performed early enough can

preserve neurocognition

• Prolongs survival

• Some somatic benefits

• No infusion needed

• Some somatic benefits

• For all disease phenotype

• As adjuvant treatment before

HSCT to improve pre-

transplant clinical condition

and engraftment

Limits • Must be done < 2 y • Does not cross BBB

• Weekly infusions for life

Disease-specific treatments in MPS I

BBB : Blood Brain Barrier

Enzyme replacement therapies in LSD

Disease ERT

Gaucher type 1 Cerezyme® (Imiglucerase)

VIPRIV® (Velaglucerase α)

Elelyso® (Taliglucerase α)

CHO cells

Human cells

Carrot cells

Fabry disease Replagal® (agalsidase α)

Fabrazyme® (agalsidase β)

Human cells

CHO cells

MPS I (H,HS,S) Aldurazyme® CHO cells

MPS II Elaprase® CHO cells

MPS VI Naglazyme® CHO cells

Pompe disease Myozyme® CHO cells

CHO cells : Chinese Hamster Ovary cells

Bacterial DNA Plasmid

Culture Medium

Protein

CHO Host Cell

Human glucocerebrosidase gene

GENZ/07-2012/C-802

Recombinant DNA technology to manifacture ERT

Disease rh-enz Nbr % Ab % patients with

reactions

Gaucher Cerezyme 1322 15 % 13,8 %

MPS I Aldurazyme 55 91% 32 %

MPS II Elaprase 11% 55 %

MPS VI Naglazyme 10 100 % 5 %

Fabry Fabrazyme

Replagal

58

55

89 %

55 %

52 %

10 %

Pompe Myozyme 3 66 % 66 %

Brooks et al. Trends Mol Med 2003, 9:450-3

ERT – Frequent immune response

• Infusing a foreign protein/enzyme not synthesized by the mutant DNA bears the risk of immune

reactions and/or enzyme activity inactivation

• Increased IgG antibody levels were detected during most treatments, but without correlation

between the occurrence of severity of adverse events and the presence of high antibody titers

• Neutralizing antibodies were (most of the time) not associated with a reduction in efficacy of

the enzyme preparation (or transient)

• Most infusion-related reactions are mild (fever, flush, tachycardia, ..)

• Hypersensitivity/anaphylactic reaction against the infused enzyme can be treated by slowing

down the infusion rates and premedication with antihistamines and/or corticosteroids

• In an ongoing strong immune response, tolerance induction by drugs such as methotrexate or

rituximab may become necessary (CRIM negative Pompe patients)

SRT are efficient if there is persistant residual

degradation activity

Synthesis Degradation

Storage SRT

• Possible application on glycosphingolipids synthesis

• Application with Gaucher disease

Substrate Reduction Therapy (SRT) in LSD - principle

Glycosphingolipids Storage Diseases

Glycosphingolipids

storage Gaucher

cell

Glucocerebrosidase

Heterogeneous group of diseases

Charrow J et al., Arch Intern Med 2000;160:2835.

Heterogeneity in clinical

presentation

Clinical diagnosis at every age

Pathologic fracture (15%)

Osteonecrosis (25%)

Osteopenia (42%)

Anemia (64%)

Thrombocytopenia (56%)

Hepatomegaly (79%)

Splenomegaly (87%)

Bone pain (63%)

Bone crisis (33%)

Joint collapse (8%)

General symptoms:

• Fatigue

• Easy bruising/bleeding

• Menorraghia

• Decreased appetite

• Abdominal pain

• Growth retardation

• Slow pubertal development

Bone marrow infiltration

(40%)

Interstitial Pulmonary

fibrosis

Erlenmeyer flask

deformity (46%)

Multisystemic symptoms in Gaucher Disease

Phenotypic continuum in Gaucher Disease

Non-neuronopathic GD

~ 95 %

Asymptomatic

Skeletal disease

Visceral disease

2e neurologic

involvement

Parkinsonian

Manifestations

Eye movement

disorder

Hydrocephalus,

cardiac valve

calcifications

Myoclonic

epilepsy

Progressive

neurological

degeneration

Congenital

icthyosis

Hydrops

fetalis

Neurological manifestations Type 1

Type 3 Type 2

Neuronopathic GD

Before treatment

girl of 8 y and 8 months

After treatment

Girl of 10 y and 10 months

Clinical Response to ERT in Gaucher Disease

n=420 n=458

Liver Volume Spleen Volume

Andersson et al, Pediatrics 2008;122(6):1182-1190S

n=768

n=771

Platelet Count Hemoglobin Level

Visceral organ and

hematologic responses to

ERT treatment in children

Clinical Response to ERT in Gaucher Disease

No brain access - tried in Gaucher type II (neurologic form)

Immune response

Limited results on bone

Limitations of ERT

Intravenous infusion therapy

NB-DNJ

Substrate Reduction Therapy in GD

Imminosucre N- butyldesoxynojirimycine (NB-DNJ) (Miglustat ®) (analogue of glucose) inhibits glucocerebroside synthesis

Indicated in mild to moderate GD type I in case ERT is not an option

Intended to treat neuronopathic GD (GD3), but not showing effectiveness in clinical trials.

Oral therapy

But inhibits also other enzymes (a-glucosidases I et II) Explains adverse effects as diarrhea (85%), weight loss (65 %). Peripheral neuropathy

Advantage : Cross the blood-brain barrier

NB-DNJ

Substrate reduction therapy in GD

Eliglustat ® , new SRT

Approved for GD type I (FDA 2004, EMEA 2015 approved)

First line oral therapy in mild to moderate GD type 1

Effects : reduction of liver and spleen volume, increase of hemoglobin and platelet count

Effects on bone (more rapid than with ERT)

Adverse effects (≥10%): arthralgia, headache, nausea, fatigue, back pain, pain in extremities.

Contraindications: Dosage adapted to metaboliser profil (poor (PM)/intermediate (IM)/extensive (EM).

Electrocardiographic changes and potential cardiac arrhythmias. Not recommended in patients with pre-existing cardiac disease, long QT syndrome, and concomitant use of class Ia and class III antiarrhythmics.

Limits : Does not Cross the blood-brain barrier

Normal « misfolding »

chaperones specific of active site

Enzyme Enhancement therapy or pharmacological « charperone »

• Novel treatment concept in development for Fabry, Pompe, Gaucher disease

• Competitive inhibitor of the enzyme at subinhibitory concentrations can act as a

chemical chaperone leading to correct conformation of the mutant enzyme

Advantage :

- Orally administered

- Better biodistribution profile in comparison with recombinant enzyme

- Binds to amenable mutant forms of enzyme and increases trafficking

to lysosomes

- Chaperons are not immunogenic

Disadvantage :

- Treatment effect is restricted to patients with amenable mutations.

- Only 10–15% Pompe patients are amenable to chaperons, 30-50%

of currently diagnosed Fabry patients are believed to have mutations

amenable to treatment (under evaluation)

Enzyme Enhancement therapy or pharmacological « charperone »

- Aminoacidopathy (PKU, tyrosinemia, …)

- Urea cycle defect (UCD)

- Organic aciduria (MMA, IVA, PA, …)

- Carbohydrate (galactosemia, hereditary fructose intolerance)

Group 2 : Intermediary metabolism – Intoxication type

Classification of IEM

- Fatty acids ß-oxidation defects

- Neoglucogenesis defects

- Ketone bodies synthesis defects

- Mitochondrial diseases (oxydative phosphorylation and Krebs cycle defects)

Group 3 : Intermediary metabolism – Energy depletion

Neurological

Deterioration

PKU

MSUD

MMA

PA, IVA

UCD

MCD

Predominant

Seizures

Pyridoxine

Pyridoxal P

Folinic acid

MCD

3PGD

GLUT-1

Jaundice

Liver failure

Galactosemia

Fructosemia

TYR-1

CDG-1b

Bile acids

LCHAD

Cardiac

FAO

Persistent

Hypoglycemia

PHHI

FAO

Glycogenosis

Hormones

Emergency treatment must be undertaken

in parallel with investigations

First think of treatable disorders

IEM (Intoxication, Energy depletion)

Neurological

Deterioration

PKU

MSUD

MMA

PA, IVA

UCD

MCD

Predominant

Seizures

Pyridoxine

Pyridoxal P

Folinic acid

MCD

3PGD

GLUT-1

Jaundice

Liver failure

Galactosemia

Fructosemia

TYR-1

CDG-1b

Bile acids

LCHAD

Cardiac

FAO

Persistent

Hypoglycemia

PHHI

FAO

Glycogenosis

Hormones

First think of treatable disorders

Anabolism

Epuration

Special diets

Glucose

Vitamins

Amino Acids

Special diet

Avoid fasting

L-carnitine

Special diet

IEM (Intoxication, Energy depletion)

PKU : Phenyketonuria

MSUD : Maple sirup disease

MMA : Methyl malonic aciduria

PA : Propionic aciduria

IVA : Isovaleric aciduria

UCD : Urea cycle defect

MCD : Multiple carboxylase deficiency

Pyridoxine responsive seizures

Pyridoxal Phosphate responsive seizures

Folinic acid responsive seizures

3PGD : 3 Phosphoglycerate dehydrogense deficiency

GLUT-1 : Glucose transporter 1

TYR 1 : Tyrosinemia type 1

CDG-1b : Congenital disorder of glycosylation type 1b

LCHAD : Long chain acylCoA dehydrogenase

FAO : Fatty Acid oxidation defects

PHHI : Persistent hyperinsulinemic hypoglycemia of infancy

Disease specific treatments – Neurologic deterioration

PKU : Phenylketonuria

• Autosomal recessive disease

• 1 / 10 000 birth

• PAH gene (chromosome 12)

• PAH - tetrameric structure

• Classic PKU is caused by a complete or near-complete

deficiency of phenylalanine hydroxylase activity (PAH) in liver.

• PAH deficiency results in intolerance to the dietary intake of the

essential amino acid phenylalanine and produces a spectrum of

disorders

Phenylalanine Hydroxylase (PAH) system

BH4, a natural

cofactor of PAH

Dried blood spot on filter paper :

Guthrie card bacterial inhibition assay

R Guthrie in early 1960s

PKU = first metabolic disease detected through Neonatal Screening - increased PHE levels

0

5

10

15

20

25

30

35

40

45

50

0 0.5 1 1.5 2 2.5 3 3.5 4 4.5

1 - 3

60 - 180

> 20

>1200

10 - 20

600 - 1200

3 - 10

180 - 600

Normal Hyperphenylalaninemia atypic PKU Classical PKU

mg/dl

µmol/l

PHE

PHE plasmatic levels and PKU classification

PAH activity PHE level Daily PHE

tolerance

Classical PKU :

0-1 % > 20 mg/dl 200 – 350 mg

Atypical PKU :

1-3 % 10 - 20 mg/dl

350 - 850 mg

Non-PKU

Hyperphenylalaninemia

3 - 5 % PHE 3 - 10 > 850 mg

Mild to severe mental retardation

Neurologic symptoms

• Microcephaly

• Gait instability, tremor

• Epilepsy

• Autistic behavior

• Auto and hetero aggressivity

Eczema

Decreased skin and hair pigmentation

(Blond hair, blue eyes)

Structural brain changes and white matter

abnormalities

Musty body odor (typical)

Untreated classical PKU

Horst Bickel (1953)

Phenylketonuria = Low-phenylalanine diet

Lancet, 1953; 2, 812-813

First dietetic treatment for an IEM

Control of natural protein intake according to PHE tolerance

Avoidance of high protein food

(milk, dairy products, meat, fish, chicken, eggs, beans and nuts,...)

Phenylalanine-free formula (amino acids mixture with vitamins and

oligoelements)

Low protein food (manufactured hypoproteic bread, pasta, biscuits, ...)

No control of protein-free food

0 9 15 11

0

6

12

20

Adult

France

15

Germany

5

GB

4

8

Phe mg/dl

USA

5

2

10 18 (y)

22

10

International recommendations for PHE control according to age and country – no universal consensus

• Enzyme is synthesized but activity is

null or decreased

• PKU as a model of « misfolding » +++

http://www.bh4.org/biopku.html

PAH gene – Importance of missense mutations

• BH4 = Natural cofactor of aromatic

amino acid hydroxylases

• Sapropterin (6R-BH4) synthetic form

of tetrahydrobiopterin

• Stabilization of the active tetramer

forms of the mutant protein

• Protection from inactivation

• Acts as a « chemical chaperone »,

preventing misfolding

• Orphan drug (FDA and EMEA)

~ 500 mutations worldwide

Responder

Different responses to oral BH4 loading test (20mg/kg)

• About 70 % of mild PKU patients proved to be responder

• About 10 % of classical PKU patient respond to BH4

• In PKU patients responsive to BH4, oral treatment could be used in addition to a

restrictive low-phenylalanine diet and might even replace the diet in some instances.

• Limited adverse effects : upper airway tract infection, headache, vomiting, abdominal pain,

diarhhea, fever, back pain.

• Limits : palatability, compliance

hours

MSUD : Maple Sirup Disease

MMA : Methylmalonic Aciduria

PA : Propionic Aciduria

IVA : Isovaleric Aciduria

Disease specific treatments – Neurologic deterioration

Acute and chronic

« intoxication »

• Metabolic decompensation precipitated by :

prolonged fasting, protein overload, infection

• Ketoacidosis

• Accumulation of abnormal organic acids or

carboxylic acids (GC/MS) in urine

• Abnormal Amino Acid chromatography (MSUD)

• Abnormal acylcarnitine profile (on paper filter)

• Neonatal form : metabolic encephalopathy : lethargy, feeding problems, dehydratation, truncal

hypotonia/limb hypertonia, myoclonic jerks, neurovegetative dysregulation cerebral oedema,

coma, multi-organ failure; unusual odor (maple sirup-like odor of urine)

• Chronic intermittent form (up to aduldhood) : recurrent episodes of ketoacidotic coma,

lethargy and ataxia, focal neurologic signs, Reye syndrome

• Chronic progressive form : failure to thrive, chronic vomiting, anorexia, osteoporosis,

hypotonia, psychomotor delay, recurrent infections (particularly candida)

Leucine Isoleucine Valine

2-oxoisocaproic Ac 2-oxo-3-methyl-N-valeric Ac 2-oxoisovaleric Ac

Isovaleryl-CoA 2-méthylbutyryl-CoA Isobutyryl-CoA

3-hydroxy-3-methylglutaryl-CoA

Tiglyl-CoA Methylacrylyl-CoA

2-methyl-3-hydroxybutyryl-CoA 3-hydroxyisobutyryl-CoA

3-hydroxy-3-methylglutaryl-CoA 2-methylacetoacetyl-CoA

3-hydroxyisobutyric ac

Methylmalonic-semialdehyde

Acetoacetate Acetoacetate

Malonyl-CoA

Acétyl-CoA

Propionyl-CoA

Methylmalonyl-CoA

Succinyl-CoA

3-methylcrotonyl-CoA

BCKD BCKD BCKD MSUD MSUD

MSUD

IVA

PA

MMA

Dehydrogenase

Carboxylase

Mutase

Branched-chain Amino Acids metabolism

B12

B8

MSUD : Maple Sirup Disease

MMA : Methylmalonic Aciduria

PA : Propionic Aciduria

IVA : Isovaleric Aciduria

Acute treatment : - Interrupt catabolic state with high dose glucose IV

- Counteract acidosis

- Stop protein intake

- Remove toxins – depending on the disease and lab findings : increase diuresis,

dialysis, haemo(dia)filtration

- Consider specific treatment (L-Carnitine, B12, B8,..)

Long term treatment :

- Diet – protein restriction (according to safe values, beware of protein deficiency in

case of overtreatment)

- Supplementation of unaffected amino acids mixture

- Supplementations of minerals, vitamins or cofactors

- L-Carnitine in all disorders that cause intramitochondrial accumulation of CoA esters

Disease specific treatments – Neurologic deterioration

Acute and chronic intoxication

Hyperammoniemia is the hallmark of UCD and responsible for severe brain damage

UCD : Urea cycle defect

Disease specific treatments

Proteins

(catabolism, diet)

Ammonia produced by protein catabolism cannot be removed by the UC because of a genetic

defect of a urea cycle enzyme (ASS, OTC, …)

- Protein restricted diet and essential amino acids supplementation

- Arginine and citrulline supplementation (intermediary metabolites of the UC are used as drugs

to avoid deficiencies (and become essential Amino Acids)

- Nitrogen scavengers : alternative excretion of nitrogen by bypassing the UC :

Sodium benzoate and sodium phenylbutyrate

Disease specific treatments – Neurologic deterioration

Administration of sodium benzoate diverts ammonia to glycine synthesis,

and the nitrogen moiety is subsequently excreted as hippurate in urine

Disease specific treatments Disease specific treatments – Neurologic deterioration

TREATMENT :

- Biotinidase deficiency : Biotin 5-10 mg/day (oral)

- HLCS : Biotin 10-20 (-40)mg/day

MCD : Multiple Carboxylase Deficiency (Biotinidase, Holocarboxylase synthetase)

Biotin

cycle Biotinidase deficiency must be ruled out in every

child with unexplained neurologic symptoms even

in absence of cutaneous or laboratory symptoms

Improvement of most symptoms with treatment

Vitaminotherapy

Disease specific treatments – Neurologic deterioration

Holocarboxylase

synthetase (HCLC)

Biotinidase

First symptoms First days of life 1w till 10 y

Hypotonia, coma,

Seizures, hypothermia

Hypotonia, ataxia,

Seizures

Cutaneous symptoms Alopecia, skin rash Alopecia, scaly perioral/facial rash

Periorificial eczema

pigmentation deficit (loss of hair color)

Complications Deafness and optic atrophy

Periventricular leucodystrophy

Or thalami abnormalities (MRI)

Spinal cord and progressive spastic paresis

Intellectual disability and developmental delay

Recurrent viral or fungal infections < immune

dysfunction

Severe metabolic acidosis Metabolic ketoacidosis

Organic aciduria : propionylglycine,

tiglylglycine, methylcitrate, 3-

hydroxypropionique, 3-methylcrotonylglycine, 3-

hydroxyisovalérique

Enzyme activity

Fibroblasts, lymphocytes DBS or plasma

MCD : Multiple Carboxylase Deficiency (Biotinidase, Holocarboxylase synthetase)

Disease specific treatments – Neurologic deterioration Disease specific treatments – Neurologic deterioration

Neurological

Deterioration

PKU

MSUD

MMA

PA, IVA

UCD

MCD

Predominant

Seizures

Pyridoxine

Pyridoxal P

Folinic acid

MCD

3PGD

GLUT-1

Jaundice

Liver failure

Galactosemia

Fructosemia

TYR-1

CDG-1b

Bile acids

LCHAD

Cardiac

FAO

Persistent

Hypoglycemia

PHHI

FAO

Glycogenosis

Hormones

First think of treatable disorders

Anabolism

Epuration

Special diets

Glucose

Vitamins

Amino Acids

Special diet

Avoid fasting

L-carnitine

Special diet

Vitamin responsive neonatal seizures :

- Pyridoxine (vit B6) -responsive seizures - Antiquitin (ALDH7A1) mutation - Pyridoxal Phosphate (PLP) -responsive seizures - PNPO (pyridox(am)ine 5-phosphate-oxidase) deficiency - Folinic acid -reponsive seizures (identical to pyridoxine-dependant epilepsy)

P5P (B6 active form) is cofactor of

> 100 transamination and decarboxylation

reactions in various pathways (serotonin and

dopamine synthesis, GABA synthesis, alanine,

glycine, threonine metabolism

Disease specific treatments – Seizures

Antiquin gene mutation (ALDH7A1)

Increased pipecolic acid

in blood, urine and CSF

Under vitamine B6 treatment :

Persistant pipecolic acid in blood and urine

Chemical inactivation of

PLP by P6C and

accumulation of

potentially neurotoxic

metabolites AASA, P6C

and pipecolic acid

Disease specific treatments – Seizures

AASA P6C

PNPO deficiency

Clinical :

• Refractory neonatal seizures not responsive to pyridoxine but to

pyridoxal phosphate

• Microcephaly

• Prematurity

• Hypotonia

Diagnosis : CSF : Alanine, threonine, glycine

urine : vanillactic acid

Gene : PNPO

Therapy : pyridoxal phosphate 30 mg/kg/day oral in 3 doses

Disease specific treatments – Seizures

Disease specific treatments – Seizures

Pyridoxine 30 mg/kg/J

Pyridoxal-phosphate 10-50 mg/kg/j

Acide folinique 10 mg/j

Biotine 10-50 mg/j

In case of neonatal seizure suspected to be an IEM

A vitaminotherapy is never contraindicated

Dimethylglycine Sarcosine

Glycine

Serine

NH3 + CO2

Serine

Glycine

Methyl THF

3-P-glycerate Glucose P-serine P-OHpyruvate

Mitochondria

Methyl THF

Methyl THF

3PGDH

3PSP

3-PGDH (phosphoglycerate dehydrogenase) deficiency or

Serine synthesis deficiency

Disease specific treatments – Seizures

Severe congenital microcephaly ++

Epileptic encephalopathy

Psychomotor delay

Spastic tetraparesis

Cataracte (sometimes)

Growth retardation

Hypogonadism

3-PGDH (phosphoglycerate dehydrogenase) deficiency or serine synthesis def

Serine/glycine given with different treatment dosages

but a favorable responses were observed :

- Major reduction in seizure frequency

- Some patients became seizure free

- Increased white matter volume

- Progress of psychomotor development in patients,

diagnosed early, and treated with a high dose of L-

serine

- Prenatal treatment and normal head

circumference at birth. Normal neurologic

development at 12 y

De Koning et al, 2013

Disease specific treatments – Seizures

Low serine in CSF and borderline in

plasma (in fasted state)

Neurological

Deterioration

MSUD

MMA

PA, IVA

UCD

MCD

PKU

Predominant

Seizures

Pyridoxine

Pyridoxal P

MCD

Folinic acid

3PGD

GLUT-1

Jaundice

Liver failure

Galactose

Fructose

TYR-1

CDG-1b

Bile acids

LCHAD

Cardiac

FAO

Persistent

Hypoglycemia

PHHI

FAO

Glycogenosis

Hormones

Inborn Metabolic Diseases

First think of treatable disorders

Anabolism

Epuration

Special diets

Glucose

Vitamins

Amino Acids

Special diet

Avoid fasting

L-carnitine

Special diet

Galactose

Disease specific treatments – Jaundice/liver failure

Classical galactosemia - Intoxication type disease

Autosomal recessive disease

1/40.000 to 1/60.000 birth (in western countries)

Galactose-1-phosphate-uridyl-transferase (GALT) deficiency

GALT gene locus 9p13.13 (11 exons)

Urgent diagnostic on DBS (neonatal screening)

In urine

First symptoms, first weeks of life

Gastrointestinal problems, feeding difficulties, failure to thrive, lethargy

Hepatomegaly

Severe hepatic insufficiency (jaundice, bleeding tendency, hypoglycemia) and death if not promptly treated

E.Coli infection frequent

Diagnosis :

– Galactose, GALT activity on DBS

– Total red blood cell (RBC) Gal-1-P concentration

– Galactitol on urine

Urgent treatment : removal of LACTOSE and GALACTOSE from diet

(soya or lactose free formula- Olac®) rapid recovery

Disease specific treatments – Jaundice/liver failure

Classical galactosemia – Intoxication type disease

Disease specific treatments – Jaundice/liver failure

Classical galactosemia and consequences

Mild growth retardation

Delayed speech development

Verbal dyspraxia

Difficulties with spatial orientation

Decreased concentration ability

Reading difficulties

Abnormal brain white matter

In women, in general puberty is normal but may be delayed

- Premature ovarian insufficiency (81%)

Despite galactose restricted diet started soon after birth Despite compliance to diet

Despite Gal-1-P concentration within normal range

Risk: cataract

Resolves under galactose restricted diet, prognosis is good

Theorical risk at adolescence when diet is released

At 2 y : 80% patients IQ > 80

At 12 y : 80 % patients IQ < 80

Disease specific treatments – Jaundice/liver failure

Tyrosinemia Type 1 – Intoxication type

• Autosomal recessive disorder

• Europe 1/100,000 ; Quebec 1/1800

• Newborn screening on DBS

Hepatic disease

Chronic hepatic insufficiency cirrhosis

Hepatocellular carcinoma (HCC)

Kidney disease

Tubulopathy Hypophosphatemic rickets

Disease specific treatments – Jaundice/liver failure

Tyrosinemia Type 1 – Intoxication type

• Autosomal recessive disorder

• Europe 1/100,000 ; Quebec 1/1800

• Newborn screening on DBS

Hepatic disease

Chronic hepatic insufficiency cirrhosis

Hepatocellular carcinoma (HCC)

Kidney disease

Tubulopathy Hypophosphatemic rickets

NTBC

Disease specific treatments – Jaundice/liver failure

Tyrosinemia Type 1 – basis of treatment

1. Dietetic : hypoproteic diet (1964) and Tyrosine/PHE free amino acid

mixture supplementation

At short term improves hepatic symptoms,

decrudescence of tubulopathy

BUT … at long term liver failure, HCC are not prevented

2. Orthotopic Liver transplantation (1976) cure hepatic and neurologic

symptoms (5-10 % mortality) – now reserved to patient with acute liver

failure and fail to respond to NTBC and patients suspected with HCC

3. Kidney and liver transplant

4. Hematin (in case of porphyric crisis before transplant)

5. NTBC (1991) – a « pharmacologic inhibitor » that inhibits Tyrosine

degradation at an early step to prevent production of toxic compounds

normalisation of hepatic function

Risk of HCC very low if NTBC started < 6 months

Disease specific treatments – Jaundice/liver failure

Congenital Disorder of glycosylation 1b

PMI :

Phosphomannose

-Isomerase

Disease specific treatments – Jaundice/liver failure

Congenital Disorder of glycosylation 1b

500

20

3

0

<10

Disease specific treatments – Jaundice/liver failure

Congenital Disorder of glycosylation 1b

IEF Tf : isoelectrofocusing of serum sialotransferins

IEF Tf studies the number of

sialic acids residues

Disease specific treatments – Jaundice/liver failure

Congenital Disorder of glycosylation 1b

• Mannose 1g/kg/day divided in 5 doses

– Clinical recovery (liver, digestive)

– ATIII normalisation

– IEF Tf pattern improves after several months

• Only one treatable CDG syndrome

• Digestive and liver symptoms – diarrhea, exsudative enteropathy (hypoalbuminemia)

– hepatomegaly, cytolysis, cirrhosis, chronic liver disease

• hypoglycemia (hyperinsulinism)

• Kidney : tubulopathy, proteinuria

• Ig deficit (infections)

• Bleeding/thrombosis (XI, ATIII, prot C, prot S)

• No neurologic symptoms

• Mild asymptomatic forms

Fatty Acids B-Oxidation Defects – Energy depletion type

Disease specific treatments – Cardiomyopathy

Cardiac arrhythmia

Acute or chronic cardiomyopathy

(LVH or dilated)

Mucle pain,

Rhabdomyolysis

Hypoketotic hypoglycemia

Energy

depletion

- Carnitine transporteur

- CPT1, translocase, CPT2

- VLCAD (Very long Chain AcylCoA

dehydrogenase),

- LCHAD (Long Chain AcylCoA

dehydrogenase ,

- MCAD (Medium Chain acylCoA

dehydrogenase),

- SCAD (short chain acylCoA

dehydrogenase), SCHAD

- ETF (electrons transfert) LCHAD

MCAD

SCHAD

Cardiomyopathy and conductive defects

Cardiomyopathy and conductive defects

• Simultaneous determination of Free fatty acids and Ketones (3-

Hydroxybutyrate) is essential for a rapid diagnosis in case of

Hypoglycemia

• Acylcarnitine profile is usually diagnostic

(plasma, filter paper)

• Organic acids analysis (dicarboxylic acids from

ω oxidation) and serum carnitine may be helpful

• Enzyme studies (leukocytes, fibroblasts)

• Molecular studies confirm diagnosis (common mutation in MCAD and

LCHAD)

• Challenge tests (fasting test, oil challenge) are indicated only in

selected, exceptional cases and should be carried out in specialised

metabolic centers (danger of acute cardiotoxicity and others)

C8

Loïc (4/11/87)

11 y : dyspnea after efforts and vomiting

• Personal history : BW 3400 g, Height 51,5 cm; HC 37 cm.

• A 10 1/2 ans : Frequent vomiting. Hypoglycemia ++

• A 10 3/4 ans : After a running competition, dizziness and transient amaurosis and vomiting.

Since then, feels tired and have muscle weakness after walking (3 km)

Regular physical activity otherwise

• Clinical Exam :

W 30 kg (P10-P25), H 135 cm (P10-P25), TA 90/50 mmHg.

Systolic murmur 2/6, Liver enlarged (3-4 cm)

Normal Peripheral pulse

Scapular muscle weakness

Case report

ECG : Sinusal arythmia, LVH Echo doppler : Dilated and hypertrophic cardiomyopathy

• Hemogram : Nl

• Hémostase : TP 48 %, TCA 37/32, Vit K dependent clotting factors deficit corrected by vit K supplement

• ASAT 34 UI/l; ALAT 20 UI/l, gamma GT 9 UI/l; Ph Alc 118 UI/l

CPK 235 UI/l (N<195)

• Redox : normal

• Plasmatic Amino Acids : N

• Urine Amino Acids : N

• Urine Organic acids : N

Case report

• Plasmatic Free Carnitine 4 uMol/l ( N 22-64)

• Plasmatic Total Carnitine 12 uMol/l ( N 34-77)

• Plasmatic Esterified Carnitine 14 uMol/l (low)

• Urine Free Carnitine 20.9 umol/mmol creat

• Urine Totale Carnitine 42.2 umol/mmol creat

• FA Oxidation on lymphocytes (Dr Brivet, Bicêtre) :

Long chain FA oxidation defect corrected after incubatiton with L-Carnitine

• Impaired Skin Fibroblast Carnitine Uptake

Patient control Normal values (n=10)

Total Uptake

(pmol/min/mg prot

0,60 1,60 1,55 - 2,90

Specific uptake

(pmol/min/mg prot)

0,0 1,20 1,19 - 2,40

Case report

• At 12 y : walk 12 km without being tired

• At 13 y : normalisation of left ventricular function. FE 65 %.

• Slight Left ventricular hypertrophy (10-11 mm).

• Stop treatment (digoxine, IEC)

• Increase of muscular mass

• Weight gain

Treatment : L-Carnitine 100 mg/kg/j/day

Case report