Inborn errors

of Metabolism
DMT 4206
DEPARTMENT OF MEDICAL TECHNOLOGY
FACULTY OF HEALTH SCIENCES
 Introduction
• Inborn errors of metabolism:

 Affect the conversion of nutrients into 1 another or into energy

 Caused by impaired activity of enzymes, transporters or cofactors

 Result in accumulation of abnormal metabolites (substrates) proximal

to the metabolic block or by lack of necessary products

 Abnormal byproducts can also be produced when alternative pathways

are used to dispose of the excess metabolites
Large
macromolecules
in nutrients are
broken down to
simple subunits
that are converted
to acetyl-CoA with
production of ATP
& NADH

Acetyl-CoA is then
completely oxidized
to C02 & H20 in
mitochondria, with
production of large
amounts of
NADH & ATP
A block in a metabolic
pathway results in:

• Accumulation of
substrate A

• Deficiency of product B

• Accumulation of
byproducts F
Classes of disorders
• Disorders of metabolism of AA, fats & CHO

• The frequency of individual diseases is rare

 1 : 10,000 to 1 : 200,000 or even rarer
 Their cumulative frequency is substantial, approaching 1 :
3000 newborn

• Themedical consequences ~ variable, ranging from failure to

thrive to acute illness ~ brain damage, coma & death in
some cases
Classes of disorders
• Many cases - acute presentation is preceded by a symptom-
free period

• Most cases - treatment available

 Consist of special diets lacking the specific nutrients that can
not be metabolized plus vitamins & other cofactors

 Treatment is effective – if it begun early before symptoms

occur b/c damage that has already occurred is usually
irreversible

 Identifying patients with metabolic disorders ~ birth

Inheritance Pattern of Metabolic Disorders
• Metabolic disorders ~ mutations in
genes that code for specific enzymes
involved in metabolic pathways

• Majority of metabolic disorders have

autosomal recessive inheritance ~
affect boys & girls equally

• Autosomal recessive disorder ~ affected

individuals have a mutation in both
alleles encoding for a specific
enzyme transporter
 Disorders
of
Amino Acids
Introduction
• Individually
rare, but collectively they affect perhaps 1 in
8,000 newborns

• Almost all are transmitted as autosomal recessive traits

• Result
from a lack of a specific enzyme in the metabolic
pathway of an AA
Introduction
• Leads to the :

 buildup of the parent AA

 its byproducts or

 catabolic products (organic acids)

~ depending on the location of the enzyme block

Groups
• Aminoacidopathies :
 Parent AA accumulates in excess in blood & spills over into
urine

 E.g. PKU, Type I tyrosinemia, Homocystinuria, Maple

syrup urine disease (MSDU)

• Organicacidemias
 Products in the catabolic pathway of certain AA accumulate

 E.g. Glutaric Acidemia Type I, Isovaleric acidemia ,

Methylmaionic acidemia, Propionic acidemia
Aminoacidopathies
Phenylketonuria (PKU)
•A disorder of phenylalanine metabolism caused by
deficiency of phenylalanine hydroxylase leading to the
accumulation of phenylalanine & production of
phenylketones that are excreted in urine

• Phenylalanine is an essential AA, constituting 4-6% of all

dietary protein

• Phenylalanine that is not used in protein synthesis is

converted to tyrosine by phenylalanine hydroxylase &
further degraded via a ketogenic pathway
PKU
• The
frequency of hyperphenylalaninemia or
phenylketonuria is 1:10,000 to 1:20,000 live births

• Classic
PKU :
 Caused by mutations in the phenylalanine hydroxylase
gene
 Represents 98% of all cases of hyperphenylalaninemia or
phenylketonuria

• Remaining 2% :
 Due to defects in biosynthesis or recycling of
tetrahydrobiopterin (BH4), the cofactor for phenylalanine
hydroxylase
PKU
• 1o or 2o (due to a deficiency of the cofactor) impairment of
phenylalanine hydroxylase results in :
 accumulation of phenylalanine, phenylketones &
phenylamines
 deficiency of tyrosine

• Elevated conc. of phenylalanine impairs brain development &

function, affecting other organs minimally

• Classic PKU – asymptomatic at birth with developmental delays

 neurological manifestations - several months of life
 brain damage already occurred
PKU
• Untreatedpatients develop :
 Microcephaly
 Eczematous skin rash
 “Mousy" odor (accumulation of phenyl acetate)
 Severe mental retardation

• Treatment :
 Low protein & phenylalanine diet
 Supplemented with tyrosine, minerals, vitamins & other
nutrients to sustain normal growth
 Continued for life
PKU
• Newborn screening leads to early detection & intervention – prevent
mental retardation

• Treatment should start before 2 weeks of age

• Pregnant women who are not on a low phenylalanine/ low protein

diet & have high conc. of phenylalanine have an increased risk of :
 Spontaneous abortions or
 Having a child with :
 Growth retardation
 Microcephaly
 Significant developmental delays
 Birth defects
PKU – Diagnosis
• Plasma
AA ~ elevated plasma phenylalanine &
phenylalanine tyrosine ratio

• Urine ~ elevated phenylketones

• Enzymatic confirmation of phenylalanine hydroxylase

deficiency is NOT performed (expressed only in the liver)

• Mutational analysis of the gene ~ b/c of correlation btw

severity of the mutation & phenylalanine tolerance
 PKU – BH4
• Children
with hyperphenylalaninemia ~ screened for defects
in BH4 synthesis or recycling

 Measuring the urinary pterins profile & by measuring the

enzyme activity of dihydropteridine reductase (DHPR) IN
BLOOD SPOTTED ON FILTER PAPER

• Cofactor
for phenylalanine hydroxylase, tyrosine
hydroxylase, tryptophan hydroxylase & nitric oxide
synthase

• Deficiency
affects the synthesis of several neurotransmitters
(dopamine & serotonin)
PKU – BH4
• Patients with a defect in BH4 synthesis or recycling
 Neurological symptoms
 Developmental regression in the 1st few months of life
 Can develop seizures
 Truncal hypotonia with hypertonia of the extremities

• Therapy with BH4 & appropriate neurotransmitters

 May or may not require low phenylalanine diet
PKU Screening
• Lossof substrate conversion from phenylalanine to tyrosine
results in formation of phenylpyruvate & metabolites and
elevated phenylalanine in blood

 The phenylketones are excreted into urine

• The semi-quantitative screening test for PKU

 Devised by Dr. Guthrie ~ 1960s

 A microbiological assay that involves the incorporation of a

bacterium (Bacillus subtilis) & a growth antagonist (beta-2-
thienvlalanine) into agar
PKU Screening
• Guthrietest screening test for PKU
 The dried blood spot punched out of the filter paper card is
placed on the agar
 Normal conc. of phenylalanine in the sample spot = bacterial
growth will be inhibited
 Excess phenylalanine will counteract the antagonist &
restore growth of the bacterium around the spot = PKU

 Sensitive to serum phenylalanine conc. >4 mg/Dl

 Simple ~ allows screening of a large no. of infants not only for

PKU but also for other disorders of AA metabolism using
different growth antagonists
PKU Screening
PKU Screening
Tyrosinemia
•A genetic disorder characterized by increased blood conc. of
tyrosine

• Types
I-III - each is caused by the deficiency of a different
enzyme

 Tyrosinemia type I (TYR-I) - most severe form, deficiency of

fumarylacetoacetate hydrolase

 Tyrosinemia type II - deficiency of tyrosine aminotransferase

 Tyrosinemia type III - deficiency of 4-hydroxyphenylpyruvate

dioxygenase
Tyrosinemia
Tyrosinemia – type 1
• Incidence is approx. 1 in 100,000 ~ Canada
• Patientspresents (before 6 months of age) with severe liver
involvement, or with :
 Chronic failure to thrive
 Mild hepatocellular dysfunction
 Renal involvement
 Rickets due to renal Fanconi syndrome
• Extremeirritability caused by peripheral neuropathy
mimicking acute intermittent porphyria

• Untreated patients - liver cirrhosis; high risk for liver cancer

Tyrosinemia – type 1
• Increased conc. of tyrosine in the plasma, but this increase
usually is NOT as marked as in patients with other forms of
tyrosinemia

• Diagnosis ~ urine organic acid testing of succinylacetone,

derived from fumarylacetoacetic acid - the intermediate
immediately upstream of the enzyme defect

• Identifiedby newborn screening only when succinylacetone is

used as the primary marker
 b/c tyrosine is not elevated in the newborn period in these
patients
Tyrosinemia- type 1
• Therapy consists of :
 Low dietary tyrosine
 Low dietary phenylalanine
 Drug therapy with 2-(2-nitro- 4-trif uoro-methylbenzoyl)-1,3-
cyclohexanedione (NTBC)
 An inhibitor of 4- hydroxyphenylpyruvate dioxygenase
 NTBC prevents the synthesis of succinylacetone

 Liver transplantation ~ patients who progress to liver failure

& liver cancer
Alkaptonuria
• Deficiency of homogentisic acid (HGA) dioxygenase leads to :

 The presence of HGA & its oxide (alkapton) in the urine

 Bluish-black pigmentation in connective tissue (ochronosis)

 Arthritis

 Urine that turns dark with standing or alkalinization

Alkaptonuria
• Accumulation of HGA in tissues causes cartilage damage in
their joints (spine) leading to low back pain at a young age

• The pigment can accumulate in cardiac valves causing their

failure

• NO treatments that reduce the complications

• Treatment with NTBC prevents formation of homogentisic acid

& is being explored as a potential therapy
Homocystinuria
• Increased conc. of the sulfur-containing AA homocysteine in
blood & urine

• Caused by at least 7 genetically different disorders

 Most common - classic homocystinuria ~ caused by reduced
activity of cystathionine β-synthase

• Incidence is approx. 1:300,000 live births

• Clinical manifestations are nonspecific at first & may include :

 failure to thrive
 developmental delay
Homocystinuria
• Patients develop lens dislocation & a body habitus
like that seen in Marfan syndrome
 Homocysteine interferes with disulfide formation
in fibrillin
 the protein defective in Marfan syndrome

• Patients whose blood homocysteine conc. continues

to increase are at risk of blood clots ~ life-
threatening complication

• Diagnosis of obtained by plasma AA analysis

showing increased plasma conc. of methionine
(children) & the presence of the disulfide
homocysteine
Homocystinuria
• Classic
Homocystinuria is detected in newborn screening by
increased conc. of methionine in whole blood spots

• Therapy requires :
 High doses of pyridoxine (the cofactor of cystathionine β-
synthase)

 Special diet low in methionine

 Administration of betaine ~ donates a methyl group to

homocysteine to generate methionine
Maple Syrup Urine Disease
• Autosomal recessive disorder with an incidence of approx.
1:250,000 live births
• Deficiency
of the branched-chain alpha-keto acid
dehydrogenase complex (BCKDC)
• Leadto a buildup of the branched-chain AA leucine,
isoleucine & valine & their toxic keto acids in the blood
& urine
• Sweet-smelling urine ~ odor similar to that of maple syrup
• Severalforms may occur, depending on the gene affected & the
severity of the mutations
Maple Syrup Urine Disease
• Diagnosis
is by measuring plasma AA & finding increased
branched chain AA & alloisoleucine (characteristic of this
disease)

• Treatment:
 diets that have a restricted content of branched-chain AA &
 supplementation with :
 high-dose thiamine
 low doses of valine & isoleucine
Organic acidemias
Glutaric Acidemia Type I
• An autosomal recessive disorder of lysine, hydroxylysine &
tryptophan metabolism caused by deficiency of glutaryl-CoA
dehydrogenase

• Glutaric acid (GA) & 3-hydroxyglutaric acid (3-OH-GA) formed in the

catabolic pathway of the AA & accumulate esp. in the urine

• Affected patients can have brain atrophy & macrocephaly with

acute dystonia btw 6 -18 months

• Identified by increased glutaryl (C5DC) carnitine on newborn

screening
Glutaric Acidemia Type I
• Urine organic acid analysis indicates the excess 3-OH-GA

• Urine
acyl carnitine profile shows glutaryl carnitine as the
major peak

• Therapy :
 Carnitine supplementation to remove glutaric acid
 A diet restricted in AA capable of producing glutaric acid
 Prompt treatment of secondary illnesses (infections)

• Earlydiagnosis & therapy reduce the risk of acute dystonia in

patients
Treatment of AA Disorders
• Special
diets restricting the compounds (AA) that result in the
formation of the abnormal organic acid / accumulation of
high conc. of AA

 supplemented with vitamins specific for each disorder,

carnitine supplements & sometimes fasting avoidance

• Some conditions - IV fluids containing glucose

Disorders of
Fatty Acid
Oxidation
Introduction
• FA are metabolized within mitochondria ~ energy

• Carnitine& the carnitine cycle are required to transfer

long-chain FA into mitochondria for β-oxidation

• β-oxidation - long-chain FA are shortened of 2 C units at each

cycle to generate Acetyl CoA
 Used by the Krebs cycle to produce energy
Disorders of FA oxidation
• Occur
when an enzyme is missing in the metabolic pathway
& FA fail to undergo oxidation to supply energy
• Usually silent & become evident only when the body needs
energy from fat during times of :
 Fasting
 Infections
 Fever
 Healthy children with these disorders become :
 Acutely sick
 Lose consciousness
 Become comatose
 Die
Disorders of FA oxidation
• Symptomatic patients:
 Develop HYPOGLYCEMIA
 Might show INCREASED SERUM TRANSAMINASES
 Liver damage

• Some disorders (LCHAD deficiency) can also affect :

 Skeletal muscle &
 Cardiac muscle
 Mother during pregnancy
Disorders of FA oxidation
• Disorders :

 Medium-chain acyl-CoA dehydrogenase (MCAD

deficiency)

 Long-chain 3-hydroxyacyl-CoA dehydrogenase deficiency

(LCHAD deficiency)

 Carnitine transporter defect

 Very long-chain acyl-CoA dehydrogenase deficiency

MCAD Deficiency
• Most common disorder of FA oxidation

• Frequency of 1 : 6000 to 1: 10,000 births among Caucasian

• Symptoms ~ variable : from asymptomatic to hypoglycemia,

lethargy, coma, & sudden death, triggered by prolonged
fasting, acute illness or both

 Majority of patients have symptoms in the 1st year of life

 But clinical symptoms can occur at any time during life & the 1st
episode is often fatal
MCAD Deficiency
• Treatment:
 Avoidance of fasting
 Consumption of low-fat foods
 Carnitine supplementation
 Institution of an emergency plan in case of illness or other
metabolic stress

• Early
diagnosis & early initiation of treatment = good
prognosis
MCAD Deficiency
• Patientsare identified by tandem mass spectrometry
(MS/MS) newborn screening b/c of the characteristic
acylcarnitine profile with increased conc. of :

 C6, C8 & C10:l carnitine

 Elevated C8/C2 & C8/C10 ratios

• Diagnosis is confirmed by urine organic acid & acylglycine

analyses
Treatment of FA Oxidation Disorders
• Consistsof :
 Avoidance of fasting
 Low-fat diet
 Carnitine supplementation

• Forsome disorders (VLCAD & LCHAD) supplementation

with medium chain triglycerides (MCT oil), that enter
mitochondria independently from carnitine & bypass the
metabolic block

• Conditions that increase catabolism (fever, vomiting &

infections) need to be aggressively treated with antibiotics &
antipyretics & with intravenous glucose
 Disorders of
Carbohydrate
Metabolism
Introduction
• Enzyme deficiency in the metabolic pathways of CHO

• Results in accumulation of sugars within organs or tissues :

 Impairing their function
 Inability to obtain energy or
 Toxicity from the excess of monosaccharides or their
derivatives (phosphorylated sugars)

• Includes :
 Glycogen storage diseases
 Glucose-6-phosphate dehydrogenase [G-6-PD] deficiency
 Classic galactosemia
Glycogen storage disorders
• Affect primarily the liver or the skeletal muscle

• The accumulation of glycogen impairs organ function &

if the liver is affected – the release of glucose is prevented ~
Hypoglycemia

• Treated with avoidance of fasting & a special diet devoid of

simple sugars & supplements with uncooked cornstarch
G-6-PD deficiency
• Affects RBCs – hemolysis ~ mild to severe jaundice in newborns
& in some cases hemolytic anemia

• Symptoms can be triggered by :

 infections
 certain drugs ~ antibiotics & medications used to treat malaria
 exposure to fava beans (favism)

• Gene for this condition is on the X chromosomes

 Affects mostly males

• Avoidance of stressors can avoid symptoms

Classic galactosemia
• Results from absence of galactose- 1-phosphate uridyl
transferase
• Galactose is derived from the disaccharide lactose found in milk
• Galactose is phosphorylated to galactose-1-phosphate, but it is
NOT further metabolized
• Increased conc. of galactose-1-phosphate in cells are TOXIC
• Infants have :
 Failure to thrive
 Jaundice
 Liver failure
 Predisposition to sometimes life-threatening infections with
E. Coli & other GNB
Classic galactosemia
• Treatment involves removal of lactose (milk) & avoidance of all
foods containing galactose

• Intervention early in life provides the best prognosis although

some long lasting effects may continue to be observed,
particularly in girls who for unknown reasons develop ovarian
failure
Galactosemia Screening
• Galactosemiais a disorder of CHO metabolism resulting in
accumulation of galactose

• Mostcommon form is caused by deficiency of galactose-1-

phosphate uridyltransferase (GALT)
 converts galactose-1-phosphate to glucose-1-phosphate

• Typical
traditional screening methods :
 Measurement of galactose & galactose-1-phosphate & assay of
GALT enzyme activity from a dried blood spot
Estimation of phenyl
pyruvate in urine
Ferric chloride method
• Examining the urine with ferric chloride
 Must be confirmed by elevated phenylalanine in blood

• 10 % Ferric chloride is added in excess to fresh urine

•A positive result is shown by a green colour that develops

within 30 seconds & fades in 2 or 3 min

• Thesame solution may be dropped directly on to a wet

diaper - a green colour indicates the presence of phenyl
pyruvic acid
Ferric chloride method
The End