M2C Unit 2

What is a biochemical genetic disease?

A problem w/ the body’s ability to regulate & utilize nutrients from food & existing body stores to create energy

What typically causes biochemical disease?

Defects in specific proteins (enzymes) that help break down (metabolize) food for energy. If the parts of food cannot be broken down, they can accumulate & can be toxic or change into other toxic chemicals OR if food cannot be broken down causing a lack of energy.

What are the main external and internal (endogenous) sources of energy?

External:

• Food

• Supplements

• Vitamins

Internal:

• Carbohydrates: stored in the liver as glycogen

• Fat: stored as triglycerides in adipose tissue

• Protein: amino acids can be synthesized endogenously (nonessential) and others must be obtained from the environment (essential)

What are the two states of metabolism?

Anabolism & catabolism

Differentiate anabolism and catabolism

Anabolism: calories in GREATER THAN calories out !!!! uses energy to GROW & BUILD MOLECULES required for normal body functioning

Catabolism: calories in LESS THAN calories out !!!! BREAKS DOWN BIG MOLECULES into smaller, easier to absorb molecules (think cats break things)

How do we use carbohydrates for energy?

Glycogen and glucose stores provide enough energy to sustain physiological function for about 24 hours

How do we use fat for energy?

Triglyceride stores allow for survival for several weeks

How do we use protein for energy?

We break it down into amino acids that are used to make new body proteins or turned into fat

What is the one gene = one enzyme hypothesis

Each gene codes for a single enzyme

What is an enzyme?

A protein that acts as a biological catalyst that facilitates a chemical reaction to change one biochemical into another

Describe how a faulty enzyme can cause a metabolic disease.

A —> B —> C —> D (A is the substrate/starting material and D is the final product)

Each arrow requires am enzyme to carry out the chemical rxn

If the enzyme that is converting substance A —> substance B is missing or not working properly, this causes three possible outcomes:

• A builds up, aka substrate accumulation bc it can’t be processed fast enough

• Too little D is made, aka product deficiency bc less B is available to make C then D

• More F is made, aka byproduct accumulation due to substrate going alternate route

How could we treat a disease caused by a faulty enzyme at the substrate?

• Dietary restriction

• Toxin removal

• Substrate reduction

How could we treat diseases caused by faulty or missing enzymes?

• Dietary restriction

• Toxin removal

• Substrate reduction

• Enzyme replacement

• Organ/cell transplant

• Chaperone therapy

• Gene therapy

• Cofactor replacement

• Product supplementation

Who coined the term “inborn errors of metabolism” in 1908?

Sir Archibald Garrod.

He also recognized “chemical individuality” and one gene = one enzyme hypothesis

What is Alkaptonuria?

An enzyme deficiency that results in dark tissue and urine due to accumulation of homogentisic acid

one of the first applications of spectroscopy!

Garrod proposed the mechanism of disease for Alkaptonuria in 1902. How long after that did it take for the deficient enzyme and gene to be discovered?

Deficient enzyme discovered 50 years later

Gene discovered 94 years later, in 1996

What is the deficient enzyme for Alkaptonuria?

Homogentisic dioxygenase

What is the pathway for Alkaptonuria?

Phenylalanine & Tyrosine degradation pathway

What geen variant causes Alkaptonuria?

HGD gene 3q13.33

What is the incidence for Alkaptonuria?

1 in 250,000 - 1,000,000

What are two symptomatic treatments for Alkaptonuria?

• NTBC (nitisinone) blocks conversion of 4-hydroxyphenylpyruvate to homogentisic acid

• Vitamin C inhibits deposition of HGA in connective tissues

Who was the first child with PKU to be treated and when was this?

2-year old Sheila Jones in 1951

Who developed the first newborn screening assay and when?

Robert Guthrie in 1961

In what year did Massachusetts mandate newborn screening for PKU?

1963

What condition causes your urine to turn black when exposed to air?

Alkaptonuria

What was the first metabolic disease that resulted in federal recommendations for newborn screening?

Phenylketonuria (PKU)

In metabolic disease, often no symptoms immediately after birth, but then infant becomes suddenly sick. Explain why.

It takes time for harmful substances to reach a dangerous level or for energy supplies to run low. Once that happens, the baby can deteriorate rapidly.

What are some signs and symptoms of metabolic disease?

• Poor feeding, vomiting, dehydration

• Abnormal breathing

• Lethargy, seizures, coma

• Multiorgan involvement

• Failure to respond to therapy for a presumed diagnosis

• Family hx of unexplained neonatal illness or death

• Unusually sweaty feet

Give an example for an acute and chronic metabolic disorder.

Acute: sudden, catastrophic illness and coma in a female carrier of OTC deficiency after labor & delivery of her first child

Chronic: progressive development of muscle weakness over time in late-onset Pompe disease

Why is there so much variability in disease severity for metabolic conditions?

It depends on the degree of enzyme deficiency! An activity threshold usually needs to be crossed to cause disease.

• no enzyme activity = infant/childhood disease

• 10-15% enzyme activity = teen/adult disease

Explain the three categories of metabolic disease.

Disorders of intoxication:

• accumulation of toxic substrates

• can present acutely in infants

• urgent need to reduce substrate accumulation

• nondysmorphic

Disorders of energy metabolism:

• product deficiency

• can present acutely in infants

• signs of running out of energy (hypoglycemia)

• nondysmorphic

Disorders of complex molecules

• deposition of byproducts around body (storage disorders)

• often progressive or later onset

• may be dysmorphic

Why is an amino acid disorder considered a disorder of intoxication?

Because Phenylalanine (Phe) is toxic to the brain, so prolonged elevated Phe interferes w/ normal brain development

Explain the basis of treatment for PKU.

Reducing the amount of Phe coming into the body by limiting Phe in the diet. Diet is lifelong.

What gene variant causes PKU?

PAH 12q23.2

What is the deficient enzyme in PKU?

Phenylalanine hydroxylase (PAH)

What causes PKU?

When Phe accumulates in the brain, which becomes neurotoxic in high amounts

What are the clinical symptoms of PKU?

• Well-appearing infant until 2-3 months old

• When irreversible developmental delay/intellectual disability become apparent

• Spasticity (tight muscles)

• +/- seizures

• Eczema/rash

• Fair skin/hair (lack of tyrosine - melanin)

• Musty odor due to phenylketones

• Non or minimally verbal

• Severe behavioral difficulties

What is the incidence for PKU? How about to be a carrier? And in Amish, Turkish, Irish, and African populations?

1 in 10,000 incidence for PKU

1 in 50 carrier frequency in East Asians/North Europeans

Amish - 1 in 1000

Turkish - 1 in 2,600

Irish - 1 in 4,500

African - 1 in 100,000 (rare!)

Explain the mechanism of disease for PKU.

PKU happens when the body cannot convert phenylalanine to tyrosine.

Faulty PAH enzyme → phenylalanine buildup → brain toxicity → developmental and neurological problems if untreated.

What are the three categories of metabolic disease?

• Disorders of intoxication

• Disorders of energy metabolism

• Disorders of complex molecules

True or false: an abnormal odor is often the first clinical sign of a metabolic disorder

False, because it takes time to accumulate the toxins that cause odor, so by the time you smell an odor, they will probably already be showing other signs such as poor feeding or lethargy.

True or false: Dietary restriction of phe in PKU is an example of substrate reduction

True, this is a classic example of substrate reduction therapy. By restricting phe in the diet, you reduce the amount of the incoming substrate, preventing its toxic accumulation in the body.

What information is important to prepare for a metabolic gc session?

• Which enzyme is deficient

• How you can summarize the pathway

• What happens to product and substrate, and how does this lead to disease

• Gene(s) and mode of inheritance

• Treatment options

• Prenatal testing options/birth plans

What are the types of disorders of protein metabolism?

• Aminoacidopathies

• Organic academias

• Urea cycle disorders

What are amino acids?

The building blocks of protein

What are the clinical symptoms of PKU?

• Severe intellectual disability

• Seizures

• Pigmentation changes

• Neurocognitive and executive functioning deficits

• Mood disorders

• Sleep disruption

What are we doing when we limit phe through a protein restricted diet for someone w/ PKU?

We are restricting the substrate to avoid/limit catabolism

How would someone w/ PKU practice a protein restricted diet?

They need to limit excess intake of phe, but just enough to prevent catabolism & support normal growth

They need to avoid meat, eggs, fish, dairy, beans, tofu, vegetarian alternatives

Limit whole grains/starchy vegetables

Consume medically modified low protein foods & UNLIMITED fruits and veggies!!!!

What are amino acid disorders?

They occur when the body cannot properly break down protein/amino acids for energy due to missing or malfunctioning enzymes

List 7 amino acid disorders.

• Alkaptonuria

• Phenylketonuria (PKU)

• Tyrosinemia

• Maple Syrup Urine Disease (MSUD)

• Homocystinuria

• Nonketotic Hyperglycinemia (NKH)

• Cystinosis vs. Cystinuria

Explain what causes disorders of intoxication.

Accumulation of toxic substrates before the block in pathway, and product deficiency after the block

What provokes acute metabolic attacks (crisis) in disorders of intoxication?

Catabolism, fever, intercurrent illness, food intake

What are some symptoms of disorders of intoxication?

• They typically present with a symptom-free interval & symptoms may be intermittent

• Most have metabolic markers & are detected by NBS

• Acute symptoms:

  • vomiting

  • coma

  • liver failure

• Chronic symptoms:

  • failure to thrive

  • developmental delay

What is the most COMMON group of biochemical disorders?

Disorders of intoxication. Most are treatable & nondysmorphic.

How are amino acids used by the body?

Protein is broken down into individual amino acids and then used to make proteins needed in the body.

Once the amino acids needed for growth are utilized, the rest are used to build other proteins or are burned for energy.

We usually eat MUCH more protein than we need!

When does the body burn protein for energy?

When you are catabolic, aka when the body is consuming fewer calories than your body is requiring.

Differentiate anabolism from catabolism.

• Anabolism = calories in > calories out

  • builds up structures and requires/consumes energy

• Catabolism = calories in < calories out

  • breaks down molecules and releases energy

What are treatment options for amino acid disorders?

• Medical “formula” that contains all amino acids except offending

• Low protein diet

• Medication (detoxifying drugs, cofactors, enzyme substitution therapy)

• Routine labs to monitor treatment efficacy

• Liver transplant

Name the two PKU enzyme treatments.

• Sapropterin dihydrochloride (Kuvan)

• Pegvaliase

Explain how sapropterin dihydrochloride (kuvan) is used to treat PKU.

It is a synthetic form of BH4, which PAH needs in order to convert phenylalanine into tyrosine.

it does NOT replace the defective PAH enzyme, it helps it function more effectively by providing that cofactor

Not everyone with PKU responds to sapropterin (kuvan). Why?

Patients must have BH4-responsive PAH activity. If their PAH enzyme has no functional activity, providing additional cofactor won’t help!

Explain how pegvaliase is used to treat PKU.

It is a pegylated enzyme replacement therapy. It provides an entirely different enzyme that can metabolize phenylalanine. It does NOT depend on PAH activity or BH4 responsiveness, so even if a patient has little or no functional PAH enzyme, it can still reduce phe concentrations.

What is maternal PKU syndrome?

It is a teratogenic effect of elevated phenylalanine during pregnancy. Because the placenta actively transports amino acids during pregnancy, fetal phe concentrations can be higher than maternal concentrations. It results in a syndrome including birth defects.

A baby can experience serious developmental complications from maternal PKU even if the baby does NOT have/inherit PKU!

How is it possible that unaffected carriers of PKU show symptoms of elevated phe?

Because the fetus is being exposed to high phe levels through the placenta. The baby’s genotype does not protect it from the effects of elevated maternal phe.

What is the phe level of a fetus with maternal PKU relative to mom’s phe level?

The phe level in fetus will be 1.5 times the amount of phe in maternal blood.

Can maternal PKU syndrome be treated?

There is NO treatment for maternal PKU syndrome POSTNATALLY.

The only strategy is PREVENTION. Prevention of maternal PKU syndrome involves lifelong education of PKU patients who can become pregnant.

What are symptoms of maternal PKU syndrome?

• Intellectual disability (92%)

• Microcephaly (73%)

• Intrauterine growth retardation (IUGR) (40%)

• Congenital heart defects (12%)

• Dysmorphic features

Case example: 3-day old female born at home is fussy and has a sweet smell. She is mennonite. What biochemical screening labs do you want?

Plasma amino acids & urine organic acids

Plasma amino acid analysis would measure concentrations of individual amino acids circulating in the blood. In MSUD, we would expect leucine, isoleucine, valine, and alloisoleucine to be elevated. This lab tells us WHICH amino acid pathways are disrupted.

We order urine organic acid labs because we want to know if abnormal organic acid metabolites are accumulating because the pathway is blocked. This test provides evidence that the downstream breakdown of branched-chain amino acids is impaired, which differentiates MSUD from other metabolic conditions.

What is the deficient enzyme in Maple Syrup Urine Disease (MSUD)? What is the purpose of this enzyme?

Branched-chain α-Ketoacid Dehydrogenase (BCKAD)

Its purpose is to break down branched-chain amino acids (BCAAs) valine, isoleucine, and leucine.

What is the inheritance pattern for Maple Syrup Urine Disease (MSUD)?

Autosomal recessive; 2 pathogenic variants can be found in all 4 subunits (locus heterogeneity)

In what population is MSUD more common/have a founder variant?

Old Order Mennonites.

1 in 1000 vs 1 in 200,000 general population.

Explain the pathway for MSUD.

The BCKD enzyme complex is defective —> Amino acids valine, isoleucine, and leucine, as well as their ketoacids, accumulate —> The elevated leucine is neurotoxic and can cause cerebral edema (brain swelling).

What does the term “pathognomonic” mean?

A finding that is highly specific to a particular disease.

What amino acid presence is a pathognomonic for MSUD?

Alloleucine

Which plasma amino acid is a neurotoxin?

Leucine.

Which urine organic acid is a neurotoxin?

2-oxyisocaproic acid.

What differentiates classic MSUD from acute MSUD?

Acute MSUD toxicity is a metabolic crisis where the toxins build up quicker and cause rapid neurological deterioration.

Classic MSUD causes long-term problems from impaired amino acid metabolism.

acuute crises can occur even in patients w/ previously well-controlled MSUD

What are some signs of acute MSUD?

• Metabolic acidosis (too much acid)

• Hypoglycemia (low blood sugar)

• Ketonuria (excess ketones in urine)

• Encephalopathy (altered brain structure/function)

• Cerebral edema (brain swelling)

What are some long-term complications for chronic MSUD?

• Developmental delays

• Intellectual disability

• Movement disorders

• Behavioral or psychiatric difficulties

Name the three branched-chain amino acids (BCAAs).

Valine, Leucine, and Isoleucine.

How can classical MSUD present in infants?

• Newborn w/ poor feeding and weak, disorganized suck - poor weight gain, failure to thrive

• Hypertonia, irritability, high-pitched cry when awake, otherwise lethargic

• Intermittent apnea, opisthotonos, “bicycling” movements

• Sometimes burnt sugar/maple syrup-y odor from diaper/urine and earwax

• Seizures, encephalopathy (due to cerebral edema), coma, death

How can classical MSUD present in older children?

• Intellectual disability

• Hyperactivity

• Sleep problems

• Focal dystonia

• Movement disorder (choreoathetosis, ataxia)

• Hallucinations

What variant forms of MSUD is NBS likely to miss?

• Intermediate

• Intermittent

• Thiamine-responsive

• E3-deficiency

Can a normal newborn screen rule out MSUD?

NO, especially mild or intermittent forms.

Why push valine and isoleucine supplementation as part of the sick day diet for MSUD treatment?

Because leucine is the most neurotoxic of the three amino acids, our goal is to lower leucine while preventing further protein breakdown.

Extra valine and isoleucine promotes protein synthesis, prevents catabolism, and supports amino acid balance.

Why would a liver transplant cure MSUD?

Because MSUD is caused by a deficient BCKD enzyme complex, a donor liver with functional BCKD enzymes will be able to metabolize branched-chain ketoacids.

remember that a liver transplant provides a functional metabolic cure, not a genetic cure

What is homocystinuria?

It is a disorder in which the body cannot properly break down homocysteine, which causes homocysteine and methionine to accumulate.

Incidence is 1-9 in 100,000

What is usually the deficient enzyme in homocystinuria?

Cystathionine β-synthase (CBS)

50% of CBS deficiencies are responsive to what?

Pyridoxine aka Vitamin B6

Why is homocystinuria more challenging to detect on NBS?

Because homocysteine is a target analyte and can be normal or borderline elevated.

What gene causes homocystinuria?

CBS 21q22.3

How can someone with homocystinuria present?

• Eyes:

  • Myopia (near-sightedness)

  • Ectopia lentis (lens dislocation)

• Skeletal system:

  • Marfanoid appearance (tall stature, long limbs)

  • Osteoporosis

  • Scoliosis

  • Ribs/chest go inward or outward

  • High-arched palate

  • High-arched foot

  • Knock-knees

• Vascular:

  • Thromboembolism (blood clots)

• CNS:

  • Developmental delays

  • Seizures

  • Psychiatric problems

  • Extrapyramidal signs (drug-induced movement disorders)

What is the treatment for homocystinuria? How about specifically for adults?

• Low protein diet

• Methionine-free medical formula and foods

• Methionine 150-200 mg/day

• Cofactors vitamin B6 and vitamin B12

• Oral betaine supplementation

For adults:

• Acute illness care - ER protocol for stroke

• Blood thinning meds

What is nonketotic hyperglycemia?

A disorder where the body cannot properly break down glycine, so it builds up to toxic levels in the body, disrupting normal brain signaling and causing severe neurological damage.

also known as glycine encephalopathy

Incidence:

• 1 in 60,000 in Finland

• British Colombia & Canada carrier frequency of 1 in 125

What is deficient in nonketotic hyperglycinemia?

The glycine cleavage system activity is deficient

How can someone with classic neonatal/infantile nonketotic hyperglycinemia (NKH) present?

• Apnea

• Hiccups (prenatal & postnatal)

• Progressive lethargy

• Seizures (onset in neonatal period or ~ 6 mo

• Hypotonia (low muscle tone)

• Myoclonic jerks leading to apnea and often death by 1 yr

• Infants that survive have severe ID, intractable seizures, and shortened lifespan

How can someone with late-onset form of nonketotic hyperglycinemia present?

• Spastic paraparesis (gradual weakness and muscle stiffness in lower limbs)

• Optic atrophy (loss of nerve fibers in optic nerve)

• +/- MR

• +/- choreoathetosis (movement disorder)

There is NO curative treatment for nonketotic hyperglycinemia (NKH), only treatment to manage/control symptoms. What are some of those treatments?

• Anti-epileptic medications

• Sodium benzoate (reduces serum glycine levels)

• Dextromethorphan (noncompetitive antagonist at the NMDA receptor)

• Ketogenic diet - new therapy - high fat, content carb diet that decreases seizures

What are the outcomes of nonketotic hyperglycinemia (NKH)?

• 80% of babies will not survive the neonatal phase (first 4 wks of life)

• Most children with severe NKH are not expected to live past their 5th bday

• Most children with mild NKH are not expected to live to adulthood

• There are a handful of people living with very mild NKH who are over 21

What is urea?

• A major organic component of urine that contains waste nitrogen

• It is a final product of a chain of reactions that break down amino acids