10, 11, 12 - Pharmacogenetics, IEM, Cancer Genetics

Pharmacokinetic vs pharmacodynamic

• What is it

• ชื่อภาษาไทย

• How the body interacts with the drug vs how the drug acts on the body

• เภสัชจลนศาสต์ vs เภสัชพลศาสต์

Pharmacogenetics vs pharmacogenomics

• A study of how a gene codes for a certain drug metabolizing enzyme or drug target

• Broader study of the entire genome to understand a patient’s drug response

Different types of metabolizers(4 main types) and what are they?

• Poor metabolizer: 2 loss of function genes for a metabolizing enzyme

• Intermediate metabolizer: 1 normal/1 LOF

• Extensive: 2 normal(wild type) genes

• Ultrarapid: Extra copies of the metabolizing gene

What is CYP450?

• It is a superfamily of enzymes in the liver that metabolize drugs

Warfarin

• Brief function

• Pharmacogenetics of this drug

• It is an drug that inhibits vitamin K action in the gamma-carboxylation reaction by stopping vitamin K from getting back to active form. It does so by inhibiting VKORC1 enzyme → Less production of vitamin K dependent functional clotting factors by liver → Anticoagulant

• It is metabolized by CYP2C9, and there is a normal variation(*1), and 2 loss of function variants(*2 and *3). If a patient has 2 normal alleles then they’ll be an extensive metabolizer, if they have one loss of function allele, they’ll be intermediate, if they have both(in any combination), they’ll be a poor metabolizer.

VKORC1

• What is it?

• Pharmacogenetics

• Enzyme that activates vitamin K; inhibited by warfarin

• Has a single nucleotide polymorphism in the promotor region, specifically if in 1639 bp upstream of the gene’s start, G becomes A, the promotor will be less efficient at transcription → Less VKORC1 produced → Inefficiency in turning vitamin K active → More vulnerability to warfarin, aka “Warfarin sensitive”

AA, GA, GG variations of VKORC1 gene and their differences?

• If a pt. has AA(aka both A in that position in both alleles), they’ll be the MOST warfarin sensitive, but if they have GG, they’ll need a higher warfarin does because they make a normal amnt of VKORC1

What is a polymorphism that is linked with the 1639 G>A SNP and how is it useful?

• 1173 C>T SNP

• It’s useful because if we can only test one of the two, then we can infer that if a pt. has one, they’ll likely have the other

What race should we monitor most carefully in terms of Warfarin?

• African ancestry

Clopidogrel

• Brief Function

• Pharmacogenetics

• Anti-platelet

• It is a PRO-DRUG metabolized by CYP2C19, meaning that it needs to be activated by CYP2C19! 1* is the normal allele, but there are loss of function variants from 2-5. If patients’ metabolism is impaired(eg. a slow metabolizer), less clopidogrel is activated → More platelet aggregation

HLA-B

• What is it?

• What drug does it relate to, and what is the pharmacogenetics?

• Human leukocytic antigen B, a gene that codes for HLA class I antigens that present on cells

• It relates to a drug called carbamazepine, an anti-epileptic drug that can cause Steven Johnson’s Syndrome if a patient has a version of the HLA-B gene called HLA-B*1502 allele because the HLA-B*1502 protein can bind carbamazepine and trigger cytotoxic T-cell attack

When is it worth it to screen for

• CYP2C9

• CYP2C19

• HLA-B*1502

• When giving higher than normal amnt of Warfarin + patients with VKORC1 A variants

• When treating patients who are in danger from platelet clumping, such as percutaneous coronary intervention(stent in coronary artery)

• Every case

Inherited metabolic disorders/Inborn errors of metabolism

• Type of genetic disease

• General pathology

• Autosomal recessive

• Gene variant leads to dysfunctional/absent enzyme → substrate builds up/deficiency in normal product/build up of accessory pathways → clinical effects

What are the types of inherited metabolic disorders and what are their key mechanisms?

1. Small molecule: Related to molecules like amino acids that we get from diet, these are small and when they build up can leave the organ with the dysfunctional enzyme(like the liver) and cross to the blood brain barrier. These often are asymptomatic at birth but show signs very fast because once the maternal enzymes no longer are connected in circulation with the newborn, the symptoms show

2. Large molecule: These usually build up in the cells like in lysosomes, and gradually cause organomegaly, bone pain, and organ dysfunction

3. Energy metabolism: Involves things like mitochondrial disorders, metabolic pathways of glucose/fatty acids → affects organs that use energy the most like the heart and brain

PKU

• Stands for

• Pathophysiology

• Result/symptoms(3)

• Phenylketouria

• Normally, phenylalanine becomes tyrosine via phenylalanine hydroxylase with the help of a cofactor called BH4, but in PKU patients, PAH is deficient

• Phe accumulates in blood and crosses the BBB → interferes with brain development and function, Phe also becomes phenylpyruvate/phenylacetate as accessory pathway → mousy/musty odor, Tyrosine deficiency → hypopigmentation and dopamine deficiency(paler skin and neurological symptoms)

Explain the symptoms of this 2y patient with PKU

• Healthy at birth, symptoms began at 5 months

• Hypopigmentation

• Seizures, microcephaly, inability to walk/talk

• Musty odor

• At first, the child has his mother’s own PAH enzyme helping clear the metabolites

• Less tyrosine → less melanin

• Lack of neurotransmitters and accumulation of Phe in the brain leads to all of these CNS symptoms

• Build up of phenylpyruvate/phenylacetate as accessory pathways

PKU Treatments

• Low Phe formula for kids, eating less Phe in diet, Pegvaliase(aka enzyme therapy that breaks down PAH)

MSUD

• Stands for

• Pathophysiology

• Results/Symptoms

• Maple Syrup Urine Disease

• Normally, BCAAs(Leucine, isoleucine, valine) are broken down by 2 enzymes, the first called branched chain aminotransferase(BCAT) who turns them into a branched chain ketoacid which gets processed by an enzyme complex called Branched Chain Alpha Ketoacid Dehydrogenase Complex(BCAKDC) and helps them enter the TCA to generate energy as a minor source. In MSUD, BCAKDC is deficient/non-functional and BCAAs and their ketoacid forms buildup.

• This disease is even more severe than PKU because it directly relates to the cell’s energy metabolism so the patient will often have reduced responsiveness/refuse to breastfeed, have spasticity(เกร็ง), extended legs(ขาเหียด), opisthotonus(หลังแอ่น)

Explain the symptoms of this patient with MSUD

• Lethargy/poor feeding/rigidity/opisthotonus

• Metabolic acidosis/ketonuria

• Lethargy/poor feeding/rigidity/opisthotonus: BCAA build up inhibits the citric acid cycle → neurons impaired, not only that, increased amino acids in the brain tissue → solutes follow → encephalopathy

• Buildup of ketoacids → Metabolic acidosis and ketonuria

OTC deficiency

• Stands for

• Pathophysiology

• Results/Symptoms(name 3)

• Ornithine transcarbamylase deficiency

• Normally, our body clears ammonia(protein metabolism waste product) by turning it into urea via the urea cycle. There are 2 steps of the cycle that occur in the mt. and the rest occur in the cytoplasm. If the OTC enzyme(Usually turns ornithine into citrulline) is deficient, citrulline will be low and the cycle can’t continue, so the cycle backs up → ammonia increases

• The ammonia increases and the brain turns it into glutamine which draws water into the brain → cerebral edema which causes vomiting, lethargy, coma

How can we tell the difference between carbamyl phosphate deficiency vs OTC deficiency, since citrulline will be low in both?

• If orotic acid is high, it’s an OTC issue

Encephalopathy meaning

Brain dysfunction

How does the brain normally detoxify ammonia, and how does OTC deficiency lead to these symptoms

• BUN low

• Glutamine high

• Orotic acid high

• Muscle spasms

• Vomiting

• Lethargy

The brain’s astrocytes turn glutamate + ammonia into glutamine, but if there’s too much, the glutamine’s osmotic pull becomes to strong and water is drawn in → cerebral edema

• Urea is unable to be produced → BUN low

• Brain turns glutamate to glutamine → Glutamine high

• Carbamoyl phosphate turns to pyrimidine synthesis due to accumulation → Orotic acid

• Glutamate is an excitatory neurotransmitter and it’s used up → Muscle spasms

• Area postrema compressed → Vomiting

• Glutamate used up + ammonia impairs mt. function → Lethargy

LSD

• Stands for

• Pathophysiology

• Results/Symptoms

• Lysosomal Storage Disease

• In these diseases, a lysosomal enzyme is deficient, which allows large molecules to accumulate inside lysosomes, causing swelling and disrupting the cell

• This leads to organomegaly due to the cells swelling, coarse facial features due to accumulation of GAGs/lipids, and skeletal abnormalities also due to the buildup of these substances

Cancer is genetic but most cancer is not inherited, what does this mean?

• It means that cancer occurs DUE to mutations in genetic information, but these mutations are usually acquired in life, not inherited

Tumor heterogeneity meaning

One tumor will not have a uniform population of cancer cells, because after dividing from the founder cell, subclones can develop their own mutations and get different behaviors

Driver vs passenger mutations

A driver mutation promotes cancer development/progression while a passenger mutation occurs in a tumor but doesn’t affect the malignancy

Oncogene vs tumor suppressor gene, and what mutations do they undergo?

• Oncogenes: A mutated proto-oncogene, a gene that was supposed to positively promote cell growth in a controlled manner that gets a mutation and is overactive, driving growth constantly. Oncogenes only need one allele to be defective, and usually receive point mutations that cause gain of function

• Tumor suppressor genes: Normally restrain the cell’s growth. These genes receive loss of function mutations and require both alleles to be lost before the effect shows up, aka the two hit hypothesis

Two-hit hypothesis

• Explains how people who inherit a mutated tumor suppressor gene are much more vulnerable to developing cancer since they need just one additional somatic mutation to knock out the remaining healthy copy

Why are things that make us suspect hereditary cancer?

1. Young age at dx.

2. Multiple affected relatives

3. Multiple primary tumors

4. Bilateral/multifocal disease

5. Rare tumor type

6. Specific tumor combinations

What are BCRA1/2 and how do they work?

• These are tumor suppressor genes that work by helping perform homologous recombination repair, where a double stranded break is held next to a homologous chromosome as the template

What is hereditary BRCA-mutant pt. vulnerable to get cancer?

• Because if the patient receives a second mutation, they can’t repair double strand breaks → high cancer chance

How do PARP inhibitors work?

• PARP is an enzyme that repairs single strand breaks. In cells, PARP and BRCA1/2 help each other. If a single strand break happens and becomes a double strand break, BRCA1/2 can fix the double strand break. PARP also tries to fix as many single strand breaks as possible before BRCA1/2 has to do anything

• A PARP inhibitor is a way we kill cancer cells. When we inhibit PARP, the cell receives many single strand breaks that become double strand breaks. Healthy cells with BRCA1/2 can fix these and survive, while the tumor cells with no BRCA will die.

What is an MSI, and finding it indicates what?

Microsatellite instability, aka a segment of small repeats like CACACACACA have different lengths that usual. This indicates an error in DNA mismatch repair because we normally fix these errors