Specific Drugs and Chromosomal tests Flashcards
Chromosomal Analysis
General Uses and Indications: Suspected abnormality of chromosome number or structure (deletion, insertion, rearrangements). Frequently obtained from pregnant women > 35 years (amniocentesis or chorionic villus sampling), from patients with congenital abnormalities (dysmorphisms, structural organ defects, mental and/or growth retardation), from families with multiple miscarriages and/or fertility problems, and directly from certain cancer biopsies.
Can Diagnose: aneuploidies (abnormal chromosome number), chromosome deletions, duplications, and insertions of moderate to large size (>3,000-5,000 kb / 3-5 Mb), and rearrangements.
Cannot Diagnose: single gene deletions, point mutations, small deletions, duplications, and insertions, methylation defects, trinucleotide repeat abnormalities.
Good for karotype stuff!
FISH
General Uses and Indications: Used to diagnose deletions, some translocations, and abnormalities of copy number. Often used to detect cytogenetic changes that are at or beyond the limits of resolution obtained by high-resolution chromosomal analysis. FISH for duplications works better on cells in interphase than metaphase (metaphase the chromatin is very compact)
Can Diagnose: recognized microdeletion syndromes, recognized chromosomal rearrangements (in cancers), and gene copy numbers (cancers). Also useful in diagnosing anueploidies (e.g. trisomy 13, 18, 21) in the prenatal setting. (100-200 kb bases?)
Cannot Diagnose: deletions, rearrangements that are not specifically tested for (i.e. FISH probes are specifically designed for each condition). FISH is not always able to detect duplications of gene region
Examples of Microdeletion Syndromes: Cri-du-chat, Smith-Magenis, DiGeorge (22qdel), Williams syndrome, Wolf-Hirschhorn, Prader-Willi syndrome, Angelman syndrome.
Sanger Sequencing
Widely used method of mutation detection based on Sanger-Gilbert method. Able to identify the genetic mutation causing many disease conditions.
General Uses and Indications: Used to identify sequence changes (mutations) in specific genes. In general you need the following:
Can Diagnose: Mutations in known genes (mutation can be previously reported or can be novel), polymorphic variants, small (1 to ~100 nucleotide) deletion/insertions. Ideal for looking at the sequence of a known disease gene
Cannot Diagnose: The technique is very specific, assaying only the region of the gene(s) for which the test has been designed. Frequently, many clinical genetic tests do NOT routinely sequence all parts of a gene (e.g. promoters, introns). This means that although the approach is often very specific, clinical sensitivity is frequently below 100% (this is an important concept to understand). This technique cannot easily detect larger deletions/insertions, rearrangements, and most chromosomal abnormalities.
Chromosomal Microarray Analysis (CMA)
Chromosomal Microarray Analysis (CMA): These have a big role in clincal genetics currently. These look for chromosomal DNA losses and gains (so called ‘deletion/duplication’ studies). Sometimes this is also called array comparative genomic hybridization (aCGH) analysis.
General Uses and Indications of CMA: CMA has become fairly standard for looking for small genomic deletions/insertions. You can think of this as a superior method to looking for chromosomal gains than losses than traditional chromosomal analysis because the resolution of the CMA is vastly superior to chromosomal analysis. The probe size used these days is between 100-200 Kb so they can pick up smaller changes than can be appreciated by chromosome analysis. Currently, some labs use >~200 Kb for deletions and >~400 Kb for duplications.
Can Diagnose: aneuploidies, unbalanced chromosomal rearrangements, chromosome deletions and duplications > 200 Kb and 400 Kb, respectively.
Cannot Diagnose: Deletions/Duplications below the resolution of CMA, nucleotide mutations, balanced chromosomal rearrangements
CYP3A:
o Substrates:
- Felodipine (Ca channel blocker)
- Cyclosporine (immunosuppresent)
o Inducer (increase activity) Rifampin (Antifungal)
o Inhibitor
♣ Ketoconazole
♣ Grape Fruit juice
CYP3A effects
If renal transplant patient has developed a fungal infection and is being treated with ketoconazole, must reduce cyclosporine dosage by 75%. Ketaconazole inhibits CYP3A activity and leads to increased levels of cyclosporine.
Rifampin
If renal transplant patient is exposed to tuberculosis, needs rifampin. Need to increase cyclosporine dosage, since rifampin is a CYP3A inducer so metabolism will be increased. Rifampin ramps.
Grapefruit juice
Inhibits CYP3A activity for 24-48 hours. E.g. if taking felodipine for hypertension could end up with hypotension since felodipine is not adequately eliminated from system.
CYP2D6
o Substrates
♣ Codeine
♣ Tricyclic antidepressants
• Gets turned into Morphine
CYP2D6 converts codeine to morphine. More copies indicate faster metabolism, so a smaller quantity can lead to intoxication. Poor metabolizers have fewer copies, so they will not do the conversion, and will therefore not receive benefits from the drug.
Warfarin
CYP2C9
o Substrate: Warfarin
♣ Will eat it up, and prevent it from targeting VKORC1
o CYP2C9 gene will eat the Warfarin, while Warfarin will try to eat the VKORC1 gene that allows blood clotting
♣ Warfarin is a blood thinner
• VKORC1
o Warfarin TARGETS VKORC1
♣ VKORC1 reduces vitamin K so that vitamin K can be recycled and used to make more coagulation factors -Single nucleotide polymorphisms
Warfarin
Warfarin is used for the prevention of thromboembolism. It functions by blocking the enzyme vitamin K epoxide reductase complex I (remember that vitamin K is needed for clotting!). This enzyme acts on vitamin K so that is can actually be used for clotting. There are two distinct genes that play a role in the metabolism of Warfarin. CYP2C9 does phase I detoxification of Warfarin. If allele is deficient, patients will require less warfarin for therapeutic use (as they are unable to metabolize as quickly).
Second element is VKORC1, which is the actual explicit target of Warfarin (remember that is makes the Vitamin K epoxide reductase complex). Alleles at non coding SNPs lead to creation of two major haplotype families A and B. AA needs 3.2 mgs/Day, AB needs 4.4, BB needs 6.1.
Take home: homozygous for reduced CYP2C9 and VKORC1 A alleles need 1/5-1/6 dose of normal CYP2C9 alleles and VKORC1 B alleles.
NAT gene
NAT gene
Substrate
Isoniazid —-> Antibiotic for tuberculosis
o NAT2 homozyous, heterozygous, and heterozygous recessive
Homozyougous or heterozygous NAT, will use phase 2 to quickly actetyl transferase the drug Isoniazid (MORE ACTIVE NAT GENE)
Homozygous Recessive NAT gene= unable to use the Isoniazid as quickly
TMPT (Thiopurine Methyltransferase)
TMPT
Substrates: treatments for Leukemia
♣ 6-mercaptopurine
♣ 6-thioguanine
o TMPT (methyltransferase) attaches a methyl group to 6-mercaptopurine and 6-thiguanine o If deficient, there are actually two potential effects: slow metabolism or increased efficacy.
CRITICAL that TMPT gene is working, otherwise 6-mercaptopurine and 6-thioguanine will run rampant and kill child as immosupressant
♣ Often presented as the classic example of a pharmacogenetic mechanism that can be fatal if ignored
G6PD (the glycolysis enzyme)
Substrates of G6PD:
- sulfonamide
- dapsone
Leads to hemolysis of the Red Blood Cell!!
10% of African American males are G6PD deficient
G6PD deficient individuals are susceptible to hemolytic anemia after drug
exposures
Favism example. G6PD is an x-linked enzyme, and most common disease producing enzyme in humans. G6PD makes NADPH, which protects cell against oxidative damage. Oxidant drugs deplete cells of glutathione, so it can no longer be reduced from its oxidized form by NADPH. This leads to oxidative damage. Favism kills due to G6PD deficiency.
