Toxicity and Genetics Flashcards
What are the different levels at which gene expression is regulated? Outline the differences between them.
- Basal gene expression: low level gene expression that is required all of the time.
- Constitutive gene expression: genes that are constantly expressed at higher levels, but need regulation. Makes use of enhancers such as TBP and HNF3.
- Reactionary gene expression: gene expression that is increased by stimuli. This is regulated by transcription factors that bind regulatory elements upstream of the gene.
How is gene expression changed in response to xenobiotics? Describe the ligand-receptor overlap and give examples.
Exposure to xenobiotics will result in changes in the expression of more than one gene.
- Ligand binding multiple receptors: Several genes have the same response elements in their promoters, i.e. activation of a single ligand receptor can lead to changes in the expression of several genes. Many ligand-activated receptors show a degree of overlap when a ligand can bind multiple substrates, or when xenobiotics can bind multiple receptors. For example, glucocorticoid can bind both the glucocorticoid receptor (GRα) and PXR, which both bind to the CYP3A4 promoter.
- Receptor being bound by multiple ligands: Many ligand-activated receptors share a response element, i.e. some genes have response elements that can be bound by more than one type of activated receptor, e.g. CYP2B6 and CYP3A4 can be bound by both PXR and CAR.
This means that there is more than one way to tackle xenobiotics that enter the body. The coordination of gene expression with xenobiotics is often termed the metabolic safety net, as it is thought to act as a survival system for the cell. Coordinated activation of alternative receptors or alternative genes will help minimise the toxic insult.
What variation exists in xenobiotic metabolism?
- Species variation
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Population variation
- Gender
- Geographical (SNPs)
Give an example of species variation in xenobiotic metabolism.
Paracetamol is metabolised to NAPQI, which is a highly reactive intermediate. Rats and rabbits have the same metabolic enzymes as mice, but mice are sensitive to paracetamol, whereas rabbits and rats are not. Paracetamol causes toxicity in mice because the rate of formation of NAPQI is greater than the rate that the liver of the mouse can remove it.
Between different species, different compounds are not necessarily metabolised using the same pathways. In rats and rabbits the phase II pathways are more reliant on glucuronide and sulphate conjugation (detoxification). Mice are reliant on glutathione conjugation which results in toxification.
Give two examples of how drug metabolism varies between men and women.
Give an example of geographical population variation in xenobiotic metabolism.
Most of the population’s response to drugs is a result of mutations in genes that code for drug metabolising enzymes, or proteins involved in the toxic response. Once a mutation reaches 1% of the population it is termed a polymorphism. Polymorphisms tend to be geographically clustered:
- 51% of the Chinese population have a polymorphism in CYP2D6, whereas 10% of the rest of the world have this mutation. Doses need to be adjusted for particular populations.
Give an example of a UDP-Glucuronosyltransferase polymorphism.
Polymorphisms in exons 2-5 of the gene encoding UGTs will result in every member of an enzyme family being affected (the subfamily members all share exons 2-5, so lots of enzymes will be affected).
- For example, one cluster of polymorphisms results in Crigler-Najjar syndrome, which is a result of bilirubin (excreted from liver to bile) not being converted into bilirubin glucuronide for excretion. This can be managed, but dosage of drugs needs to readjusted.
- Another example is in Gilberts syndrome, where there are higher than normal levels of bilirubin, and changes in enzyme levels may affect how these people metabolise drugs. Again, this needs to be taken into consideration for correct drug dosage.
Give an example of a Sulphotransferase polymorphism.
Polymorphisms have been identified in all of the human SULT genes, and functional polymorphisms have been identified in SULT1A1 and SULT1A2 genes. These result in a decrease in enzyme activity, but not a total reduction. There is less of an issue with these polymorphisms, as glucuronide metabolism is able to pick up the slack.
Give an example of a Glutathione Transferase polymorphism.
There are a number of clinically relevant human GSH polymorphisms, and they are present in high frequencies within the population. They can in some cases involved entire gene deletions, which lead to an increase risk of cancer. Glutathione conjugation is the biological ‘hoover’ to get rid of the most reactive intermediates, so polymorphisms in this pathway in particular will lead to formation of cancer. Colon cancer in particular because a lot of toxins/carcinogens are ingested in food that will then not be removed through this pathway.
Significant interindividual variation in CYP450 enzymes can be seen in populations. Which enzymes vary the most?
Significant interindividual variation in basal rates of P450-mediated drug metabolism have been observed, including up to 30- to 40-fold variation for CYP3A enzymes, 100-fold variation for CYP2D6, 50- to 60-fold variation for CYP2B6, and 40- to 50-fold variation for CYP2C9.
Some pharmacogenetic variation affects metabolism of certain drugs in particular, so are clinically relevant. Give two examples of such cases.
Clinically relevant examples of this pharmacogenetic variation have been observed with:
- CYP2C9 (for warfarin)
- CYP2C19 (for clopidogrel and omeprazole)
- CYP2D6 (for tamoxifen and codeine)
- CYP3A5 (for tacrolimus)
In some instances, the resulting protein is still functional but exhibits reduced activity (e.g., CYP2C9); in other cases, the resulting variant protein may be completely inactive or is not expressed (e.g., CYP2D6, CYP2C19, and CYP3A5).
Depending on whether metabolism produces an active or inactive metabolite, the consequences of these polymorphisms can result in significant alternations in therapeutic effect.
What type of gene expression do many of the CYP450 enzymes display? What can cause this?
The concentrations of many of the CYPs that are
involved in xenobiotic metabolism can be increased or
‘induced’ by xenobiotics via reactionary gene expression. Frequently, the inducer is also
a substrate for the induced CYP, which provides a
mechanism for amplifying the body’s detoxification
response during prolonged periods of xenobiotic
challenge. Once the chemical exposure is gone, CYP
levels can return to normal.
Two of the prototypical
inducers of CYP gene expression are the synthetic
steroid pregnenolone 16α-carbonitrile (PCN) and the
barbiturate phenobarbital (PB).
How is reactionary gene expression important in drug-drug interactions? For what drugs does this represent a problem in particular?
Although the induction of CYP gene expression represents an adaptive response to protect against chemical exposure, this physiological response is also the basis for an important class of drug–drug interactions.Many of the xenobiotics that induce CYP genes are widely used prescription drugs, such as the antibiotic rifampicin, the anti-inflammatory glucocorticoid dexamethasone, and the diabetes drug troglitazone.
The induction of these CYPs, in turn, accelerates the metabolism of other medications. CYP3A4 alone is responsible for the metabolism of 50–60% of all prescription drugs. So, drugs that induce CYP3A4 gene expression accelerate the metabolism of many other medications.
This phenomenon can be a serious problem in these
days of ‘polypharmacy’, in which a patient is likely to
be taking several medications. Ideally, drugs
should be developed that are neutral with respect to
their effects on CYP expression.
What nuclear receptor is a transcription factor that regulates CYP3A expression?
Pregnane X receptor (PXR)
Outline the nuclear receptor PXR.
Pregnane X receptor (PXR), which functions as a
heterodimer with the orphan retinoid X receptor
(RXR), is activated by a naturally occurring progesterone metabolite (5β-pregnane-3,20-dione26), and is expressed in the liver and intestine.
PXR is activated by micromolar concentrations of a surprisingly wide range of chemicals, including glucocorticoids, antiglucocorticoids, macrocyclic antibiotics, antifungals and herbal extracts, which have no obvious structural features in common. However, PXR ligands have a common biological property which is the activation of CYP3A activity.
