Genetics of Cystic Fibrosis Flashcards
Defne Cystic fibrosis
Multi-organ genetic disorder of the CFTR gene
Autosomal recessive
- Developed world: lung disease main cause of morbidity and mortatlity*
- Less-developed: Intestinal main cause of mortality (aless availbility of pancreatic enzymes)*
Describe the function of the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) protein
Lets Cl- ions flow into the lumen.
Inhibits eNaC (so ↓Na movement from lumen so that H2O follows Cl-)
Describe the consequences of a faulty CFTR
Less Cl- movement into the lumen
ENaC is active, so Na moves into the cell from lumen
H2O follows Na
Mucus dries up and becomes thick (Low volume hypothesis)
Cilia are not standing up straight
↓Mucociliary clearance
Name two hypotheses to explain the pathology of CF
- Low Volume Hypothesis
- Bicarbonate hypothesis
- High salt hypothesis - old and disproven*
- Aletered macrophage and neutrophil activity - jury still out (leads to impaired bacterial killing and clearance)*
Explain the bicarbonate hypothesis in the pathology of CF
CFTR allows HCO3- movement into the lumen to raise the pH of the mucus - created optimum pH environment
Mutated CFTR leads to ↓HCO3- into the lumen, therefore ↓pH (more acidic) and denaturing anti-bacterial proteins
There is dysregulation of the Airway Surface Liquid (ASL) Compartment
Explain the Low Volume Hypothesis in the pathology of CF
Mutated CFTR leads to no Cl- movement into lumen and dysregulation of ENaC (activates ENaC).
Na moves in from lumen into cell and H2O follows
Mucus dries up and there is inadequate airway surface liquid for the cilia to beat
Apart from potential low volume hypothesis, what else could be making the mucus thicker?
Infections are cleared by neutrophils mainly in CF.
Neutrophil death releases genomic DNA which makes mucus thick and sticky,
DNAse degrades the genetic material but it is expensive
Describe the risk of CF mutation in caucasians
Currently >1800 mutations identified for CF
1 in 25 (30)
Name some types of mutations
Missense
Frameshift
Splicing
Nonsense
In frame insertino/deletion
Large insertion/deletion
Promotor
Sequence Variation
Explain a missense mutation
Changing of a nucleotide in a sequence causing an amino acid change
Explain a frameshift mutation
An insertion or deletion causes a shift in the reading frame so that the codons are read in different triplets, coding for different amino acids
Explain a splicing mutation
The insertion, deletion of exons/introns to form a completely different amino acid sequence
Explain a nonsense mutation
The incorporation of a STOP codon
Sickel cell anaemia give a heterozygous advantage against malaria. What does CF give a heterozygous advantage to?
Cholera (potentiatlly)
Due to the involvement of Cl- channels in the pathogenesis of the watery diarrhoea
What is the most common mutation for CF and explain what the notation means
ΔF508
Deletion of three nucleotides at position 508, leading to a frameshift mutation (phenylalanine)
NBD1 and NBD2 involved in hydrolysis of ATP -> needed to transport ions across channel
Describe the normal processing of CFTR
Transcription from DNA
Translation by ribosomes
Post-translational modifications in golgi
Packaged into vesicles
Incorporated into the cell membrane
Describe and explain the 6 molecular consequences of CFTR mutations
- Class I
- No synthesis
- Big deletions
- Class II
- Block in processing
- Protein made in ER, but does NOT get to golgi
- ΔF508
- Class III
- Block in regulation
- R-domain is mutated
- Ivacaftor (potentiator)
- Class IV
- Altered conductance
- Gets to membrane but no ion movement
- Class V
- Reduced synthesis
- TF not binding too well?
- Class VI
- Increased recycling
- Functional protein, but recycled too quickly
ΔF508 has a 79% prevelance and G551D has 2.17%. Other mutations are < 1%. Explain the implicatoins of studying a disease with mutation prevelance like CF.
Less cases and therefore, RCT would not be useful to evaluate the effects of novel treatment.
Currently can use proof that the novel treatment treats a type of class mutations.
Describe the founder effect in terms of population genetics and CF.
The loss of genetic variation when a new population is established by a very small group from the whole population
W1282X is a nonsense (STOP) mutation found in 51% of Ashkazi Jews who have CF.
Describe the mechanistic basis of the sweat test in CF
Non-functioning CFTR in duct cells
Less re-absorption of Cl- from the gland secretions
Hypertonic sweat
FEV1 is often used as a primary outcome in respiratory research. Describe why it isn’t the most useful?
Effort-dependent
We use FEV1 anyway as it is the best we have at the moment
Ivacaftor (Kalydeco) is a good CFTR _____ but is only effective in 5% of patients. It’s main positive effects were:
- ↓_____
- ↓time to first pulmonary exacerbation
- ↑_____
- ↑_____
One method looked at taking _____ biopsy and observe en ex-vivo effect on the Cl- conductance. This is used for class _____ mutations.
Ivacaftor (Kalydeco) is a good CFTR potentiator but is only effective in 5% of patients. It’s main positive effects were:
- ↓Sweat chloride
- ↓time to first pulmonary exacerbation
- ↑10-12% FEV1
- ↑Weight
One method looked at taking intestinal biopsy and observe en ex-vivo effect on the Cl- conductance. This is used for class III mutations.
_____ is a CFTR protein corrector, which aims to treat class _____ mutations by increasing modificationand processing of proteins in the golgi. Trials showed changes in sweat chloriide but no differences in lung function.
Lumacaftor is a CFTR protein corrector, which aims to treat class II mutations by increasing modificationand processing of proteins in the golgi. Trials showed changes in sweat chloriide but no differences in lung function.
Ivacafotr and Lumacaftor was observed in combination and the results is as shown below. Interpret the data from the Phase IIa study and conclude whether it is useful or not.
Combination therapy shows significant increase in the change in FEV1 compared to placebo
No difference between high and low dose Lumacaftor
Not useful as the increase in FEV1 was only 2%
Describe the mechanism of action of ataluren
Targets genetic transcription to ignore a mutated stop codon
Especially useful for Jewish population due to high prevalance of Stop codon mutations
Describe two other alternative therapies for CF
- ENaC inhibitors
- Calcium activated Cl- channels
Describe whether there is a good genotype-phenotype correlation in:
Lungs
Pancreas
Lungs - NO
Pancreas - YES (Class I-III severe, Class IV-VI mild-moderate)
The same genetic mutation can occur but with different lung disease severity. What can affect the disease severity of CF?
- Other genetic factors
- Modifier genes
- Lower baseline of sodium transport
- Environmental
- Smoking
- Recurrent infections
- Socio-economic
- Adherance
- Access
Describe the phenotype seen in CFTR KO mice
Intestinal disease but no lung disease
