Scott: Cystic Fibrosis Flashcards
What type of genetic disease is CF?
Autosomal RECESSIVE
What causes CF?
INACTIVATING MUTATIONS in CF transmembrane CFTR gene
What is the second most frequent life-shortening genetic disease in hte US?
CF
What is CFTR?
chloride ion channel
Where is CFTR found?
on the apical surface of epithelial cells lining:
AIRWAYS
PANCREATIC DUCTS
INTESTINES
What are the most serious consequences and common COD from CF?
Pulmonary
Progressive lung disease causes death in 90% of pts
What are the clinical consequences of CF?
- Pulmonary
- GI- exocrine pancreas, diabetes, intestine
- Male infertility
Where is the CFTR gene found?
long arm of chromosome 7
What ethnic group is most commonly affected by CF?
Caucasians
Hispanics
African americans
Asian americans
What is a class I CFTR mutation?
NO PROTEIN PRODUCED
Nonsense mutation creates stop codon, Often mRNA is degraded
G542X, 5% of CF alleles
What is a class 2 CFTR mutation?
DEFECTIVE PROTEIN FOLDING
Activates ER quality control,
degradation of protein
F508del, 70% of CF alleles
What is a class 3 CFTR mutation?
DEFECTIVE GATING OR REGULATION OF CHANNEL OPENING
G551D, 4% of CF alleles
What is a class 4 CFTR mutation?
defective in ion transport
What is a class 5 mutation?
Normal CFTR produced but decreased amounts
What are more severe mutations? Less severe?
More severe- 1-3
Less severe- 4 and 5
What characterizes severe CFTR mutations?
≤ 1% CFTR activity remaining Diagnosed in FIRST YEAR Median survival 37.4 y Pancreatic INSUFFICIENT At risk for CF-related diabetes, liver disease
What characterizes LESS severe CFTR mutations?
~5% CFTR activity remaining
May have LATE presentation
Survival to 50 not uncommon
Pancreatic SUFFICIENT
5 mutations occur in > 1% of US CF patients- F508Δ, G542X, G551D, W1282X,
N1303K. What category are they in?
SEVERE
What is the structure of the CFTR chloride ion channel?
The MSD forms a PORE for the chloride ion channel.
Nucleotide binding domains NBD and R domain provide REGULATORY SITES that promote opening of channel.
What do the NPD and R domains do?
NBD binds ATP
R has phosphorylation site for PKA
What is the F508del folding defect?
Principle folding defect
Where is the CFTR protein complex synthesized and what does it require?
ER
INTRA domain folding and INTER domain interactions
chaperones
What causes the F508del defect?
Incorrect folding of the NBD1 domain
Incorrect interaction of NBD1 with other domains
What happens when there is incorrect folding?
Altered interaction w/ chaperones
- RETENTION in ER and activation of quality control pathways
- DEGRADATION by the proteasome
Which mutations cause the most severe disease?
Mutations that result in little or NO functional protein
What outcomes are commonly associated w/ SEVERE mutations?
- Defect in pancreatic duct> pancreatic enzyme insufficiency
- Worse pulmonary disease
- Diabetes and Liver disease (severe mutations necessary to cause these but not all do)
What are the most frequent CF mutations in the US?
G542X
G551D
F508del
What is the G542X mutation?
Mutation creates stop codon at amino acid 542–>
Decreased mRNA/ non-functional protein
What is G551D?
Mutation makes ion channel unable to respond to ATP opening signal> greatly increase time in CLOSED conformation
What the F508del mutation?
Most common CFTR mutation
Almost complete LOSS of functional protein
- Folding defect–> degradation of most protein
- small fraction of protein at cell surface is defective in gating
What happens when the CFTR is functioning normally?
When the channel is OPEN ion channels flow through it “downhill” in the direction of the EC gradient>
EC gradient across the airway epithelial apical cell membrane favors flow of Cl ions OUT of the cell and into the EXTRACELLULAR space>
Increased concentration of Cl ions OUTSIDE the cell>
Creates osmotic conditions for flow of water out of the cell
What does the opening and closing of CFTR channel control?
Movement of water to apical surface of epithelial cells
What happens if the CFTR is NOT function or not regulated correctly?
Insufficient water is delivered to the cell surface
What is required to open the chloride channel?
Two signals:
- Phorsphorylation of R domain by PKA (sets conditions for opening making it possible for ATP to communicate w/ channel). PKA is activated by cAMP
- Binding of ATP to two NBD domains–> conformational change
What determines the balance of water between the cell surface and the cell interior?
How long the CFTR stays open
What closes channels?
Absence of environmental signals>
dephosphorylation of R domain
Hydrolyzation of ATP
Why is CFTR chloride ion channel essential for respiratory function?
Airway surface liquid volume (ASL) is dependent on CFTR
What is ASL?
A layer of liquid about 7 um thick overlying airway epithelium that is essential for the clearance of mucus (activity of cilia and hydration of mucus)
What happens to ASL if CFTR is defective?
ASL layer is NOT maintained
What happens if there is a decreased ASL layer?
Chronic infection–> permanent damage
What are the 2 ways that a decreased ASL layer can lead to chronic infection?
- Build up of viscous mucous> failure to clear bacteria
2. Persistent colonization and desicated mucus> inflammation
What are characteristic CF bacterial infections?
Stapholococcus aureus,
Pseudomonas aeruginosa, Burkholderia cepacia
What are the 2 ways that a decreased ASL can lead to permanent damage?
- Infection, inflammation and mucus plugging> long term structural damage (bronchioletasis, bronchiectasis)
- Unless stopped ultimate outcome is respiratory failure
What is the major COD for most individuals w/ CF?
Pulmonary failure d/t airway infection
What are the major organisms that infect the airways of pts w/ CF?
Haemophilus influenzae Staphylococccus aureus Pseudomonas aeruginosa (pediatric pts) Burkholderia cepacia Aspergillus fummigatus
*these pathogens are usually NOT found in non-CF population
What is the goal of treatment for pt’s w/ pulmonary related CF problems?
LIMIT mucus buildup
PREVENT/TREAT infections
What is the mainstay tx for pulmonary sxs of CF?
Chest physiotherapy to improve drainage
Aggressive treatment of infection- antibiotics
What is additional tx for pulmonary sxs of CF?
Hypertonic saline and Dnase aerosols to make MUCUS LESS VISCUS
Anti-inflammatories
Where is CFTR located in the pancreas?
Epithelial cells lining PANCREATIC DUCT express CFTR
Base of the duct secretes high concentration of proteins, especially proteolyctic enzymes
What do the CFTR expressing cells of the pancreas secrete?
HCO3
What does the secretion of HCO3 in the pancreas do?
recruits luminal water and neutralizes secretions
prevents formation of protein plugs
What happens in the pancreas if the CFTR lose function?
BLOCKAGE OF DUCT by secreted protein plugs>
damage to cells>
leakage of proteolytic enzymes>
destruction of tissue
What is the pancreatic phenotype of CFTR dependent on?
severity of the genetic mutation
Where is CFTR expressed in the intestine?
Crypts of intestinal epithelium
What happens if the intestinal CFTR lose function?
fail to secrete water>
dehydration of the lumen and viscus mucus
What are the consequences of loss of function of CFTR in the intestine?
- Obstruction (Meconium illeus-infants, and DIOS-older)
- Inability to absorb protein/fats
- Susceptibility to GI cancers
How does CF affect male fertility?
Vas deferens is ABSENT
Becomes OBSTRUCTED in the fetus/infancy and is REABSORBED
How does CFTR act in sweat glands?
Promotes uptake of Cl ions from extracellular space w/ Na following
How does CF affect sweat glands?
Decreased ability to absorb Cl and Na> increased NaCl in sweat
NO TISSUE DAMAGE–> other ion channels compensate
What is a definitive diagnostic test for CF?
Sweat test
How does CF affect the pancreas?
- Pancreatic exocrine insufficiency
2. CF related diabetes
What is pancreatic exocrine insufficiency? How does his affect individuals w/ two “severe” alleles?
Malabsorption of lipids
Malabsorption of fat based vitamins
All individuals with two “Severe” alleles are pancreatic insufficient and require pancreatic enzyme replacement therapy to maintain nutrition
What is CF related diabetes and who does it affect??
Only occurs in homozygous for “severe” alleles but not all
What are characteristics of both T1D and T2D associated w/ CF related diabetes?
Loss of pancreatic cell mass
AND
insulin resistance
How id molecular genetic knowledge used for better testing and treatment?
- widespread screening
2. new txs to correct specific mut defects
Why is screening for CF feasible?
Molecular genetics!
CFTR gene mutation is KNOWN
DNA seq of most muts is KNOWN
What is the ACOG recommendation for genetic carrier screening?
Offer to ALL individuals planning a pregnancy or seeking prenatal care
When do you offer prenatal testing for CF?
When 25% chance of CF
Both parents are carriers or they have a previous child w/ CF
Do we screen newborns for CF?
YES
required in ALL st
What is the value of carrier screening?
Allows parents to know which group they fall into
Two carriers: 1/4
1 carrier: 1/1000
Neither: 1/250,000
What is the carrier frequency for individuals of northern european caucasian ethnic background?
1/25
How is prenatal testing for CF conducted?
Chorionic villus sampling 10-12 weeks for DNA mutation testing
Amniotic fluid 16-17 weeks for DNA mutation testing
Also ultrasound for echogenic bowel
Why does newborn screening for CF and early detection make a difference?
Nutrition- pancreatic enzyme replacement
Pulmonary function- aggressive tx to prevent obstruction, infection
What is the typical protocol for newborn screening of CF?
- IRT immunoreactive trypsin- test of pancreatic function. Part of panel done on heel stick blood sample.
(screening step 1) - DNA mutation analysis based on panel representing vast majority of disease causing mutations
(screening step 2) - sweat test
test of CFTR function
(diagnostic)
What is the definitive test for CF?
Sweat test
Induce sweat w/ chemical treatment>
sweat released over 30 min is absorbed>
Cl content measured
Is DNA testing for CF done?
Yes , b/c of new treatments that are mutation specific
How do we treat CF?
Aggressive, meticulous treatment of all possible manifestations
What are the major objectives when treating CF?
- Lung- clear secretions, control infections
- Maintain nutrition
- Prevent intestinal obstruction
What is a curative intervention for pulmonary sxs from CF?
lung transplant
What are the two new strategies to treat CF?
- Lung transplant- 60% survival after 2 years
2. Drugs to target specific deficiencies of CFTR mutations
What drug targets G551D (represents 4% of CFTR mutations and is a defect in ATP regulation of gating)?
IVACAFTOR (Kalydeco)-
corrector for G551D
keeps channel in open conformation for higher percent of time, even without ATP gating
IMPROVES FEV1 BY 10%
FDA approved Jan. 2012
What drug targets F508 mutation (70% of alleles, Defect in folding leads to degradation before reaching cell surface
Small percent that reaches surface is defective in gating)?
IVACAFTOR + LUMACAFTOR
Lumacaftor is a corrector for F508 and promtes traffic to cell surface instead of degradation
Combination of the two–> 4-7% improvement of FEV1
What drug targets G542X (Premature stop codon, 5% of alleles)?
ATALUREN
corrector for premature stop codon mutations
causes ribosome to be less sensitive to stop codons
phase 3 clinical trials show some improvement, but not statistically significant
