Airway Inflammation Affects pH, Response to CFTR Therapies

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by Steve Bryson, PhD |

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Inflammation in the airways of people with cystic fibrosis (CF) regulates the pH of the surface liquid and increases the response to approved CFTR modulators, a cell-based study suggested.

Airway inflammation may be a key determinant in response to CF medicines, the researchers noted.

“To advance personalized CFTR modulation, it may be relevant to study the airway inflammatory milieu and its interactions with approved modulators,” they wrote.

The study, “Inflammatory cytokines TNFα and IL-17 enhance the efficacy of cystic fibrosis transmembrane conductance regulator modulators,” was published in the Journal of Clinical Investigation.

CF is caused by mutations in the gene that carries instructions for the cystic fibrosis transmembrane conductance regulator (CFTR), a cell membrane protein that allows the passage of chloride ions in and out of cells.

Chloride secretion helps balance water movement and regulates the volume of airway surface liquid (ASL), which is essential for the production of mucus. In the absence of functional CFTR, mucus becomes thick and sticky, damaging the lungs and increasing the risk of infection.

CFTR also transports bicarbonate ions (HCO3-) that control the pH — acid-alkaline balance — of the airway surface liquid. The loss of CFTR decreases alkaline bicarbonate secretion, leading to increased acidity (lower pH) of the airways, altering mucus properties, and impairing the ability to fight infections.

People with CF and animal models of the disease show abnormally acidic airways soon after birth. However, a few months later, the pH of the airway surface liquid increases to that seen in individuals without CF, even without normalized CFTR activity.

Studies suggest inflammation, which occurs in the weeks and months after birth, may influence airway pH by increasing bicarbonate secretion.

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Here, researchers based at the University of Iowa and their collaborators conducted a series of experiments in cells derived from CF patients to determine the impact of inflammation on the secretion of bicarbonate and pH of airway surface liquid.

Epithelial cells, which line the airways, were treated with TNF-alpha and interleukin-17 (IL-17), two pro-inflammatory proteins (cytokines) elevated in CF airways. After 24 hours, the pH of the surface liquid increased with either of the cytokine treatments and even more with a combination of TNF-alpha plus IL-17.

pH after TNF-alpha plus IL-17 peaked after two days and persisted up to day seven. Withdrawal of the cytokines caused the pH to return to pre-treatment levels, indicating that TNF-alpha plus IL-17 “synergistically increase [airways surface liquid pH], and this response is time-dependent, long-lasting, and reversible,” the team wrote.

A comparison of cytokine treatment in both CF and non-CF cells confirmed the inflammatory response was not affected by the presence or absence of CFTR activity. Further, experiments showed the increase in pH was not due to calcium-activated chloride channels, which also transport bicarbonate.

Analysis identified a gene called SLC26A4, which had low activity before treatment, and increased more than 100 times after TNF-alpha plus IL-17. This gene provides instructions to make pendrin, a membrane protein that transports chloride and bicarbonate.

Lowering pendrin reduced the elevated pH in treated epithelial cells, which suggested that “pendrin alkalinizes CF ASL, and might be particularly relevant to inflamed CF airways,” the researchers wrote.

The F508del mutation is the most common disease-causing mutation in CFTR, which leads to a misfolded and degraded CFTR protein. This defect can be modulated with Trikafta, a combination of three CFTR modulators: elexacaftor, tezacaftor, and ivacaftor.

TNF-alpha plus IL-17 modestly increased CFTR activity without these modulators, but after adding Trikafta, the combination significantly increased CFTR. An additional experiment showed that cytokine treatment increased CFTR gene activity by 3.5 times and CFTR by protein twofold, which indicated that TNF-alpha plus IL-17 “increased the CF epithelial response to CFTR modulators by increasing CFTR amount, biosynthesis, and function,” the researchers added.

Another common mutation — G551D — is a gating mutation that results in a CFTR protein that does not open correctly. Treatment with the so-called CFTR potentiator ivacaftor (sold as Kalydeco) can be used to keep the channel open longer.

TNF-alpha plus IL-17 increase the activity of CFTR with G551D in response to ivacaftor, which suggested that cytokine “enhancement of modulator efficacy is not specific to a particular class of mutations or modulators,” the scientists wrote.

Finally, to determine whether airway inflammation correlated with modulator-induced early lung function improvements in CF patients, the team analyzed data from a group of participants in a previous clinical trial with mutations affected by ivacaftor treatment.

Before treatment (baseline), all participants had inflammatory markers in their sputum. Lung function was assessed at baseline and two days after ivacaftor treatment, when any modulator-induced changes in inflammation would be minimal. A positive correlation was found between the presence of inflammatory markers before treatment and lung function improvement.

The researchers also found significant positive correlations between lung function and the inflammatory markers IL-1-beta and IL-8.

“In summary, the inflammatory milieu in CF airways regulates [airways surface liquid pH], and increases the response to CFTR modulators,” they wrote.