Clinically approved CFTR modulators rescue Nrf2 dysfunction in cystic fibrosis airway epithelia
Dana C. Borcherding, … , Scott M. Plafker, Assem G. Ziady
Dana C. Borcherding, … , Scott M. Plafker, Assem G. Ziady
Published May 30, 2019
Citation Information: J Clin Invest. 2019;129(8):3448-3463. https://doi.org/10.1172/JCI96273.
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Clinically approved CFTR modulators rescue Nrf2 dysfunction in cystic fibrosis airway epithelia

Abstract

Cystic fibrosis (CF) is a multiorgan progressive genetic disease caused by loss of functional cystic fibrosis transmembrane conductance regulator (CFTR) channel. Previously, we identified a significant dysfunction in CF cells and model mice of the transcription factor nuclear factor E2–related factor-2 (Nrf2), a major regulator of redox balance and inflammatory signaling. Here we report that the approved F508del CFTR correctors VX809 and VX661 recover diminished Nrf2 function and colocalization with CFTR in CF human primary bronchial epithelia by proximity ligation assay, immunoprecipitation, and immunofluorescence, concordant with CFTR correction. F508del CFTR correctors induced Nrf2 nuclear translocation, Nrf2-dependent luciferase activity, and transcriptional activation of target genes. Rescue of Nrf2 function by VX809/VX661 was dependent on significant correction of F508del and was blocked by inhibition of corrected channel function, or high-level shRNA knockdown of CFTR or F508del CFTR. Mechanistically, F508del CFTR modulation restored Nrf2 phosphorylation and its interaction with the coactivator CREB-binding protein (CBP). Our findings demonstrate that sufficient modulation of F508del CFTR function corrects Nrf2 dysfunction in CF.

Authors

Dana C. Borcherding, Matthew E. Siefert, Songbai Lin, John Brewington, Hesham Sadek, John P. Clancy, Scott M. Plafker, Assem G. Ziady

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Figure 5

F508del knockdown in CFhBE cells blocks CFTR modulator–mediated activation of Nrf2.

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F508del knockdown in CFhBE cells blocks CFTR modulator–mediated activati...
(A–D) CFhBE cells were infected with CFTR shRNA or scrambled control (Scr Con) lentivirus for 4 days, then treated with vehicle (DMSO) control or 1–10 μM VX809 for 48 hours. Gene expression was determined for CFTR (A), HMOX1 (B), NQO1 (C), and GCLC (D), by real-time qPCR, with mRNA levels shown as fold changes versus Scr Con cells incubated with vehicle control (DMSO). Data for 4 independent experiments from 3 CF donors with 3 replicates per treatment per donor are expressed as box-and-whisker plots. Horizontal bars indicate the median, box borders indicate 25th and 75th percentiles, and whiskers indicate 5th and 95th percentiles. *P < 0.05, **P < 0.01, ***P < 0.001 vs. Scr Con cells treated with DMSO control, or #P < 0.05 vs. Scr Con cells treated with the same dose of VX809, by mixed-effects ANOVA and Dunnett’s multiple-comparisons test. (E and F) Primary CFhBE cells were incubated with vehicle control (DMSO) or the indicated doses of VX809 or VX661, with or without 1 μM VX770, for 48 hours, and gene expression of HMOX1 (E) or NQO1 (F) was determined by qPCR as above. Data for 3–4 independent experiments from 3 CF donors with 3–4 replicates per treatment per donor are expressed as box-and-whisker plots. Horizontal bars indicate the median, box borders indicate 25th and 75th percentiles, and whiskers indicate 5th and 95th percentiles. *P < 0.05, **P < 0.01, ***P < 0.001 vs. DMSO control cells by 1-way ANOVA (Control group) or 2-way ANOVA (VX770 group) and Dunnett’s multiple-comparisons test; #P < 0.05 vs. control cells treated with the same dose of VX809/VX661 by 1-way ANOVA.