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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Research Article Pulmonology

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 4

VX809 increases colocalization of CFTR and Nrf2 in CFhBE cells, which is blocked by coincubation with inhibitors of CFTR.

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VX809 increases colocalization of CFTR and Nrf2 in CFhBE cells, which is...
(A and B) Representative photographs, at ×60 original magnification, of IF for Nrf2 (green), CFTR (red), and nuclei (DAPI, blue) in primary NhBE (A) and CFhBE (B) cells treated with indicated doses of VX809 and/or CFTRinh-172 (Inh172) for 72 hours. Colocalization of CFTR and Nrf2 is in yellow, and arrows indicate apical (A) and basolateral (B) sides. Cells permeabilized as in Figure 2. (C and D) Quantification of Nrf2-CFTR colocalization in NhBE and CFhBE cells after incubation with 1–10 μM VX809 and/or 20 μM CFTRinh-172 or GlyH-101 for 72 hours (11 independent experiments with 6 CF and 5 non-CF donors, with 3 technical replicates per treatment) (C), or 1–10 μM VX809 or VX661 and/or 1 μM VX770 for 48 hours (4 independent experiments with 4 CF and 3 non-CF donors, with 3 replicates per treatment per donor) (D). Data 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. For C, *P < 0.05, ***P < 0.001 vs. NhBE vehicle control; †P < 0.05, †††P < 0.001 CFhBE treatments vs. CFhBE vehicle control; #P < 0.05 vs. same donor cells with same dose of VX809 by mixed-effects ANOVA and Dunnett’s multiple-comparisons test. For D, *P < 0.05, **P < 0.01, ***P < 0.001 vs. CFhBE vehicle control by mixed-effects ANOVA and Dunnett’s multiple-comparisons test. Scale bars: 10 μm.