Fig. 5.

Potentiator combinations rescue the function of CFTR gating mutants in CFBE cells. (a) The effect of the indicated single potentiators or potentiator combinations on the I_sc of G551D-CFTR. I_sc was measured in presence of a basolateral-to-apical chloride gradient after basolateral permeabilization with amphotericin B (100 μM) and inhibition of the epithelial sodium channel ENaC with 100 μM amiloride. (b) Effect of acute addition of forskolin (20 μM) followed by VX-770 (10 μM) or ABBV-974 (10 μM) and apigenin (50 μM), bDMC (10 μM) or P7 (50 μM) as percent of forskolin-activated WT-CFTR (n = 3). (c) Comparison between the acute and chronic (24 hours, VX-770 – 3 μM, ABBV-974 – 3 μM, bDMC - 10 μM, apigenin and P7 – 50 μM) potentiator effect on the acute forskolin-stimulated G551D I_sc (n = 3). (d) The fractional PM activity of G551D after forskolin-stimulation alone or in combination with single or dual potentiator treatment expressed as percentage of WT (n = 3). (e) Effect of acute addition of forskolin (20 μM) followed by VX-770 (10 μM) or ABBV-974 (10 μM) and subsequent apigenin (50 μM), bDMC (10 μM) or A15 (50 μM) on the I_sc of R352Q, S549R, S549N, G1244E and S1251N expressed as percentage of WT-CFTR function (n = 3). (f) Fractional PM activity of R352Q, S549R, S549N, G1244E and S1251N-CFTR upon forskolin alone or in combination with single or dual potentiator treatment, expressed as percentage of WT (n = 3). (g) Effect of order-of-addition on the efficacy of G551D potentiation (10 μM VX-770, 50 μM apigenin) (left panel, n = 3) and potency of VX-770 in the presence or absence of apigenin (50 μM) (right panel, n = 3). Data in b–g are means ± SEM of the indicated number of independent experiments. * P < 0.05, ** P < 0.01, *** P < 0.001 by unpaired, two-tailed Student’s t-test.