Patients vary in their responses to CFTR modulators in vitro HBE cells (Matthes et al., 2018). To examine the impact of CFTR deficiency on lipid profiles and cytokines in cells with strong and weak drug responses, cells were studied from the CF patients used in lipid analysis, three of which are homozygous and two heterozygous for F508del-CFTR, after selection based on CFTR dependent responses in Ussing chamber experiments (Table 1). Well-differentiated monolayers were pretreated for 24 h at 37°C with vehicle (0.1% DMSO) or under conditions designed to mimic the clinically-prescribed combination drugs lumacaftor/ivacaftor (1 μM VX809 for 24 h, followed by acute treatment with 300 nM VX770 or 50 μM genistein) or ivacaftor/tezacaftor/elexacaftor (3 μM VX445-1 plus 3 μM VX661 with or without 10 nM VX770 for 24 h). Monolayers were mounted in modified Ussing chambers and short-circuit current (I sc in μA·cm−2) was measured to assay of CFTR function as described previously (Matthes et al., 2018). Transepithelial voltage was clamped at 0 mV except for 2 s bipolar pulses to ±1 mV at 100 s intervals to monitor transepithelial resistance. A basolateral-to-apical NaCl chloride gradient was imposed to ensure a driving force for Cl secretion: Apical (in mM): 1.2 NaCl, 115 Na-gluconate, 25 NaHCO3, 1.2 MgCl2, 4 CaCl2, 2.4 KH2PO4, 1.24 K2HPO4, 10 Glucose; basolateral (in mM): 115 NaCl, 25 NaHCO3, 1.2 MgCl, 1.2 CaCl2, 2.4 KH2PO4, 1.24 K2HPO4, 10 Glucose. After the short-circuit current had stabilized (typically 2–3 min), Fsk (10 μM) was added to both sides to raise intracellular cAMP, and this was followed by sequential addition of a potentiator (50 μM genistein or 300 nM VX-770), the CFTR inhibitor CFTRinh-172 (10 μM) and the purinergic agonist ATP (100 μM, apical) to stimulate Ca2+-activated Cl channels as a positive control and to confirm cell viability (Matthes et al., 2016).

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