Cystic Fibrosis Transmembrane Regulator Inhibitors CFTR_inh-172 and GlyH-101 Target Mitochondrial Functions, Independently of Chloride Channel Inhibition

Abstract

Two highly potent and selective cystic fibrosis (CF) transmembrane regulator (CFTR) inhibitors have been identified by high-throughput screening: the thiazolidinone CFTR inh -172 [3-[(3-trifluoromethyl)phenyl]-5-[(4-carboxyphenyl)methylene]-2-thioxo-4-thiazolidinone] and the glycine hydrazide GlyH-101 [N-(2-naphthalenyl)-((3,5-dibromo-2,4-dihydroxyphenyl)methylene)glycine hydrazide]. Inhibition of the CFTR chloride channel by these compounds has been suggested to be of pharmacological interest in the treatment of secretory diarrheas and polycystic kidney disease. In addition, functional inhibition of CFTR by CFTR inh -172 has been proposed to be sufficient to mimic the CF inflammatory profile. In the present study, we investigated the effects of the two compounds on reactive oxygen species (ROS) production and mitochondrial membrane potential in several cell lines: the CFTR-deficient human lung epithelial IB3-1 (expressing the heterozygous F508del/ W1282X mutation), the isogenic CFTR-corrected C38, and HeLa and A549 as non-CFTR-expressing controls. Both inhibitors were able to induce a rapid increase in ROS levels and depolarize mitochondria in the four cell types, suggesting that these effects are independent of CFTR inhibition. In HeLa cells, these events were associated with a decrease in the rate of oxygen consumption, with GlyH-101 demonstrating a higher potency than CFTR inh -172. The impact of CFTR inhibitors on inflammatory parameters was also tested in HeLa cells. CFTR inh -172, but not GlyH-101, induced nuclear translocation of nuclear factor-B (NF-B). CFTR inh -172 slightly decreased interleukin-8 secretion, whereas GlyH-101 induced a slight increase. These results support the conclusion that CFTR inhibitors may exert nonspecific effects regarding ROS production, mitochondrial failure, and activation of the NF-B signaling pathway, independently of CFTR inhibition.

Divergent CFTR orthologs respond differently to the channel inhibitors CFTRinh-172, glibenclamide, and GlyH-101

“…CFTR inh -172 does not function as an open channel blocker since there is no rectification of current during block (32). Several different mechanisms of block have been proposed (24,27). For GlyH-101, no site-directed mutagenesis studies have been performed.…”

“…CFTR inh -172 inhibits sodium transport in sweat glands (62). Both CFTR inh -172 and GlyH-101 affect mitochondrial function, independent of their action on CFTR (24).…”

Disease-relevant proteostasis regulation of cystic fibrosis transmembrane conductance regulator

“…In spite of the fact that short-time CFTR-unrelated effects of CFTR inh-172 have been described in CFTR lacking HeLa cells, 48 We also investigated the effects of CFTR depletion on EGF-induced trafficking of the EGFR. In 16HBE14o-cells transfected with scrambled siRNAs, EGF (and its receptor) was efficiently degraded after 30 and 60 min of stimulation.…”

Interaction with CREB binding protein modulates the activities of Nrf2 and NF-κB in cystic fibrosis airway epithelial cells

“…These data are consistent with our previous studies showing that inhibition of CFTR with CFTR inh -172 was sufficient to inhibit Nrf2 activity and thereby increase oxidant-mediated elevations in cytokine levels (7). Recently, Kelly and colleagues (14) concluded that increases in oxidants and inflammatory signaling in CF cell are independent of CFTR channel function loss, as CFTR inh -172 elicited these changes in cells that lack detectable levels of CFTR. However, these studies did show elevated levels of intracellular oxidants in the absence of CFTR inh -172 in CF vs. non-CF controls.…”