Correctors and Potentiators Rescue Function of the Truncated W1282X-Cystic Fibrosis Transmembrane Regulator (CFTR) Translation Product

Abstract

Edited by Thomas Sö llnerW1282X is the fifth most common cystic fibrosis transmembrane regulator (CFTR) mutation that causes cystic fibrosis. Here, we investigated the utility of a small molecule corrector/ potentiator strategy, as used for ⌬F508-CFTR, to produce functional rescue of the truncated translation product of the W1282X mutation, CFTR 1281 , without the need for readthrough. In transfected cell systems, certain potentiators and correctors, including VX-809 and VX-770, increased CFTR 1281 activity. To identify novel correctors and potentiators with potentially greater efficacy on CFTR 1281 , functional screens were done of ϳ30,000 synthetic small molecules and drugs/nutraceuticals in CFTR 1281 -transfected cells. Corrector scaffolds of 1-arylpyrazole-4-arylsulfonyl-piperazine and spiro-piperidine-quinazolinone classes were identified with up to ϳ5-fold greater efficacy than VX-809, some of which were selective for CFTR 1281 , whereas others also corrected ⌬F508-CFTR. Several novel potentiator scaffolds were identified with efficacy comparable with VX-770; remarkably, a phenylsulfonamide-pyrrolopyridine acted synergistically with VX-770 to increase CFTR 1281 function ϳ8-fold over that of VX-770 alone, normalizing CFTR 1281 channel activity to that of wild type CFTR. Corrector and potentiator combinations were tested in primary cultures and conditionally reprogrammed cells generated from nasal brushings from one W1282X homozygous subject. Although robust chloride conductance was seen with correctors and potentiators in homozygous ⌬F508 cells, increased chloride conductance was not found in W1282X cells despite the presence of adequate transcript levels. Notwithstanding the negative data in W1282X cells from one human subject, we speculate that corrector and potentiator combinations may have therapeutic efficacy in cystic fibrosis caused by the W1282X mutation, although additional studies are needed on human cells from W1282X subjects.

Positional effects of premature termination codons on the biochemical and biophysical properties of CFTR

“…However, the expression of this shorter CFTR can be increased by incubation with VX-809 ( Fig. 1C), as reported previously (Haggie et al 2017). Although the full-length read-through protein was not detected, even with G418 treatment (Fig.…”

“…; Haggie et al . ; Mutyam et al . ; ClinicalTrials.gov Identifier: NCT03624101 (https://clinicaltrials.gov/ct2/show/NCT03624101)).…”

“…Lumacaftor and Tezacaftor) to enhance the function of the truncated protein. However, it is unclear if these CFTR modulators, which are now clinical medicines, can be universally applied to PTCs since the relationship between the position of the PTC and the function of the truncated protein has yet to be established (Wang et al 2007;Haggie et al 2017;Mutyam et al 2017;Xue et al 2017;Yeh et al 2019b; ClinicalTrials.gov Identifier: NCT03624101 (https://clinicaltrials.gov/ct2/show/NCT03624101)). In theory, reagents that can promote read-through have the greatest potential for overcoming PTCs, and indeed compounds such as Ataluren and G418 have been shown to increase the expression of read-through CFTR in vitro (Howard et al 1996;Roy et al 2016) and in vivo (Wilschanski et al 2003;Du et al 2008).…”

ΔF508-CFTR Modulator Screen Based on Cell Surface Targeting of a Chimeric Nucleotide Binding Domain 1 Reporter

“…Also, a panel of known CFTR modulators was tested, including correctors VX-809, VX-661, C3, and C18, which target the NBD1-MSD interface, 9 C4, which targets NBD2, 16 CoPo-22, a CFTR modulator with dual corrector and potentiator function, 12 and W1282X corr -A23, which corrects CFTR 1281 , the protein product generated by the W1282X-CFTR nonsense mutation. 17 None of these correctors increased ΔF508-NBD1 cell surface expression; nor did previously described potentiators VX-770 and PG-01.…”

CFTR potentiators: from bench to bedside

“…A recent study showed that VX-770 acts additively with VX-809 to restore CFTR function in chronically treated R117H/ ΔF508 cells [52]. Surprisingly however, increased chloride conductance was not seen in W1282X cells despite the presence of adequate transcript levels when treated with both VX-770 and VX-809 [53]. As the combination of VX-770 and VX-809 (i.e., Orkambi) is now used clinically for treating patients carrying the ΔF508 mutation, it seems fitting to rigorously examine its effects on other CF mutations to potentially extend its application.…”