Abstract
Background: Pharmacotherapies for people with cystic fibrosis (pwCF) who have premature termination codons (PTCs) in the cystic fibrosis transmembrane conductance regulator (CFTR) gene are under development. Thus far, clinical studies focused on compounds that induce translational readthrough (RT) at the mRNA PTC location. Recent studies using primary airway cells showed that PTC functional restoration can be achieved through combining compounds with multiple mode-of-actions. Here, we assessed induction of CFTR function in PTC-containing intestinal organoids using compounds targeting RT, nonsense mRNA mediated decay (NMD) and CFTR protein modulation. Methods: Rescue of PTC CFTR protein was assessed by forskolin-induced swelling of 12 intestinal organoid cultures carrying distinct PTC mutations. Effects of compounds on mRNA CFTR level was assessed by RT-qPCRs. Results: Whilst response varied between donors, significant rescue of CFTR function was achieved for most donors with the quintuple combination of a commercially available pharmacological equivalent of the RT compound (ELX-02-disulfate or ELX-02ds), NMD inhibitor SMG1i, correctors VX-445 and VX-661 and potentiator VX-770. The quintuple combination of pharmacotherapies reached swelling quantities higher than the mean swelling of three VX-809/VX-770-rescued F508del/F508del organoid cultures, indicating level of rescue is of clinical relevance as VX-770/VX-809-mediated F508del/F508del rescue in organoids correlate with substantial improvement of clinical outcome. Conclusions: Whilst variation in efficacy was observed between genotypes as well as within genotypes, the data suggests that strong pharmacological rescue of PTC requires a combination of drugs that target RT, NMD and protein function.
Highlights
Cystic fibrosis (CF) is a monogenic, autosomal-recessive disease caused by mutations in the CFTR gene [1]
RT-agent nonsense mRNA mediated decay (NMD)-inhibitor CFTR amplifier CFTR potentiator CFTR corrector CFTR co-potentiator NMD-inhibitor NMD-inhibitor at the place of the premature termination codons (PTCs) results in a pool of full-length transcripts of which function could be enhanced with CFTR modulation therapy (VX-770, VX-661, VX-445 and ASP-11)
Combining these small molecules acting on different steps along the CFTR biosynthesis pathway, channel trafficking and channel gating might collectively result in CFTR function restoration of PTCs in general, to clinically relevant levels
Summary
Cystic fibrosis (CF) is a monogenic, autosomal-recessive disease caused by mutations in the CFTR gene [1]. We assessed induction of CFTR function in PTC-containing intestinal organoids using compounds targeting RT, nonsense mRNA mediated decay (NMD) and CFTR protein modulation. Methods: Rescue of PTC CFTR protein was assessed by forskolin-induced swelling of 12 intestinal organoid cultures carrying distinct PTC mutations. Effects of compounds on mRNA CFTR level was assessed by RT-qPCRs. Results: Whilst response varied between donors, significant rescue of CFTR function was achieved for most donors with the quintuple combination of a commercially available pharmacological equivalent of the RT compound (ELX-02disulfate or ELX-02ds), NMD inhibitor SMG1i, correctors VX-445 and VX-661 and potentiator VX-770. Conclusions: Whilst variation in efficacy was observed between genotypes as well as within genotypes, the data suggests that strong pharmacological rescue of PTC requires a combination of drugs that target RT, NMD and protein function
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