Cystic fibrosis is caused primarily by deletion of Phe508. An exciting discovery was that CFTR’s sister protein, the P-glycoprotein (P-gp) containing the equivalent mutation (ΔY490), could be repaired by a drug-rescue approach. Drug substrates showed specificity, and their mechanism involves direct binding to the transmembrane domains (TMDs) since arginine suppressor mutations were identified in TMDs that mimicked drug-rescue to promote maturation. We tested the possibility of rescuing CFTR processing mutants with a drug-rescue approach. 1) Arginine mutagenesis was performed on TM6, 8, and 12. 2) Correctors were tested for specificity. 3) Truncation mutants were used to map the VX-809 rescue site. Correctors 5a, 5c, and VX-809 were specific for CFTR. VX-809 appeared to specifically rescue CFTR by stabilizing TMD1. Therefore, the TMDs are potential targets to rescue CFTR. Rescue of P-gp and CFTR appeared to occur by different mechanisms since no arginine suppressor mutations were identified in CFTR.

Cystic fibrosis is caused primarily by deletion of Phe508. An exciting discovery was that CFTR’s sister protein, the P-glycoprotein (P-gp) containing the equivalent mutation (ΔY490), could be repaired by a drug-rescue approach. Drug substrates showed specificity, and their mechanism involves direct binding to the transmembrane domains (TMDs) since arginine suppressor mutations were identified in TMDs that mimicked drug-rescue to promote maturation. We tested the possibility of rescuing CFTR processing mutants with a drug-rescue approach. 1) Arginine mutagenesis was performed on TM6, 8, and 12. 2) Correctors were tested for specificity. 3) Truncation mutants were used to map the VX-809 rescue site. Correctors 5a, 5c, and VX-809 were specific for CFTR. VX-809 appeared to specifically rescue CFTR by stabilizing TMD1. Therefore, the TMDs are potential targets to rescue CFTR. Rescue of P-gp and CFTR appeared to occur by different mechanisms since no arginine suppressor mutations were identified in CFTR.

Cystic fibrosis (CF) is a rare genetic disease caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR), an epithelial anion channel expressed in several vital organs. Absence of functional CFTR results in imbalanced osmotic equilibrium and subsequent mucus build up in the lungs-which increases the risk of infection and eventually causes death. CFTR is an ATP-binding cassette (ABC) transporter family protein composed of two transmembrane domains (TMDs), two nucleotide binding domains (NBDs), and an unstructured regulatory domain. The most prevalent patient mutation is the deletion of F508 (F508del), making F508del CFTR the primary target for current FDA approved CF therapies. However, no experimental multi-domain F508del CFTR structure has been determined and few studies have modeled F508del using multi-domain WT CFTR structures. Here, we used cryo-EM density data and Rosetta comparative modeling (RosettaCM) to compare a F508del model with published experimental data on CFTR NBD1 thermodynamics. We then apply this modeling method to generate multi-domain WT and F508del CFTR structural models. These models demonstrate the destabilizing effects of F508del on NBD1 and the NBD1/TMD interface in both the inactive and active conformation of CFTR. Furthermore, we modeled F508del/R1070W and F508del bound to the CFTR corrector VX-809. Our models reveal the stabilizing effects of VX-809 on multi-domain models of F508del CFTR and pave the way for rational design of additional drugs that target F508del CFTR for treatment of CF.