K8/F508del-CFTR, a new target for corrector therapy in cystic fibrosis – CORCF

Cystic Fibrosis (CF) is a recessive autosomic life-limiting disease which affects more than 70000 patients around the world. It is caused by mutations in the Cystic Fibrosis Transmembrane Regulator (CFTR) gene coding for the CFTR protein. The most frequent mutation is the deletion of the 508th aminoacid (F508del), which results in a complex defect affecting both the folding of the protein and its trafficking to the membrane. Recent proof of concept studies have demonstrated that CFTR mutation-targeted therapies constitute an efficient therapy for CF. Correctors of F508del-CFTR identified so far have limited efficacy in vitro and disappointing bioactivity.
Our previous results support a novel strategy for F508del-CFTR correction. We have demonstrated a direct binding between F508del-CFTR and keratin 8 and found in different models (cell lines, F508del mice and primary respiratory cells of CF patients) that CFTR disruption of this interaction restores F508del-CFTR function. We then mapped the molecular target by computational modelization of K8/F508del-NBD1 interaction site and designed in silico by molecular docking 12 compounds targeting the site of interaction. Four of them corrected efficiently F508del CFTR trafficking defects and restored partially F508del CFTR function (correctors hereafter). In addition, we demonstrated that anti K8 antibodies also allowed to correct F508del-CFTR function. These results provide a novel specific target for corrector therapy in CF. We propose now to further develop this research by focusing on the 2 most active correctors and on implementation of specific anti-K8 antibodies.
The first objective of this project is to investigate in deep the mechanisms underlying action of the two most active correctors. The site of interaction between the correctors and F508del will be mapped using biophysical technics. Corrector binding sites on F508del-NBD1 will be further investigated in epithelial cells by site-directed mutagenesis of aminoacids located in the K8/NBD1 interaction site. Impact of the correctors on biogenesis of F508del-CFTR will be evaluated by studying change in protein half life, ubiquitination status, proteolysis and maturation. The second objective is to optimize the efficacy correctors by a structure-activity strategy. For both first generation and optimized correctors, we will investigate change in CFTR function (ion transport and cellular inflammation profile).
The third objective is to implement antibodies against the region of K8 identified by hydrogen deuterium exchange as interacting with F508del-NBD1 and initiate the studies of cellular mechanisms underlying their action. The fourth objective is to perform a dose-finding approach in WT mice followed by pharmacokinetic/pharmacodynamic studies in F508del homozygous mice.
The results of this project should provide mechanism-based and target-specific novel compounds for mutation therapy in F508del homozygous patients.