Pharmacogenomic discovery of therapeutic targets for corrector-refractory cystic fibrosis

ABSTRACT Chronic lung disease is the main cause of morbidity and mortality in cystic fibrosis (CF), a fatal genetic disorder caused by mutations in CFTR, a Cl– ion channel in the plasma membrane (PM) of the airway mucosa. Over 80% of people with CF (pwCF) have mutations that interfere with folding of CFTR in the endoplasmic reticulum (ER), causing the ion channel to be triaged by protein quality control (PQC) machinery and destroyed by the proteasome. Small-molecule “correctors” that bind directly to and promote the folding of CFTR have become the standard of care for people with ~180 CFTR variants, including the most common, F508del. Despite this success, thousands of pwCF are not helped by the FDA-approved correctors, either because they have ineligible variants that are unresponsive to the drugs, or they have eligible variants that, for unknown reasons, are non-responsive. Moreover, correction is inefficient because corrected CFTR variants that escape triage in the ER become substrates for PQC machinery in post-ER compartments. The lack of effective mechanism-based therapies for thousands of pwCF that do not respond to correctors is a critical unmet need demanding the development of novel therapeutics. Development of new treatments is hampered by a profound gap in knowledge of the cellular PQC machinery that promotes CFTR trafficking and degrades CFTR in the ER and post-ER compartments. The proposed studies will exploit leading-edge CRISPR-based pharmacogenomic analysis to fill this gap by identifying the PQC machinery that underlies CFTR variant triage in the presence and absence of correctors. Specific Aim 1 will use pooled genome-wide CRISPR analyses to comprehensively identify genes that govern the stability of CFTR-F508del in ER and post-ER compartments and to dissect their epistatic interactions with pharmacological CFTR correctors. Specific Aim 2 will modulate candidate PQC targets in human airway epithelial cells from pwCF who carry the F508del variant. Specific Aim 3 will leverage the methodology from Aims 1 and 2 to identify drug targets for corrector-ineligible CFTR variants using CRISPR screens and CFTR functional assays in patient-derived airway epithelia. The proposed studies will test the hypothesis that targeting cellular PQC systems is an effective approach to CF management that is orthogonal and complementary to existing and emerging CFTR modulators. Our findings are significant because they will help advance new targets for treating corrector-resistant CF.