POSSIBLE USES OF CYSTIC FIBROSIS DRUGS IN PROTEIN PROCESSING IN THE ER

PROTEINS ARE ESSENTIAL FOR CELLULAR FUNCTION, AND THEIR CORRECT FOLDING WITHIN THE ER IS CRUCIAL FOR MAINTAINING CELL HOMEOSTASIS. WHEN PROTEIN QUALITY CONTROL FAILS, MISFOLDED PROTEINS ACCUMULATE, TRIGGER ER STRESS, AND ACTIVATE THE UNFOLDED PROTEIN RESPONSE (UPR), WHICH CAN ULTIMATELY LEAD TO CELL DEATH. THIS CONDITION UNDERLIES SEVERAL CONFORMATIONAL DISORDERS, INCLUDING CYSTIC FIBROSIS (CF), CAUSED BY MUTATIONS IN THE CFTR GENE. CF THERAPY HAS BEEN REVOLUTIONIZED BY MODULATORS, PARTICULARLY CORRECTORS, WHICH IMPROVE THE AMOUNT AND STABILITY OF MISFOLDED CFTR AT THE PLASMA MEMBRANE. HOWEVER, THEIR PRECISE MECHANISM OF ACTION IN THE CELLULAR ENVIRONMENT REMAINS UNCLEAR, AND A DIRECT BINDING TO CFTR HAS NOT BEEN DEMONSTRATED. ON THIS BASIS, WE HYPOTHESIZED THAT CFTR CORRECTORS MAY EXERT BROADER CYTOPROTECTIVE EFFECTS AND COULD BE REPURPOSED FOR DISEASES CHARACTERIZED BY IMPAIRED PROTEIN PROCESSING IN THE ER. WE FIRST INVESTIGATED LUMACAFTOR (VX-809), THE FIRST CLINICALLY APPROVED CORRECTOR, IN LUNG ADENOCARCINOMA AND MELANOMA CELL MODELS IN WHICH ER STRESS WAS INDUCED BY THAPSIGARGIN, A SERCA INHIBITOR THAT DISRUPTS ER CA²⁺ HOMEOSTASIS. VX-809 SIGNIFICANTLY MODULATED KEY ER STRESS-RELATED PATHWAYS, INCLUDING UPR SIGNALING, APOPTOSIS, OXIDATIVE STRESS, AND INFLAMMATION. CONSIDERING THE HIGH BURDEN AND LIMITED THERAPEUTIC OPTIONS OF NEURODEGENERATIVE DISEASES, WE THEN TESTED VX-809 IN A NEUROBLASTOMA MODEL COMMONLY USED TO STUDY THESE DISORDERS AND CONFIRMED A BROAD PROTECTIVE EFFECT AGAINST ER STRESS. SINCE THE CORRECTOR CURRENTLY USED IN CLINICAL PRACTICE IS ELEXACAFTOR (VX-445), WE NEXT PERFORMED A DIRECT COMPARISON BY REPEATING THE FULL IN VITRO PANEL IN LUNG ADENOCARCINOMA AND NEUROBLASTOMA CELLS. FUNCTIONAL PROTEOMIC ANALYSES IN NEUROBLASTOMA CELLS REVEALED THAT VX-445 LARGELY RECAPITULATES AND EXTENDS VX-809 EFFECTS, WITH A PROMINENT ROLE IN THE REGULATION OF CA²⁺ HOMEOSTASIS AND OXIDATIVE STRESS. FINALLY, BY ASSESSING MITOCHONDRIAL APOPTOSIS, MEMBRANE POTENTIAL, OXIDATIVE STRESS, AND CALCIUM HANDLING, WE SHOWED THAT VX-445 ALSO MODULATES ALL THESE MITOCHONDRIAL PROCESSES, INDICATING A SPECIFIC MITOCHONDRIAL-DIRECTED ACTION. ON THE BASIS OF THE RESULTS OBTAINED, IT IS WELL-FOUNDED TO HYPOTHESIZE THAT THE CORRECTORS INVESTIGATED IN THIS THESIS COULD ALSO BE EMPLOYED FOR THE TREATMENT OF OTHER DISEASES ASSOCIATED WITH ALTERED PROTEOSTASIS IN THE ER.