The cereblon E3 ligase modulator, CC-90009, rescues CFTR nonsense variants by promoting NMD suppression and PTC readthrough in rectal organoids

Approximately 10% of the worldwide cystic fibrosis (CF) population has premature termination codon (PTC) variants, that prevent translation of cystic fibrosis transmembrane conductance regulator (CFTR) full-length protein and trigger nonsense-mediated mRNA decay (NMD). CFTR modulators approval opened a new era for CF precision medicine, but to date no therapies are still available to restore PTC variants. Readthrough agents that target distinct components of the translation machinery represent a promising and feasible treatment strategy. Analysis of primary cells representative of different variants and patients is necessary to evaluate and compare their effectiveness for a potential clinical evaluation in CF. Here we investigate the efficacy of the cereblon (CRBN) E3 ubiquitin ligase modulator, CC-90009, in restoring CFTR function and expression using patient-derived rectalorganoids homozygous for W1282X and G542X variants. Drugs testing were performed on 3D model by Forskolin Induced Swelling (FIS) assay and analysed with an in-house developed AI-based software (SOFTD). CFTR mRNA expression was characterized in response to all compounds by quantitative PCR. Our data show that CC-90009, as stand alone, induces significant increase of FIS rates compared to vehicle in our cultures, comparable to Vx809-Vx770-treated F508del/F508del carrying organoids. Combination with the aminoglycosides G418 or ELX-02 significantly enhances their mutual effects, dramatically increasing the PTC readthrough response. Furthermore, the mRNA level is incremented by CC-90009 treatment and correlates with functional recovery. Our results demonstrate that the eRF3a degrader, CC-90009, alone or in combination with other compounds, could represent a promising therapeutic approach to rescue CFTR nonsense variants.