We established 14 CF-CRC cultures derived from affected patients and carriers with different CFTR defects. We validated all cultures for the presence of the patient CFTR gene defect, AESC phenotype and properties. CFTR transcript and protein analysis revealed low levels of CFTR in the CRC cells and higher levels in CRC-derived ALI-differentiated cells, therefore the latter may constitute a highly suitable CF model for in vitro CFTR experimentation. We optimized a second CF model consisting in 3D CRC-derived organoids mimicking.
the respiratory tissue in terms of structure and properties. The two optimized CRC-derived CF models proved highly suitable for the project aims of testing CFTR variants activity and pharmacological response. ALI model was validated for the pharmacological correction assays using positive control cells carrying F508del pathogenic variant (responsive patients) that proved highly responsive to Lumacaftor and Tezacaftor. We set up the organoid FIS assay in responsive carrier cells (marked swelling) and are currently optimizing the procedures to evaluate and quantify the ability of correctors to rescue defective CFTR function. For gene editing evaluation, a Droplet Digital PCR assay able to detect 1 wild type copy among 20,000 F508del mutated copies of DNA has been set up. These approaches represent powerful tools toward the introduction of more effective, patient-specific therapeutic options as well as to predict response to specific therapies for patients with rare mutations. The possibility to test cell response from each CF patient, to pharmacologic and genetic approaches may be of great translational impact, in the direction of theratyping. The new project FFC#14/2019 will continue these studies.

Congress abstracts

– Mazio C, Scognamiglio LS, Casale C et al. “Development of a 3D Full thickness cystic fibrosis model on chip” 41st European Cystic Fibrosis Conference, Belgrade, Serbia, 6-9 June 2018
– Scognamiglio LS, Mazio C, Casale C et al. “Development of a 3D full thickness cystic fibrosis model on chip” Thermschool2018, University of Trento, 18-22 June 2018