High-throughput screening for modulators of CFTR activity applying an organotypic functional assay based on genetically engineered Cystic Fibrosis disease-specific iPSCs

Organotypic culture systems from disease-specific induced pluripotent stem cells (iPSCs) exhibit obvious advantages compared to immortalized cell lines and primary cell cultures but implementation of iPSC-based high throughput (HT) assays is still technically challenging. Here we demonstrate the development and conduction of an organotypic HT Cl-/I- exchange assay using Cystic Fibrosis (CF) disease-specific iPSCs. The introduction of a halide sensitive YFP variant enabled automated quantitative measurement of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) function in iPSC-derived intestinal epithelia. CFTR function was partially rescued by treatment with VX-770 and VX-809, and seamless gene correction of the p.Phe508del mutation resulted in full restoration of CFTR function. The identification of a series of validated primary hits that improve the function of p.Phe508del CFTR from a library of ~ 42.500 chemical compounds demonstrates that the advantages of complex iPSC-derived culture systems for disease modelling can also be utilized for drug screening at a true HT format. For detailed analysis of the differentiated hiPSC cell populations on day 15 of differentiation 32 samples in total were analyzed. Three independent donor lines were utilized (donor 1 and 6 CFTR WT, donor 2 (p.Phe508del)) and one isogenic gene corrected control line (donor 2 gene corrected-CFTR WT.) Samples from undifferentiated and differentiated cells represent biological replicates (n=3). As controls RNA from adult intestine, liver and colon was. Tissue samples are represented as technical replicates.