Modelling gene therapy efficiency in mosaic airway organoids for treatment of cystic fibrosis

Abstract

Worldwide over 100,000 people suffer from cystic fibrosis (CF), a recessive genetic disease caused by functional defects of the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) protein. This disease is characterized by damaging symptoms in multiple organs with respiratory failure as the cause of death for most patients. Gene replacement therapy allows for the introduction of a functional copy of CFTR into the genome and is therefore a very promising treatment option. Due to delivery limitations of this treatment option to reach all targeted cells, its use has yet to be translated to in vivo. Fortunately, from previous research it is clear that a 100% replacement efficiency is seemingly not necessary for beneficial effects. However, a clear, exact consensus on what efficiency is required is still missing. Therefore, the aim of this research is to determine the number of CFTR-deficient cells required to be effectively treated with gene replacement therapy for total functional recovery of CFTR to relieve or cure patients from their symptoms. For this purpose, this report shows the establishment of a mosaic airway organoid model, modelling gene replacement efficiency by combining differently labelled (green and red) cells representing CFTR-deficient and -proficient cells. With the use of forskolin-induced swelling of these organoids as an indicator of presence of functional CFTR, in relation to counted cell numbers with computational segmentation, an estimation of the required number of treated cells can be made. With further optimizations and expansions, the organoid model representing gene therapy efficiency can be finalized and used for the estimation of efficiency needed for restoring CFTR function, moving closer to effective and life-changing gene therapy for patients.