Cystic Fibrosis (CF) is the most prevalent lethal genetic disease in Caucasians caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. The triple combination of modulators (TRIKAFTA) was approved for ~80% of CF patients. Still, many patients carrying rare mutations are not eligible to this treatment, due to limited accessibility to patients’ samples and lack of clinical trials. The use of ex-vivo models, such as rectal organoids, enables testing of CFTR modulators in these individuals, predicting the possibility of a clinical benefit. Spatial CFTR structures, modeling the binding of the different TRIKAFTA components, shed light on the functional effects of the modulators on the mutated CFTR. These structural predictions enable the identification of candidate rare mutations that may benefit from the treatment. Our current aim is to use this new data to prioritize the assessment of the response of patients carrying rare mutations to TRIKAFTA and further shed light on the basis of the CFTR response to various modulators.

Cystic Fibrosis (CF) is the most prevalent lethal genetic disease in Caucasians caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. We are studying different molecular mechanisms acting as modifiers of disease severity and response to therapy.

Our goal is to develop personalized medicine for CF patients carrying rare CFTR mutations including splicing and nonsense mutations (see below). For this, we are collecting patient-derived respiratory and intestinal epithelial cells that recapitulate essential features of the in vivo epithelium and exhibit CFTR-dependent functional readout. These patient-derived cellular models serve as a pre-clinical tool in the development of personalized therapies.

A significant fraction of CF-causing mutations (10-15%) affects pre-mRNA splicing, leading to the generation of aberrantly spliced CFTR transcripts. We are now developing a targeted modulation approach aimed to correct the splicing pattern of CFTR transcripts, according to the specific mutation. This strategy, which is based on splice-switching Antisense Oligonucleotides (ASO) is aimed to inhibit or activate specific splicing events by a steric blockade of the recognition of specific splicing elements. Together with the drug development company SpliSense, we have demonstrated the potential of this strategy in human respiratory cells (HNEs, HBEs) derived from CF patients carrying the 3849+10kb C-to-T splicing mutation. A highly potent ASO was able to significantly increase the level of correctly spliced CFTR mRNA and restore CFTR function (Oren et al. Journal of Cystic Fibrosis 2021). We are currently expanding our personalized medicine research, implementing the ASO approach in patient-derived intestinal organoids which serve as biomarkers predicting clinical benefit from CFTR modulator therapies.

10-15% of CF-causing mutations are nonsense mutations which lead to premature termination codon (PTC). Transcripts carrying PTCs undergo degradation by the Nonsense Mediated mRNA Decay (NMD) pathway. Various small molecules, which can read-through PTCs, permit translation of full-length proteins. However, this approach is dependent on NMD efficiency and thus may limit the response of patients to the treatment. Using various cellular models, among them patient-derived epithelial cells, we are investigating a novel therapeutic strategy aimed to stabilize NMD-prone CFTR transcripts by epigenetic modulation of m6A RNA methylation (in collaboration with Prof. Rotem Karni).

Methylation of Adenine on position 6 in RNA molecules (m6A) plays a key role in RNA biogenesis. M6A marks the translation termination codon and helps the NMD machinery to distinguish it from premature termination codons (PTCs). It was recently discovered that m6A modulation can inhibit NMD and stabilize unstable mRNA molecules carrying PTCs.