Mapping the Impact of ETI-mediated CFTR Reactivation On Development: a zebrafish model study

Abstract

CFTR therapy with elexacaftor/tezacaftor/ivacaftor (ETI) has transformed the life of individuals with cystic fibrosis (CF), also enhancing women's fertility. While adverse effects regarding ETI during pregnancy have not been reported, the evidence remains limited. However, it has been demonstrated that ETI can cross the placenta, therefore reaching the foetus due to placental transfer processes. The hyperactivation of CFTR through ETI administration during pregnancy could have an impact on foetus development, potentially leading to significant undesired effects. Indeed, all the functional studies conducted so far have focused on analysing the effects of CFTR loss of function/mutation, while the hyperactivation of CFTR has never been considered. In this study, we compared the effects of CFTR loss-of-function and its overexpression on zebrafish development. This model is particularly suitable for developmental studies and offers the unique opportunity to be “humanized” by injecting human genes. We compared cftr loss-of-function zebrafish embryos, which showed bilateral asymmetry of internal organs, increased basal inflammation levels, and delayed innate inflammatory response, with CFTR overexpressed embryos (whether in the human wild-type form or the F508del mutant form), which showed significant defects along the anteriorposterior body axis. This phenotype is reminiscent of defects in the WNT-beta signalling regulation, a pathway crucial for early developmental differentiation, such as mesoderm specification. Indeed, CFTR protein directly interacts with Dishevelled, a WNT-positive regulator, preventing it from lysosomal degradation. Importantly, this pathway can be pharmacologically modulated, opening the possibility that the undesired effects of ETI-mediated CFTR hyperactivation during development can be mitigated or avoided. In addition, since ETI has been shown to accumulate in various foetal tissues, including the developing brain, we are testing possible off-targets effects of ETI using molecular and behavioural assays during zebrafish development. To improve the translational potential of our findings and validate the results obtained in zebrafish, we will overexpress wild-type and F508del CFTRs in human iPS and we will treat them with ETI. This project aims to shed light on CFTR role during the first stages of life and could be essential to ensure that pregnant women with CF can safely undergo therapy with ETI