Investigating the STING signalling pathway as an alternative therapeutic strategy in Cystic Fibrosis

Introduction Cystic Fibrosis (CF) is a life-shortening rare disease mainly affecting the lungs. The principal cause of morbidity and mortality is the airways obstruction by viscous mucus and inflammation with recurrent bacterial infections and colonization. CF is caused by more than 2000 mutations in the gene coding for the CF Transmembrane Conductance Regulator (CFTR), a chloride channel at the apical membrane of epithelial cells. The vast majority (70-90%) of CF patients bear the most common class II F508del mutation, where F508delCFTR is retained in the endoplasmic reticulum (ER) and degraded before it reaches the plasma membrane. Even if a promising therapeutic proposal (Trikafta®, Vertex) is available for the F508del mutation, aimed at directly target the CFTR, there are many CF mutations that Vertex compounds are not approved for. Alternative approaches aimed at targeting the cellular environment perturbed by the lack of a functional CFTR recently emerged. In particular, tissue transglutaminase (TG2) upregulation is due to the presence of reactive oxygen species (ROS), produced by both F508delCFTR degradation and ER stress. TG2 depletes the essential autophagy-related protein Beclin1, leading to secondary accumulation of the autophagic substrate SQSTM1/p62 which, in turn, increases CFTR degradation, through the phosphorylation of p62 S403 by the protein kinase TBK1. Aim of the study Our experimental approach aims to study the oxidative and autophagy imbalance in CF models from a biochemical and pharmacological point of view. The advantage of such an approach is to pave the way for a therapeutic proposal which, not aiming at directly targeting the channel, would be independent of a specific mutation on the CFTR. In particular, we investigate the cGAS-cGAMP-STING pathway, which recruits and activates TBK1, as a potential target for the development of novel treatments in CF. Methods Techniques for the management and maintenance of specific cell cultures, for the extraction, quantification and identification of proteins, as well as spectroscopic and fluorescence techniques were used during the experimental approach. Results The experimental study allowed the isolation of a series of compounds with the ability to recover a mature F508delCFTR in the plasma membrane. Moreover, we investigated the mechanism of action of these compounds, identifying their ability both to influence the autophagy mechanism through the modulation of the STING pathway, and to partially invert the redox imbalance, acting on the levels of the Nrf2 protein. Conclusions The project granted some preliminary data to support the initial hypothesis; these results require further confirmation in particular through functional assays of F508delCFTR channel. Subsequent studies of great interest will be planned with particular reference to investigate our active molecules in models bearing other CFTR mutations, and to combine them with drugs currently approved for CF.