Pirfenidone anti-fibrotic effects are partially mediated by the inhibition of MUC1 bioactivation.

Pirfenidone is a pleiotropic molecule approved to treat idiopathic pulmonary fibrosis (IPF). Pirfenidone has demonstrated to downregulate transforming growth factor-β1 (TGF-β1) cellular effects. However, its anti-fibrotic mechanism remains unclear. Here, we aim to analyze the effects of pirfenidone on the TGF-β1 canonical and non-canonical pathways, as well as, on the most characteristic IPF cellular processes. Results observed in this work showed that TGF-β1-induced canonical SMAD3 and non-canonical ERK1/2 phosphorylations were not inhibited by pirfenidone in alveolar A549 and lung fibroblasts MRC5 cells. In contrast, pirfenidone inhibited TGF-β1-induced MUC1-CT Thr41 (1224) and Tyr46 (1229) phosphorylations, thus reducing the β-catenin activation. Additionally, immunoprecipitation and immunofluorescence studies in ATII cells and lung fibroblasts showed that pirfenidone inhibited the formation and nuclear translocation of the transcriptional fibrotic TGF-β1-induced phospho-SMAD3/MUC1-CT/active-β-catenin complex, and consequently the SMAD-binding element activation (SBE). This study provided also evidence of the inhibitory effect of pirfenidone on the TGF-β1-induced ATII to mesenchymal and fibroblast to myofibroblast transitions, fibroblast proliferation and ATII and fibroblast senescence. Therefore, it indicates that pirfenidone's inhibitory effect on TGF-β1-induced fibrotic cellular processes is mediated by the inhibition of MUC1-CT phosphorylation, β-catenin activation, nuclear complex formation of phospho-SMAD3/MUC1-CT/active β-catenin and SBE activation, which may be of value to further develop anti-fibrotic IPF therapies.