Carlo Vancheri

Idiopathic pulmonary fibrosis: a disease with similarities and links to cancer biology

Several clinical trials have recently targeted specific pathways implicated in the pathogenesis of idiopathic pulmonary fibrosis (IPF). However, IPF remains plagued by a median survival of 3 yrs and emphasises the need for further research with new insights and perspectives. The prevailing pathogenic hypotheses assume that either an inflammatory process or an independent epithelial/fibroblastic disorder may propagate the disease process. Based on knowledge developed with considerable scientific evidence, we provide our perspectives with an alternative point of view that IPF be considered as a neoproliferative disorder of the lung. Genetic alterations, response to growth and inhibitory signals, resistance to apoptosis, myofibroblast origin and behaviour, altered cellular communications, and intracellular signalling pathways are all fundamental pathogenic hallmarks of both IPF and cancer. The concept of IPF as a lethal malignant disorder of the lung might extend beyond the pathogenic link between these two diseases and disclose new pathogenic mechanisms leading to novel therapeutic options, adopted from cancer biology. Moreover, this vision might dawn the awareness of the public, political and scientific community of this devastating disease from an angle different from the current perception and provoke new ideas and studies to get a better understanding to control the otherwise relentless progressive disease.

TGF-β1 targets the GSK-3β/β-catenin pathway via ERK activation in the transition of human lung fibroblasts into myofibroblasts

Transforming growth factor-beta1 (TGF-beta1) is known to induce the transition of human lung fibroblasts to myofibroblasts, a primary event in the pathogenesis of idiopathic pulmonary fibrosis. The molecular pathways involved in myofibroblast transformation are only partially identified. We found that a 24-h treatment with TGF-beta1 (10 ng/ml) induced alpha-smooth actin (SMA) expression and collagen production in human lung fibroblasts. These effects were abrogated by PD98059, a specific inhibitor of the mitogen-activated protein kinase (MAPK) pathway. TGF-beta1 treatment activated the MAPK pathway, as shown by an increased phosphorylation of extracellular-regulated kinases (ERK)1/2 after 30 min of exposure. TGF-beta1 also increased the expression of the Ser-9-phosphorylated inactive form of glycogen synthase kinase-3beta (GSK-3beta), an effect that was largely attenuated by PD98059. A nuclear translocation of beta-catenin in human lung fibroblasts was observed 2h after TGF-beta1 addition both by confocal microscopy and nuclear protein analysis. At this time, TGF-beta1 also increased the total levels of beta-catenin, an effect that was prevented by PD98059. Similarly to TGF-beta1, the GSK-3beta inhibitor lithium chloride (10mM), increased the total levels of beta-catenin and promoted alpha-SMA expression and collagen production. This study demonstrates that TGF-beta1 induces alpha-SMA expression and collagen production in human lung fibroblasts via ERK1/2 activation, GSK-3beta inhibition and nuclear beta-catenin translocation. The evidence that the silencing of beta-catenin by siRNAs was able to prevent the induction of alpha-SMA expression in TGF-beta1-treated fibroblasts further supports the hypothesis of a contribution of the GSK-3beta/beta-catenin pathway in the pathogenesis of idiopathic pulmonary fibrosis.

Effect of rosiglitazone and 15-deoxy-Δ12,14-prostaglandin J2 on bleomycin-induced lung injury

Thiazolidinedione rosiglitazone and 15-deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2), are two peroxisome proliferator-activated receptor (PPAR)-γ ligands. The aim of this study was to investigate the effect of rosiglitazone and 15d-PGJ2 on the lung injury caused by bleomycin administration. Mice subjected to intratracheal administration of bleomycin developed significant lung injury. An increase in immunoreactivity to nitrotyrosine, poly(ADP ribose) polymerase (PARP) and inducible nitric oxide synthase as well as a significant loss of body weight and mortality was observed in the lung of bleomycin-treated mice. Administration of the two PPAR-γ agonists rosiglitazone (10 mg·kg−1 i.p.) and 15d-PGJ2 (30 µg·kg−1 i.p.) significantly reduced the: 1) loss of body weight, 2) mortality rate, 3) infiltration of the lung with polymorphonuclear neutrophils (myeloperoxidase activity), 4) oedema formation, and 5) histological evidence of lung injury. Administration of rosiglitazone and 15d-PGJ2 also markedly reduced the nitrotyrosine, PARP and inducible nitric oxide synthase formation. In addition, treatment with the PPAR-γ antagonist bisphenol A diglycidyl ether (1 mg·kg−1 i.p. 30 min before the rosiglitazone or 15d-PGJ2) significantly antagonised the effect of the two PPAR-γ agonists. These results demonstrate that the two peroxisome proliferator-activated receptor-γ agonists, rosiglitazone and 15-deoxy-Δ12,14-prostaglandin J2, significantly reduce lung injury induced by bleomycin in mice.

Effect of pirfenidone on proliferation, TGF-β-induced myofibroblast differentiation and fibrogenic activity of primary human lung fibroblasts

Pirfenidone is an orally active small molecule that has been shown to inhibit the progression of fibrosis in animal models and in patients with idiopathic pulmonary fibrosis. Although pirfenidone exhibits well documented antifibrotic and antiinflammatory activities, in vitro and in vivo, its molecular targets and mechanisms of action have not been elucidated. In this study, we investigated the effects of pirfenidone on proliferation, TGF-β-induced differentiation and fibrogenic activity of primary human lung fibroblasts (HLFs). Pirfenidone reduced fibroblast proliferation and attenuated TGF-β-induced α-smooth muscle actin (SMA) and pro-collagen (Col)-I mRNA and protein levels. Importantly, pirfenidone inhibited TGF-β-induced phosphorylation of Smad3, p38, and Akt, key factors in the TGF-β pathway. Together, these results demonstrate that pirfenidone modulates HLF proliferation and TGF-β-mediated differentiation into myofibroblasts by attenuating key TGF-β-induced signaling pathways.

Reactive oxygen species are required for maintenance and differentiation of primary lung fibroblasts in idiopathic pulmonary fibrosis.

Background Idiopathic pulmonary fibrosis (IPF) is a progressive and fatal illness whose pathogenesis remains poorly understood. Recent evidence suggests oxidative stress as a key player in the establishment/progression of lung fibrosis in animal models and possibly in human IPF. The aim of the present study was to characterize the cellular phenotype of fibroblasts derived from IPF patients and identify underlying molecular mechanisms. Methodology/Principal Findings We first analyzed the baseline differentiation features and growth ability of primary lung fibroblasts derived from 7 histology proven IPF patients and 4 control subjects at different culture passages. Then, we focused on the redox state and related molecular pathways of IPF fibroblasts and investigated the impact of oxidative stress in the establishment of the IPF phenotype. IPF fibroblasts were differentiated into alpha-smooth muscle actin (SMA)-positive myofibroblasts, displayed a pro-fibrotic phenotype as expressing type-I collagen, and proliferated lower than controls cells. The IPF phenotype was inducible upon oxidative stress in control cells and was sensitive to ROS scavenging. IPF fibroblasts also contained large excess of reactive oxygen species (ROS) due to the activation of an NADPH oxidase-like system, displayed higher levels of tyrosine phosphorylated proteins and were more resistant to oxidative-stress induced cell death. Interestingly, the IPF traits disappeared with time in culture, indicating a transient effect of the initial trigger. Conclusions/Significance Robust expression of α-SMA and type-I collagen, high and uniformly-distributed ROS levels, resistance to oxidative-stress induced cell death and constitutive activation of tyrosine kinase(s) signalling are distinctive features of the IPF phenotype. We suggest that this phenotype can be used as a model to identify the initial trigger of IPF.

Combination therapy: the future of management for idiopathic pulmonary fibrosis?

Findings from recently published placebo-controlled trials in idiopathic pulmonary fibrosis have established that pirfenidone and nintedanib prevent about 50% of the decline in forced vital capacity typically seen in this disease; future trials are therefore unlikely to use placebo as a control group for ethical reasons. Future clinical assessment will probably include add-on trials in which a new drug is combined with an intervention with established efficacy; this development is in turn likely to herald the use of combination regimens in clinical practice. Personalised medicine (the selection of monotherapies on the basis of individualised biomarker signal) is an intrinsically attractive alternative approach, but is unlikely to be useful in routine management of idiopathic pulmonary fibrosis in the medium-term future because of the complex nature of the diseases pathogenesis. In this Personal View, we review the pleiotropic nature of disease pathogenesis in idiopathic pulmonary disease, the use of combination regimens in other selected chronic lung diseases, and the conceptual basis for combination therapies in interstitial lung disorders other than idiopathic pulmonary fibrosis. On the basis of these considerations, and the emergence of data from add-on trials, we believe that the future of management for idiopathic pulmonary fibrosis lies in the development of combination regimens.

Inhibition of PI3K Prevents the Proliferation and Differentiation of Human Lung Fibroblasts into Myofibroblasts: The Role of Class I P110 Isoforms

Idiopathic pulmonary fibrosis (IPF) is a progressive fibroproliferative disease characterized by an accumulation of fibroblasts and myofibroblasts in the alveolar wall. Even though the pathogenesis of this fatal disorder remains unclear, transforming growth factor-β (TGF-β)-induced differentiation and proliferation of myofibroblasts is recognized as a primary event. The molecular pathways involved in TGF-β signalling are generally Smad-dependent yet Smad-independent pathways, including phosphatidylinositol-3-kinase/protein kinase B (PI3K/Akt), have been recently proposed. In this research we established ex-vivo cultures of human lung fibroblasts and we investigated the role of the PI3K/Akt pathway in two critical stages of the fibrotic process induced by TGF-β: fibroblast proliferation and differentiation into myofibroblasts. Here we show that the pan-inhibitor of PI3Ks LY294002 is able to abrogate the TGF-β-induced increase in cell proliferation, in α- smooth muscle actin expression and in collagen production besides inhibiting Akt phosphorylation, thus demonstrating the centrality of the PI3K/Akt pathway in lung fibroblast proliferation and differentiation. Moreover, for the first time we show that PI3K p110δ and p110γ are functionally expressed in human lung fibroblasts, in addition to the ubiquitously expressed p110α and β. Finally, results obtained with both selective inhibitors and gene knocking-down experiments demonstrate a major role of p110γ and p110α in both TGF-β-induced fibroblast proliferation and differentiation. This finding suggests that specific PI3K isoforms can be pharmacological targets in IPF.

Effect of immunotherapy on asthma progression, BHR and sputum eosinophils in allergic rhinitis

Background:  Bronchial hyperresponsiveness (BHR) and airway inflammation are frequently associated with allergic rhinitis, and may be important risk factors for the development of asthma. Specific immunotherapy (SIT) reduces symptom in subjects with allergic rhinitis, but the mechanisms are not clear.

Pirfenidone in idiopathic pulmonary fibrosis: expert panel discussion on the management of drug-related adverse events.

Pirfenidone is currently the only approved therapy for idiopathic pulmonary fibrosis, following studies demonstrating that treatment reduces the decline in lung function and improves progression-free survival. Although generally well tolerated, a minority of patients discontinue therapy due to gastrointestinal and skin-related adverse events (AEs). This review summarizes recommendations based on existing guidelines, research evidence, and consensus opinions of expert authors, with the aim of providing practicing physicians with the specific clinical information needed to educate the patient and better manage pirfenidone-related AEs with continued pirfenidone treatment. The main recommendations to help prevent and/or mitigate gastrointestinal and skin-related AEs include taking pirfenidone during (or after) a meal, avoiding sun exposure, wearing protective clothing, and applying a broad-spectrum sunscreen with high ultraviolet (UV) A and UVB protection. These measures can help optimize AE management, which is key to maintaining patients on an optimal treatment dose.

Common pathways in idiopathic pulmonary fibrosis and cancer

Idiopathic pulmonary fibrosis (IPF) is marked by a very disappointing survival rate and still represents a clinical dilemma. According to the current pathogenic hypothesis, chronic damage of the alveolar epithelium is followed by abnormal tissue repair and impairment of the alveolar structure. This process is driven by pathogenic events very similar to cancer, including epigenetic and genetic changes, altered response to regulatory signals, abnormal expression of microRNAs and activation of specific signalling pathways. IPF also resembles cancer with regard to its poor response to medical treatment and prognosis, which is very often worse than many cancers. We have hypothesised that IPF might be assimilated to a neoproliferative disorder of the lung. Viewing IPF as a cancer-like disease may satisfy the need for a better understanding of the pathogenesis of IPF by exploiting the large amount of knowledge that cancer biology evokes. The recognition of common pathogenic pathways between the two diseases may stimulate new clinical trials with cancer drugs, different drug combinations and different lines of drugs, as already experimented in oncology. Moreover, the concept of IPF as a cancer-like disorder may improve the attention given to this dreadful disease on a public, political and healthcare level.