Mutational activation of BRAF confers sensitivity to transforming growth factor beta inhibitors in human cancer cells

Recent data implicate elevated transforming growth factor-β (TGFβ) signalling in BRAF inhibitor drug-resistance mechanisms, but the potential for targeting TGFβ signalling in cases of advanced melanoma has not been investigated. We show that mutant BRAFV600E confers an intrinsic dependence on TGFβ/TGFβ receptor 1 (TGFBR1) signalling for clonogenicity of murine melanocytes. Pharmacological inhibition of the TGFBR1 blocked the clonogenicity of human mutant BRAF melanoma cells through SMAD4-independent inhibition of mitosis, and also inhibited metastasis in xenografted zebrafish. When investigating the therapeutic potential of combining inhibitors of mutant BRAF and TGFBR1, we noted that unexpectedly, low-dose PLX-4720 (a vemurafenib analogue) promoted proliferation of drug-naïve melanoma cells. Pharmacological or pharmacogenetic inhibition of TGFBR1 blocked growth promotion and phosphorylation of SRC, which is frequently associated with vemurafenib-resistance mechanisms. Importantly, vemurafenib-resistant patient derived cells retained sensitivity to TGFBR1 inhibition, suggesting that TGFBR1 could be targeted therapeutically to combat the development of vemurafenib drug-resistance

Inflammatory and mucociliary dysfunction based endotypes across the spectrum of chronic airways diseases

Introduction: There is substantial overlap between COPD, asthma, bronchiectasis(BE) and cystic fibrosis(CF) and each is characterised by inflammation and mucociliary dysfunction.Hypothesis: Biology, rather than disease labels, may stratify patients into therapeutically relevant subtypes.Methods: Patients were categorized by primary disease and clinical characteristics, spontaneous sputum was collected and inflammatory characteristics (neutrophil elastase(NE) and 19 cytokines) and sputum properties (DNA content, mucins, rheology, dry weight) measured. K-means clustering was performed and parameters compared between and within disease groups. Smokers without respiratory disease were used as controls.Results: The study included patients with asthma(76), COPD(91), BE(54), CF(24) and controls(26). 10 cytokines, NE, dry weight, mucins and multiple sputum parameters were different between disease groups and healthy controls (p<0.05). K means clustering identified 2 clusters defined by neutrophilic (N) or eosinophilic (E) inflammation. The E cluster was associated with lower dry weight, DNA content and higher mucins, particularly MUC5B. The rheology parameters G’ and G* were significantly higher in the E group while Tan(delta) was higher in the N group (p<0.05 all comparisons). Both clusters were present in all disease groups with more neutrophilic inflammation in CF and BE (42% of COPD patients and 46% of asthma patients were neutrophilic vs 78% of BE and 87% of CF,p<0.0001).Conclusion: Airways diseases have heterogenous inflammatory and mucus parameters. Assessment based on disease labels may be aided by endotyping using inflammatory and mucociliary clearance parameters

Inflammatory and mucociliary dysfunction based endotypes across the spectrum of chronic airways diseases

Introduction: There is substantial overlap between COPD, asthma, bronchiectasis(BE) and cystic fibrosis(CF) and each is characterised by inflammation and mucociliary dysfunction.Hypothesis: Biology, rather than disease labels, may stratify patients into therapeutically relevant subtypes.Methods: Patients were categorized by primary disease and clinical characteristics, spontaneous sputum was collected and inflammatory characteristics (neutrophil elastase(NE) and 19 cytokines) and sputum properties (DNA content, mucins, rheology, dry weight) measured. K-means clustering was performed and parameters compared between and within disease groups. Smokers without respiratory disease were used as controls.Results: The study included patients with asthma(76), COPD(91), BE(54), CF(24) and controls(26). 10 cytokines, NE, dry weight, mucins and multiple sputum parameters were different between disease groups and healthy controls (p<0.05). K means clustering identified 2 clusters defined by neutrophilic (N) or eosinophilic (E) inflammation. The E cluster was associated with lower dry weight, DNA content and higher mucins, particularly MUC5B. The rheology parameters G’ and G* were significantly higher in the E group while Tan(delta) was higher in the N group (p<0.05 all comparisons). Both clusters were present in all disease groups with more neutrophilic inflammation in CF and BE (42% of COPD patients and 46% of asthma patients were neutrophilic vs 78% of BE and 87% of CF,p<0.0001).Conclusion: Airways diseases have heterogenous inflammatory and mucus parameters. Assessment based on disease labels may be aided by endotyping using inflammatory and mucociliary clearance parameters

Airway IL-1β is related to disease severity and mucociliary function in bronchiectasis

Rationale The inflammasome is a key regulatory complex of the inflammatory response leading to interleukin-1β (IL-1β) release and activation. IL-1β amplifies inflammatory responses and induces mucus secretion and hyperconcentration in other diseases. The role of IL-1β in bronchiectasis has not been investigated. Objectives To characterise the role of airway IL-1β in bronchiectasis, including the association with mucus properties, ciliary function, airway inflammation, microbiome and disease severity. Methods Stable bronchiectasis patients were enrolled in an international cohort study (n=269). IL-1β was measured in sputum supernatant. A validation cohort also had sputum rheology and hydration measured (n=53). For analysis, patients were stratified according to the median value of IL-1β in the population (high versus low) to compare disease severity, airway infection, microbiome (16S rRNA sequencing), inflammation and caspase-1 activity. Primary human nasal epithelial cells grown in air–liquid interface culture were used to study the effect of IL-1β on cilia function. Results Patients with high sputum IL-1β had more severe disease, increased caspase-1 activity and an increased T-helper type 1, T-helper type 2 and neutrophil inflammatory response compared with patients with low IL-1β. The active-dominant form of IL-1β was associated with increased disease severity. High IL-1β was related to higher relative abundance of Proteobacteria in the microbiome and increased mucus solid content and viscoelastic properties. Chronic IL-1β treatment reduced the functionality of cilia and tight junctions of epithelial cells in vitro. Conclusions A subset of stable bronchiectasis patients show increased airway IL-1β, suggesting pulmonary inflammasome activation is linked with more severe disease, airway infection, mucus dehydration and epithelial dysfunction.</p