Decreased serpin C1 in extracellular vesicles predicts response to methotrexate treatment in patients with pulmonary sarcoidosis.

BACKGROUND: Sarcoidosis is a systemic granulomatous disease of unknown etiology primarily affecting the lungs. Treatment is needed when disease symptoms worsen and organ function deteriorates. In pulmonary sarcoidosis, prednisone and methotrexate (MTX) are the most common anti-inflammatory therapies. However, there is large inter-patient variability in response to treatment, and predictive response markers are currently lacking. OBJECTIVE: In this study, we investigated the predictive potential of biomarkers in extracellular vesicles (EVs) isolated from biobanked serum of patients with pulmonary sarcoidosis stored prior to start of therapy. METHODS: Protein concentrations of a four-protein test panel of inflammatory proteins were measured in a discovery (n = 16) and replication (n = 129) cohort of patients with sarcoidosis and 47 healthy controls. Response to therapy was defined as an improvement of the absolute score of > 5% forced vital capacity (FVC) and/or > 10% diffusion lung of carbon monoxide (DLCO) after 24 weeks compared to baseline (before treatment). RESULTS: Serum protein levels differed between EV fractions and serum, and between sarcoidosis cases and controls. Serpin C1 concentrations in the low density lipid particle EV fraction were lower at baseline in the group of patients with a good response to MTX treatment in both the discovery cohort (p = 0.059) and in the replication cohort (p = 0.032). EV Serpin C1 showed to be a significant predictor for response to treatment with MTX (OR 0.4; p = 0.032). CONCLUSION: This study shows that proteins isolated from EVs harbor a distinct signal and have potential as new predictive therapy response biomarkers in sarcoidosis.

A new variant in the ZCCHC8 gene: diverse clinical phenotypes and expression in the lung.

Introduction: Pulmonary fibrosis is a severe disease which can be familial. A genetic cause can only be found in ∼40% of families. Searching for shared novel genetic variants may aid the discovery of new genetic causes of disease. Methods: Whole-exome sequencing was performed in 152 unrelated patients with a suspected genetic cause of pulmonary fibrosis from the St Antonius interstitial lung disease biobank. Variants of interest were selected by filtering for novel, potentially deleterious variants that were present in at least three unrelated pulmonary fibrosis patients. Results: The novel c.586G>A p.(E196K) variant in the ZCCHC8 gene was observed in three unrelated patients: two familial patients and one sporadic patient, who was later genealogically linked to one of the families. The variant was identified in nine additional relatives with pulmonary fibrosis and other telomere-related phenotypes, such as pulmonary arterial venous malformations, emphysema, myelodysplastic syndrome, acute myeloid leukaemia and dyskeratosis congenita. One family showed incomplete segregation, with absence of the variant in one pulmonary fibrosis patient who carried a PARN variant. The majority of ZCCHC8 variant carriers showed short telomeres in blood. ZCCHC8 protein was located in different lung cell types, including alveolar type 2 (AT2) pneumocytes, the culprit cells in pulmonary fibrosis. AT2 cells showed telomere shortening and increased DNA damage, which was comparable to patients with sporadic pulmonary fibrosis and those with pulmonary fibrosis carrying a telomere-related gene variant, respectively. Discussion: The ZCCHC8 c.586G>A variant confirms the involvement of ZCCHC8 in pulmonary fibrosis and short-telomere syndromes and underlines the importance of including the ZCCHC8 gene in diagnostic gene panels for these diseases.

New Insights via RNA Profiling of Formalin-Fixed Paraffin-Embedded Lung Tissue of Pulmonary Fibrosis Patients.

In sporadic idiopathic pulmonary fibrosis (sIPF) and pulmonary fibrosis caused by a mutation in telomere (TRG-PF) or surfactant related genes (SRG-PF), there are a number of aberrant cellular processes known that can lead to fibrogenesis. We investigated whether RNA expression of genes involved in these processes differed between sIPF, TRG-PF, and SRG-PF and whether expression levels were associated with survival. RNA expression of 28 genes was measured in lung biopsies of 26 sIPF, 17 TRG-PF, and 6 SRG-PF patients. Significant differences in RNA expression of TGFBR2 (p = 0.02) and SFTPA2 (p = 0.02) were found between sIPF, TRG-PF, and SRG-PF. Patients with low (

Simultaneous Assessment of mTORC1, JAK/STAT, and NLRP3 Inflammasome Activation Pathways in Patients with Sarcoidosis.

The unknown etiology of sarcoidosis, along with the variability in organ involvement and disease course, complicates the effective treatment of this disease. Based on recent studies, the cellular inflammatory pathways involved in granuloma formation are of interest regarding possible new treatment options, such as the mechanistic (formerly mammalian) target of rapamycin complex 1 (mTORC1) pathway, the Janus kinase/signal transducers and activators of transcription (JAK/STAT) pathway, and the nucleotide-binding domain, leucine-rich-containing family, pyrin domain-containing-3 (NLRP3) inflammasome pathway. The aim of this study was to explore the potential coexpression of these three inflammatory pathways in patients with sarcoidosis and see whether possible differences were related to disease outcome. The tissue of 60 patients with sarcoidosis was used to determine the activity of these three signaling pathways using immunohistochemistry. The activation of NLRP3 was present in 85% of all patients, and the activation of mTORC1 and JAK/STAT was present in 49% and 50% of patients, respectively. Furthermore, the presence of NLRP3 activation at diagnosis was associated with a chronic disease course of sarcoidosis. Our finding of different new conceptual inflammatory tissue phenotypes in sarcoidosis could possibly guide future treatment studies using the available inhibitors of either NLRP3, JAK-STAT, and mTORC1 inhibitors in a more personalized medicine approach.

Improved Survival of IPF patients Treated With Antifibrotic Drugs Compared With Untreated Patients.

PURPOSE: Pirfenidone and nintedanib unequivocally inhibit FVC decline, but have been inconsistently linked to reduced mortality in phase III studies. On the contrary, real-world data show a survival benefit of antifibrotic drugs. However, it is unknown what this benefit is across different Gender, Age, and Physiology (GAP) stages. RESEARCH QUESTIONS: Is there a difference in transplant-free (TPF) survival of IPF patients receiving antifibrotic drugs (IPFAF) compared with an untreated cohort (IPFnon-AF)? Is this different for patients with GAP stage I, II, or III. METHODS: This is a single-center observational cohort study using prospectively included patients diagnosed with IPF between 2008-2018. Primary outcomes were TPF survival difference and 1-, 2-, and 3-year cumulative mortality for IPFAF and IPFnon-AF. This was repeated after stratification for GAP stage. RESULTS: In total, 457 patients were included. The median transplant-free survival was 3.4 years in IPFAF (n = 313) and 2.2 years in IPFnon-AF (n = 144, p = 0.005). For GAP stage II, a median survival of 3.1 and 1.7 years was noted for IPFAF (n = 143) and IPFnon-AF (n = 59, p < 0.001), respectively. A significantly lower 1-, 2-, and 3- year cumulative mortality was found for IPFAF with GAP stage II (1 yr: 7.0% vs 35.6%, 2 yr: 26.6% vs 55.9%, and 3 yr: 46.9% vs 69.5%). The 1-year cumulative mortality of IPFAF with GAP III was also significantly lower (19.0% vs 65.0%). CONCLUSION: This large real-world study showed a survival benefit in IPFAF compared with IPFnon-AF. This especially holds true for patients with GAP stage II and III.

Delineating associations of progressive pleuroparenchymal fibroelastosis in patients with pulmonary fibrosis.

Background: Computer quantification of baseline computed tomography (CT) radiological pleuroparenchymal fibroelastosis (PPFE) associates with mortality in idiopathic pulmonary fibrosis (IPF). We examined mortality associations of longitudinal change in computer-quantified PPFE-like lesions in IPF and fibrotic hypersensitivity pneumonitis (FHP). Methods: Two CT scans 6-36 months apart were retrospectively examined in one IPF (n=414) and one FHP population (n=98). Annualised change in computerised upper-zone pleural surface area comprising radiological PPFE-like lesions (Δ-PPFE) was calculated. Δ-PPFE >1.25% defined progressive PPFE above scan noise. Mixed-effects models evaluated Δ-PPFE against change in visual CT interstitial lung disease (ILD) extent and annualised forced vital capacity (FVC) decline. Multivariable models were adjusted for age, sex, smoking history, baseline emphysema presence, antifibrotic use and diffusion capacity of the lung for carbon monoxide. Mortality analyses further adjusted for baseline presence of clinically important PPFE-like lesions and ILD change. Results: Δ-PPFE associated weakly with ILD and FVC change. 22-26% of IPF and FHP cohorts demonstrated progressive PPFE-like lesions which independently associated with mortality in the IPF cohort (hazard ratio 1.25, 95% CI 1.16-1.34, p<0.0001) and the FHP cohort (hazard ratio 1.16, 95% CI 1.00-1.35, p=0.045). Interpretation: Progression of PPFE-like lesions independently associates with mortality in IPF and FHP but does not associate strongly with measures of fibrosis progression.

Decreased Survival and Lung Function in Progressive Pulmonary Fibrosis.

Background and Objectives: Progressive pulmonary fibrosis (PPF) is a recently described term reserved for patients with fibrotic ILD other than idiopathic pulmonary fibrosis (IPF) with fast clinical deterioration. Here, survival and prognostic biomarkers at the time of diagnosis for PPF are investigated in a fibrotic ILD other than IPF cohort (non-IPF). Materials and Methods: Patients diagnosed during the period of 2012-2018 at the ILD Center of Excellence (St. Antonius Hospital, Nieuwegein, The Netherlands) with a fibrotic ILD were included in this study. The presence of PPF was investigated using the criteria from the updated IPF/PPF guideline during the first year after diagnosis. Logistic regression analysis was used to determine risk factors for PPF. A Kaplan-Meier survival analysis with log-rank test was conducted to analyze survival in patients with and without PPF. Results: This study included 304 non-IPF patients and, for comparison, 379 IPF patients. In non-IPF patients, 146 (46%) fulfilled ≥2 criteria for PPF. These patients had a median transplant-free survival rate of 2.9 ± 0.4 years, which was worse than non-IPF patients without PPF (10.1 ± 1.8 years, p < 0.001). The risk for PPF was increased in patients with FVC < 50% (odds ratio (OR) of 2.50, 95% CI = 1.01-6.17, p = 0.047) or DLCOc ≤ 35% (OR = 2.57, 95% CI = 1.24-5.35, p = 0.011). In the first 3 years after diagnosis, survival in PPF and IPF is the same, while in the following years IPF has a significantly worse survival. Conclusions: The non-IPF cohort with PPF had a significantly worse transplant-free survival compared with the non-IPF cohort without PPF. Independent risk factors for PPF in non-IPF were FVC < 50% and DLCOc ≤ 35%.

MUC5B rs35705950 minor allele associates with older age and better survival in idiopathic pulmonary fibrosis.

BACKGROUND AND OBJECTIVE: The minor T-allele of the MUC5B promoter polymorphism rs35705950 is strongly associated with idiopathic pulmonary fibrosis (IPF). However, conflicting results have been reported on the relationship between the MUC5B minor allele and survival and it is unknown whether a specific subgroup of IPF patients might benefit from MUC5B minor allele carriage. We investigated the association between MUC5B rs35705950, survival and patient characteristics in a real-world population of European IPF patients. METHODS: In this retrospective study, 1751 patients with IPF from 8 European centres were included. MUC5B rs35705950 genotype, demographics, clinical characteristics at diagnosis and survival data were analysed. RESULTS: In a multi-variate Cox proportional hazard model the MUC5B minor allele was a significant independent predictor of survival when adjusted for age, sex, high resolution computed tomography pattern, smoking behaviour and pulmonary function tests in IPF. MUC5B minor allele carriers were significantly older at diagnosis (p = 0.001). The percentage of MUC5B minor allele carriers increased significantly with age from 44% in patients aged <56 year, to 63% in patients aged >75. In IPF patients aged <56, the MUC5B minor allele was not associated with survival. In IPF patients aged ≥56, survival was significantly better for MUC5B minor allele carriers (45 months [CI: 42-49]) compared to non-carriers (29 months [CI: 26-33]; p = 4 × 10-12 ). CONCLUSION: MUC5B minor allele carriage associates with a better median transplant-free survival of 16 months in the European IPF population aged over 56 years. MUC5B genotype status might aid disease prognostication in clinical management of IPF patients.

Progressive Disease With Low Survival in Adult Patients With Pulmonary Fibrosis Carrying Surfactant-Related Gene Mutations: An Observational Study.

BACKGROUND: In some patients with progressive fibrosing interstitial lung disease (ILD), disease is caused by carriage of a mutation in a surfactant-related gene (SRG) such as SFTPC, SFTPA2, or ABCA3. However, no aggregated data on disease evolution and treatment outcome have been presented for these patients. RESEARCH QUESTION: In adult patients with ILD with an SRG mutation, what is the course of lung function after diagnosis and during treatment and the survival in comparison with patients with sporadic idiopathic pulmonary fibrosis (sIPF) and familial pulmonary fibrosis (FPF)? STUDY DESIGN AND METHODS: We retrospectively examined the clinical course of a cohort of adults with an SRG mutation by screening 48 patients from 20 families with an SRG mutation for availability of clinical follow-up data. For comparison, 248 patients with FPF and 575 patients with sIPF were included. RESULTS: Twenty-three patients with ILD (median age: 45 years; 11 men) with an SRG mutation fulfilled criteria. At diagnosis, patients with an SRG mutation were younger and less often male, but had lower FVC (72% predicted) and diffusing capacity of the lungs for carbon monoxide (46% predicted) compared with patients with FPF or sIPF. In the SRG mutation group, median FVC decline 6 months after diagnosis was -40 mL and median transplant-free survival was 44 months and not different from patients with FPF or sIPF. FVC course was not different among the three cohorts; however, a significantly larger decrease in FVC was found while patients received immunomodulatory or antifibrotic treatment compared with those receiving no treatment. Subsequent analysis in the SRG group showed that patients with a surfactant mutation (n = 7) treated for 6 months with antifibrotic drugs showed stable lung function with a median change in FVC of +40 mL (interquartile range, -40 to 90 mL), whereas patients with an SRG mutation treated with immunomodulatory drugs showed a variable response dependent on the gene involved. INTERPRETATION: This study showed that patients with ILD carrying an SRG mutation experience progressive loss of lung function with severely reduced survival despite possible beneficial effects of treatment.

Prevalence and clinical associations of myositis antibodies in a large cohort of interstitial lung diseases.

BACKGROUND: Serologic testing for autoantibodies is recommended in interstitial lung diseases (ILDs), as connective tissue diseases (CTDs) are an important secondary cause. Myositis antibodies are associated with CTD-ILD, but clinical associations with other ILDs are unclear. In this study, associations of myositis antibodies in various ILDs were evaluated. METHODS: 1463 ILD patients and 116 healthy subjects were screened for myositis antibodies with a line-blot assay on serum available at time of diagnosis. Additionally, bronchoalveolar lavage fluid (BALf) was analysed. RESULTS: A total of 394 patients demonstrated reactivity to at least one antibody, including anti-Ro52 (36.0%), anti-Mi-2ß (17.3%) and anti-Jo-1 (10.9%). Anti-Jo-1 (OR 6.4; p<0.100) and anti-Ro52 (OR 6.0; p<0.001) were associated with CTD-ILD. Interestingly, anti-Mi-2ß was associated with idiopathic pulmonary fibrosis (IPF; OR 5.3; p = 0.001) and hypersensitivity pneumonitis (HP; OR 5.9; p<0.001). Furthermore, anti-Mi-2ß was strongly associated with a histological usual interstitial pneumonia (UIP) pattern (OR 6.5; p < 0.001). Moreover, anti-Mi-2ß reactivity was identified in BALf and correlated with serum anti-Mi-2ß (r = 0.64; p = 0.002). No differences were found in survival rates between ILD patients with and without serum Mi-2ß reactivity (hazard ratio 0.835; 95% CI 0.442-1.575; p = 0.577). CONCLUSION: In conclusion, novel associations of antibody Mi-2ß with fibrotic ILD were found. Furthermore, serum anti-Mi-2ß was associated with a histological UIP pattern and presence of anti-Mi-2ß in BALf. Possibly, anti-Mi-2ß could be implemented as a future diagnostic biomarker for fibrotic ILD.

Genetic testing in interstitial lung disease: An international survey.

BACKGROUND AND OBJECTIVE: Genetic analysis is emerging for interstitial lung diseases (ILDs); however, ILD practices are not yet standardized. We surveyed patients', relatives' and pulmonologists' experiences and needs on genetic testing in ILD to evaluate the current situation and identify future needs. METHODS: A clinical epidemiologist (MT) together with members of the ERS taskforce and representatives of the European Idiopathic Pulmonary Fibrosis and related disorders Federation (EU-IPFF) patient organisation developed a survey for patients, relatives and pulmonologists. Online surveys consisted of questions on five main topics: awareness of hereditary ILD, the provision of information, genetic testing, screening of asymptomatic relatives and clinical impact of genetic analysis in ILD. RESULTS: Survey respondents consisted of 458 patients with ILD, 181 patients' relatives and 352 pulmonologists. Most respondents think genetic testing can be useful, particularly for explaining the cause of disease, predicting its course, determining risk for developing disease and the need to test relatives. Informing patients and relatives on genetic analysis is primarily performed by the pulmonologist, but 88% (218) of pulmonologists identify a need for more information and 96% (240) ask for guidelines on genetic testing in ILD. A third of the pulmonologists who would offer genetic testing currently do not offer a genetic test, primarily because they have limited access to genetic tests. Following genetic testing, 72% (171) of pulmonologists may change the diagnostic work-up and 57% (137) may change the therapeutic approach. CONCLUSION: This survey shows that there is wide support for implementation of genetic testing in ILD and a high need for information, guidelines and access to testing among patients, their relatives and pulmonologists.

ANXA11 rs1049550 Associates with Löfgren's Syndrome and Chronic Sarcoidosis Patients.

Sarcoidosis is an immune mediated granulomatous disease commonly affecting the lungs. Genome wide association studies identified many genomic regions that are shared among multiple immune mediated diseases. However, ANXA11 gene polymorphism rs1049550 is exclusively associated with sarcoidosis, making it a key gene of interest for sarcoidosis disease pathogenesis. However, sarcoidosis is a heterogeneous disease and contradictory findings for ANXA11 have been reported for disease phenotypes. We performed a case-control association study to investigate if ANXA11 associates with benign (Löfgren's syndrome (LS)) or chronic sarcoidosis and performed a meta-analysis on previously reported findings. A total of 262 sarcoidosis patients, of which 149 had LS and 113 chronic sarcoidosis, and 363 controls were genotyped for rs1049550. Meta-analysis included allele findings for rs1049550 from 6 additional studies. We found a significantly lower T allele frequency in sarcoidosis patients than in healthy controls (0.30 vs. 0.41, respectively, odds ratio (OR) 0.61, 95% confidence interval (CI) 0.48-0.77, p = 3 × 10-5). In LS the T allele frequency of 0.33, and in chronic sarcoidosis the T allele frequency of 0.26 were significantly lower than in healthy controls (OR 0.69, 95% CI 0.52-0.92, p = 0.01 and OR 0.51, 95% CI 0.36-0.70, p = 4 × 10-5, respectively). Meta-analysis including previously published European, African American and Asian cohorts confirmed the association of rs1049550 with sarcoidosis and resulted in a pooled OR of 0.70 (CI 0.66-0.75, p = 3.58 × 10-29). Presence of the T allele of rs1049550 in ANXA11 is protective for sarcoidosis, including benign and chronic phenotypes of the disease.

Clustering of lung diseases in the family of interstitial lung disease patients.

BACKGROUND: The presence of familial interstitial lung disease (ILD) has been found to predict development of progressive pulmonary fibrosis. However, the role of non-ILD lung diseases in ILD patients' families has not yet been investigated. We aimed to identify associations between ILDs and non-ILD lung diseases from ILD patients' self-reported family health history. METHODS: We analysed questionnaires on family health history of 1164 ILD patients for the occurrence of ILD and non-ILD lung disease in relatives. Logistic regression analysis was used to study associations with diagnosis groups. RESULTS: Familial pulmonary fibrosis was reported by 20% of patients with idiopathic pulmonary fibrosis (IPF; OR 9.2, 95% CI 4.7-17.9), and 15% of patients with unclassifiable pulmonary fibrosis (OR 4.1, 95% CI 2.0-8.2). Familial occurrence was reported by 14% of patients with sarcoidosis (OR 3.3, 95% CI 1.9-5.8). Regarding non-ILD lung disease, significantly more patients with IPF (36%) reported lung cancer in their family (OR 2.3, 95% CI 1.4-3.5), and patients with hypersensitivity pneumonitis (18%) mostly reported COPD (OR 2.3, 95% CI 1.3-4.2). Comparison of sporadic and familial ILD patients' reports showed that emphysema (OR 4.6, 95% CI 1.8-11.6), and lung cancer (OR 2.4, 95% CI 1.2-4.9) were predictive for familial pulmonary fibrosis, particularly when reported both in a family (OR 16.7, 95% CI 3.2-86.6; p < 0.001). CONCLUSIONS: Our findings provide evidence for clustering of ILD and non-ILD lung diseases in families and show that self-reported emphysema and lung cancer of relatives in this population predicts familial pulmonary fibrosis.

Extrapulmonary manifestations of a telomere syndrome in patients with idiopathic pulmonary fibrosis are associated with decreased survival.

BACKGROUND AND OBJECTIVE: Idiopathic pulmonary fibrosis (IPF) is a heterogenous disease with a median survival of 3-4 years. Patients with mutations in telomere-related genes exhibit extrapulmonary signs and symptoms. These patients represent a distinct phenotype of IPF with worse survival. As genetic analyses are not available for most patients with IPF, we sought to determine the predictive value of extrapulmonary signs and symptoms of a telomere syndrome in patients with IPF. METHODS: We retrospectively studied 409 patients with IPF. Clinical characteristics, laboratory results and family history suggestive of a telomere syndrome were related to leukocyte telomere length measured by quantitative PCR and patient outcomes. RESULTS: The cohort included 293 patients with sporadic IPF and 116 patients with a background of familial pulmonary fibrosis. Any or a combination of a clinical history (haematological disease, liver disease, early greying of hair, nail dystrophy, skin abnormalities), a family history or haematological laboratory abnormalities (macrocytosis, anaemia, thrombopenia or leukopenia) suggestive of a telomere syndrome was present in 27% of IPF patients and associated with shorter leukocyte telomere length and shorter survival (p = 0.002 in a multivariate model). In sporadic IPF, having either a clinical history, family history or haematological laboratory abnormalities was not significantly associated with decreased survival (p = 0.07 in a multivariate model). CONCLUSION: Taking a careful clinical and family history focused on extrapulmonary manifestations of a telomere syndrome can provide important prognostic information in patients with IPF, as this is associated with shorter survival.

Telomere dysfunction implicates POT1 in patients with idiopathic pulmonary fibrosis.

Exonic sequencing identified a family with idiopathic pulmonary fibrosis (IPF) containing a previously unreported heterozygous mutation in POT1 p.(L259S). The family displays short telomeres and genetic anticipation. We found that POT1(L259S) is defective in binding the telomeric overhang, nuclear accumulation, negative regulation of telomerase, and lagging strand maintenance. Patient cells containing the mutation display telomere loss, lagging strand defects, telomere-induced DNA damage, and premature senescence with G1 arrest. Our data suggest POT1(L259S) is a pathogenic driver of IPF and provide insights into gene therapy options.

Evidence for shared genetic risk factors between lymphangioleiomyomatosis and pulmonary function.

INTRODUCTION: Lymphangioleiomyomatosis (LAM) is a rare low-grade metastasising disease characterised by cystic lung destruction. The genetic basis of LAM remains incompletely determined, and the disease cell-of-origin is uncertain. We analysed the possibility of a shared genetic basis between LAM and cancer, and LAM and pulmonary function. METHODS: The results of genome-wide association studies of LAM, 17 cancer types and spirometry measures (forced expiratory volume in 1 s (FEV1), forced vital capacity (FVC), FEV1/FVC ratio and peak expiratory flow (PEF)) were analysed for genetic correlations, shared genetic variants and causality. Genomic and transcriptomic data were examined, and immunodetection assays were performed to evaluate pleiotropic genes. RESULTS: There were no significant overall genetic correlations between LAM and cancer, but LAM correlated negatively with FVC and PEF, and a trend in the same direction was observed for FEV1. 22 shared genetic variants were uncovered between LAM and pulmonary function, while seven shared variants were identified between LAM and cancer. The LAM-pulmonary function shared genetics identified four pleiotropic genes previously recognised in LAM single-cell transcriptomes: ADAM12, BNC2, NR2F2 and SP5. We had previously associated NR2F2 variants with LAM, and we identified its functional partner NR3C1 as another pleotropic factor. NR3C1 expression was confirmed in LAM lung lesions. Another candidate pleiotropic factor, CNTN2, was found more abundant in plasma of LAM patients than that of healthy women. CONCLUSIONS: This study suggests the existence of a common genetic aetiology between LAM and pulmonary function.

Connective Tissue Growth Factor Single Nucleotide Polymorphisms in (Familial) Pulmonary Fibrosis and Connective Tissue Disease Associated Interstitial Lung Disease.

PURPOSE: Connective tissue growth factor (CTGF) is an important mediator in fibrotic disease. Single nucleotide polymorphisms (SNPs) in CTGF have been found to be associated with different fibrotic diseases and CTGF protein was found to be upregulated in lung tissue, bronchoalveolar lavage cells, and plasma of idiopathic pulmonary fibrosis (IPF) patients. We investigated whether genetic variants predispose to sporadic IPF (spIPF), familial pulmonary fibrosis (FPF), and connective tissue disease associated ILD (CTD-ILD). METHODS: In total, 294 patients with spIPF and 294 healthy individuals were genotyped for CTGF rs12526196, rs9402373, rs6918698, and rs9399005. For replication of CTGF rs6918698 findings in pulmonary fibrosis, 128 patients with FPF, 125 with CTD-ILD, and an independent control cohort of 130 individuals were included. Lung tissue of 6 IPF patients was stained for CTGF to assess pulmonary localization. RESULTS: Of the four SNPs, only the minor allele frequency (MAF) of CTGF rs6918698 deviated between spIPF (MAF 0.41) and controls (MAF 0.47; OR 0.774 (0.615-0.975); p = 0.030). Further comparison of CTGF rs6918698G showed a difference between FPF (MAF 0.33) and controls (MAF 0.48; OR 0.545 (0.382-0.778); p = 0.001), but not with CTD-ILD. CTGF was localized in alveolar and bronchiolar epithelium, alveolar macrophages, myofibroblasts and endothelium and highly expressed in the basal cell layer of sandwich foci. CONCLUSION: CTGF rs6918698G associates with spIPF and with FPF, but not with CTD-ILD in a Dutch cohort. CTGF is localized in lung tissue involved in IPF pathogenesis. Further research into the role of this SNP on CTGF expression and fibrogenesis is warranted.

Pleuroparenchymal fibroelastosis in idiopathic pulmonary fibrosis: Survival analysis using visual and computer-based computed tomography assessment.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) and pleuroparenchymal fibroelastosis (PPFE) are known to have poor outcomes but detailed examinations of prognostic significance of an association between these morphologic processes are lacking. METHODS: Retrospective observational study of independent derivation and validation cohorts of IPF populations. Upper-lobe PPFE extent was scored visually (vPPFE) as categories of absent, moderate, marked. Computerised upper-zone PPFE extent (cPPFE) was examined continuously and using a threshold of 2·5% pleural surface area. vPPFE and cPPFE were evaluated against 1-year FVC decline (estimated using mixed-effects models) and mortality. Multivariable models were adjusted for age, gender, smoking history, antifibrotic treatment and diffusion capacity for carbon monoxide. FINDINGS: PPFE prevalence was 49% (derivation cohort, n = 142) and 72% (validation cohort, n = 145). vPPFE marginally contributed 3-14% to variance in interstitial lung disease (ILD) severity across both cohorts.In multivariable models, marked vPPFE was independently associated with 1-year FVC decline (derivation: regression coefficient 18·3, 95 CI 8·47-28·2%; validation: 7·51, 1·85-13·2%) and mortality (derivation: hazard ratio [HR] 7·70, 95% CI 3·50-16·9; validation: HR 3·01, 1·33-6·81). Similarly, continuous and dichotomised cPPFE were associated with 1-year FVC decline and mortality (cPPFE ≥ 2·5% derivation: HR 5·26, 3·00-9·22; validation: HR 2·06, 1·28-3·31). Individuals with cPPFE ≥ 2·5% or marked vPPFE had the lowest median survival, the cPPFE threshold demonstrated greater discrimination of poor outcomes at two and three years than marked vPPFE. INTERPRETATION: PPFE quantification supports distinction of IPF patients with a worse outcome independent of established ILD severity measures. This has the potential to improve prognostic management and elucidate separate pathways of disease progression. FUNDING: This research was funded in whole or in part by the Wellcome Trust [209,553/Z/17/Z] and the NIHR UCLH Biomedical Research Centre, UK.

Histamine signaling and metabolism identify potential biomarkers and therapies for lymphangioleiomyomatosis.

Inhibition of mTOR is the standard of care for lymphangioleiomyomatosis (LAM). However, this therapy has variable tolerability and some patients show progressive decline of lung function despite treatment. LAM diagnosis and monitoring can also be challenging due to the heterogeneity of symptoms and insufficiency of non-invasive tests. Here, we propose monoamine-derived biomarkers that provide preclinical evidence for novel therapeutic approaches. The major histamine-derived metabolite methylimidazoleacetic acid (MIAA) is relatively more abundant in LAM plasma, and MIAA values are independent of VEGF-D. Higher levels of histamine are associated with poorer lung function and greater disease burden. Molecular and cellular analyses, and metabolic profiling confirmed active histamine signaling and metabolism. LAM tumorigenesis is reduced using approved drugs targeting monoamine oxidases A/B (clorgyline and rasagiline) or histamine H1 receptor (loratadine), and loratadine synergizes with rapamycin. Depletion of Maoa or Hrh1 expression, and administration of an L-histidine analog, or a low L-histidine diet, also reduce LAM tumorigenesis. These findings extend our knowledge of LAM biology and suggest possible ways of improving disease management.