89Zr-pembrolizumab imaging as a non-invasive approach to assess clinical response to PD-1 blockade in cancer.

BACKGROUND: Programmed cell death protein 1 (PD-1) antibody treatment is standard of care for melanoma and non-small-cell lung cancer (NSCLC). Accurately predicting which patients will benefit is currently not possible. Tumor uptake and biodistribution of the PD-1 antibody might play a role. Therefore, we carried out a positron emission tomography (PET) imaging study with zirconium-89 (89Zr)-labeled pembrolizumab before PD-1 antibody treatment. PATIENTS AND METHODS: Patients with advanced or metastatic melanoma or NSCLC received 37 MBq (1 mCi) 89Zr-pembrolizumab (∼2.5 mg antibody) intravenously plus 2.5 or 7.5 mg unlabeled pembrolizumab. After that, up to three PET scans were carried out on days 2, 4, and 7. Next, PD-1 antibody treatment was initiated. 89Zr-pembrolizumab tumor uptake was calculated as maximum standardized uptake value (SUVmax) and expressed as geometric mean. Normal organ uptake was calculated as SUVmean and expressed as a mean. Tumor response was assessed according to (i)RECIST v1.1. RESULTS: Eighteen patients, 11 with melanoma and 7 with NSCLC, were included. The optimal dose was 5 mg pembrolizumab, and the optimal time point for PET scanning was day 7. The tumor SUVmax did not differ between melanoma and NSCLC (4.9 and 6.5, P = 0.49). Tumor 89Zr-pembrolizumab uptake correlated with tumor response (P trend = 0.014) and progression-free (P = 0.0025) and overall survival (P = 0.026). 89Zr-pembrolizumab uptake at 5 mg was highest in the spleen with a mean SUVmean of 5.8 (standard deviation ±1.8). There was also 89Zr-pembrolizumab uptake in Waldeyer's ring, in normal lymph nodes, and at sites of inflammation. CONCLUSION: 89Zr-pembrolizumab uptake in tumor lesions correlated with treatment response and patient survival. 89Zr-pembrolizumab also showed uptake in lymphoid tissues and at sites of inflammation.

The relation between age and airway epithelial barrier function.

BACKGROUND: The prevalence of age-associated diseases, such as chronic obstructive pulmonary disease (COPD), is increasing as the average life expectancy increases around the world. We previously identified a gene signature for ageing in the human lung which included genes involved in apical and tight junction assembly, suggesting a role for airway epithelial barrier dysfunction with ageing. AIM: To investigate the association between genes involved in epithelial barrier function and age both in silico and in vitro in the airway epithelium. METHODS: We curated a gene signature of 274 genes for epithelial barrier function and tested the association with age in two independent cohorts of bronchial brushings from healthy individuals with no respiratory disease, using linear regression analysis (FDR < 0.05). Protein-protein interactions were identified using STRING©. The barrier function of primary bronchial epithelial cells at air-liquid interface and CRISPR-Cas9-induced knock-down of target genes in human bronchial 16HBE14o-cells was assessed using Trans epithelial resistance (TER) measurement and Electric cell-surface impedance sensing (ECIS) respectively. RESULTS: In bronchial brushings, we found 55 genes involved in barrier function to be significantly associated with age (FDR < 0.05). EPCAM was most significantly associated with increasing age and TRPV4 with decreasing age. Protein interaction analysis identified CDH1, that was negatively associated with higher age, as potential key regulator of age-related epithelial barrier function changes. In vitro, barrier function was lower in bronchial epithelial cells from subjects > 45 years of age and significantly reduced in CDH1-deficient 16HBE14o-cells. CONCLUSION: The significant association between genes involved in epithelial barrier function and age, supported by functional studies in vitro, suggest a role for epithelial barrier dysfunction in age-related airway disease.

Impact of COVID-19 pandemic on diagnostic pathology in the Netherlands.

BACKGROUND: The COVID-19 pandemic has a huge impact on healthcare provided. The nationwide pathology registry of the Netherlands, PALGA, offers an outstanding opportunity to measure this impact for diseases in which pathology examinations are involved. METHODS: Pathology specimen numbers in 2020 were compared with specimen numbers in 2019 for 5 periods of 4 weeks, representing two lockdowns and the periods in between, taking into account localization, procedure and benign versus malignant diagnosis. RESULTS: The largest decrease was seen during the first lockdown (spring 2020), when numbers of pathology reports declined up to 88% and almost all specimen types were affected. Afterwards each specimen type showed its own dynamics with a decrease during the second lockdown for some, while for others numbers remained relatively low during the whole year. Generally, for most tissue types resections, cytology and malignant diagnoses showed less decrease than biopsies and benign diagnoses. A significant but small catch-up (up to 17%) was seen for benign cervical cytology, benign resections of the lower gastro-intestinal tract, malignant skin resections and gallbladder resections. CONCLUSION: The COVID-19 pandemic has had a significant effect on pathology diagnostics in 2020. This effect was most pronounced during the first lockdown, diverse for different anatomical sites and for cytology compared with histology. The data presented here can help to assess the consequences on (public) health and provide a starting point in the discussion on how to make the best choices in times of scarce healthcare resources, considering the impact of both benign and malignant disease on quality of life.

The Microbiome in Bronchial Biopsies from Smokers and Ex-Smokers with Stable COPD - A Metatranscriptomic Approach.

Current knowledge about the respiratory microbiome is mainly based on 16S ribosomal RNA gene sequencing. Newer sequencing approaches, such as metatranscriptomics, offer the technical ability to measure the viable microbiome response to environmental conditions such as smoking as well as to explore its functional role by investigating host-microbiome interactions. However, knowledge about its feasibility in respiratory microbiome research, especially in lung biopsies, is still very limited. RNA sequencing was performed in bronchial biopsies from clinically stable smokers (n = 5) and ex-smokers (n = 6) with COPD not using (inhaled) steroids. The Trinity assembler was used to assemble non-human reads in order to allow unbiased taxonomical and microbial transcriptional analyses. Subsequently, host-microbiome interactions were analyzed based on associations with host transcriptomic data. Ultra-low levels of microbial mass (0.009%) were identified in the RNA-seq data. Overall, no differences were identified in microbiome diversity or transcriptional profiles of microbial communities or individual microbes between COPD smokers and ex-smokers in the initial test dataset as well as a larger replication dataset. We identified an upregulated host gene set, related to the simultaneous presence of Bradyrhizobium, Roseomonas, Brevibacterium.spp., which were related to PERK-mediated unfolded protein response (UPR) and expression of the microRNA-155-5p. Our results show that metatranscriptomic profiling in bronchial biopsy samples from stable COPD patients yields ultra-low levels of microbial mass. Further, this study illustrates the potential of using transcriptional profiling of the host and microbiome to gain more insight into their interaction in the airways.

Human lung extracellular matrix hydrogels resemble the stiffness and viscoelasticity of native lung tissue.

Chronic lung diseases such as idiopathic pulmonary fibrosis (IPF) and chronic obstructive pulmonary disease (COPD) are associated with changes in extracellular matrix (ECM) composition and abundance affecting the mechanical properties of the lung. This study aimed to generate ECM hydrogels from control, severe COPD [Global Initiative for Chronic Obstructive Lung Disease (GOLD) IV], and fibrotic human lung tissue and evaluate whether their stiffness and viscoelastic properties were reflective of native tissue. For hydrogel generation, control, COPD GOLD IV, and fibrotic human lung tissues were decellularized, lyophilized, ground into powder, porcine pepsin solubilized, buffered with PBS, and gelled at 37°C. Rheological properties from tissues and hydrogels were assessed with a low-load compression tester measuring the stiffness and viscoelastic properties in terms of a generalized Maxwell model representing phases of viscoelastic relaxation. The ECM hydrogels had a greater stress relaxation than tissues. ECM hydrogels required three Maxwell elements with slightly faster relaxation times (τ) than that of native tissue, which required four elements. The relative importance (Ri) of the first Maxwell element contributed the most in ECM hydrogels, whereas for tissue the contribution was spread over all four elements. IPF tissue had a longer-lasting fourth element with a higher Ri than the other tissues, and IPF ECM hydrogels did require a fourth Maxwell element, in contrast to all other ECM hydrogels. This study shows that hydrogels composed of native human lung ECM can be generated. Stiffness of ECM hydrogels resembled that of whole tissue, while viscoelasticity differed.

Impact of acute exposure to cigarette smoke on airway gene expression.

BACKGROUND: Understanding effects of acute smoke exposure (ASE) on airway epithelial gene expression and their relationship with the effects of chronic smoke exposure may provide biological insights into the development of smoking-related respiratory diseases. METHODS: Bronchial airway epithelial cell brushings were collected from 63 individuals without recent cigarette smoke exposure and before and 24 h after smoking three cigarettes. RNA from these samples was profiled on Affymetrix Human Gene 1.0 ST microarrays. RESULTS: We identified 91 genes differentially expressed 24 h after ASE (false discovery rate < 0.25). ASE induced genes involved in xenobiotic metabolism, oxidative stress, and inflammation and repressed genes related to cilium morphogenesis and cell cycle. While many genes altered by ASE are altered similarly in chronic smokers, metallothionein genes are induced by ASE and suppressed in chronic smokers. Metallothioneins are also suppressed in current and former smokers with lung cancer relative to those without lung cancer. CONCLUSIONS: Acute exposure to as little as three cigarettes and chronic smoking induce largely concordant changes in airway epithelial gene expression. Differences in short-term and long-term effects of smoking on metallothionein expression and their relationship to lung cancer requires further study given these enzymes' role in the oxidative stress response.

Serum periostin does not reflect type 2-driven inflammation in COPD.

Although Th2 driven inflammation is present in COPD, it is not clearly elucidated which COPD patients are affected. Since periostin is associated with Th2 driven inflammation and inhaled corticosteroid (ICS)-response in asthma, it could function as a biomarker in COPD. The aim of this study was to analyze if serum periostin is elevated in COPD compared to healthy controls, if it is affected by smoking status, if it is linked to inflammatory cell counts in blood, sputum and endobronchial biopsies, and if periostin can predict ICS-response in COPD patients.Serum periostin levels were measured using Elecsys Periostin immunoassay. Correlations between periostin and inflammatory cell count in blood, sputum and endobronchial biopsies were analyzed. Additionally, the correlation between serum periostin levels and treatment responsiveness after 6 and 30 months was assessed using i.e. ΔFEV1% predicted, ΔCCQ score and ΔRV/TLC ratio. Forty-five COPD smokers, 25 COPD past-smokers, 22 healthy smokers and 23 healthy never-smokers were included. Linear regression analysis of serum periostin showed positive correlations age (B = 0.02, 95%CI 0.01-0.03) and FEV1% predicted (B = 0.01, 95%CI 0.01-0.02) in healthy smokers, but not in COPD patients In conclusion, COPD -smokers and -past-smokers have significantly higher periostin levels compared to healthy smokers, yet periostin is not suitable as a biomarker for Th2-driven inflammation or ICS-responsiveness in COPD.

Prenatal exposure to tobacco smoke sex dependently influences methylation and mRNA levels of the Igf axis in lungs of mouse offspring.

Prenatal smoke exposure is a risk factor for abnormal lung development and increased sex-dependent susceptibility for asthma and chronic obstructive pulmonary disease (COPD). Birth cohort studies show genome-wide DNA methylation changes in children from smoking mothers, but evidence for sex-dependent smoke-induced effects is limited. The insulin-like growth factor (IGF) system plays an important role in lung development. We hypothesized that prenatal exposure to smoke induces lasting changes in promoter methylation patterns of Igf1 and Igf1r, thus influencing transcriptional activity and contributing to abnormal lung development. We measured and compared mRNA levels along with promoter methylation of Igf1 and Igf1r and their protein concentrations in lung tissue of 30-day-old mice that had been prenatally exposed to cigarette smoke (PSE) or filtered air (control). Body weight at 30 days after birth was measured as global indicator of normal development. Female PSE mice showed lower mRNA levels of Igf1 and its receptor (Igf1: P = 0.05; Igf1r: P = 0.03). Furthermore, CpG-site-specific methylation changes were detected in Igf1r in a sex-dependent manner and the body weight of female offspring was reduced after prenatal exposure to smoke, while protein concentrations were unaffected. Prenatal exposure to smoke induces a CpG-site-specific loss of Igf1r promoter methylation, which can be associated with body weight. These findings highlight the sex-dependent and potentially detrimental effects of in utero smoke exposure on DNA methylation and Igf1 and Igf1r mRNA levels. The observations support a role for Igf1 and Igf1r in abnormal development.

Combining genomewide association study and lung eQTL analysis provides evidence for novel genes associated with asthma.

BACKGROUND: Genomewide association studies (GWASs) of asthma have identified single-nucleotide polymorphisms (SNPs) that modestly increase the risk for asthma. This could be due to phenotypic heterogeneity of asthma. Bronchial hyperresponsiveness (BHR) is a phenotypic hallmark of asthma. We aim to identify susceptibility genes for asthma combined with BHR and analyse the presence of cis-eQTLs among replicated SNPs. Secondly, we compare the genetic association of SNPs previously associated with (doctor's diagnosed) asthma to our GWAS of asthma with BHR. METHODS: A GWAS was performed in 920 asthmatics with BHR and 980 controls. Top SNPs of our GWAS were analysed in four replication cohorts, and lung cis-eQTL analysis was performed on replicated SNPs. We investigated association of SNPs previously associated with asthma in our data. RESULTS: A total of 368 SNPs were followed up for replication. Six SNPs in genes encoding ABI3BP, NAF1, MICA and the 17q21 locus replicated in one or more cohorts, with one locus (17q21) achieving genomewide significance after meta-analysis. Five of 6 replicated SNPs regulated 35 gene transcripts in whole lung. Eight of 20 asthma-associated SNPs from previous GWAS were significantly associated with asthma and BHR. Three SNPs, in IL-33 and GSDMB, showed larger effect sizes in our data compared to published literature. CONCLUSIONS: Combining GWAS with subsequent lung eQTL analysis revealed disease-associated SNPs regulating lung mRNA expression levels of potential new asthma genes. Adding BHR to the asthma definition does not lead to an overall larger genetic effect size than analysing (doctor's diagnosed) asthma.

Shifted T-cell polarisation after agricultural dust exposure in mice and men.

RATIONALE: A low prevalence of asthma and atopy has been shown in farmers and agricultural workers. However, in these workers, a higher prevalence of respiratory symptoms has been reported, in which T helper 1 (Th1) and/or Th17 responses may play a role. AIM: We investigated the effect of exposure to dust extracts (DEs) from different farms on airway inflammation and T-cell polarisation in a mouse model and assessed T-cell polarisation in agricultural workers from the same farms. METHODS: DEs were prepared from settled dust collected at cattle and pig farms and bulb and onion industries. Mice were exposed to phosphate-buffered saline (PBS), DEs, house dust mite (HDM) or HDM+DE via nasal instillation, four times per week during 5 weeks. Hyperresponsiveness, airway inflammation, IgE levels and T-cell polarisation were assessed. Th-cell and T cytotoxic (Tc)-cell subsets were investigated in peripheral blood samples from 33 agricultural workers and 9 non-exposed controls. RESULTS: DEs induced interleukin(IL)-17, IL-1ß and IL-6 in mouse lung homogenates. DE-exposed mice had more mixed inflammatory infiltrates in the lungs, and more neutrophils compared with PBS-exposed mice. DEs protected against the HDM-induced Th2 response and methacholine hyperresponsiveness. Interestingly, occupationally exposed humans had higher frequencies of Th cells spontaneously expressing IL-17 and interferon γ compared with controls. CONCLUSION: Chronic exposure to different types of farm dust induces a Th/Tc-17 inflammatory response in mice and agricultural workers. This may contribute to the low prevalence of Th2-related diseases but may constitute a risk for other chronic respiratory diseases.

Regional differences in predictive biomarker testing rates for patients with metastatic NSCLC in the Netherlands.

BACKGROUND: Predictive biomarker testing has a key role in the treatment decision-making for patients with non-small cell lung cancer (NSCLC) and is mandated by (inter)national guidelines. The aim of this study was to establish guideline-adherent biomarker testing rates in the Netherlands in 2019 and to examine associations of demographical, clinical, and environmental factors with guideline-adherent testing. METHODS: This study involved the integration of clinical data of the Netherlands Cancer Registry with pathology reports of the Dutch Nationwide Pathology Databank. Data extracted from these reports included sample type, diagnosis, and molecular testing status of predictive biomarkers. The study population comprised all patients diagnosed with metastatic non-squamous NSCLC in the Netherlands in 2019. RESULTS: In the cohort of 3877 patients with metastatic non-squamous NSCLC under investigation, overall molecular testing rates for non-fusion predictive biomarkers (EGFR, KRAS, BRAF, ERBB2, MET) ranged from 73.9 to 89.0 %, while molecular testing for fusion-drivers (ALK, ROS1, RET, NTRK) ranged from 12.6 % to 63.9 %. Guideline-adherent testing of EGFR, KRAS, and ALK was performed in 85.2 % of patients, with regional rates spanning from 76.0 % to 90.8 %. Demographical and clinical factors associated with guideline-adherent biomarker testing included lower age (OR = 1.05 per one year decrease; p < 0.001), female sex (OR = 1.36; p = 0.002), diagnosis of adenocarcinoma (OR = 2.48; p < 0.001), availability of histological tumor material (OR = 2.46; p < 0.001), and clinical stage of metastatic disease (p = 0.002). Other factors associated with guideline-adherent biomarker testing included diagnosis at academic center (OR = 1.87; p = 0.002) and patient's region of residence (p < 0∙001). CONCLUSION: Optimization of the chain-of-care of predictive biomarker testing in patients with NSCLC in the Netherlands is needed to provide adequate care for these patients.

Circulating tumor cells in small-cell lung cancer: a predictive and prognostic factor.

BACKGROUND: Initial response of small-cell lung cancer (SCLC) to chemotherapy is high, and recurrences occur frequently, leading to early death. This study investigated the prognostic value of circulating tumor cells (CTCs) in patients with SCLC and whether changes in CTCs can predict response to chemotherapy. Patients and methods In this multicenter prospective study, blood samples for CTC analysis were obtained from 59 patients with SCLC before, after one cycle, and at the end of chemotherapy. CTCs were measured using CellSearch systems. RESULTS: At baseline, lower numbers of CTCs were observed for 21 patients with limited SCLC (median = 6, range 0-220) compared with 38 patients with extensive stage (median = 63, range 0-14,040). Lack of measurable CTCs (27% of patients) was associated with prolonged survival (HR 3.4; P ≤ 0.001). CTCs decreased after one cycle of chemotherapy; this decrease was not associated with tumor response after four cycles of chemotherapy. CTC count after the first cycle of chemotherapy was the strongest predictor for overall survival (HR 5.7; 95% CI 1.7-18.9; P = 0.004). CONCLUSION: Absolute CTCs after one cycle of chemotherapy in patients with SCLC is the strongest predictor for response on chemotherapy and survival. Patients with low initial CTC numbers lived longer than those with higher CTCs.

Obesity in asthma: more neutrophilic inflammation as a possible explanation for a reduced treatment response.

BACKGROUND: The incidence of asthma and obesity is increasing worldwide, and reports suggest that obese patients have more severe asthma. We investigated whether obese asthma patients have more severe airway obstruction and airway hyper-responsiveness and a different type of airway inflammation than lean asthmatics. Furthermore, we assessed the effect of obesity on corticosteroid treatment response. METHODS: Patient data from four well-documented asthma cohorts were pooled (n = 423). We evaluated FEV(1) , bronchial hyper-responsiveness (PC(20) ) to either methacholine/histamine or adenosine 5'-monophosphate (AMP) (differential) cell counts in induced sputum and blood and corticosteroid treatment response in 118 patients. RESULTS: At baseline, FEV(1) , PC(20) methacholine or histamine, and PC(20) AMP values were comparable in 63 obese (BMI ≥ 30 kg/m(2) ) and 213 lean patients (BMI <25 kg/m(2) ). Obese patients had significantly higher blood neutrophils. These higher blood neutrophils were only seen in obese women and not in obese men. After a two-week treatment with corticosteroids, we observed less corticosteroid-induced improvement in FEV(1) %predicted in obese patients than in lean patients (median 1.7% vs 6.3% respectively, P = 0.04). The percentage of sputum eosinophils improved significantly less with higher BMI (P = 0.03), and the number of blood neutrophils increased less in obese than in lean patients (0.32 x10(3) /µl vs 0.57 x10(3) /µl, P = 0.046). CONCLUSIONS: We found no differences in asthma severity between obese and nonobese asthmatics. Interestingly, obese patients demonstrated more neutrophils in sputum and blood than nonobese patients. The smaller improvement in FEV(1) and sputum eosinophils suggests a worse corticosteroid treatment response in obese asthmatics.

Antinuclear autoantibodies are more prevalent in COPD in association with low body mass index but not with smoking history.

BACKGROUND: Chronic obstructive pulmonary disease (COPD) is associated with a higher prevalence of antinuclear autoantibodies (ANAs). However, a significant subgroup of patients is ANA negative. It remains to be determined which patient groups carry autoantibodies. METHODS: The association of smoking behaviour, disease status, gender, age and body mass index (BMI) with the presence of autoantibodies in the serum was determined in 124 patients with COPD and 108 non-COPD control subjects. In addition, the role of B cells in autoantibody generation in COPD was investigated by sequencing the antibody repertoire of B cells in the lungs of patients with COPD and of ex-smoking and never-smoking control subjects. RESULTS: Patients with COPD had a significantly higher risk of being serum positive for ANAs (OR 3.12, 95% CI 1.68 to 5.76, p<0.001). ANAs were not significantly associated with age, smoking status, gender or pack-years of smoking. Within the COPD population, subjects with BMI <22 kg/m2 had a significantly higher risk of ANAs (OR 4.93, 95% CI 1.50 to 16.50, p=0.009) than those with normal or high BMI. The antibody repertoire of B cells in the lungs of patients with COPD had a high frequency of positively charged CDR3 residues, a feature which is associated with self-reactive antibodies. CONCLUSION: The results show that COPD is a heterogeneous disease with respect to the prevalence of ANAs. ANAs are primarily associated with the presence of COPD and with low BMI, but not with smoking and forced expiratory volume in 1 s.

E-cadherin gene polymorphisms in asthma patients using inhaled corticosteroids.

E-cadherins form intercellular junctions that maintain epithelial integrity. Epithelial integrity is impaired in asthma and can be restored by inhaled corticosteroids (ICSs). Our aim was to investigate the association of CDH1 gene polymorphisms (single-nucleotide polymorphisms (SNPs)) with airway remodelling, inflammation and forced expiratory volume in 1 s (FEV1) decline in asthma patients and assess whether ICSs modulate these effects. Bronchial biopsies of 138 asthmatics were available (population 1). Associations of 17 haplotype-tagging SNPs with epithelial E-cadherin expression, biopsy parameters and FEV1/vital capacity (VC) ratio were tested. FEV1 and VC data were collected in 281 asthmatics with 30-yr follow-up (population 2). Linear mixed-effect models were used to assess associations of SNPs with FEV1 decline. Seven out of the 17 SNPs were associated with airway remodelling, three with CD8+ T-cell counts, two with eosinophil counts and seven with FEV1 decline. All associations occurred only in patients using ICS. In general, alleles associated with less remodelling correlated with less FEV1 decline and higher FEV1/VC. Decreased epithelial E-cadherin expression was associated with five SNPs in non-ICS users. In conclusion, our data show that CDH1 polymorphisms are associated with epithelial E-cadherin expression and suggest that epithelial adhesion is an important contributor to airway remodelling and lung function in asthma. These effects are modified by the use of inhaled corticosteroids.

SERPINE1 -675 4G/5G polymorphism is associated with asthma severity and inhaled corticosteroid response.

Asthma is characterised by chronic airway inflammation and remodelling, which can be (partially) suppressed by inhaled corticosteroids (ICSs). Plasminogen activator inhibitor-1, encoded by the SERPINE1 gene, is the key inhibitor of the plasminogen activator system, which affects tissue repair and remodelling. We studied associations between a functional SERPINE1 -675 4G/5G promoter polymorphism and asthma development, severity and response to ICSs. Longitudinal cohorts of 281 asthmatics and their nonasthmatic spouses, and the general population (n=1,390) were studied. No significant associations were found with asthma development and immunoglobulin (Ig)E levels, or with forced expiratory volume in 1 s (FEV1) in nonasthmatic controls. Asthmatic subjects carrying the SERPINE1 5G allele had higher IgE and lower lung function levels at follow-up, lower maximally attained lung function levels, and faster lung function decline compared with individuals with the 4G/4G genotype. ICS treatment showed an immediate improvement in FEV1 in asthmatics carrying the 5G allele. However, these asthmatics still had the fastest rate of FEV1 decline after initiating ICS treatment. Finally, the 5G allele was associated with a lower prevalence of complete asthma remission at follow-up. These findings suggest that SERPINE1 is not an asthma susceptibility gene, but rather affects the severity, progression and long-term ICS response in asthma.

Prognostic impact of KRAS mutation status for patients with stage IV adenocarcinoma of the lung treated with first-line pembrolizumab monotherapy.

OBJECTIVES: Monotherapy with pembrolizumab is the preferred first-line treatment for metastatic non-small cell lung cancer with programmed death-ligand 1 (PD-L1) expression ≥50 %, without targetable oncogenic drivers. Although targeted therapies are in development for patients with specific Kirsten rat sarcoma (KRAS) mutations, these are not available in daily care yet. It is not clear whether there is a difference in survival on first-line pembrolizumab for patients with a high PD-L1 status with or without a KRAS mutation. We aim to compare this survival based on real-world data. MATERIALS AND METHODS: This is a real-world retrospective population-based study using data from the Netherlands Cancer Registry. We selected patients with stage IV lung adenocarcinoma with PD-L1 expression ≥50 % diagnosed between January 2017 and December 2018, treated with first-line pembrolizumab. Patients with EGFR mutations, ALK translocations or ROS1 rearrangements were excluded. The primary outcome parameter was overall survival. RESULTS: 388 (57 %) of 595 patients had a KRAS mutation. KRAS was seen more frequently in women than in men (65 % versus 49 % respectively, p < 0.001). The median overall survival was 19.2 months versus 16.8 months for patients with and without KRAS mutation, respectively (p = 0.86). Multivariable analysis revealed WHO performance score, number of organs with metastases and PD-L1 percentage as independent prognostic factors. KRAS mutation status had no prognostic influence (hazard ratio = 1.03, 95 % CI 0.83-1.29). CONCLUSION: The survival of KRAS mutated versus KRAS wild-type lung adenocarcinoma patients, treated with first-line pembrolizumab monotherapy, is similar, suggesting that KRAS has no prognostic value with respect to treatment with pembrolizumab.

Airway inflammation and remodeling in two mouse models of asthma: comparison of males and females.

BACKGROUND: Asthma and especially severe asthma affect women more frequently than men. Since asthma severity correlates with remodeling changes in the lung, a female propensity to remodeling could be expected. We studied whether our previous observation that female mice have more pronounced airway inflammation than males is associated with more pronounced remodeling in two models of chronic allergic asthma. METHODS: Male and female BALB/c mice were (1) sensitized and subsequently challenged with ovalbumin (OVA) for 4 weeks, or (2) exposed to house dust mite (HDM) for 5 weeks. In both models, allergic inflammation, remodeling, antigen-specific IgE and methacholine (MCh) responsiveness were assessed. RESULTS: Females had higher antigen-specific serum IgE levels, higher numbers of eosinophils and were more responsive to MCh. In the OVA model, females also had higher levels of Th2 cytokines in lung tissue than males. Both sexes developed similar airway remodeling (smooth muscle layer thickness, collagen III deposition and goblet cell hyperplasia) in the two models. CONCLUSIONS: Combining results of an OVA- and a HDM-induced mouse model of allergic airway inflammation, we have shown that more severe allergic inflammation in females is not accompanied with more pronounced airway remodeling.

Determinants of SARS-CoV-2 receptor gene expression in upper and lower airways.

The recent outbreak of the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), which causes coronavirus disease 2019 (COVID-19), has led to a worldwide pandemic. One week after initial symptoms develop, a subset of patients progresses to severe disease, with high mortality and limited treatment options. To design novel interventions aimed at preventing spread of the virus and reducing progression to severe disease, detailed knowledge of the cell types and regulating factors driving cellular entry is urgently needed. Here we assess the expression patterns in genes required for COVID-19 entry into cells and replication, and their regulation by genetic, epigenetic and environmental factors, throughout the respiratory tract using samples collected from the upper (nasal) and lower airways (bronchi). Matched samples from the upper and lower airways show a clear increased expression of these genes in the nose compared to the bronchi and parenchyma. Cellular deconvolution indicates a clear association of these genes with the proportion of secretory epithelial cells. Smoking status was found to increase the majority of COVID-19 related genes including ACE2 and TMPRSS2 but only in the lower airways, which was associated with a significant increase in the predicted proportion of goblet cells in bronchial samples of current smokers. Both acute and second hand smoke were found to increase ACE2 expression in the bronchus. Inhaled corticosteroids decrease ACE2 expression in the lower airways. No significant effect of genetics on ACE2 expression was observed, but a strong association of DNA- methylation with ACE2 and TMPRSS2- mRNA expression was identified in the bronchus.