Introduction Persistent inflammation and immune activation in the lungs are associated with adverse outcomes such as radiation pneumonitis (RP) and poor survival in non-small-cell lung cancer (NSCLC) patients. However, it is unknown how this is reflected by leukocyte activation markers in serum. Objective The aim was to evaluate the serum levels of activation of different leukocyte subsets and to examine those in relation to the pathogenesis of RP and survival in NSCLC. Methods We analyzed the serum levels of MPO, sCD25, sTIM-3, sPD-L1, sCD14, sCD163, CCL19 and CCL21 in 66 inoperable NSCLC patients with stage IA-IIIA disease. The patients were treated with stereotactic body radiation therapy (SBRT) or concurrent chemoradiation therapy (CCRT), followed by regular blood sampling for 12 months after treatment and for 5 years for survival. Results Nineteen (29%) patients developed RP, which occurred more frequently and earlier in patients receiving CCRT than in those receiving SBRT. Increases in sCD25, sTIM-3 and CCL21 levels were observed at the last 6 months of follow-up in patients who had RP after SBRT. Patients who had RP after CCRT had higher sTIM-3 levels during the first 3 months of follow-up. Baseline sCD25 was independently associated with both 2- and 5-year mortality outcomes, while baseline sTIM-3 was independently associated with 2-year mortality. Conclusion We showed that T cell activation and exhaustion markers such as sCD25 and sTIM-3 are enhanced in patients developing RP and are associated with poor survival in NSCLC.

Background Genetic alterations are common in non-small cell lung cancer (NSCLC), and DNA mutations and translocations are targets for therapy. Copy number aberrations occur frequently in NSCLC tumors and may influence gene expression and further alter signaling pathways. In this study we aimed to characterize the genomic architecture of NSCLC tumors and to identify genomic differences between tumors stratified by histology and mutation status. Furthermore, we sought to integrate DNA copy number data with mRNA expression to find genes with expression putatively regulated by copy number aberrations and the oncogenic pathways associated with these affected genes. Methods Copy number data were obtained from 190 resected early-stage NSCLC tumors and gene expression data were available from 113 of the adenocarcinomas. Clinical and histopathological data were known, and EGFR -, KRAS - and TP53 mutation status was determined. Allele-specific copy number profiles were calculated using ASCAT, and regional copy number aberration were subsequently obtained and analyzed jointly with the gene expression data. Results The NSCLC tumors tissue displayed overall complex DNA copy number profiles with numerous recurrent aberrations. Despite histological differences, tissue samples from squamous cell carcinomas and adenocarcinomas had remarkably similar copy number patterns. The TP53 -mutated lung adenocarcinomas displayed a highly aberrant genome, with significantly altered copy number profiles including gains, losses and focal complex events. The EGFR -mutant lung adenocarcinomas had specific arm-wise aberrations particularly at chromosome7p and 9q. A large number of genes displayed correlation between copy number and expression level, and the PI(3)K-mTOR pathway was highly enriched for such genes. Conclusions The genomic architecture in NSCLC tumors is complex, and particularly TP53 -mutated lung adenocarcinomas displayed highly aberrant copy number profiles. We suggest to always include TP53 -mutation status when studying copy number aberrations in NSCLC tumors. Copy number may further impact gene expression and alter cellular signaling pathways.

IntroductionThe present study explores changes in pulmonary function, symptoms and radiological signs of pneumonitis after curatively intended stereotactic body radiation therapy (SBRT).MethodsAll inoperable, early-stage non-small cell lung cancer patients treated with stereotactic body radiation therapy (SBRT) from 2014-2017 were included in this single-centre study. They were followed regularly for 12 months after treatment. The patients were classified into three groups based on radiology and symptomatology: no radiation pneumonitis, asymptomatic and symptomatic radiation pneumonitis.ResultsForty-four patients with stage IA-IIB disease were treated with 45–56 Gy in 3–8 fractions. The median age was 75 years, 43% of the patients were female; 60% of the patients had a COPD in GOLD grade of 2-4, and 95.5% were active or former smokers. Symptomatic radiation pneumonitis occurred in 18% of the patients and asymptomatic pneumonitis as defined by radiology, in 39%. The mean of forced expiratory volume in 1 second (FEV1) and diffusion capacity for carbon monoxide (DLCO) decreases for all patients during the first years were higher than one would expect from physiologic ageing. FEV1 and DLCO in percent decrease 7-8% at 1-1.5 months in the symptomatic radiation pneumonitis group. CT scan findings consistent with radiation pneumonitis occurred after a median of 2.9 months in the symptomatic and 5.4 months in the asymptomatic radiation pneumonitis groups. In the group with symptomatic radiation pneumonitis, symptoms, as measured by the Clinical COPD questionnaire score, significantly increased at 3 and 6 months. Significant higher maximum doses to the critical lung volumes DC1000cm3(1000 cm3of lung receiving a given dose or less) and DC 1500cm3(1500 cm3of lung receiving a given dose or less) were observed in patients who developed radiation pneumonitis.ConclusionEarly decrease in measured FEV1 and DLCO occurred before imaging changes and symptoms and might indicate the development of symptomatic radiation pneumonitis. The dose to critical lung volumes of DC1000 cm3and DC1500 cm3may predict the risk for the development of symptomatic radiation pneumonitis.

The implementation of immune checkpoint inhibitors (ICI) into the standard care of advanced non-small cell lung cancer (NSCLC) has improved prognosis for this group of patients. However, long-term survival is rare. The aim of the study was to identify predictors of response and, especially, to investigate the impact radiotherapy might have on duration of response. The association between pretreatment patient/tumor characteristics and progression-free survival (PFS), overall survival (OS), and lung cancer-specific survival was investigated in 78 patients receiving an ICI as ≥2nd line treatment for advanced NSCLC, using Cox regression analysis. Due to competing risk, cause-specific deaths were also examined with cumulative incidence plots. Median OS was 12.6 months (95% CI 7.8–18.2) and median PFS 4.1 months (95% CI 3.0–6.2), after median follow-up time of 49.7 months (range 20.9–51.5). Increasing CRP and neutrophil/lymphocyte ratio (NLR), were associated with poor PFS (CRP: HR 1.49, 95% CI 1.12–1.98; NLR: HR 1.59, 95% CI 1.22–1.85) and OS (CRP: HR 1.94, 95% CI 1.47–2.56; NLR: HR 1.54, 95% CI 1.27–1.87). Radiotherapy prior to immunotherapy was not significantly associated with patient outcome. However, when the dataset was split at 6 months of follow-up, to be able to identify early and late predictors of prognosis, we found that patients receiving radiotherapy <6 months prior to immunotherapy had better PFS (HR: 0.27, 95% CI 0.09–0.84) and lung cancer-specific survival (HR: 0.41, 95% CI 0.18–0.95) after the first 6 months of follow-up, while increasing CRP (PFS: HR1.61, 95% CI 1.21–2.14; OS: HR2.04, 95% CI 1.51–2.74) and NLR (PFS: HR 1.57, 95% CI 1.29–1.91; OS: HR 1.63, 95% CI 1.35–1.97) were predictors of poor short-term prognosis. Radiotherapy may be of importance to achieve a long-lasting response to immunotherapy, while indicators of systemic inflammation can help in identifying patients with poor short-term prognosis.

e21065 Background: RP is a clinically challenging side effect following SBRT. The reported incidence of RP varies from 2.2% to 46.8%, depending on study design and different grading systems. We evaluated the grading systems for RP and identified possible predictive tests for RP. Furthermore, we estimated the correlation between RP and dosimetric measurements from the radiation therapy. Methods: Medically inoperable patients, n = 44, with peripherally located NSCLC stage I-II, were treated with SBRT, with a total dose of 45–56 Gy in 3–8 fractions. Median age was 75 years, 43.2% were female and 60% had moderate to very severe COPD. Follow up included physical examination by pulmonologist, spirometry and single-breath lung diffusing capacity (DLCO) and CT evaluation at baseline and 1.5, 3, 6, 9 and 12 months after SBRT. Results: We constructed three groups for grading RP based on CTCAE version 5.0 and imaging changes according to EORTC (LENT-SOMA) Non-RP (asymptomatic or mild symptoms and slight imaging changes, n = 19, 43%), Asymptomatic, radiology-only RP (asymptomatic or mild symptoms, increased density imaging changes, n = 17, 39%) and Symptomatic and radiology-detected RP (moderate and severe symptoms and increased density imaging changes, n = 8, 18%). Active smokers and patients with emphysema were overrepresented in non-RP group. In the symptomatic RP group, DLCO and FEV1 dropped almost 5% 4-6 weeks after SBRT, before symptoms and imaging changes, and DLCO, FVC and FEV1 dropped significantly at 3 months (table). Development of RP was associated with critical dose-volume parameter 1000ccm and 1500ccm. Several dosimetric parameters significantly negative correlate with FVC-drop at 1 and 3 months only in symptomatic RP. Conclusions: Active smokers and patients with emphysema have less propensity of RP. Symptomatic RP is seen with a presymptomatic drop in FEV1 and DLCO occurring 4-6 weeks after SBRT. Critical dose-volume parameter is important parameters for RP. Correlation between dosimetric parameters and FVC-drop in symptomatic RP. Clinical trial information: NCT02428049 . [Table: see text]

IntroductionProtein expression is deregulated in cancer, and the proteomic changes observed in lung cancer may be a consequence of mutations in essential genes. The purpose of this study was to identify protein expression associated with prognosis in lung cancers stratified by smoking status, molecular subtypes, and EGFR-, TP53-, and KRAS-mutations.MethodsWe performed profiling of 295 cancer-relevant phosphorylated and non-phosphorylated proteins, using reverse phase protein arrays. Biopsies from 80 patients with operable lung adenocarcinomas were analyzed for protein expression and association with relapse free survival (RFS) were studied.ResultsSpearman’s rank correlation analysis identified 46 proteins with significant association to RFS (p<0.05). High expression of protein kinase C (PKC)-α and the phosporylated state of PKC-α, PKC-β, and PKC-δ, showed the strongest positive correlation to RFS, especially in the wild type samples. This was confirmed in gene expression data from 172 samples. Based on protein expression, unsupervised hierarchical clustering separated the samples into four subclusters enriched with the molecular subtypes terminal respiratory unit (TRU), proximal proliferative (PP), and proximal inflammatory (PI) (p=0.0001). Subcluster 2 contained a smaller cluster (2a) enriched with samples of the subtype PP, low expression of the PKC isozymes, and associated with poor RFS (p=0.003) compared to the other samples. Low expression of the PKC isozymes in the subtype PP and a reduced relapse free survival was confirmed with The Cancer Genome Atlas (TCGA) lung adenocarcinoma (LUAD) samples.ConclusionThis study identified different proteins associated with RFS depending on molecular subtype, smoking- and mutational-status, with PKC-α, PKC-β, and PKC-δ showing the strongest correlation.

Personalised cancer treatment depends on identification of therapeutically relevant biological subgroups of patients for assessing effect of treatment and to discover new therapeutic options. By analyses in heterogeneous patient populations, the effects may be lost in noise. Squamous cell carcinoma of the lung is a major killer worldwide. Despite recent advances, mortality is high and response to therapies varies greatly from patient to patient. Target search in biologically relevant subgroups may identify treatment options not so far discovered. A total of 198 patients undergoing surgery for squamous cell carcinomas of the lung were included in the study. The tumours were analysed for copy number alterations (n = 152) and gene expression from tumour (n = 188) and normal lung (n = 21), with both data levels present in 140 patients. We studied alterations in tumours harbouring mutations in TP53 and in previously published gene expression subtypes. Genes with consistent alterations in both genomic levels were identified as putative biomarkers. Results were validated in TCGA. The most convincing biomarker in TP53 mutated squamous cell carcinomas of the lung was BIRC5 with amplification in 36% of mutated samples, 5% in wild-type samples and a 17%-fold change of expression between TP53 mutated tumours and normal lung tissue. BIRC5 was significantly altered in the classical and primitive subtypes. We suggest BIRC5 as a putative predictive biomarker and putative druggable target in squamous cell lung carcinomas harbouring TP53 mutation or classified as classical and primitive subtypes.

Dysregulation of microRNAs is a common mechanism in the development of lung cancer, but the relationship between microRNAs and expression subtypes in non‐small‐cell lung cancer (NSCLC) is poorly explored. Here, we analyzed microRNA expression from 241 NSCLC samples and correlated this with the expression subtypes of adenocarcinomas (AD) and squamous cell carcinomas (SCC) to identify microRNAs specific for each subtype. Gene set variation analysis and the hallmark gene set were utilized to calculate gene set scores specific for each sample, and these were further correlated with the expression of the subtype‐specific microRNAs. In ADs, we identified nine aberrantly regulated microRNAs in the terminal respiratory unit (TRU), three in the proximal inflammatory (PI), and nine in the proximal proliferative subtype (PP). In SCCs, 1, 5, 5, and 9 microRNAs were significantly dysregulated in the basal, primitive, classical, and secretory subtypes, respectively. The subtype‐specific microRNAs were highly correlated to specific gene sets, and a distinct pattern of biological processes with high immune activity for the AD PI and SCC secretory subtypes, and upregulation of cell cycle‐related processes in AD PP, SCC primitive, and SCC classical subtypes were found. Several in silico predicted targets within the gene sets were identified for the subtype‐specific microRNAs, underpinning the findings. The results were significantly validated in the LUAD (n = 492) and LUSC (n = 380) TCGA dataset (False discovery rates‐corrected P‐value < 0.05). Our study provides novel insight into how expression subtypes determined with discrete biological processes may be regulated by subtype‐specific microRNAs. These results may have importance for the development of combinatory therapeutic strategies for lung cancer patients.

Abstract Background Protein expression is deregulated in cancer, and the proteomic changes observed in lung cancer may be a consequence of mutations in essential genes. The purpose of this study was to identify protein expression associated with prognosis in lung cancers stratified by smoking status, molecular subtypes, and EGFR-, TP53- and KRAS-mutations. Methods We performed profiling of 295 cancer-relevant phosphorylated and non-phosphorylated proteins, using reverse phase protein arrays. Biopsies from 80 patients with operable lung adenocarcinomas were analyzed for protein expression and association with progression free survival (PFS) were studied. Results Spearman rank correlation analysis identified 56 proteins with significant association to PFS (p<0.05). High expression of protein kinase C (PKC)-α and the phosporylated state of PKC-α, PKC-β and PKC-δ, showed the strongest positive correlation to PFS, especially in the wild type samples. This was confirmed in gene expression data from 186 samples. Based on protein expression, unsupervised hierarchical clustering separated the samples into four subclusters enriched with the molecular subtypes TRU, PI or PP (p=0.0001). Subcluster 2 contained a smaller cluster (2a) enriched with samples of the subtype PP, low expression of the PKC isozymes, and associated with poor PFS (p=0.003) compared to the other samples. Subcluster 2a revealed increased expression of neuroendocrine markers, supporting the aggressive behavior. Low expression of the PKC isozymes in the subtype PP and a reduced relapse free survival was confirmed with the TCGA LUAD samples. Conclusion This study identified different proteins associated with PFS depending on molecular subtype, smoking- and mutational-status, with PKC-α, PKC-β and PKC-δ showing the strongest correlation. Cluster analysis detected a subgroup of samples enriched for samples of the PP subtype and poor PFS, which may benefit from a more aggressive treatment regimen.

The impact of the tumor immune microenvironment on overall survival in non‐small cell lung cancer (NSCLC) has been studied, but there is little information on its relevance for risk of relapse after surgery. Understanding more about the immune microenvironment in previously untreated NSCLC could help in identifying high‐risk patients and patients more likely to benefit from neoadjuvant/adjuvant immunotherapy. Here, we examined gene expression in 399 surgically derived NSCLC samples and 47 samples from normal lung, using Agilent microarray and RNA sequencing. In 335 of the tumor samples, programmed death‐ligand 1 (PD‐L1) expression was evaluated by immunohistochemistry. Gene expression was used to estimate content of immune cells and to calculate an immune score. Properties of the immune microenvironment, and its impact on prognosis, were compared in histological subgroups and gene expression subtypes. Tumors with an active immune microenvironment were found for both adenocarcinomas (AD) and squamous cell carcinomas (SCC). In AD, high immune score and high estimates of several immune cell types belonging to the adaptive immune system were associated with better progression‐free survival (PFS), while in SCC, no association between immune characteristics and PFS was found. The immune microenvironment, including PD‐L1 expression, and its impact on prognosis showed clear differences in AD and SCC gene expression subtypes. In conclusion, the NSCLC immune microenvironment is predictive of prognosis after surgery. Lung AD and SCC gene expression subtypes should be investigated as potential prognostic biomarkers in patients treated with immune checkpoint inhibitors.

Background: The introduction of immune check-point inhibition in non-small cell lung cancer (NSCLC) therapy represents improved prospects for the patients. The response rates to check-point inhibitors are approximately 20% in unselected NSCLC patients. Increasing levels of tumor PD-L1 expression are associated with higher response rates. However, patients with low PD-L1 levels may also have durable responses, and improved strategies for patient stratification are needed. Material and methods: In this study, we investigated circulating microRNAs aiming to identify circulating predictive biomarkers associated with increased overall survival after immune check-point treatment. Using next generation sequencing, we performed microRNA profiling in serum from NSCLC patients (n=20) treated with nivolumab. Serum samples from 31 patients were used for validation using qPCR assays. Serum samples were collected prior to immune therapy initiation. Results: Based on multivariate regression analysis, we identified a signature of seven microRNAs (miR-215-5p, miR-411-3p, miR-493-5p, miR-494-3p, miR-495-3p, miR-548j-5p and miR-93-3p) significantly associated with overall survival (OS)>6 months in discovery cohort (p=.0003). We further validated this in another similar set of samples (n=31) and the model was significantly associated with overall survival (OS)>6 months (p=.001) with sensitivity and specificity of 71% and 90%, respectively. Conclusions: In this study of circulating microRNAs, we have identified a 7-miR signature associated with survival in nivolumab-treated NSCLC patients. This signature may lead to better treatment options for patients with NSCLC, but a validation in an independent cohort is needed to confirm the predicted potential.

Introduction: We aimed to investigate the characteristics of radiation pneumonitis (RP) and lung function after stereotactic body radiotherapy (SBRT) and concomitant radiotherapy (CRT). These are preliminary data from our prospective study. Material and Methods: We have serial measurements up to 12 months after treatment from 32 inoperable patients with non-small cell lung cancer, treated with SBRT (n=24) and CRT (60-66 Gy with platinum-based chemotherapy) (n=8). The patients were followed with spirometry, total lung capacity measurements, clinical COPD Questionnaire (CCQ) and CT scans. Results: 10 patients in the SBRT group (42%) and 5 patients in the CRT group (63%) developed RP grade 2-4. In our cohort, RP grade 2-4 was diagnosed 6 months (mean) after SBRT and 3 months after CRT. In patients who developed RP after SBRT, the lung function varied over time. Some patients experienced improvement in lung function after treatment, whereas some patients had a reduction. By using T-test, both FEV1% and FVC% were significantly reduced at follow-ups after SBRT. In patients who developed RP after CRT, there was interindividual variation in lung function, and by using t-test, DLCO% were significantly reduced during the follow-up time. Patients without RP and with RP grade 1 after SBRT (n=14) had no significant changes in pulmonary function values. No significant changes were observed in CCQ scores. Conclusion: Pneumonitis after SBRT occur later than after CRT. Although a high percentage of the patients receiving curatively intended radiotherapy developed radiation pneumonitis, this was not reflected in the health related quality of life as measured by CCQ.

Purpose: Proteins are the functional players driving both normal and disease physiology. The proteomic changes observed in lung cancer may be a consequence of mutations in essential driver-genes. The purpose of this study was to identify proteins in lung cancer biopsies associated with survival in groups stratified by smoking status, and EGFR-, TP53- and KRAS-mutations. We have performed a profiling of 295 cancer relevant proteins, of which 60 were in a phosphorylated state, using reverse phase protein arrays (RPPA). We analyzed biopsies from 80 lung cancer patients (adenocarcinoma) and correlated the protein expression pattern with progression free survival (PFS) based on mutational- and smoking-status. Spearman correlation analysis revealed a higher number of proteins with significant association to PFS (p<0.05) among the wild type samples compared to the mutated samples. High expression of protein kinase C (PKC) and the isoforms alpha, beta and delta, included the phosporylated state, showed the strongest association with better PFS, especially in the wild type samples. Ten proteins were associated with PFS in never-smokers, where eight of these were unique for this group. Unsupervised hierarchical clustering separated the samples into four subclusters, each enriched with one of the three molecular subtypes TRU, PI and PP (Comprehensive molecular profiling of lung adenocarcinoma, Nature, 2014). Subcluster 2 contained two smaller clusters (2a and 2b) enriched with samples of subtype PP, where subcluster 2a was associated with poor PFS (p=0.003, Figure 1). As of today, we do not have any effective treatment targeting KRAS- and TP53- mutated cells. This study shows that expression of specific proteins and phospho-proteins associated with PFS differ depending on molecular subtype, and mutational- and smoking-status. Proteins associated with PFS may serve as therapeutic targets to circumvent treatment resistance.

PURPOSE:Radiation therapy effectively kills cancer cells and elicits local effects in the irradiated tissue. The aim of this study was to investigate the kinetics of cytokines in the serum of patients with lung cancer undergoing radiation therapy and to identify associations with metabolic tumor burden as determined by 2-deoxy-2-fluoro-D-glucose (18F-FDG) positron emission tomography (PET). METHODS AND MATERIALS:Forty-five patients with advanced non-small cell lung cancer were included in a phase 2 clinical trial and randomized between fractionated thoracic radiation therapy alone or concurrent with an epidermal growth factor receptor inhibitor. Blood was sampled at 4 different time points: prior to treatment, midtherapy, at the end of therapy, and 6 to 8 weeks after the start of treatment. The serum concentrations of 48 cytokines and 9 matrix metalloproteinases were measured with multiplex immunoassays. A subset of patients was examined by 18F-FDG PET/computed tomography before, during, and after radiation therapy. The maximum standardized uptake values (SUVmax) of the primary lung tumor, whole-body metabolic tumor volume, and total lesion glycolysis were calculated, and correlations between the PET parameters and cytokines were investigated. RESULTS:The SUVmax decreased from baseline through midtherapy to posttherapy 18F-FDG PET/computed tomography (P = .018). The serum levels of C-C motif chemokine ligand (CCL) 23, CCL24, C-X3-C motif chemokine ligand 1, and interleukin-8 (C-X-C motif ligand [CXCL]8) were significantly correlated to SUVmax, metabolic tumor volume, and total lesion glycolysis before, during, and after radiation therapy. CXCL2 (P = .030) and CXCL6 (P = .010) decreased after the start of therapy and changed significantly across the sample time points. Serum concentrations of CCL15 (P = .031), CXCL2 (P = .028), and interleukin-6 (P = .007) were positively correlated to the irradiated volume during the second week of treatment. CONCLUSIONS:Cytokine serum levels vary and correlate with metabolic tumor burden in patients with advanced non-small cell lung cancer undergoing palliative thoracic radiation therapy.

BACKGROUND:The development of both chronic obstructive pulmonary disease (COPD) and lung cancer (LC) is influenced by smoking related chronic pulmonary inflammation caused by an excessive innate immune response to smoke exposure. In addition, the smoking induced formation of covalent bonds between the carcinogens and DNA and the accumulation of permanent somatic mutations in critical genes are important in the carcinogenic processes, and can also induce inflammatory responses. How chronic inflammation is mirrored by serum markers in COPD and LC and if these markers reflect prognosis in patients with LC is, however, largely unknown.METHODS:Serum levels of 18 markers reflecting inflammation, endothelial activation and extracellular matrix remodelling were analysed in 207 patients with non-small lung carcinoma (NSCLC) before surgery and 42 COPD patients. 56% of the LC patients also suffered from COPD. The serum samples were analysed by enzyme immunoassays.RESULTS:Serum levels of OPG, PTX3, AXL, ALCAM, sCD163, CD147, CatS and DLL1 were significantly higher in patients with COPD as compared to patients with LC. High sTNFR1 levels were associated with improved progression free survival (PFS) and overall survival (OS) in LC patients with (PFS hazard ratio (HR) 0.49, OS HR 0.33) and without COPD (OS HR 0.30). High levels of OPG were associated with improved PFS (HR 0.17) and OS (HR 0.14) for LC with COPD. CRP was significantly associated with overall survival regardless of COPD status.CONCLUSION:Several markers reflecting inflammation, endothelial activation and extracellular matrix remodelling are elevated in serum from patients with COPD compared to LC patients. Presence of COPD might influence the levels of circulating biomarkers. Some of these markers are also associated with prognosis.

To investigate the tumor microenvironment in NSCLC and try to understand more about how, and why, it differs in histological and expression subtypes of NSCLC . Samples were collected from 399 patients who underwent surgery for stage I-IV NSCLC at Oslo university hospital 2006-16. Gene expression was assessed by using Agilent microarray on samples from adenocarcinomas (n=184) and squamous cell carcinomas (n=183) separately. RNA sequencing was performed of 32 samples. Of these 53 % (n=17) were adenocarcinomas (AD) and 47 % (n=15) were squamous cell carcinomas (SCC). Xcell(1) was used to find the proportion of different cells in the tumor microenvironment and to calculate an immunescore. All samples were assigned a gene expression subtype by using previously described nearest centroid classifiers for AD and SCC respectively (2, 3) For AD we found significant differences in progression free survival (PFS) between the three expression subtypes with better prognosis for TRU versus non-TRU and worse for PP versus non-PP, adjusted for stage. We found no differences in prognosis between SCC expression subtypes. Higher immune score was associated with better PFS in AD but not in SCC. The composition of both stromal and immune cells in the tumor microenvironment will be further investigated. Differential gene expression analysis will be preformed to better understand what differentiates immunologically active from immunologically silent tumors. 1. Aran D, Hu Z, Butte AJ. xCell: digitally portraying the tissue cellular heterogeneity landscape. Genome Biol. 2017;18(1):220. 2. Wilkerson MD, Yin X, Walter V, Zhao N, Cabanski CR, Hayward MC, et al. Differential pathogenesis of lung adenocarcinoma subtypes involving sequence mutations, copy number, chromosomal instability, and methylation. PLoS One. 2012;7(5):e36530. 3. Wilkerson MD, Yin X, Hoadley KA, Liu Y, Hayward MC, Cabanski CR, et al. Lung squamous cell carcinoma mRNA expression subtypes are reproducible, clinically important, and correspond to normal cell types. Clin Cancer Res. 2010;16(19):4864-75.

Chronic inflammation plays an important role in lung carcinogenesis and in chronic obstructive pulmonary disease (COPD), and is accompanied with alterations in specific serum-proteins. Both COPD and lung cancer are associated with smoking behavior, and 40-70% of lung cancer patients have COPD. The aim of the study is to compare levels of specific serum markers related to inflammation, extracellular matrix remodeling (ECM) and endothelial cell activation in patients with lung cancer and COPD. Blood samples were collected from 208 lung cancer patients with stage I-IIIA disease before surgery in addition to blood samples from 47 COPD patients, stage I-IV (4 patients in stage I, 16 in II, 19 in III and 8 in IV). Six of COPD-patients used oral steroids, 28 used inhaled corticosteroids. Serum levels of various markers were measured by enzyme immunoassays. Of 17 proteins (table 1), 9 were significantly elevated in the COPD group compared to lung cancer group including proteins associated with lung cancer in other studies as OPG, PTX3, ePCR, GDF15 and endostatin. Only 3 proteins, CRP, vWF og GDF15 reflecting systemic inflammation and endothelial cell activation, were more abundant in serum from lung cancer patients, and one of these (CRP) significantly so.Table 1Serum proteins measured in our studyProtein short nameProtein full nameOPGOsteoprotegrinePCREndothelial cell protein C receptorvWFVon Willebrand factorPTX3Pentraxin 3AxlTyrosine-protein kinase receptorCXCL16C-X-C motif chemokine ligand 16DLL1Delta-like protein 1CatsCathepsin S (Chloramphenicol acetyl transferase)GDF15Growth differentiation factor-15EndostatinCD147Cluster of differentiation 147 (Basigin. EMMPRIN)sTNFR1Tumor necrosis factor receptor 1CRPC-reactive proteinAlcam (CD166)Activated leukocyte cell adhesion moleculePARCp53-associated parkin-like cytoplasmic proteinsCD163Cluster of differentiation 163Gal3BPGalectin-3-binding protein Open table in a new tab Chronic inflammation plays an important role in both diseases: lung cancer and COPD. However, it seems that inflammation as determined by these selected markers is more pronounced in patients with COPD as most of the biomarkers levels were significantly higher in these patients than lung cancer group.

Yes-associated protein (YAP) is a downstream target of the Hippo pathway and has been found to be oncogenic driving many cancers into developing metastatic phenotypes leading to poor survival outcomes. This study investigated if YAP expression is associated with drug resistance in two non-small cell lung cancer (NSCLC) lines (HCC827 and H1975) generated to become resistant to the EGFR tyrosine kinase inhibitors (EGFR TKI) erlotinib, gefitinib or the T790M-specific osimertinib. We found that acquired EGFR TKI resistance was associated with YAP over-expression (osimertinib-resistant cells) or YAP amplification (erlotinib- and gefitinib-resistant cells) along with EMT phenotypic changes. YAP was localized in the nucleus, indicative of active protein. siRNA-mediated silencing of YAP resulted in re-sensitizing the drug-resistant cells to EGFR TKI compared to the negative siRNA controls (p = <0.05). These results suggest YAP is a potential mechanism of EGFR-TKI resistance in NSCLC and may presents itself as a viable therapeutic target.

Proteins are the functional players driving both normal and disease physiology. The proteomic changes observed in lung cancer can be a consequence of mutations in cancer associated genes. Some of the most common types of mutations in lung cancer are EGFR, TP53 and KRAS mutations, known to influence treatment response. The purpose of this study was to identify proteins associated with smoking status and EGFR, TP53 and KRAS mutations in lung cancer biopsies which may serve as novel targets to overcome treatment resistance. We have performed a profiling of 300 cancer relevant proteins, of which 60 were in a phosphorylated state, using reverse phase protein arrays (RPPA). We analyzed biopsies from 80 non small-cell lung cancer biopsies and correlated the protein expression pattern with mutation status of EGFR, TP53 and KRAS, in addition to smoking status. The protein expression was correlated with mRNA expression analyzed on hybridization arrays. Ten of the samples were EGFR mutated, 9 were never-smokers, 31 TP53 mutated and 32 KRAS mutated. With a FDR < 0.001, we detected 7 proteins (ATP5A, EIF4EBP1, CCNB1, CDK1, CASP3, FN1 and CDKN2A) differentially expressed between TP53 mutated and wild type samples. Further, 4 proteins (RPS6K, CAV1, YBX1 and VIM) were higher expressed in KRAS mutated samples compared with KRAS wild type samples. For EGFR mutated samples, only the protein EGFR was upregulated. Three proteins were overexpressed in never smokers (EGFR, KRT19 and TUBA1A). A significant correlation between protein expression and mRNA expression were found for all the proteins except YBX1, ATP5A and TUBA1A (p<0.05). These results demonstrate that key driver mutations in lung cancer affect several proteins linked to the same pathway. We observed significant upregulation of the TP53 cell cycle target genes CDK1 and CCNB1 in samples with TP53 mutations. KRAS mutated samples showed an overexpression of cavolin 1 (CAV1), an important regulator of cell proliferation and metastasis, and an increased expression of the mesenchymal protein vimentin (VIM). As of today, we do not have any effective treatment targeting KRAS- and TP53- mutated cells. However, proteins associated with the mutation status may be new potential therapeutic targets to circumvent treatment resistance. In the present study, we identified both known and novel proteins associated with TP53 and KRAS mutated samples.

Ovarian cancer patients are recognized with poor prognosis. This study aimed to identify microRNAs in plasma for predicting response to treatment and outcome. We have investigated microRNAs in plasma from ovarian cancer patients enrolled in a large multicenter study (ICON7), investigating the effect of adding bevacizumab to standard chemotherapy in patients diagnosed with epithelial ovarian cancer. Patients with different histology, grade, and FIGO stages were included (n = 207) in this study. Screening of 754 unique microRNAs was performed in the discovery phase (n = 91) using TaqMan Low Density Arrays. The results were validated using single assays and RT-qPCR. Low levels of miR-200b, miR-1274A (tRNALys5), and miR-141 were significantly associated with better survival, confirmed with log-rank test in the validation set. Thelevel of miR-1274A (tRNA(Lys5)) correlated with outcome was especially pronounced in the high-grade serous tumors. Interestingly, low level of miR-200c was associated with 5month prolongation of PFS when treated with bevacizumab compared to standard chemotherapy. We found prognostic significance of miR-200b, miR-141, and miR-1274A (tRNA(Lys5)) in all histological types, where miR-1274A (tRNA(Lys5)) may be a specific marker in high-grade serous tumors. The level of miR-200cmay be predictive of effect of treatment with bevacizumab. However, this needs further validation.