Nm23/Nucleoside Diphosphate Kinase-A as a Potent Prognostic marker in Invasive Pancreatic Ductal Carcinoma Identified by Proteomic Analysis of Laser Micro-dissected Formalin-Fixed Paraffin-Embedded Tissue

Background

Pancreatic cancer is among the most lethal malignancies worldwide. This study aimed to identify a novel prognostic biomarker, facilitating treatment selection, using mass spectrometry (MS)-based proteomic analysis with formalin-fixed paraffin-embedded (FFPE) tissue.

Results

The two groups with poor prognosis (n = 4) and with better prognosis (n = 4) had been carefully chosen among 96 resected cases of pancreatic cancer during 1998 to 2007 in Tohoku University Hospital. Although those 2 groups had adjusted background (UICC-Stage IIB, Grade2, R0, gemcitabine adjuvant), there was a significant difference in postoperative mean survival time (poor 21.0 months, better 58.1 months, P = 0.0067). Cancerous epithelial cells collected from FFPE tissue sections by laser micro-dissection (LMD) were processed for liquid chromatography-tandem mass spectrometry (LC-MS/MS). In total, 1099 unique proteins were identified and 6 proteins showed different expressions in the 2 groups by semi-quantitative comparison. Among these 6 proteins, we focused on Nm23/Nucleoside Diphosphate Kinase A (NDPK-A) and immunohistochemically confirmed its expression in the cohort of 96 cases. Kaplan-Meier analysis showed high Nm23/NDPK-A expression to correlate with significantly worse overall survival (P = 0.0103). Moreover, in the multivariate Cox regression model, Nm23/NDPK-A over-expression remained an independent predictor of poor survival with a hazard ratio of 1.97 (95% CI 1.16-3.56, P = 0.0110).

Conclusions

We identified 6 candidate prognostic markers for postoperative pancreatic cancer using FFPE tissues and immunohistochemically demonstrated high Nm23/NDPK-A expression to be a useful prognostic marker for pancreatic cancer.     

Selected Reaction Monitoring (SRM) Analysis of Epidermal Growth Factor Receptor (EGFR) in Formalin Fixed Tumor Tissue

Background

Analysis of key therapeutic targets such as epidermal growth factor receptor (EGFR) in clinical tissue samples is typically done by immunohistochemistry (IHC) and is only subjectively quantitative through a narrow dynamic range. The development of a standardized, highly-sensitive, linear, and quantitative assay for EGFR for use in patient tumor tissue carries high potential for identifying those patients most likely to benefit from EGFR-targeted therapies.

Methods

A mass spectrometry-based Selected Reaction Monitoring (SRM) assay for the EGFR protein (EGFR-SRM) was developed utilizing the Liquid Tissue®-SRM technology platform. Tissue culture cells (n = 4) were analyzed by enzyme-linked immunosorbent assay (ELISA) to establish quantitative EGFR levels. Matching formalin fixed cultures were analyzed by the EGFR-SRM assay and benchmarked against immunoassay of the non-fixed cultured cells. Xenograft human tumor tissue (n = 10) of non-small cell lung cancer (NSCLC) origin and NSCLC patient tumor tissue samples (n = 23) were microdissected and the EGFR-SRM assay performed on Liquid Tissue lysates prepared from microdissected tissue. Quantitative curves and linear regression curves for correlation between immunoassay and SRM methodology were developed in Excel.

Results

The assay was developed for quantitation of a single EGFR tryptic peptide for use in FFPE patient tissue with absolute specificity to uniquely distinguish EGFR from all other proteins including the receptor tyrosine kinases, IGF-1R, cMet, Her2, Her3, and Her4. The assay was analytically validated against a collection of tissue culture cell lines where SRM analysis of the formalin fixed cells accurately reflects EGFR protein levels in matching non-formalin fixed cultures as established by ELISA sandwich immunoassay (R² = 0.9991). The SRM assay was applied to a collection of FFPE NSCLC xenograft tumors where SRM data range from 305amol/μg to 12,860amol/μg and are consistent with EGFR protein levels in these tumors as previously-reported by western blot and SRM analysis of the matched frozen tissue. In addition, the SRM assay was applied to a collection of histologically-characterized FFPE NSCLC patient tumor tissue where EGFR levels were quantitated from not detected (ND) to 670amol/μg.

Conclusions

This report describes and evaluates the performance of a robust and reproducible SRM assay designed for measuring EGFR directly in FFPE patient tumor tissue with accuracy at extremely low (attomolar) levels. This assay can be used as part of a complementary or companion diagnostic strategy to support novel therapies currently under development and demonstrates the potential to identify candidates for EGFR-inhibitor therapy, predict treatment outcome, and reveal mechanisms of therapeutic resistance.     

The CIN4 Chromosomal Instability qPCR Classifier Defines Tumor Aneuploidy and Stratifies Outcome in Grade 2 Breast Cancer

Purpose

Quantifying chromosomal instability (CIN) has both prognostic and predictive clinical utility in breast cancer. In order to establish a robust and clinically applicable gene expression-based measure of CIN, we assessed the ability of four qPCR quantified genes selected from the 70-gene Chromosomal Instability (CIN70) expression signature to stratify outcome in patients with grade 2 breast cancer.

Methods

AURKA, FOXM1, TOP2A and TPX2 (CIN4), were selected from the CIN70 signature due to their high level of correlation with histological grade and mean CIN70 signature expression in silico. We assessed the ability of CIN4 to stratify outcome in an independent cohort of patients diagnosed between 1999 and 2002. 185 formalin-fixed, paraffin-embedded (FFPE) samples were included in the qPCR measurement of CIN4 expression. In parallel, ploidy status of tumors was assessed by flow cytometry. We investigated whether the categorical CIN4 score derived from the CIN4 signature was correlated with recurrence-free survival (RFS) and ploidy status in this cohort.

Results

We observed a significant association of tumor proliferation, defined by Ki67 and mitotic index (MI), with both CIN4 expression and aneuploidy. The CIN4 score stratified grade 2 carcinomas into good and poor prognostic cohorts (mean RFS: 83.8±4.9 and 69.4±8.2 months, respectively, p = 0.016) and its predictive power was confirmed by multivariate analysis outperforming MI and Ki67 expression.

Conclusions

The first clinically applicable qPCR derived measure of tumor aneuploidy from FFPE tissue, stratifies grade 2 tumors into good and poor prognosis groups.     

Circulating levels of copeptin predict outcome in patients with pulmonary arterial hypertension

Objective

To determine the levels of circulating copeptin in patients with pulmonary arterial hypertension (PAH), and to evaluate its relation with disease severity, outcome and response to treatment.

Background

Vasopressin is a key regulator of body fluid homeostasis. The co-secreted protein copeptin serves as surrogate for plasma vasopressin levels and increases in acute and chronic left ventricular dysfunction. Copeptin has not been studied in PAH.

Methods

Serum copeptin levels were evaluated in a retrospective cohort of 92 treatment-naïve patients with PAH, 39 patients with normal right ventricular hemodynamics (diseased controls) and 14 apparently healthy individuals (healthy controls). In a second prospective cohort of 15 patients with PAH, serial changes of copeptin levels after initiation of PAH treatment were measured. Copeptin levels were compared with clinical, biochemical and hemodynamic parameters as well as response to treatment and clinical outcome.

Results

Circulating copeptin levels were elevated in PAH patients compared to diseased controls (20.1 pmol/l vs. 5.1 pmol/l; p = 0.001). Baseline levels of copeptin correlated with NYHA functional class (r = 0.46; p = 0.01), 6 minute walking distance (r = -0.26; p = 0.04), NT-proBNP (r = 0.49, p = 0.01), creatinine (r = 0.39, p = 0.01) and estimated glomerular filtration rate (r = -0.32, p = 0.01). Copeptin levels did not correlate with hemodynamics but decreased after initiation of PAH therapy (p = 0.001). Elevated copeptin levels were associated with shorter survival (p < 0.001) and independent predictors of mortality in a multiple Cox regression analysis (HR1.4; 95% confidence interval 1.1-2.0; p = 0.02).

Conclusions

Patients with PAH had elevated copeptin levels. High circulating levels of copeptin were independent predictors of poor outcome, which makes copeptin a potentially useful biomarker in PAH.     

Increased human Ca2+-activated Cl- channel 1 expression and mucus overproduction in airway epithelia of smokers and chronic obstructive pulmonary disease patients

Background

The mechanisms underlying the association between smoking and mucus overproduction remain unknown. Because of its involvement in other airway diseases, such as asthma, we hypothesized that Ca²⁺-activated Cl^- channel 1 (CLCA1) was associated with overproduction of mucus in the airways of smokers and COPD patients.

Methods

Using real-time quantitative PCR analyses, we compared the CLCA1 mRNA expression levels in induced-sputum cells from COPD patients (n = 20), smokers without COPD (n = 5), and non-smokers (n =13). We also examined the relationship between CLCA1 protein expression and mucus production in lung airway epithelia of COPD patients (n = 6), smokers without COPD (n = 7), and non-smokers (n = 7).

Results

CLCA1 mRNA expression was significantly up-regulated in the induced-sputum cells of COPD patients compared with cells of non-smokers (p = 0.02), but there was no significant difference compared with cells of smokers without COPD. Using immunostaining with an anti-CLCA1 antibody, semi-quantitative image analyses of airway epithelium demonstrated significantly increased CLCA1 expression in smokers without COPD (p = 0.02) and in COPD patients (p = 0.002) compared with non-smokers. There were significant negative correlations between CLCA1 protein expression and FEV₁/FVC (r = −0.57, p = 0.01) and %predicted FEV₁ (r = −0.56, p = 0.01). PAS staining for mucus showed that there was a significant positive correlation between CLCA1 protein expression and mucus production (r = 0.67, p = 0.001). These markers were significantly increased in smokers without COPD (p = 0.04) and in COPD patients (p = 0.003) compared with non-smokers (non-smokers < smokers ≤ COPD).

Conclusions

CLCA1 expression is significantly related to mucus production in the airway epithelia of smokers and COPD patients, and may contribute to the development and pathogenesis of COPD by inducing mucus production.     

The relationship between lateral meniscus shape and joint contact parameters in the knee: a study using data from the Osteoarthritis Initiative

Introduction

The meniscus has an important role in force transmission across the knee, but a detailed three-dimensional (3D) morphometric shape analysis of the lateral meniscus to elucidate subject-specific function has not been conducted. The aim of this study was to perform 3D morphometric analyses of the lateral meniscus in order to correlate shape variables with anthropometric parameters, thereby gaining a better understanding of the relationship between lateral meniscus shape and its load-bearing function.

Methods

The lateral meniscus (LM) was manually segmented from magnetic resonance images randomly selected from the Osteoarthritis Initiative (OAI) non-exposed control subcohort. A 3D statistical shape model (SSM) was constructed to extract the principal morphological variations (PMV) of the lateral meniscus for 50 subjects (25 male and 25 female). Correlations between the principal morphological variations and anthropometric parameters were tested. Anthropometric parameters that were selected included height, weight, body mass index (BMI), femoral condyle width and axial rotation.

Results

The first principal morphological variation (PMV) was found to correlate with height (r = 0.569), weight (r = 0.647), BMI (r = 0.376), and femoral condyle width (r = 0.622). The third PMV was found to correlate with height (r = 0.406), weight (r = 0.312), and femoral condyle width (r = 0.331). The percentage of the tibial plateau covered by the lateral meniscus decreases as anthropometric parameters relating to size of the subject increase. Furthermore, when the size of the subject increases, the posterior and anterior horns become proportionally longer and wider.

Conclusion

The correlations discovered suggest that variations in meniscal shape can be at least partially explained by the levels of loads transmitted across the knee on a regular basis. Additionally, as the size of the subject increases and body weight rises, the coverage percentage of the meniscus is reduced, suggesting that there would be an increase in the load-bearing by the cartilage. However, this reduced coverage percentage is compensated by the proportionally wider and longer meniscal horn.     

Potential role of the lectin pathway of complement in the pathogenesis and disease manifestations of systemic sclerosis: a case-control and cohort study

Introduction

Repetitive episodes of ischemia and reperfusion (I/R) are a cardinal feature of the pathogenesis of systemic sclerosis (SSc), which precedes tissue fibrosis. The complement system is a key mediator of tissue damage after I/R, primarily by activation of the lectin pathway. This study investigated whether serum levels and polymorphisms of mannose-binding lectin (MBL) and ficolin-2 (FCN2), two pattern recognition receptors of the lectin pathway, are associated with the predisposition to and clinical features of SSc.

Methods

A case-control study was undertaken involving 90 patients with SSc from a single SSc outpatient clinic and 90 age- and sex-matched blood donors. MBL and FCN2 levels and polymorphisms were measured in both groups, and in cases correlated with clinical data.

Results

MBL levels and genotypes were equally distributed in cases and controls while there were some significant differences in FCN2 polymorphisms. Median MBL levels were higher in SSc cases with diffuse disease compared with controls (2.6 versus 1.0 μg/ml, P <0.001).In cases, higher MBL levels were associated with the presence of clinical findings associated with vascular dysfunction and local tissue damage (digital ulcers, calcinosis and pitting). Moreover, MBL levels were associated with fibrotic disease manifestations as evidenced by the presence of diffuse disease (median 2.6 versus 0.8 μg/ml, P = 0.002), the modified Rodnan skin score (r = 0.39, P <0.001), and interstitial lung disease as measured by forced vital capacity (r = −0.33, P = 0.001). Importantly, MBL levels also correlated with the Scleroderma Health Assessment Questionnaire scores (r = 0.33, P = 0.002). The results for FCN2 levels were less striking. Phenotypic MBL results were largely confirmed by analysis of MBL polymorphisms. MBL levels were not associated with the presence of autoantibodies or hypocomplementaemia.

Conclusions

Overall, predisposition to SSc was not influenced by the lectin pathway of complement in our matched case-control study. However, our preliminary data suggest that MBL, and to a lesser extent FCN2, may modulate disease manifestations of SSc, particularly in diffuse cutaneous disease.

Electronic supplementary material

The online version of this article (doi:10.1186/s13075-014-0480-6) contains supplementary material, which is available to authorized users.     

Association between individual quadriceps muscle volume/enthesis and patello femoral joint cartilage morphology

Introduction

The aim of this study was to determine the association between individual quadriceps muscle volumes and the quadriceps enthesis structures and cartilage morphology at the patellofemoral joint (PFJ).

Methods

We studied 12 cadavers (age 75 ± 5 years). For both legs, individual quadriceps muscles (vastus lateralis (VL), rectus femoris (RF), vastus intermedialis (VI) and vastus medialis (VM)) were dissected and their volumes measured. Cartilage areas at the PFJ were classified using the International Cartilage Repair Society (ICRS) score. Histological sections were evaluated at the quadriceps tendon enthesis (laterally, centrally and medially). Several variables were calculated on the binary images based on two-dimensional analysis. These were apparent bone area (BA) and apparent trabecular thickness (TH). A Spearman rank test was used to determine the strength of correlation between individual quadriceps muscles volume, the structure of the quadriceps tendon enthesis and the ICRS score.

Results

The thickness of calcified fibrocartilage tissue was significantly greater in the central part of the enthesis than both medially (P = 0.03) and laterally (P = 0.04). Uncalcified fibrocartilage was significantly thicker laterally (P = 0.04) and centrally (P = 0.02) than medially. Muscle volume was highest (P <0.05) for the VL, followed by the VI, VM and RF. There was no association between total and individual muscle volumes and ICRS or BA. However, there was a strong positive correlation (r = 0.81) between the VL/VM volume ratio and BA ratio (bone volume at the lateral part divided by bone volume at the medial part). There was a moderate positive correlation between VL/VM and ICRS (r = 0.65) and between ICRS and BA ratio (lateral/medial; r = 0.74).

Conclusions

Individual and total quadriceps volumes were not correlated with cartilage loss at the PFJ or fibrocartilage thickness. However, both VL/VM and BA ratio (lateral/medial) were positively correlated with ICRS scoring and therefore could be a tool for predicting degree of PFJ osteoarthritis severity.     

Efficacy and safety of once-daily fluticasone furoate 50 mcg in adults with persistent asthma: a 12-week randomized trial

Background

Fluticasone furoate (FF) is a novel, once-daily inhaled corticosteroid (ICS) that has been shown to improve lung function vs. placebo in asthma patients. This study evaluated the efficacy and safety of FF 50 mcg compared with placebo in asthma patients uncontrolled by non-ICS therapy.

Methods

This 12-week, multicentre, randomized, double-blind, placebo-controlled, parallel-group, phase III study randomized 248 patients (aged ≥12 years) to once-daily FF 50 mcg administered via the ELLIPTA™^a dry powder inhaler or placebo. The primary endpoint was change from baseline in pre-dose evening trough forced expiratory volume in one second (FEV₁). Secondary endpoints were change from baseline in percentage of rescue-free 24-h periods (powered), evening and morning peak expiratory flow, symptom-free 24-h periods and withdrawals due to lack of efficacy. Other endpoints included Asthma Control Test™, Asthma Quality of Life Questionnaire and ELLIPTA ease of use questions. Safety was assessed throughout the study.

Results

There was a significant difference in evening trough FEV₁ between FF 50 mcg and placebo (treatment difference: 120 mL; p = 0.012). There was also a significant difference in rescue-free 24-h periods (11.6%; p = 0.004) vs. placebo. There were numerically greater improvements with FF vs. placebo for all remaining secondary endpoints. The incidence of adverse events was lower with FF (31%) than with placebo (38%); few were treatment-related (FF 50 mcg: n = 1, <1%; placebo: n = 4, 3%).

Conclusion

FF 50 mcg once daily significantly improved FEV₁ and percentage of rescue-free 24-h periods experienced over 12 weeks vs. placebo, and was well tolerated.

Trial registration

www.clinicaltrials.gov, registration number: {"type":"clinical-trial","attrs":{"text":"NCT01436071","term_id":"NCT01436071"}}NCT01436071     

Study of 320-Slice Dynamic Volume CT Perfusion in Different Pathologic Types of Kidney Tumor: Preliminary Results

Objective

To investigate microcirculatory differences between pathologic types of kidney tumor using 320-slice dynamic volume CT perfusion.

Methods

Perfusion imaging with 320-slice dynamic volume CT was prospectively performed in 85 patients with pathologically proven clear cell renal cell carcinoma (RCC) (n = 66), papillary RCC (n = 7), chromophobe RCC (n = 5), angiomyolipoma (AML) with minimal fat (n = 7), or RCC (n = 78). Equivalent blood volume (Equiv BV), permeability surface-area product (PS; clearance/unit volume = permeability), and blood flow (BF) of tumor and normal renal cortex were measured and analyzed. Effective radiation dose was calculated.

Results

There was a significant difference in all three parameters between tumor and normal renal cortex (P<0.001). Equiv BV was significantly different between RCC and AML with minimal fat (P = 0.038) and between clear cell RCC and AML with minimal fat (P<0.001). Mean Equiv BV and BF were significantly higher in clear cell RCC than in papillary RCC (P<0.001 for both) and mean Equiv BV was higher in clear cell RCC than in chromophobe RCC (P<0.001). The effective radiation dose of the CT perfusion protocol was 18.5 mSv.

Conclusion

Perfusion imaging using 320-slice dynamic volume CT can be used to evaluate hemodynamic features of the whole kidney and kidney tumors, which may be useful in the differential diagnosis of these four pathologic types of kidney tumor.     

Increased levels of soluble interleukin-6 receptor and CCL3 in COPD sputum

Background

COPD patients have increased numbers of macrophages and neutrophils in the lungs. Interleukin-6 (IL-6) trans-signaling via its soluble receptor sIL-6R, governs the influx of innate immune cells to inflammatory foci through regulation of the chemokine CCL3. We hypothesized that there would be enhanced levels of IL-6, sIL-6R and CCL3 in COPD sputum.

Methods

59 COPD patients, 15 HNS and 15 S underwent sputum induction and processing with phosphate buffered saline to obtain supernatants for IL-6, sIL-6R and CCL3 analysis. Cytoslides were produced for differential cell counting and immunocytochemistry (COPD; n = 3) to determine cell type surface expression of the CCL3 receptors CCR5 and CCR1.

Results

COPD patients expressed higher levels (p < 0.05) of sIL-6R and CCL3 compared to controls (sIL-6R medians pg/ml: COPD 166.4 vs S 101.1 vs HNS 96.4; CCL3 medians pg/ml: COPD 117.9 vs S 0 vs HNS 2.7). COPD sIL-6R levels were significantly correlated with sputum neutrophil (r = 0.5, p < 0.0001) and macrophage (r = 0.3, p = 0.01) counts. Immunocytochemical analysis revealed that CCR5 and CCR1 were exclusively expressed on airway macrophages.

Conclusion

Enhanced airway generation of sIL-6R may promote IL-6 trans-signaling in COPD. Associated upregulation of CCL3 may facilitate the recruitment of macrophages into the airways by ligation of CCR1 and CCR5.

Electronic supplementary material

The online version of this article (doi:10.1186/s12931-014-0103-4) contains supplementary material, which is available to authorized users.     

Gene expression profiles from formalin fixed paraffin embedded breast cancer tissue are largely comparable to fresh frozen matched tissue

Background and Methods

Formalin Fixed Paraffin Embedded (FFPE) samples represent a valuable resource for cancer research. However, the discovery and development of new cancer biomarkers often requires fresh frozen (FF) samples. Recently, the Whole Genome (WG) DASL (cDNA-mediated Annealing, Selection, extension and Ligation) assay was specifically developed to profile FFPE tissue. However, a thorough comparison of data generated from FFPE RNA and Fresh Frozen (FF) RNA using this platform is lacking. To this end we profiled, in duplicate, 20 FFPE tissues and 20 matched FF tissues and evaluated the concordance of the DASL results from FFPE and matched FF material.

Methodology and Principal Findings

We show that after proper normalization, all FFPE and FF pairs exhibit a high level of similarity (Pearson correlation >0.7), significantly larger than the similarity between non-paired samples. Interestingly, the probes showing the highest correlation had a higher percentage G/C content and were enriched for cell cycle genes. Predictions of gene expression signatures developed on frozen material (Intrinsic subtype, Genomic Grade Index, 70 gene signature) showed a high level of concordance between FFPE and FF matched pairs. Interestingly, predictions based on a 60 gene DASL list (best match with the 70 gene signature) showed very high concordance with the MammaPrint® results.

Conclusions and Significance

We demonstrate that data generated from FFPE material with the DASL assay, if properly processed, are comparable to data extracted from the FF counterpart. Specifically, gene expression profiles for a known set of prognostic genes for a specific disease are highly comparable between two conditions. This opens up the possibility of using both FFPE and FF material in gene expressions analyses, leading to a vast increase in the potential resources available for cancer research.     

Impact of exacerbations of cystic fibrosis on muscle strength

Background

Adult patients with cystic fibrosis have peripheral muscle weakness, which is related to exercise intolerance and poor prognosis. The influence of acute exacerbations on muscle strength has been poorly studied. This study aimed to investigate whether quadriceps force (QF), as assessed with an involuntary technique, changes during intravenous antibiotics therapy (IVAT) for an exacerbation.

Methods

QF was measured in 20 patients using twitch stimulation of the femoral nerve at the day of hospitalization (day 1) and at termination (day 14) of the IVAT. Physical activity was monitored during IVAT using a SenseWear armband. Ten stable patients served as control subjects.

Results

QF did not change during exacerbation (potentiated twitch force at day 1: 140 ± 42 N, at day 14: 140 ± 47 N), but a decrease was observed in individual patients. Changes in twitch force during exacerbation were correlated with time spent in activities of at least moderate intensity (r = 0.61, p = 0.007).

Conclusions

QF does not systematically decrease during exacerbations of cystic fibrosis. Individual changes in QF are well correlated with daily time spent in activities of at least moderate intensity.     

TNF-α is associated with loss of lean body mass only in already cachectic COPD patients

Background

Systemic inflammation may contribute to cachexia in patients with chronic obstructive pulmonary disease (COPD). In this longitudinal study we assessed the association between circulating C-reactive protein (CRP), tumor necrosis factor (TNF)-α, interleukin (IL)-1ß, and IL-6 levels and subsequent loss of fat free mass and fat mass in more than 400 COPD patients over three years.

Methods

The patients, aged 40–76, GOLD stage II-IV, were enrolled in 2006/07, and followed annually. Fat free mass and fat mass indexes (FFMI & FMI) were calculated using bioelectrical impedance, and CRP, TNF-α, IL-1ß, and IL-6 were measured using enzyme immunoassays. Associations with mean change in FFMI and FMI of the four inflammatory plasma markers, sex, age, smoking, FEV₁, inhaled steroids, arterial hypoxemia, and Charlson comorbidity score were analyzed with linear mixed models.

Results

At baseline, only CRP was significantly (but weakly) associated with FFMI (r = 0.18, p < 0.01) and FMI (r = 0.27, p < 0.01). Univariately, higher age, lower FEV₁, and use of beta2-agonists were the only significant predictors of decline in FFMI, whereas smoking, hypoxemia, Charlson score, and use of inhaled steroids predicted increased loss in FMI. Multivariately, high levels of TNF-α (but not CRP, IL-1ß or IL-6) significantly predicted loss of FFMI, however only in patients with established cachexia at entry.

Conclusion

This study does not support the hypothesis that systemic inflammation is the cause of accelerated loss of fat free mass in COPD patients, but suggests a role for TNF-α in already cachectic COPD patients.     

Blood-based lung cancer biomarkers identified through proteomic discovery in cancer tissues,cell lines and conditioned medium

Background

Support for early detection of lung cancer has emerged from the National Lung Screening Trial (NLST), in which low-dose computed tomography (LDCT) screening reduced lung cancer mortality by 20 % relative to chest x-ray. The US Preventive Services Task Force (USPSTF) recently recommended annual screening for the high-risk population, concluding that the benefits (life years gained) outweighed harms (false positive findings, abortive biopsy/surgery, radiation exposure). In making their recommendation, the USPSTF noted that the moderate net benefit of screening was dependent on the resolution of most false-positive results without invasive procedures. Circulating biomarkers may serve as a valuable adjunctive tool to imaging.

Results

We developed a broad-based proteomics discovery program, integrating liquid chromatography/mass spectrometry (LC/MS) analyses of freshly resected lung tumor specimens (n = 13), lung cancer cell lines (n = 17), and conditioned media collected from tumor cell lines (n = 7). To enrich for biomarkers likely to be found at elevated levels in the peripheral circulation of lung cancer patients, proteins were prioritized based on predicted subcellular localization (secreted, cell-membrane associated) and differential expression in disease samples. 179 candidate biomarkers were identified. Several markers selected for further validation showed elevated levels in serum collected from subjects with stage I NSCLC (n = 94), relative to healthy smoker controls (n = 189). An 8-marker model was developed (TFPI, MDK, OPN, MMP2, TIMP1, CEA, CYFRA 21–1, SCC) which accurately distinguished subjects with lung cancer (n = 50) from high risk smokers (n = 50) in an independent validation study (AUC = 0.775).

Conclusions

Integrating biomarker discovery from multiple sample types (fresh tissue, cell lines and conditioned medium) has resulted in a diverse repertoire of candidate biomarkers. This unique collection of biomarkers may have clinical utility in lung cancer detection and diagnoses.

Electronic supplementary material

The online version of this article (doi:10.1186/s12014-015-9090-9) contains supplementary material, which is available to authorized users.     

Target renal damage: the microvascular associations of increased aortic stiffness in patients with COPD

Background

Although renal impairment has been described in COPD, there is opportunity to evaluate further to determine nature and consider optimal management. Increased aortic stiffness, as seen in COPD, leads to reduced buffering of pulsatile flow. We hypothesised that urinary albumin creatinine ratio (UACR) would reflect glomerular damage related to aortic stiffness.

Methods

Patients with COPD and controls underwent spirometry, blood pressure, arterial stiffness - aortic pulse wave velocity (PWV) and provided a spot urine sample for UACR, with other renal biomarkers measured.

Results

The UACR was increased in patients (n = 52): 0.80 mg/mmol compared to controls (n = 34): 0.46 mg/mmol, p < 0.05. Aortic PWV was related to log₁₀ UACR in all subjects (r = 0.426, p < 0.001) and COPD patients alone. Aortic PWV was a significant variable for UACR with oxygen saturations, after accounting for potential confounders. Eight subjects (7 patients) reached a defined clinical microalbuminuria threshold, with aortic PWV greater in these patients compared to those patients without, although albuminuria is a continuum. Proximal tubular damage biomarkers, unlike the glomerular marker, were not different between patients and controls.

Conclusions

There is glomerular damage in patients with COPD evidenced by increased UACR, related to increased aortic stiffness. Besides the macrovascular prognostic implications of increased aortic stiffness, the microvascular state in COPD management should be considered.