High expression of Cathepsin E in tissues but not blood of patients with Barrett’s esophagus and adenocarcinoma

Background Cathepsin E (CTSE), an aspartic proteinase, is differentially expressed in the metaplasia–dysplasia–neoplasia sequence of gastric and colon cancer. We evaluated CTSE in Barrett’s esophagus (BE) and cancer because increased CTSE levels are linked to improved survival in several cancers, and other cathepsins are up-regulated in BE and esophageal adenocarcinoma (EAC). Methods A total of 273 pretreatment tissues from 199 patients were analyzed [31 normal squamous esophagus (NE), 29 BE intestinal metaplasia, 31 BE with dysplasia (BE/D), 108 EAC]. CTSE relative mRNA expression was measured by real-time polymerase chain reaction, and protein expression was measured by immunohistochemistry. CTSE serum levels were determined by enzyme-linked immunosorbent assay. Results Median CTSE mRNA expression levels were ≥1,000-fold higher in BE/intestinal metaplasia and BE/D compared to NE. CTSE levels were significantly lower in EAC compared to BE/intestinal metaplasia and BE/D, but significantly higher than NE levels. A similar expression pattern was present in immunohistochemistry, with absent staining in NE, intense staining in intestinal metaplasia and dysplasia, and less intense EAC staining. CTSE serum analysis did not discriminate patient groups. In a uni- and multivariable Cox proportional hazards model, CTSE expression was not significantly associated with survival in patients with EAC, although CTSE expression above the 25th percentile was associated with a 41 % relative risk reduction for death (hazard ratio 0.59, 95 % confidence interval 0.27–1.26, p = 0.17). Conclusions CTSE mRNA expression is up-regulated more than any known gene in Barrett intestinal metaplasia and dysplasia tissues. Protein expression is similarly highly intense in intestinal metaplasia and dysplasia tissues

Gene expression alterations in formalin-fixed, paraffin-embedded Barrett's esophagus and esophageal adenocarcinoma tissues

Background and aim: Widespread applicability of tissue-based mRNA expression screening for Barrett's esophagus (BE) is likely to require (1) accurate methods for assaying archival formalin-fixed, paraffin-embedded (FFPE ) histopathology specimens taken at endoscopy, and (2) validation studies of promising biomarkers in different patient cohorts. Results: 30 genes were significantly differentially expressed by histopathology tissue type. The direction and magnitude of the differences were very similar to those found in previous studies using fresh frozen tissues. Novel upregulated genes were TSPA N8 (also known as CO-029), TSPA N24 (CD151), EGR1 and TCIRG1. Novel downregulated genes were DPYD, TSPA N29 (CD9) and Ets1. Strong associations between histopathology type and gene expression were observed with the overexpressed genes: cyclo-oxygenase-2 (COX-2), for which histopathology type explained 77.7% of the variation in expression, TSPA N1 (73.5%), TSPA N8 (62.9%), SPA RC (62.1%), MMP7 (50.8%); and the under-expressed genes ADH7 (53.7%), AP C (58.2%), RAR-gamma (51.3%). Methods: mRNA was isolated from 54 FFPE small endoscopic biopsies from patients with Barrett's intestinal metaplasia (BE), esophageal adenocarcinoma (EA C), or control patients with a normal squamous-lined esophagus. Multiplexed tandem PCR (MT-PCR) was used to quantitate 50 selected candidate genes for BE and control genes in duplicate. Principal component analysis (PCA) was conducted to explore the presence of global differences in gene expression profiles. Oneway analysis of variance (ANOVA) of the transformed data was used to identify genes that were differentially expressed between different histological subtypes. Differentially expressed genes with a fold change of ≥2 (upregulated) or ≤-2 (downregulated) are reported with the p value for each comparison (EA C vs. normal, EA C vs. BE and BE vs. normal). The Benjamini-Hochberg method was used to control the false discovery rate at 0.01 for all comparisons. Conclusions: Alterations in expression of select genes are strongly associated with BE or EA C or both. This study's findings for many highly differentially expressed genes are very similar to those previously reported, suggesting that these genes should be tested further in longitudinal studies for their potential role as biomarkers of progression to more advanced Barrett's disease

Anti-inflammatory and anti-remodelling effects of ISU201, a modified form of the extracellular domain of human BST2, in experimental models of asthma: association with inhibition of histone acetylation.

There are few alternatives to glucocorticosteroids for treatment of asthma. We assessed the activity of a novel protein drug designated ISU201, the extracellular domain of the human cell surface protein BST2, stabilised by fusion with the Fc region of IgG, in mouse models of mild chronic asthma and an acute exacerbation of asthma. The ability of ISU201 to suppress airway inflammation and remodelling was compared with that of dexamethasone. Female BALB/c mice were systemically sensitised with ovalbumin, then received controlled low-level challenge with aerosolised ovalbumin for 6 weeks, which induced lesions of mild chronic asthma, and were treated with drugs during the final 2 weeks. Alternatively, sensitised mice received 4 weeks of chronic low-level challenge and were treated 24 and 2 hours before a final single moderate-level challenge, which triggered acute airway inflammation simulating an asthmatic exacerbation. Inflammation and remodelling were quantified, as was the expression of pro-inflammatory cytokines in bronchoalveolar lavage fluid and tissues. To identify cellular targets of ISU201, we assessed the effects of the drug on activated lymphocytes, macrophages and airway epithelial cells. In the model of mild chronic asthma, ISU201 was as effective as dexamethasone in suppressing airway inflammation and most changes of remodelling. In the model of an allergen-induced acute exacerbation of chronic asthma, ISU201 was also an effective anti-inflammatory agent, although it was less active than dexamethasone. The drug acted on multiple cellular targets, suppressing production of pro-inflammatory cytokines by lymphocytes and macrophages. ISU201 significantly reduced acetylation of histone H4 in airway epithelial cells, suggesting at least one potential mechanism of action. We conclude that in these models of asthma, ISU201 is a broad-spectrum inhibitor of both airway inflammation and remodelling. Thus, unlike drugs which target specific mediators, it could potentially be an alternative or an adjunct to glucocorticoids for the treatment of asthma

Immunoreactivity for acetylated histone H4 in nuclei of airway epithelial cells.

<p>(A) Photomicrograph of immunopositive nuclei (brown) in epithelial cells lining the trachea, original magnification ×400. (B) Percent positive cells in the trachea in the model of an allergen-induced acute exacerbation of chronic asthma. OVA-exposed animals treated with vehicle alone are compared to unexposed animals or to animals treated with ISU201 or dexamethasone. Data are mean ± SEM (<i>n</i> = 6 samples per group). Significant differences relative to the naïve group are shown as **(p<0.01); relative to the vehicle-treated group are shown as #(p<0.05).</p

Effects of drug treatment on expression of cytokine mRNA by AM from the acute exacerbation model.

<p>Values are fold expression relative to naïve animals, shown as mean ± SEM (<i>n</i> = 6). Significant differences compared to naïve animals are shown as ***(p<0.001); compared to the vehicle-treated group as ^#(p<0.05), ^##(p<0.01) and ^###(p<0.001).</p

Effects of drug treatment on cytokine concentrations in BAL fluid in the acute exacerbation model.

<p>Values are pg/mL, shown as mean ± SEM (<i>n</i> = 8 animals per group). Significant differences compared to naïve animals are shown as *(p<0.05), **(p<0.01) and ***(p<0.001); compared to the vehicle-treated group as ^#(p<0.05), ^##(p<0.01) and ^###(p<0.001).</p

Effects of drug treatment in vitro on cytokine concentrations in supernatants of restimulated PBLN cells.

<p>Values are pg/mL, shown as mean ± SEM (<i>n</i> = 4 samples per group, pooled pairs from 8 animals). Significant differences relative to cells cultured in medium only are shown as * (p<0.05), ** (p<0.01) and *** (p<0.001); relative to cells restimulated with OVA are shown as ^#(p<0.05) and ^##(p<0.01).</p

Effects of drug treatment on expression of cytokine mRNA in airway tissue in the chronic challenge model.

<p>Values are fold expression relative to naïve animals, shown as mean ± SEM (<i>n</i> = 8). Significant differences compared to naïve animals are shown as *(p<0.05), **(p<0.01) and ***(p<0.001); compared to the vehicle-treated group as ^#(p<0.05), ^##(p<0.01) and ^###(p<0.001).</p