We hypothesize that a cell-type-specific absence of C/EBPalpha is responsible for the enhanced proliferation of bronchial smooth-muscle cells derived from subjects with asthma and that it explains the failure of glucocorticoids to inhibit proliferation in vitro.

Somatic STAT3 mutations present in a subset of inflammatory hepatocellular adenomas result in the generation of constitutively active STAT3 proteins that homodimerize independently of IL-6 stimulation.

Transforming growth factor (TGF)-␤1 induces extracellular matrix deposition and proliferation of mesenchymal cells. We recently reported that interleukin (IL)-6 is an essential mediator of growth factor-induced proliferation of lung fibroblasts. Here, we demonstrate by reverse transcriptase polymerase chain reaction and enzyme-linked immunoassay that TGF-␤1 is a potent inducer of IL-6 mRNA and protein in primary human lung fibroblasts. Transient transfections of fibroblasts with a luciferase reporter gene construct containing nucleotides ؊651 to ؉1 of the human IL-6 promoter revealed that TGF-␤1 also potently activated IL-6 promoter activity. Progressive 5-deletions and sitedirected mutagenesis of the parental construct located the TGF-␤1-responsive cis-regulatory element to a known activating protein-1 (AP-1) sequence (nucleotides ؊284 to ؊276). Gel shift analyses revealed that AP-1 DNA binding activity in nuclear extracts was increased 30 min after stimulation with TGF-␤1. In contrast, neither CCAAT enhancer-binding protein-␤, NF-B, nor Sp1 were activated by TGF-␤1. Supershift analyses demonstrated that the AP-1 complex induced by TGF-␤1 was composed of Jun isoforms and absent of Fos isoforms. Moreover, this complex was found to be a JunD homodimer. Our data thus demonstrate that TGF-␤1 is a potent inducer of IL-6 in primary human lung fibroblasts. The TGF-␤1-activated JunD homodimer may be essential for a majority of the biological effects induced by TGF-␤1 in this cell type, such as proliferation and extracellular matrix synthesis.

Background: A small subset (10-15%) of gastrointestinal stromal tumours (GISTs) lack mutations in KIT and PDGFRA (wild-type GIST). Recently, a novel BRAF exon 15 mutation (V600E) was detected in imatinib-naive wildtype high-risk intestinal GISTs (4%). However, the frequency and distribution of BRAF mutations within the spectrum of GISTs, and whether they might represent secondary events acquired during tumour progression, remain unknown. Methods: 69 GISTs (39 KIT mutants, 2 PDGFRA mutants and 28 wild-type) were analysed for mutations in BRAF exon 15 and KRAS exon 2. To assess the stage at which these mutations might occur in GIST, a considerable number of incidental gastric (n = 23) and intestinal (n = 2) tumours were included. Results: BRAF mutations (V600E) were detected in 2 of 28 wild-type GISTs (7%), but in none of the 41 KIT/ PDGFRA mutants. No KRAS mutation was detected. The two BRAF-mutated GISTs measured 4 mm in diameter and originated in the gastric body and the jejunum in two men (mean age, 76 years). Both tumours were mitotically inactive KIT-positive spindle-cell GISTs that were indistinguishable histologically from their more common KITmutated counterparts. Conclusion: BRAF mutations represent an alternative molecular pathway in the early tumorigenesis of a subset of KIT/PDGFRA wild-type GISTs and are per se not associated with a high risk of malignancy. Mutations in KIT, PDGFRA and BRAF were mutually exclusive in this study. Results from this and a previous study indicate that BRAF-mutated GISTs show a predilection for the small bowel (four of five tumours), but this needs further evaluation in larger studies.

Background:The tumour-host interaction at the invasive front of colorectal cancer, including the epithelial–mesenchymal transition and its hallmark ‘tumour budding', is an important area of investigation in terms of prognosis. The aim of this study was to determine the prognostic impact of a ‘pro-/anti-tumour' approach defined by an established ‘pro-tumour' (tumour budding) and host-related ‘anti-tumour' factor of the adaptive immunological microenvironment (CD8+ lymphocytes).Methods:Double immunostaining for CK22/CD8 on whole tissue sections (n=279; Cohort 1) and immunohistochemistry for CD8+ using tissue microarrays (n=191; Cohort 2) was carried out. Tumour buds, CD8+ and CD8+ T-lymphocytes : tumour buds indices were evaluated per high-power field.Results:In Cohort 1, a low-CD8+/ buds index was associated with lymph node metastasis (P<0.001), vascular invasion (P=0.009), worse survival in univariate (P<0.001) and multivariable (P<0.001) analysis, and furthermore in lymph node-negative patients (P=0.002). In Cohort 2, the CD8+/ buds index was associated with T stage (P<0.001), N stage (P=0.041), vascular invasion (P=0.005) and survival in patients with TNM stage II (P=0.019), stage III (P=0.004), and adjuvantly untreated (P=0.009) and treated patients (P<0.001).Conclusion:The CD8+ lymphocyte : tumour-budding index is an independent prognostic factor in colorectal cancer and a promising approach for a future prognostic score for patients with this disease.

Recent evidence highlights the potential prognostic and predictive value of BRAF and K‐RAS gene alterations in patients with colorectal cancer. However, a comprehensive evaluation of BRAF and K‐RAS mutations and their specific clinicopathological features, histomorphological presentation and effect on protein expression have not been systematically analyzed. The aim of this study was to characterize the clinicopathological, histomorphological and protein expression profiles of BRAF‐ and K‐RAS‐mutated colorectal cancers and determine their impact on patient survival. Molecular analysis for microsatellite instability (MSI), K‐RAS and BRAF was carried out on paraffin‐embedded samples from 404 patients with primary colorectal cancer. Using tissue microarrays, 36 tumor‐associated and 14 lymphocyte/inflammatory‐associated markers were evaluated by immunohistochemistry. BRAF mutation was associated with right‐sided tumor location (p < 0.001), higher tumor grade (p = 0.029), absence of peritumoral lymphocytic inflammation (p = 0.026) and MSI‐H (p < 0.001). In right‐sided tumors, loss of CDX2 expression was observed in 23 of 24 cases (95.8%). BRAF mutation was a poor prognostic indicator in patients with right‐sided disease (p = 0.01). This result was maintained in multivariable analysis (p < 0.001; HR = 2.82; 95% CI: 1.5–5.5) with pT, pN and vascular invasion and independent of CDX2 expression. K‐RAS mutation, in contrast, was not associated with any of the features analyzed. BRAF gene mutation is an adverse prognostic factor in right‐sided colon cancer patients independent of MSI status and, moreover, in patients with lymph node‐negative disease. These results indicate that molecular analysis for BRAF may be a useful biomarker for identifying patients with right‐sided colon cancer with poor outcome who may benefit from a more individualized course of therapy.

Epidermal growth factor receptor (EGFR) gene mutations and increased copy numbers are considered as predictors of response to EGFR tyrosine kinase inhibitors (EGFR-TKI) in non-small-cell lung cancer (NSCLC). Lung cancer diagnosis is often based on cytology alone. However, almost all published data on EGFR gene analyses were obtained from biopsies. This study tested the feasibility of EGFR gene analyses on cytological specimens. Eighty-four cytological specimens from NSCLCs were prospectively analysed for EGFR gene mutation in exons 18 -21 and EGFR gene copy numbers were evaluated by fluorescence in situ hybridisation (FISH). A FISHpositive result was defined according to the criteria by Cappuzzo et al established for biopsies of NSCLCs. Fluorescence in situ hybridisation results of cytological specimens were compared to the FISH results on matching biopsies (n ¼ 33). Initial diagnosis of NSCLC was solely based on cytology in 37 out of 84 (44.0%) patients. Out of 80 NSCLCs, 6 (7.5%) showed EGFR gene mutations. Out of 67 cancers, 45 (67.2%) were FISH positive on cytological specimens. Comparison of FISH showed a FISH-positive result in 21 out of 33 (63.6%) cytological specimens but in only 8 out of 33 (24.2%) matched biopsies. Epidermal growth factor receptor gene analyses are well applicable to cytological specimens. The high FISH-positive rate of NSCLC on cytological specimens contrasts with the low rate on biopsies when previously suggested criteria are used. New criteria for a positive EGFR FISH status to predict response to therapy with EGFR-TKI need to be defined for cytological specimens.

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