Oncological results of extended resection for locally advanced rectal cancer: the value of postirradiation MRI in predicting local recurrence.

AIM: The primary purpose of this study was to analyse the overall survival and local recurrence rate after extended resection of locally advanced rectal cancer. The second aim was to determine the ability of the response to radiological irradiation to predict R0 resection. METHOD: A retrospective study was performed of 94 consecutive patients with locally advanced rectal cancer operated on at the Helsinki University Hospital, Helsinki, Finland between 2005 and 2013. Data were collected from patient records. All patients were treated with an en bloc resection. Sixty-two patients received preoperative long-term chemoradiotherapy. RESULTS: The 30-day mortality was 3.2%. Local recurrence occurred in 10 (10.6%) patients. The cumulative 1-, 3- and 5-year overall survival to each year was 89.4%, 68.3% and 51.8%. The most important prognostic factor for both local recurrence (P = 0.006) and survival (P = 0.003) was an R0 resection. A poor or no response seen on posttreatment MRI predicted local recurrence (P = 0.045) and decreased disease-free survival in patients treated curatively (P = 0.052). The histological tumour regression grade was not associated with local recurrence or survival. CONCLUSION: Multivisceral resection offers a 5-year survival of over 50% and local control of advanced rectal cancer in nearly 90% of carefully selected patients.

Diminished salivary epidermal growth factor secretion: a link between Sjögren's syndrome and autoimmune gastritis?

OBJECTIVES: Healthy human labial salivary glands produce epidermal growth factor (EGF). In Sjögren's syndrome (SS), EGF staining is diminished. SS is also associated with chronic autoimmune corpus gastritis. We therefore hypothesized that EGF secretion would be diminished in SS and that this could affect gastric target cells. METHODS: Salivary EGF secretion in SS was compared to that in healthy controls using an enzyme-linked immunosorbent assay (ELISA). EGF receptor (EGFR) immunoreactive cells in the gastric corpus of healthy human subjects were analysed using immunostaining. RESULTS: Salivary secretion of EGF was diminished in SS patients (232.4, range 52.6-618.4, vs. 756.6, range 105.3-1631.6 pg/min, p = 0.002). Proton-pump positive parietal cells were mostly EGFR immunoreactive whereas very few pepsinogen I (PGI)-positive cells were EGFR positive. CONCLUSIONS: As EGF is relatively acid resistant, salivary gland-derived EGF might participate in an exo/endocrine mode of parietal cell maintenance in the gastric corpus. Deficiency of salivary gland-derived EGF in SS patients may cause impairment of gastric parietal cells resulting in exposure of immunogenic cryptic antigens and loss of immunological self-tolerance.

Combination of microsatellite instability and BRAF mutation status for subtyping colorectal cancer.

BACKGROUND: The objective of the study was to examine the role of microsatellite instability (MSI) and BRAF(V600E)mutation in colorectal cancer (CRC) by categorising patients into more detailed subtypes based on tumour characteristics. METHODS: Tumour samples from 762 population-based patients with sporadic CRC were analysed for MSI and BRAF(V600E) by immunohistochemistry. Patient survival was followed-up for a median of 5.2 years. RESULTS: Compared with microsatellite stable (MSS) CRC, MSI was prognostic for better disease-free survival (DFS; 5 years: 85.8% vs 75.3%, 10 years: 85.8% vs 72.9%, P=0.027; HR 0.49, CI 0.30-0.80, P=0.005) and disease-specific survival (DSS; 5 years: 83.2% vs 70.5%; 10 years: 83.2 vs 65.0%, P=0.004). Compared with BRAF wild type, BRAF(V600E) was a risk for poor survival (overall survival; 5 years: 62.3% vs 51.6%, P=0.014; HR 1.43, CI 1.07-1.90, P=0.009), especially in rectal cancer (for DSS, HR: 10.60, CI: 3.04-36.92, P<0.001). The MSS/BRAF(V600E) subtype was a risk for poor DSS (HR: 1.88, CI: 1.06-3.31, P=0.030), but MSI/BRAF(V600E) was a prognostic factor for DFS (HR: 0.42, CI: 0.18-0.96, P=0.039). Among stage I-II patients, the MSS/BRAF(V600E) subtype was independently associated with poor DSS (HR: 5.32, CI: 1.74-16.31, P=0.003). CONCLUSIONS: Microsatellite instable tumours were associated with better prognosis compared with MSS. BRAF(V600E) was associated with poor prognosis unless it occurred together with MSI. The MSI/BRAF(V600E) subtype was a favourable prognostic factor compared with the MSS/BRAF wild-type subtype. BRAF(V600E) rectal tumours showed particularly poor prognosis. The MSS/BRAF(V600E) subtype was associated with increased disease-specific mortality even in stage I-II CRC.

Management of precancerous conditions and lesions in the stomach (MAPS): guideline from the European Society of Gastrointestinal Endoscopy (ESGE), European Helicobacter Study Group (EHSG), European Society of Pathology (ESP), and the Sociedade Portuguesa de Endoscopia Digestiva (SPED).

Atrophic gastritis, intestinal metaplasia, and epithelial dysplasia of the stomach are common and are associated with an increased risk for gastric cancer. In the absence of guidelines, there is wide disparity in the management of patients with these premalignant conditions. The European Society of Gastrointestinal Endoscopy (ESGE), the European Helicobacter Study Group (EHSG), the European Society of Pathology (ESP) and the Sociedade Portuguesa de Endoscopia Digestiva (SPED) have therefore combined efforts to develop evidence-based guidelines on the management of patients with precancerous conditions and lesions of the stomach (termed MAPS). A multidisciplinary group of 63 experts from 24 countries developed these recommendations by means of repeat online voting and a meeting in June 2011 in Porto, Portugal. The recommendations emphasize the increased cancer risk in patients with gastric atrophy and metaplasia, and the need for adequate staging in the case of high grade dysplasia, and they focus on treatment and surveillance indications and methods.

Prognostic role of CIP2A expression in serous ovarian cancer.

BACKGROUND: Cancerous inhibitor of protein phosphatase 2A (CIP2A) is an oncoprotein expressed in several solid cancers. Our purpose was to study its role in serous ovarian cancer patients, and the association to clinicopathological variables and molecular markers. METHODS: We collected retrospectively 562 consecutive serous ovarian cancer patients treated at the Helsinki University Central Hospital. We stained tumour tissue microarrays for CIP2A by immunohistochemistry and constructed survival curves according to the Kaplan-Meier method. Associations to clinicopathological and molecular markers were assessed by the χ(2)-test. RESULTS: We found strong cytoplasmic CIP2A immunoreactivity in 212 (40.4%) specimens, weak positivity in 222 (42.4%) specimens, and negative in 90 (17.2%). Immunopositive CIP2A expression was associated with high grade (P<0.0001), advanced stage (P=0.0005), and aneuploidy (P=0.001, χ(2)-test). Cancerous inhibitor of protein phosphatase 2A overexpression was also associated with EGFR protein expression (P=0.006) and EGFR amplification (P=0.043). Strong cytoplasmic CIP2A immunopositivity predicted poor outcome in ovarian cancer patients (P<0.0001, log-rank test). CONCLUSION: Our results show that CIP2A associates with reduced survival and parameters associated with high grade in ovarian cancer patients, and may thus be one of the factors that identify aggressive subtype (type II) of this disease.

High CIP2A immunoreactivity is an independent prognostic indicator in early-stage tongue cancer.

BACKGROUND: No reliable prognostic markers exist for squamous cell carcinoma of the tongue, and its prognosis can even in early stages be unpredictable and survival poor despite treatment. A potential marker is oncoprotein cancerous inhibitor of PP2A (CIP2A), which acts as a prognostic marker in gastric and non-small cell lung cancers. METHODS: We collected specimens of 73 stage T1N0M0 and T2N0M0 oral squamous cell carcinomas of the tongue, as well as samples from normal oral mucosa, dysplastic lesions, and invasive carcinomas (n=39). All samples were stained for CIP2A by immunohistochemistry. Survival curves were constructed according to the Kaplan-Meier method. The Cox proportional hazard model served for univariate and multivariate survival analysis. RESULTS: High CIP2A immunoreactivity predicted poor survival in tongue cancer patients (P=0.027, logrank test). In multivariate survival analysis, CIP2A was an independent prognostic factor (HR 2.02, 95% confidence interval 1.07-3.82, P=0.030). Cytoplasmic CIP2A expression was higher in severe dysplasia than in mild dysplasia. CONCLUSION: Our results suggest that high CIP2A expression characterises aggressive disease. Acting as a prognostic marker it might be of help when choosing patients for adjuvant treatment in tongue cancer patients.

Risk of cancer in patients with chronic pouchitis after restorative proctocolectomy for ulcerative colitis.

AIM: The aim of this study was to evaluate the consequences of chronic pouchitis after restorative proctocolectomy for ulcerative colitis. METHOD: Forty-two patients with chronic pouchitis underwent pouch endoscopy with biopsies after a median of 8.3 years of postoperative follow up. The pouchitis disease activity index (PDAI) was calculated. Morphological changes were recorded. Immunohistochemical analyses for cyclooxygenase 2 (COX-2), Ki-67 and p53 were performed, as was DNA flow cytometry. Endoscopy was also carried out in 10 patients without pouchitis and in nine healthy subjects. RESULTS: In patients with chronic pouchitis, the PDAI was 6 (standard error of the mean ± 4). Eighteen (43%) patients used continuous medication. The PDAI correlated positively with villous atrophy (P < 0.05). None of the pouch biopsies showed dysplasia. COX-2 immunostaining was detected in 35 (83.3%) patients with chronic pouchitis, in five (50%) without pouchitis, but in none of the normal controls. COX-2 expression correlated with mucosal atrophy (P < 0.01). In 15 (35.7%) of 42 patients with chronic pouchitis, Ki-67 immunostaining was increased, but no increase was observed in either control group (P < 0.002). No p53 immunopositivity was found, and DNA flow cytometry was normal in all pouches. One of the patients developed adenocarcinoma at the anal anastomosis. CONCLUSION: No dysplastic changes were detected during the first decade after surgery. Routine follow up of patients with chronic pouchitis with a hand-sewn anastomosis may not be necessary, although a small risk of cancer seems to remain at the anal anastomosis. The follow up should be focused on at-risk groups.

Treatment response of colorectal cancer liver metastases to neoadjuvant or conversion therapy: a prospective multicentre follow-up study using MRI, diffusion-weighted imaging and 1H-MR spectroscopy compared with histology (subgroup in the RAXO trial).

BACKGROUND: Colorectal cancer liver metastases respond to chemotherapy and targeted agents not only by shrinking, but also by morphologic and metabolic changes. The aim of this study was to evaluate the value of advanced magnetic resonance imaging (MRI) methods in predicting treatment response and survival. PATIENTS AND METHODS: We investigated contrast-enhanced MRI, apparent diffusion coefficient (ADC) in diffusion-weighted imaging and 1H-magnetic resonance spectroscopy (1H-MRS) in detecting early morphologic and metabolic changes in borderline or resectable liver metastases, as a response to first-line neoadjuvant or conversion therapy in a prospective substudy of the RAXO trial (NCT01531621, EudraCT2011-003158-24). MRI findings were compared with histology of resected liver metastases and Kaplan-Meier estimates of overall survival (OS). RESULTS: In 2012-2018, 52 patients at four Finnish university hospitals were recruited. Forty-seven patients received neoadjuvant or conversion chemotherapy and 40 liver resections were carried out. Low ADC values (below median) of the representative liver metastases, at baseline and after systemic therapy, were associated with partial response according to RECIST criteria, but not with morphologic MRI changes or histology. Decreasing ADC values following systemic therapy were associated with improved OS compared to unchanged or increasing ADC, both in the liver resected subgroup (5-year OS rate 100% and 34%, respectively, P = 0.022) and systemic therapy subgroup (5-year OS rate 62% and 23%, P = 0.049). 1H-MRS revealed steatohepatosis induced by systemic therapy. CONCLUSIONS: Low ADC values at baseline or during systemic therapy were associated with treatment response by RECIST but not with histology, morphologic or detectable metabolic changes. A decreasing ADC during systemic therapy is associated with improved OS both in all patients receiving systemic therapy and in the resected subgroup.

Prolonged systemic circulation of chimeric oncolytic adenovirus Ad5/3-Cox2L-D24 in patients with metastatic and refractory solid tumors.

Eighteen patients with refractory and progressive solid tumors were treated with a single round of triple modified oncolytic adenovirus (Ad5/3-Cox2L-D24). Ad5/3-Cox2L-D24 is the first non-Coxsackie-adenovirus receptor-binding oncolytic adenovirus used in humans. Grades 1-2 flu-like symptoms, fever, and fatigue were seen in most patients, whereas transaminitis or thrombocytopenia were seen in some. Non-hematological grades 3-5 side effects were seen in one patient with grade 3 ileus. Treatment resulted in high neutralizing antibody titers within 3 weeks. Virus appeared in serum 2-4 days after treatment in 83% of patients and persisted for up to 5 weeks. One out of five radiologically evaluable patients had partial response (PR), one had minor response (MR), and three had progressive disease (PD). Two patients scored as PD had a decrease in tumor density. Tumor reductions not measurable with Response Evaluation Criteria In Solid Tumors (RECIST) were seen in a further four patients. PR, MR, stable disease, and PD were seen in 12, 23.5, 35, and 29.5% of tumor markers analyzed, respectively (N=17). Ad5/3-Cox2L-D24 appears safe for treatment of cancer in humans and extended virus circulation results from a single treatment. Objective evidence of anti-tumor activity was seen in 11/18 (61%) of patients. Clinical trials are needed to extend these findings.

Ad5/3-9HIF-Delta24-VEGFR-1-Ig, an infectivity enhanced, dual-targeted and antiangiogenic oncolytic adenovirus for kidney cancer treatment.

Despite good safety data in clinical trials, oncolytic adenoviruses have not been efficient enough to make them a viable treatment alternative for cancers. As more potent viruses are being made, transcriptional and transductional targeting to tumor tissues becomes increasingly appealing. To improve antitumor efficacy, oncolytic adenoviruses can be armed with therapeutic transgenes, such as the antiangiogenic soluble vascular endothelial growth factor receptor 1-Ig fusion protein. We hypothesized that an infectivity enhanced, targeted, vascular endothelial growth factor receptor 1-Ig armed oncolytic adenovirus would exhibit improved specificity and antitumor effect in murine kidney cancer models. Two hypoxia inducible factor-sensitive promoters were evaluated for renal cancer specificity using a novel in vivo dual luciferase-imaging system. Earlier data had shown usefulness of the 5/3-serotype chimera capsid modification for kidney cancer. Therefore, we constructed Ad5/3-9HIF-Delta24-VEGFR-1-Ig, which showed good specificity and oncolytic effect on renal cancer cells in vitro and resulted in antitumor efficacy in a subcutaneous in vivo model, in which vascular endothelial growth factor receptor 1-Ig expression and a concurrent antiangiogenic effect were confirmed. In an intraperitoneally disseminated kidney cancer model, significantly enhanced survival was observed when compared with control viruses. These results suggest that a targeted, antiangiogenic, oncolytic adenovirus might be a valuable agent for testing in kidney cancer patients.

The Mutational Profile of Unicystic Ameloblastoma.

BRAF V600E is the most common mutation in conventional ameloblastoma (AM) of the mandible. In contrast, maxillary AMs appear to harbor more frequently RAS, FGFR2, or SMO mutations. Unicystic ameloblastoma (UAM) is considered a less aggressive variant of ameloblastoma, amenable to more conservative treatment, and classified as a distinct entity. The aim of this study was to characterize the mutation profile of UAM ( n = 39) and to compare it to conventional AM ( n = 39). The associations between mutation status and recurrence probability were also analyzed. In the mandible, 94% of UAMs (29/31, including 8/8 luminal, 6/8 intraluminal, and 15/15 mural subtypes) and 74% of AMs (28/38) revealed BRAF V600E mutations. Among the BRAF wild-type cases, 1 UAM showed a missense SMO mutation (p.L412F), whereas 2 NRAS (p.Q61R), 2 HRAS (p.Q61R), and 2 FGFR2 (p.C383R) activating mutations were identified in AM. Of the 3 maxillary UAMs, only 1 revealed a BRAF V600E mutation. Taken together, our findings demonstrate high frequency of activating BRAF V600E mutations in both UAM and AM of the mandible. In maxillary UAMs, the BRAF V600E mutation prevalence appears to be lower as was shown for AM previously. It could therefore be argued that UAM and AM are part of the spectrum of the same disease. AMs without BRAF V600E mutations were associated with an increased rate of local recurrence ( P = 0.0003), which might indicate that routine mutation testing also has an impact on prognosis.

Cyclooxygenase-1 and -2 expression in rheumatoid synovial tissues. Effects of interleukin-1 beta, phorbol ester, and corticosteroids.

High levels of immunoreactive cyclooxygenase (Cox; prostaglandin H synthase) are present in synovia from patients with rheumatoid arthritis (RA). We now show that the recently identified inducible isoform of Cox, Cox-2, is expressed in synovia from patients with RA. To further explore modulation of the Cox isoforms in RA synovial tissues, we examined the expression and modulation of Cox-1 and -2 in rheumatoid synovial explant cultures and cultured rheumatoid synovial fibroblast-like cells (synoviocytes). Immunoprecipitation of in vitro labeled proteins and Western blot analysis demonstrated the presence of both Cox-1 and -2 under basal conditions in freshly explanted rheumatoid synovial tissues. De novo synthesis of Cox-2 polypeptide was enhanced by IL-1 beta or PMA, and dramatically suppressed by dexamethasone (dex). Cox-1 expression, under the same conditions, showed only minor variation. Since mRNA for Cox-2 is highly unstable, we examined the regulation of Cox-2 transcripts in cultured rheumatoid synoviocytes. Under basal conditions both Cox-1 and -2 mRNAs were present at low levels, but Cox-2 mRNA was markedly increased by treatment with IL-1 beta or PMA. dex markedly suppressed the induction of Cox-2 mRNA. In sharp contrast, Cox-1 transcripts were not modulated by IL-1 beta or dex. These data suggest that modulation of Cox-2 expression by IL-1 beta and corticosteroids may be an important component of the inflammatory process in synovial tissues from patients with RA.

Non-steroidal anti-inflammatory drugs and the risk of oral cancer: a nested case-control study.

BACKGROUND: Non-steroidal anti-inflammatory drugs (NSAIDs) seem to prevent several types of cancer, but could increase the risk of cardiovascular complications. We investigated whether use of NSAIDs was associated with a change in the incidence of oral cancer or overall or cardiovascular mortality. METHODS: We undertook a nested case-control study to analyse data from a population-based database (Cohort of Norway; CONOR), which consisted of prospectively obtained health data from all regions of Norway. People with oral cancer were identified from the 9241 individuals in CONOR who were at increased risk of oral cancer because of heavy smoking (15 pack-years), and matched controls were selected from the remaining heavy smokers (who did not have cancer). FINDINGS: We identified and analysed 454 (5%) people with oral cancer (279 men, 175 women, mean [SD] age at diagnosis 63.3 [13.2] years) and 454 matched controls (n=908); 263 (29%) had used NSAIDs, 83 (9%) had used paracetamol (for a minimum of 6 months), and 562 (62%) had used neither drug. NSAID use (but not paracetamol use) was associated with a reduced risk of oral cancer (including in active smokers; hazard ratio 0.47, 95% CI 0.37-0.60, p<0.0001). Smoking cessation also lowered the risk of oral cancer (0.41, 0.32-0.52, p<0.0001). Additionally, long-term use of NSAIDs (but not paracetamol) was associated with an increased risk of cardiovascular-disease-related death (2.06, 1.34-3.18, p=0.001). NSAID use did not significantly reduce overall mortality (p=0.17). INTERPRETATION: Long-term use of NSAIDs is associated with a reduced incidence of oral cancer (including in active smokers), but also with an increased risk of death due to cardiovascular disease. These findings highlight the need for a careful risk-benefit analysis when the long-term use of NSAIDs is considered.

Cyclooxygenase-2 expression correlates with poor prognosis in pancreatic cancer.

BACKGROUND: Cyclooxygenase-2 (COX-2) overexpression is related to poor outcome in several cancers. COX-2 is upregulated in 42-90% of pancreatic ductal adenocarcinomas and is a potential target for chemotherapy. Earlier studies have not shown the expression of COX-2 to be a prognostic factor in pancreatic cancer. OBJECTIVE: To evaluate the prognostic value of COX-2 in a series of patients with pancreatic adenocarcinoma. METHODS: 128 patients operated on for pancreatic adenocarcinoma at Helsinki University Central Hospital between 1974 and 1998 provided sections from primary tumours which were immunohistochemically stained with a COX-2-antihuman monoclonal antibody. RESULTS: Cytoplasmic COX-2 reactivity (>5%) occurred in 46 specimens (36%), correlating neither with age, sex, stage, size, tumour stage, nodal metastases, nor grade. Lack of COX-2 expression correlated with distant metastases (p = 0.026). In univariate survival analysis, COX-2 expression (p = 0.0114), stage (p = 0.0002), grade (p = 0.0001), and age (p = 0.042) had prognostic significance. One, two, and five year survival rates were 51%, 32%, and 8% in the COX-2 negative groups compared with 34%, 5%, and 5% in the COX-2 positive groups (p = 0.011). Prognostic significance was especially high for patients operated on with curative intent (p = 0.004). In multivariate analysis, COX-2 was an independent prognostic factor (hazard ratio = 1.6 (95% confidence interval, 1.1 to 2.3)). CONCLUSIONS: Expression of COX-2 was associated with poor outcome from pancreatic ductal adenocarcinoma and was independent of tumour stage, grade, or age in multivariate analysis.

MMP-2 but not MMP-9 associated with COX-2 and survival in gastric cancer.

BACKGROUND AND AIM: Matrix metalloproteinases (MMPs) MMP-2 and MMP-9 can degrade type IV collagen of extracellular matrix and basal membranes. As cyclo-oxygenase-2 (COX-2) has been shown to activate MMPs, creating one of the COX-2-promoted pathways of tumour growth and metastasis, the prognostic role of MMP-2 and MMP-9 in gastric cancer was assessed and their association with COX-2 expression was evaluated. MATERIALS AND METHODS: Samples were collected from 342 consecutive patients operated on for gastric cancer, of which 315 were acceptable for MMP-2, MMP-9 and COX-2 immunohistochemistry. Specimens were stained with specific antibodies, evaluated and categorised by two interpreters, and then correlated with clinical data and survival. RESULTS: Epithelial MMP-2 immunoreactivity was associated with male sex, high stage, advanced penetration depth, non-curative surgery, high COX-2 expression and poor survival. Stromal MMP-2 expression correlated with high stage, intestinal type and non-curative surgery whereas MMP-9 correlated only with intestinal type. Stage, intent of surgery and COX-2 were independent prognostic factors. CONCLUSIONS: Epithelial MMP-2 expression in gastric cancer is associated with aggressive forms, COX-2 and poor survival, although MMP-2 was not an independent prognostic factor. In gastric cancer tumour growth is apparently induced by COX-2, and invasion is mediated by MMP-2.

Decreased xanthine oxidoreductase is a predictor of poor prognosis in early-stage gastric cancer.

BACKGROUND: Xanthine oxidoreductase (XOR) is a key enzyme in the degradation of DNA, RNA and high-energy phosphates. About half of the patients with breast cancer have a decrease in XOR expression. Patients with breast cancer with unfavourable prognosis are independently identified by the loss of XOR. AIM: To assess the clinical relevance of XOR expression in gastric cancer. METHODS: XOR levels were studied by immunohistochemistry in tissue microarray specimens of 337 patients with gastric cancer and the relation between XOR expression and a series of clinicopathological variables, as well as disease-specific survival, was assessed. RESULTS: XOR was moderately decreased in 41% and was undetectable in another 14% of the tumours compared with the corresponding normal tissue. Decreased XOR was associated with advanced stage, deep tumour penetration, diffusely spread tumour location, positive lymph node status, large tumour size, non-curative disease, cellular aneuploidy, high S-phase fraction and high cyclooxygenase-2 expression, but not with p53 expression or Borrmann classification. Down regulation of XOR was associated with unfavourable outcome, and the cumulative 5-year gastric cancer-specific survival in patients with strong XOR expression was 47%, compared with 22% in those with moderate to negative expression (p<0.001). This was also true in patients with stage I-II (p = 0.01) and lymph node-negative (p = 0.02) disease, as well as in patients with smaller (< or =5 cm) tumours (p = 0.02). CONCLUSION: XOR expression in gastric cancer may be a new marker for a more aggressive gastric cancer biology, similar to that previously reported for breast cancer.

Expression of cyclooxygenase-2 in human gastric carcinoma.

Epidemiological studies suggest that the use of aspirin decreases the incidence of and mortality from gastrointestinal cancers. The best known target of aspirin and other nonsteroidal anti-inflammatory drugs is cyclooxygenase (Cox), the rate-limiting enzyme in the conversion of arachidonic acid to prostanoids. Two Cox genes have been cloned, of which Cox-2 is an inducible immediate-early gene. It is still unknown how nonsteroidal anti-inflammatory drugs act as chemopreventive agents, but they may target Cox-2. Cox-2 mRNA and protein were recently found to be expressed in human colon carcinoma. We have now studied the expression of Cox-2 in human gastric adenocarcinoma tissues which contained significantly higher levels of Cox-2 mRNA when compared with paired gastric mucosal specimens devoid of cancer cells. In contrast, Cox-1 mRNA levels were not elevated in the carcinoma. However, Cox-2 mRNA was not expressed in mucinous ovarian carcinoma samples as detected by Northern blot hybridization. Immunohistological detection of Cox-2 protein showed cytoplasmic staining in the gastric carcinoma cells but not in the surrounding stroma. Some hyperplastic glands showed intense staining, whereas glands of normal morphology were negative. Our data thus suggest that Cox-2 is expressed by human gastric adenocarcinoma.

Expression of cyclooxygenase-2 in human lung carcinoma.

Epidemiological studies indicate that the use of aspirin decreases incidence of and mortality from gastrointestinal cancers. A major target of aspirin and other nonsteroid anti-inflammatory drugs is cyclooxygenase (Cox), the rate-limiting enzyme in the conversion of arachidonic acid to prostanoids. Two Cox genes have been cloned (Cox-1 and Cox-2), of which Cox-2 has recently been found to be expressed in several human carcinomas. We have now studied the expression of Cox-2 mRNA and protein in human lung adenocarcinoma, squamous cell carcinoma, and small cell lung cancer. Cox-2 mRNA steady-state levels were high in well-differentiated adenocarcinoma samples, but low in poorly differentiated adenocarcinoma, squamous cell carcinoma, and small cell lung cancer, as detected by Northern blot analysis. Immunohistochemistry showed Cox-2 staining in 19 of 21 adenocarcinomas. However, well-differentiated adenocarcinomas contained more Cox-2 staining than the poorly differentiated ones. Expression of the Cox-2 protein was also seen in all 11 squamous cell carcinomas studied, although the level of staining seemed to be less than that in the adenocarcinomas. Small cell lung cancer specimens (n = 4) stained with a relatively weak intensity. Interestingly, atypical alveolar epithelium, which associates with asbestosis and idiopathic fibrosing alveolitis and is considered to be a precursor lesion for lung cancer, expressed the Cox-2 protein. Our data, thus, suggest that Cox-2 is expressed in human lung carcinomas and in precursor lesions leading to this malignancy.

Tumor-associated trypsin participates in cancer cell-mediated degradation of extracellular matrix.

We have recently demonstrated that many cancer cell lines produce a novel trypsinogen isoenzyme called tumor-associated trypsinogen 2 (TAT-2). It was found during a search of the target protease for tumor-associated trypsin inhibitor (TATI). We now show that degradation of subendothelial cell extracellular matrix (ECM) by four different cell lines (COLO 205 colon carcinoma, K-562 erythroleukemia, CAPAN-1 pancreatic carcinoma, and HT 1080 fibrosarcoma) can be partially inhibited by TATI or neutralizing trypsin antibodies. When cells were cultured in serum-free medium on ECM, TATI and trypsin antibodies inhibited the release of immunoreactive fibronectin fragments from ECM by 47-54 and 40%, respectively. Degradation of isotopically labeled ([3H]serine, [3H]proline, and [35S]sulfate) ECM was also significantly prevented by TATI. At its maximum, it exerted a 57% inhibition on the degradation of [3H]serine-labeled ECM. Plasminogen added exogenously to the culture medium further potentiated the proteolysis of ECM. Interestingly, addition of enteropeptidase, an activator of TAT-2, also enhanced cell-mediated proteolysis as assessed by degradation of purified fibronectin coated onto the surface of wells. Immunoblot analysis showed that enteropeptidase-mediated proteolysis generated a pattern of fibronectin fragments similar to that obtained by digestion of purified fibronectin by TAT-2. These results demonstrate the existence of a proteolytic system in tumor cells which is dependent on the activation of TAT-2. We suggest that TAT-2 is involved in a protease cascade-stimulating tumor cell invasion and degradation of extracellular matrix.

The Bmx tyrosine kinase is activated by IL-3 and G-CSF in a PI-3K dependent manner.

Cytoplasmic protein tyrosine kinases play crucial roles in signaling via a variety of cell surface receptors. The Bmx tyrosine kinase, a member of the Tec family, is expressed in hematopoietic cells of the granulocytic and monocytic lineages. Here we show that Bmx is catalytically activated by interleukin-3 (IL-3) and granulocyte-colony stimulating factor (G-CSF) receptors. Activation of Bmx required phosphatidylinositol 3-kinase (PI-3K) as demonstrated by the ability of PI-3K inhibitors to block the activation signal. A green fluorescent protein (GFP) tagged Bmx was translocated to cellular membranes upon co-expression of a constitutively active form of PI-3K, further indicating a role for PI-3K in signaling upstream of Bmx. The expression of wild type Bmx in 32D myeloid progenitor cells resulted in apoptosis in the presence of G-CSF, while cells expressing a kinase dead mutant of Bmx differentiated into mature granulocytes. However, Bmx did not modulate IL-3-dependent proliferation of the cells. These results demonstrate distinct effects of Bmx in cytokine induced proliferation and differentiation of myeloid cells, and suggest that the stage specific expression of Bmx is critical for the differentiation of myeloid cells. Oncogene (2000) 19, 4151 - 4158