[Infection status of Helicobacter pylori with antibody values of 3.0U/mL or more and less than 10.0U/mL in the LZ test "Eiken" H. pylori antibody].

The LZ test "Eiken" for H. pylori antibody (LZ test) was examined. In patients with an antibody titer of ≥3.0U/mL and <10.0U/mL among 698 patients who underwent breath tests, the status of H. pylori infection was assessed from the results of the 13C urea breath test and the findings of gastric mucosal atrophy by performing an upper gastrointestinal endoscopy. A positive 13C urea breath test was observed in 22.3% of these patients (156/698, 95% confidence interval 19.4-25.6%), and gastric mucosal atrophy of C-2 or greater was observed in 39.7%. We presumed that 156 (22.3%) patients had present H. pylori infections, 141 (20.2%) patients had experienced a previous infection, and 401 (57.5%) patients were uninfected. The infection rate of H. pylori (current infection+previous infection) was treated as a so-called "risk of gastric cancer" and was 42.6% (297/698, 95% confidence interval 38.9-46.3%). In the LZ test, the concept of a negative high value should be understood. A receiver operating characteristic curve plotted depending on whether the 13C urea breath test was positive or not gave a positive cutoff value of 5.6U/mL;values greater than the cutoff value were taken as indicative of the need to investigate the status of H. pylori infection. Even without gastric mucosal atrophy, 2.0% of these patients had a positive breath test. For gastritis localized in the antrum (C-1), 17.8% of the patients had positive breath test results.

Diversity of epidermal growth factor receptor-mediated activation of downstream molecules in human lung carcinomas.

The correlations among epidermal growth factor receptor (EGFR) gene amplification, gene mutation, overexpression/phosphorylation of EGFR protein and activation of its downstream molecules, signal transducers and activators of transcription 3 (Stat-3), Akt and extracellular signal-related protein kinase 1/2 (Erk1/2) were investigated in 28 cases of human lung carcinomas. In five cases of carcinomas with EGFR amplification, EGFR expression and phosphorylation levels were higher than other cases, and Stat-3 was activated in all five cases. Point mutations in the tyrosine kinase domain of EGFR were detected in five cases, one of which was also associated with gene amplification. In these five cases, both EGFR expression and phosphorylation were enhanced, and Akt was activated in four cases. In the remaining 19 cases, EGFR protein expression was upregulated in eight cases and phosphorylated in four cases, but neither EGFR nor phosphorylated-EGFR expression levels specifically correlated with activation of particular downstream molecules. In general, either Stat-3 or Akt, but not both, was activated reciprocally and complementarily to each other, as indicated by their phosphorylation. However, Erk1/2 was activated regardless of the status of Stat-3, Akt or EGFR proteins. The current data suggest that persistent Stat-3 activation may be a critical event downstream of EGFR that has been overexpressed by gene amplification. In contrast, tumor cells harboring the EGFR mutation may persistently activate a cascade via Akt. Finally, in the majority of cases that have no aberration of the EGFR, its downstream molecules function in reciprocal and/or complementary manner in the maintenance and/or progression of carcinomas. These overall results could provide novel insights into potential chemotherapeutic regimens for lung carcinomas, such as inhibitors of Stat-3, Akt and Erk1/2.

EGFR protein overexpression and gene amplification in squamous cell carcinomas of the esophagus.

Overexpression of epidermal growth factor receptor (EGFR) is observed in many cancers, sometimes accompanied by gene amplification. Recently, several clinical therapies targeting EGFR were developed, but the eligibility criteria for these therapies is not fully established. To develop such eligibility criteria for esophageal squamous cell carcinoma (ESCC), we sought to clarify: (i) the exact frequency of EGFR overexpression, (ii) the relationship between protein overexpression and gene amplification, (iii) the relationship between gene amplification and specific gene mutations and (iv) the correlation between the status of EGFR and clinical or pathological features. Immunohistochemistry revealed that EGFR protein is overexpressed in 53 (50%) of the 106 ESCC examined. Fluorescence in situ hybridization (FISH) indicated clear EGFR gene amplification in 15 of the 53 tumors, somewhat higher EGFR copy in 32 cases, and no increase in 6 cases. Gene amplification was significantly associated with high level overexpression. Direct sequencing of exons 19 and 21 of EGFR revealed no mutations in 15 tumors exhibiting gene amplification, and no mutations in 25 tumors not exhibiting gene amplification. Overexpression of EGFR was significantly correlated with depth of invasion of the tumor. In conclusion, anti-EGFR therapies may be appropriate for patients with ESCC. We assume that combined analyses by immunohistochemistry/FISH would clarify aberrations in protein and gene function, and could help to identify those patients who may benefit from anti-EGFR therapy.

Trastuzumab-mediated antibody-dependent cellular cytotoxicity against esophageal squamous cell carcinoma.

PURPOSE: In the present study, we investigated the degree of protein expression and gene amplification of HER-2 in esophageal squamous cell carcinoma (SCC) cell lines and freshly isolated tumors, and trastuzumab-mediated biological activity, in particular antibody-dependent cellular cytotoxicity (ADCC) against HER-2-expressing esophageal SCC cell lines. EXPERIMENTAL DESIGN: Ten different SCC cell lines with various levels of HER-2 status evaluated by flow cytometry, immunocytochemistry (HercepTest), and fluorescence in situ hybridization were evaluated for ADCC, growth inhibitory, or apoptosis-inducing activities mediated by trastuzumab. RESULTS: Trastuzumab induced ADCC against HER-2-expressing esophageal SCC and the activities reflected the degree of HER-2 expression analyzed by flow cytometric analysis, but not by HercepTest nor fluorescence in situ hybridization analysis. Furthermore, trastuzumab-mediated ADCC against transforming growth factor-beta-producing SCC was enhanced by the treatment with SB-431542, which is a selective inhibitor of the phosphorylation induced by transforming growth factor-beta. There were very marginal effects of anti-proliferative or apoptosis-inducing activities mediated by trastuzumab for HER-2-expressing esophageal SCC. CONCLUSION: HER-2-expressing esophageal SCC cells could be killed by trastuzumab-mediated ADCC and the activity reflected the degree of HER-2 expression detected by flow cytometry.

Protein overexpression and gene amplification of epidermal growth factor receptor in nonsmall cell lung carcinomas. An immunohistochemical and fluorescence in situ hybridization study.

BACKGROUND: Recently, molecular therapies targeting epidermal growth factor receptor (EGFR) have been developed for clinical use. The current study was conducted to determine 1) the exact frequency of EGFR protein overexpression, 2) the correlation between protein overexpression and EGFR amplification, and 3) the correlation between the status of the genetic and clinicopathologic features in nonsmall cell lung carcinomas (NSCLC). METHODS: In total, 181 NSCLC samples were examined immunohistochemically using an antibody against EGFR, and tumor cells that exhibited overexpression were examined further for EGFR amplification by fluorescence in situ hybridization. RESULTS: Overexpression of EGFR protein was found in 34% of the tumors. Among these, EGFR amplification was demonstrated in 74%. High-level gene amplification was found exclusively in tumors cells with high protein expression. In most of these tumors, cells that exhibited EGFR overexpression and gene amplification were distributed heterogeneously, even within a single tumor nodule. Statistically, EGFR overexpression was correlated significantly with lymph node metastasis and with a more advanced pathologic stage. Moreover, in adenocarcinomas, gene amplification was correlated significantly with lymph node metastasis and tended to be correlated with a more advanced pathologic stage. CONCLUSIONS: The overexpression of EGFR in NSCLC was accompanied predominantly, but not exclusively, by gene amplification. It is important to evaluate not only protein overexpression but also the EGFR status to design adjuvant therapies for patients with NSCLC, because specimens that exhibit both protein overexpression and gene amplification may predict eventual lymph node metastasis and, possibly, aggressive tumor behavior.

Aberration of epidermal growth factor receptor expression in bone and soft-tissue tumors: protein overexpression, gene amplification and activation of downstream molecules.

In order to evaluate the involvement of epidermal growth factor receptor, and to analyze the correlation between gene aberration and protein expression in mesenchymal tumors, we examined protein expression by immunohistochemistry in 125 cases of bone and soft-tissue tumors. Furthermore, amplification of epidermal growth factor receptor gene was determined by fluorescence in situ hybridization. Positive immunostaining was found in 23 cases (18.4%). Among these 23 cases, one of malignant fibrous histiocytoma showed the highest degree (3+) of protein overexpression and gene amplification as clusters of hybridization signals, indicating homogeneously staining regions. The second case of malignant fibrous histiocytoma also showed a higher degree (2+) of overexpression and coamplification of the epidermal growth factor receptor gene with the centromeric regions, indicating polysomy of chromosome 7. The levels of expression observed in immunohistochemistry were confirmed by immunoblotting and found to be comparable. Moreover, although expression of phosphorylated epidermal growth factor receptor was detected in those two cases of malignant fibrous histiocytoma, constitutive activation of extracellular signal-related protein kinase 1/2 was not observed, suggesting that activation of epidermal growth factor receptor does not necessarily and constantly lead to signal transduction to the downstream molecules. In the remaining 123 cases, including 21 cases exhibiting weak (1+) immunoreactivity, no gene amplification nor polysomy was found. Collectively, expression of epidermal growth factor receptor was observed not infrequently in mesenchymal tumors, but 'overexpression' is rare and can be attributed to an increase in gene copy number, resulting from amplification or polysomy. Although cases that scored positive for protein expression and/or gene amplification could be qualified candidates for antiepidermal growth factor receptor therapies, further examination of the status of downstream molecules in the signal cascade, such as phosphorylated epidermal growth factor receptor and extracellular signal-related protein kinase 1/2, may be required as the process of therapeutic strategy.