O6-Methylguanine-DNA methyltransferase protein expression by immunohistochemistry in brain and non-brain systemic tumours: systematic review and meta-analysis of correlation with methylation-specific polymerase chain reaction

Background: The DNA repair protein O6-Methylguanine-DNA methyltransferase (MGMT) confers resistance to alkylating agents. Several methods have been applied to its analysis, with methylation-specific polymerase chain reaction (MSP) the most commonly used for promoter methylation study, while immunohistochemistry (IHC) has become the most frequently used for the detection of MGMT protein expression. Agreement on the best and most reliable technique for evaluating MGMT status remains unsettled. The aim of this study was to perform a systematic review and meta-analysis of the correlation between IHC and MSP. Methods A computer-aided search of MEDLINE (1950-October 2009), EBSCO (1966-October 2009) and EMBASE (1974-October 2009) was performed for relevant publications. Studies meeting inclusion criteria were those comparing MGMT protein expression by IHC with MGMT promoter methylation by MSP in the same cohort of patients. Methodological quality was assessed by using the QUADAS and STARD instruments. Previously published guidelines were followed for meta-analysis performance. Results Of 254 studies identified as eligible for full-text review, 52 (20.5%) met the inclusion criteria. The review showed that results of MGMT protein expression by IHC are not in close agreement with those obtained with MSP. Moreover, type of tumour (primary brain tumour vs others) was an independent covariate of accuracy estimates in the meta-regression analysis beyond the cut-off value. Conclusions Protein expression assessed by IHC alone fails to reflect the promoter methylation status of MGMT. Thus, in attempts at clinical diagnosis the two methods seem to select different groups of patients and should not be used interchangeably

Gene amplification and protein expression of EGFR and HER2 by chromogenic in situ hybridisation and immunohistochemistry in atypical adenomatous hyperplasia and adenocarcinoma of the lung

Aims: To investigate the importance of gene amplification and EGFR (epidermal growth factor receptor) and HER2 protein expression during the progression of adenocarcinoma of the lung. Methods: EGFR and HER2 gene amplification was examined in atypical adenomatous hyperplasia (AAH), bronchioloalveolar carcinoma (BAC), and adenocarcinoma with mixed subtypes (MX) by chromogenic in situ hybridisation (CISH), and protein expression was examined by immunohistochemistry using paraffin wax embedded tissues. Results: EGFR and HER2 gene amplification was found in four and two of 86 cases, respectively, and was detected only in the invasive components of MX. EGFR and HER2 protein expression was seen in 24 and 18 of 86 cases, respectively. EGFR and HER2 proteins were not expressed in AAH but were expressed in one BAC case each. EGFR and HER2 proteins were expressed in 23 and 17 of 55 adenocarcinomas with MX. EGFR and HER2 protein expression was seen more often in the invasive components than in the BAC components of MX, and increased significantly as lesions progressed from AAH to BAC, early MX, and overt MX. Because EGFR and HER2 protein expression was frequently seen without gene amplification, other mechanisms apart from gene amplification may be associated with protein expression. Conclusions: EGFR and HER2 gene amplification may be a late event and EGFR and HER2 protein expression may be associated with the development of adenocarcinoma of the lung

Immunohistochemical marker panels for distinguishing between epithelioid mesothelioma and lung adenocarcinoma

The distinction between epithelioid mesothelioma and lung adenocarcinoma remains an important diagnostic challenge for surgical pathologists. The aim of the present study was to select a limited and appropriate panel of antibodies that can differentiate between epithelioid mesothelioma and lung adenocarcinoma. Specimens of 90 epithelioid mesotheliomas and 51 lung adenocarcinomas obtained from Japanese cases were examined using calretinin, WT1, AE1/AE3, CAM5.2, cytokeratin (CK) 5/6, vimentin, epithelial membrane antigen (EMA), thrombomodulin, CEA, CA19-9, and CA125. Ninety-six percent of epithelioid mesotheliomas were positive for calretinin; 99% for WT1; 100% for AE1/AE; 97% for CAM5.2; 70% for CK 5/6; 91% for vimentin; 96% for EMA; 71% for thrombomodulin; 77% for mesothelin; 7% for CEA; 17% for CA19-9; and 85% for CA125. In contrast, 33% of lung adenocarcinomas were positive for calretinin; 16% for WT1; 100% for AE1/AE3, CAM5.2, and EMA; 41% for CK 5/6; 47% for vimentin; 20% for thrombomodulin; 69% for mesothelin; 98% for CEA; 73% for CA19-9; and 80% for CA125. For distinguishing between epithelioid mesothelioma and lung adenocarcinoma, the combination of CEA, calretinin and each WT1 or thrombomodulin was suggested to be the best panel of immunohistochemical markers

Suppression of plasminogen activator inhibitor-1 by RNA interference attenuates pulmonary fibrosis

Background and aim: There is a growing body of evidence demonstrating that plasminogen activator inhibitor-1 (PAI-1) is involved in the progression of pulmonary fibrosis. In fact, PAI-1 knockout mice are protected from bleomycin-induced pulmonary fibrosis. This study was conducted to determine whether the intrapulmonary administration of small interfering RNA (siRNA) targeting PAI-1 (PAI-1-siRNA) limits the development of bleomycin-induced pulmonary fibrosis. Methods: Lung biopsies from patients with idiopathic pulmonary fibrosis (IPF) were stained for PAI-1. The distribution of siRNA in the lung, the PAI-1 level in bronchoalveolar (BAL) fluid and the extent of fibrotic changes in the lung were evaluated following the intranasal administration of PAI-1-siRNA in a mouse model of bleomycin-induced pulmonary fibrosis. The effect of PAI-1-siRNA on the epithelial to mesenchymal transition (EMT) was also evaluated using a mouse lung epithelial cell line, LA-4. Results: PAI-1 was overexpressed in the hyperplastic type 2 pneumocytes lining the honeycomb lesions of patients with IPF. The single intranasal instillation of PAI-1-siRNA resulted in the diffuse uptake of siRNA into the epithelial cells lining the dense fibrotic lesions. The repeated administration of PAI-1-siRNA initiated during either the inflammatory or the fibrotic phase into bleomycin-injured mice reduced the PAI-1 level in BAL fluid and limited the accumulation of collagen in the lungs. EMT induced by transforming growth factor β (TGFβ) in LA-4 cells was inhibited by transfection with PAI-1-siRNA. Conclusions: The direct suppression of PAI-1 in the lung by the intrapulmonary administration of PAI-1-siRNA attenuated the development and progression of pulmonary fibrosis. The inhibition of EMT may be, at least in part, involved in this effect

Genetic ablation of the Bach1 gene reduces hyperoxic lung injury in mice: Role of IL-6

Bach1 is a transcriptional repressor of the heme oxygenase (HO)-1 gene. Bach1-null (Bach1(-/-)) mice are reported to be protected from myocardial ischemia/reperfusion injury; however, the effect of Bach1 disruption on another oxidative stress model of hyperoxic lung injury has yet to be determined. To investigate the role of Bach1 in hyperoxic lung injury, Bach1(-/-) mice and wild-type (WT) mice were exposed to 90% O-2. During hyperoxic exposure, the survival of Bach1(-/-) mice was significantly longer than that of WT mice. However, the administration of zinc protoporphyrin, an inhibitor of HO-1 activity, did not change the mortality in either of the mice. thus suggesting that this protective effect was not mediated by an HO-1 overexpression in Bach1(-/-) mice. The indices of lung injury in the lungs of Bach1(-/-) mice were lower than those of WT mice; unexpectedly, however, the levels of IL-6 in bronchoalveolar lavage (BAL) fluid from Bach1(-/-) mice were significantly higher than those of WT mice. Interestingly, the intrapulmonary administration of small interfering RNA against IL-6 was shown to reduce the IL-6 levels in BAL fluids and shorten the survival in Bach1(-/-) mice during hyperoxic exposure. In addition, a chromatin immunoprecipitation analysis revealed the binding of Bach1 to the IL-6 promoter and its detachment after oxidative stress. Considering the previous observation that the transgenic mice overexpressing IL-6 are protected from hyperoxic lung injury, these results therefore indicate that IL-6 mediates an increased survival in Bach1(-/-) mice during hyperoxic exposure