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p53 and K-ras gene mutations in lung cancer of workers exposed to silica and asbestos
Pathology low differentiated squamous cell carcinoma (C-57) were cultured by 0.1 I~M all-tran retinoicacid (AT-RA) to induce differentiation and observed on the change of nm23-H1 protein with immunocytochemistry and enzyme-link-immunosorption assay (ELISA). The nm23-H1 DNA segment was infected to nm23-H1 protein negative PLA-801 cells through recombinant retrovirus expressig vector pLXSN-nmhl. The transfected nm23-H1-PLA-801 cells were implanted into BALB/c nude mice. Results showed: Only embryonic tissue developing to the stage of definite configuration (organ structure) accumulated nm23-H1 protein more constantly; Positive rate of nm23-H1 in 49 cases of human lung carcinoma was 34.8%, Among them, squamous cell carcinoma 27%, Adenocarcinoma 50%. All 5 cases of giant cell lung carcinoma were negative; Except no expression of nm23-H1 protein in PLA-801 cells all the other three cell trains expressed nm23-H1 protein. After AT-RA treatment the level of nm23-H1 of the four strain cells was increased especially in PLA-801 cells; The positive rate of nm23-H1 protein in 27 cases of hilar lymph node metastatic foci was 27%, compared with 34.8% in primary tumor foci, not significant, also no difference of lymph node metastatic rate between nm23-H1 positive group and negative group. Even the lung adenocarcinoma with more positive rate of nm23H1 protein have much hight hilar lymph node metastatic rate than the squamous cell carcinoma. The tumorigenesis and aggressiveness of transfected nm23-H1- PLA-801 implanted into nude mice had no significant difference compared with the control group. These findings suggested: There are positive corelation between accumulation of nm23-H1 protein and the differentiation of lung tissue and organoid or tissue structure of lung carcinoma; AT-RA and nm23-H1 gent transfection could increase the expression of nm23-H1 protein in lung carcinoma cells, but it can't seems to inhibit metastasis of the cancer cells directly.
~-2-~ p53 and K-ras gene mutations in lung cancer of workers exposed to silica and asbestos B.C. Liu, R. Guan, P.H. Zhou, D.C. Fu, B.H. Huang, Q. Miao, H.H. Wang, B.R. You. Institute of Occupational Medicine, Chinese
Academy of Preventive Medicine, Beijing, China Pneumoconiosis is the most serious occupational disease in China. Asbestos and crystalline silica are concluded by IARC as human carcinogens. To reveal the mutational spectra of lung cancer related to these dusts, we analyzed the p53 and K-ras gene mutations in lung cancer of worker exposed to silica (36 case) and asbestos (10 case). 8 cases of lung cancer without occupational dust exposure were also studied. DNA was extracted from paraffin-embedded or fresh lung cancer tissue. PCR-SSCP analysis on exons 5, 7 and 8 of p53 gene revealed that the mutational frequencies were 70% and 41.7% for asbestos and silica related lung cancer, respectively. More than 40% of mutations clustered on exon 8 for dust related lung cancer, but only around 20% mutations were found on exon 8 for nonoccupational lung cancer as reported. The histological spectrum of mutations of pneumoconiosis related lung cancer also differs from that of non-occupational lung cancer, which is with the highest prevalence (70%) in SCLC and the lowest in adenocarcinoma (33%). In our study, adenocarcinoma had the highest mutation rates (57% and 54%) and SCLC had the lowest (14% and 31%) for asbestos and silica related lung cancer. K-ras gene mutations were high in asbestos and silica related lung cancer. Unexpectively, no codon 12 mutation, which is the predominant mutation in K-ras gene in non-occupational lung cancer, was detected in 36 silica-related lung cancer either by PCR-RFLP, or by DNA sequencing. The mutations in silica-related lung cancer frequently distributed in codons 13 and 15, principally a G-C transvertion, instead of G-T transversion in non-occupational lung cancer. Codon 12 mutations were found in asbestos related lung cancer (29%) and nonoccupational lung cancer (67%). The distinct mutational spectra of p53 and K-ras genes in silicosis related lung cancer provides a DNA molecular level evidence implicating the carcinogen of silica in causing lung cancers.
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role of matrix metalloproteinase 2 and laminin-5~2 chain in developmental and invasive processes of peripheral lung adenocarcinoma
H. Kitamura I , Y. Kagesato 2,3, Y. Oosawa 1, N. Kawano 1, H. Hayashi 1, K. Okudela 1, T. Yazawa 1, T. Ito I , M. Kanisawa 1, Y. Inayama 1, Y. Nakatani 1, H. Mizushima 3, K. Miyazaki 3. Departments of
1Pathology; 20torhinolaryngology, School of Medicine; 3Division of Cell Biology, Kihara Institute for Biological Research, Yokohama City University, Yokohama, Japan Adenocarcinoma (AC) of the peripheral lung largely develops in a multistep manner from atypical adenomatous hyperplasia (AAH), through in situ cancer, into invasive/metastatic cancer. Although many investigators have studied this process including central fibrosis, basement membrane (BM) patterns, and/or expression of matrix degradative proteases, the precise mechanisms have not been fully clarified yet. The present study elucidated the process and mechanisms of the development and progression of peripheral lung AC. Using immunohistochemistry, we investigated the relationship among stromal fibrosis, the patterns of BM components (laminin1, laminin-5 (Lm5) ~,2 chain, type IV collagen), and the expressions of matrix metalloproteinase 2 (MMP-2) and tissue inhibitor of metalloproteinases-2 (TIMP-2) in the lesions at various developmental steps of peripheral lung AC including 33 lesions of AAH, 24 lesions of bronchioloalveolar carcinoma (BAC), and 24 lesions of non-BAC AC. The alveolar BM was intact in all the 33 lesions of AAH and 11 lesions of nonsclerosing BAC (NSC-BAC) that did not form a central scar. The preexistent BM was disrupted or even absent in the area of central scar of some sclerosing BAC (SC-BAC) with collapse fibrosis alone (2/4) and in those of all of the AC lesions with desmoplastic stromal fibrosis (9 SC-BAC and 24 non-BAC tumors), suggesting that destruction of the BM was correlated with the formation of a central scar, particularly with desmoplasia. Expressions of MMP-2 and TIMP2 were associated with the central scar formation as well as with destruction of the BM components. Both the neoplastic and stromal cells expressed MMP-2 and TIMP-2. Cytoplasmic staining for Lm5 y2 chain of neoplastic cells was negative or only weak in 22/23 lesions of AAH and 12/13 lesions of NSC-BAC, whereas moderate staining was observed in and around the central scar in 6/10 lesions of SCBAC and diffuse, moderate or strong staining was observed in 12/15 lesions of moderately or poorly differentiated (non-BAC) AC. AAH and NSC-BAC are early-stage intraepithelial neoplasia, while AC with a central scar are advanced and invasive cancers. It is likely that MMP-2 and TIMP-2 play an important role in tumor cell invasion. The intracytoplasmic Lm5 y2 chain in neoplastic cells may be associated with tumor invasion, particularly with tumor cell migration.
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Evaluation of HER2/neu expression in lung tumors by immunohistochemistry and fluorescence in situ hybridization (FISH)
F,R. Hirsch, R. Veve, L. Varella-Garcia, P.A. Bunn, W.A. Franklin.
Colorado Cancer Center, Denver, Colorado; Dept of Pathology, Denver, Colorado; Dept of Oncology, Denver, Colorado, USA Herceptin, a humanized monoclonal antibody to HER2/neu oncoprotein, was originally developed as a treatment for breast carcinoma but is now being evaluated in the treatment of lung carcinoma. We have compared results of the Herceptest (Dako Corp.) performed on paraffin sections to an alkaline phosphatase-antialkaline phosphatase (APAAP) immunohistochemical test on frozen sections using the monoclonal antibody 4D5 (a gift from Genentech Corp.) in 36 lung carcinomas including 14 adenocarcinomas, 17 squamous carcinomas, 3 large cell carcinomas (LCLC) and 2 small cell lung carcinomas. Adenocarcinomas and LCLCs were more frequently strongly positive than squamous tumors in the Herceptest (65% vs. 29%) but not in the APAAP test (65% vs. 82%). Seventeen tumors were tested by twocolor FISH to evaluate levels of gene amplification. Moderate level amplification (1.78-2.1) of HER2/neu was detected in 2/17 tumors and low level amplification (1.17-1.23) was detected in 5/17 tumors
~-2-~ p53 and K-ras gene mutations in lung cancer of workers exposed to silica and asbestos B.C. Liu, R. Guan, P.H. Zhou, D.C. Fu, B.H. Huang, Q. Miao, H.H. Wang, B.R. You. Institute of Occupational Medicine, Chinese
Academy of Preventive Medicine, Beijing, China Pneumoconiosis is the most serious occupational disease in China. Asbestos and crystalline silica are concluded by IARC as human carcinogens. To reveal the mutational spectra of lung cancer related to these dusts, we analyzed the p53 and K-ras gene mutations in lung cancer of worker exposed to silica (36 case) and asbestos (10 case). 8 cases of lung cancer without occupational dust exposure were also studied. DNA was extracted from paraffin-embedded or fresh lung cancer tissue. PCR-SSCP analysis on exons 5, 7 and 8 of p53 gene revealed that the mutational frequencies were 70% and 41.7% for asbestos and silica related lung cancer, respectively. More than 40% of mutations clustered on exon 8 for dust related lung cancer, but only around 20% mutations were found on exon 8 for nonoccupational lung cancer as reported. The histological spectrum of mutations of pneumoconiosis related lung cancer also differs from that of non-occupational lung cancer, which is with the highest prevalence (70%) in SCLC and the lowest in adenocarcinoma (33%). In our study, adenocarcinoma had the highest mutation rates (57% and 54%) and SCLC had the lowest (14% and 31%) for asbestos and silica related lung cancer. K-ras gene mutations were high in asbestos and silica related lung cancer. Unexpectively, no codon 12 mutation, which is the predominant mutation in K-ras gene in non-occupational lung cancer, was detected in 36 silica-related lung cancer either by PCR-RFLP, or by DNA sequencing. The mutations in silica-related lung cancer frequently distributed in codons 13 and 15, principally a G-C transvertion, instead of G-T transversion in non-occupational lung cancer. Codon 12 mutations were found in asbestos related lung cancer (29%) and nonoccupational lung cancer (67%). The distinct mutational spectra of p53 and K-ras genes in silicosis related lung cancer provides a DNA molecular level evidence implicating the carcinogen of silica in causing lung cancers.
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role of matrix metalloproteinase 2 and laminin-5~2 chain in developmental and invasive processes of peripheral lung adenocarcinoma
H. Kitamura I , Y. Kagesato 2,3, Y. Oosawa 1, N. Kawano 1, H. Hayashi 1, K. Okudela 1, T. Yazawa 1, T. Ito I , M. Kanisawa 1, Y. Inayama 1, Y. Nakatani 1, H. Mizushima 3, K. Miyazaki 3. Departments of
1Pathology; 20torhinolaryngology, School of Medicine; 3Division of Cell Biology, Kihara Institute for Biological Research, Yokohama City University, Yokohama, Japan Adenocarcinoma (AC) of the peripheral lung largely develops in a multistep manner from atypical adenomatous hyperplasia (AAH), through in situ cancer, into invasive/metastatic cancer. Although many investigators have studied this process including central fibrosis, basement membrane (BM) patterns, and/or expression of matrix degradative proteases, the precise mechanisms have not been fully clarified yet. The present study elucidated the process and mechanisms of the development and progression of peripheral lung AC. Using immunohistochemistry, we investigated the relationship among stromal fibrosis, the patterns of BM components (laminin1, laminin-5 (Lm5) ~,2 chain, type IV collagen), and the expressions of matrix metalloproteinase 2 (MMP-2) and tissue inhibitor of metalloproteinases-2 (TIMP-2) in the lesions at various developmental steps of peripheral lung AC including 33 lesions of AAH, 24 lesions of bronchioloalveolar carcinoma (BAC), and 24 lesions of non-BAC AC. The alveolar BM was intact in all the 33 lesions of AAH and 11 lesions of nonsclerosing BAC (NSC-BAC) that did not form a central scar. The preexistent BM was disrupted or even absent in the area of central scar of some sclerosing BAC (SC-BAC) with collapse fibrosis alone (2/4) and in those of all of the AC lesions with desmoplastic stromal fibrosis (9 SC-BAC and 24 non-BAC tumors), suggesting that destruction of the BM was correlated with the formation of a central scar, particularly with desmoplasia. Expressions of MMP-2 and TIMP2 were associated with the central scar formation as well as with destruction of the BM components. Both the neoplastic and stromal cells expressed MMP-2 and TIMP-2. Cytoplasmic staining for Lm5 y2 chain of neoplastic cells was negative or only weak in 22/23 lesions of AAH and 12/13 lesions of NSC-BAC, whereas moderate staining was observed in and around the central scar in 6/10 lesions of SCBAC and diffuse, moderate or strong staining was observed in 12/15 lesions of moderately or poorly differentiated (non-BAC) AC. AAH and NSC-BAC are early-stage intraepithelial neoplasia, while AC with a central scar are advanced and invasive cancers. It is likely that MMP-2 and TIMP-2 play an important role in tumor cell invasion. The intracytoplasmic Lm5 y2 chain in neoplastic cells may be associated with tumor invasion, particularly with tumor cell migration.
•]
Evaluation of HER2/neu expression in lung tumors by immunohistochemistry and fluorescence in situ hybridization (FISH)
F,R. Hirsch, R. Veve, L. Varella-Garcia, P.A. Bunn, W.A. Franklin.
Colorado Cancer Center, Denver, Colorado; Dept of Pathology, Denver, Colorado; Dept of Oncology, Denver, Colorado, USA Herceptin, a humanized monoclonal antibody to HER2/neu oncoprotein, was originally developed as a treatment for breast carcinoma but is now being evaluated in the treatment of lung carcinoma. We have compared results of the Herceptest (Dako Corp.) performed on paraffin sections to an alkaline phosphatase-antialkaline phosphatase (APAAP) immunohistochemical test on frozen sections using the monoclonal antibody 4D5 (a gift from Genentech Corp.) in 36 lung carcinomas including 14 adenocarcinomas, 17 squamous carcinomas, 3 large cell carcinomas (LCLC) and 2 small cell lung carcinomas. Adenocarcinomas and LCLCs were more frequently strongly positive than squamous tumors in the Herceptest (65% vs. 29%) but not in the APAAP test (65% vs. 82%). Seventeen tumors were tested by twocolor FISH to evaluate levels of gene amplification. Moderate level amplification (1.78-2.1) of HER2/neu was detected in 2/17 tumors and low level amplification (1.17-1.23) was detected in 5/17 tumors