Cyclin D1 is overexpressed in atypical teratoid/rhabdoid tumor with <Emphasis Type="Italic">hSNF5/INI1</Emphasis> gene inactivation

Object: Although atypical teratoid/rhabdoid tumor (AT/RT) is known to generate through inactivation of the hSNF5/INI1 gene on chromosome 22q, the downstream molecular mechanism remains unclear. We histologically and molecularly reviewed our pediatric brain tumors for unrecognized AT/RTs and evaluated the role of cyclin D1, a potential molecular target of hSNF5/INI1. Methods: We analyzed 16 tumors under three years of age: seven medulloblastomas, three anaplastic ependymomas (E IIIs), two each of supratentorial primitive neuroectodermal tumors (sPNETs) and choroid plexus carcinomas (CPCs), and one each of neuroblastoma and pineoblastoma. Immunohistochemistry for glial fibrillary acidic protein, vimentin, epithelial membrane antigen, smooth muscle actin and cyclin D1 was performed. Polymerase chain reaction (PCR)-single-strand conformation polymorphism analysis with direct sequencing, differential PCR and microsatellite analysis were conducted for hSNF5/INI1mutation, homozygous deletion and loss of heterozygosity (LOH) on 22q, respectively. Because of the presence of rhabdoid cells and the polyimmunophenotypic features, the diagnosis was revised to AT/RT in five (31%) tumors, namely, two E IIIs and one each of medulloblastoma, CPC and pineoblastoma. Three of them harbored such hSNF5/INI1 aberrations as germline single base deletion (492/6 delC) and missense mutation (C157T) together with LOH 22q or homozygous deletion. Cyclin D1 was overexpressed in those three tumors but not in the two that lacked hSNF5/INI1 inactivation. Conclusion: AT/RT can be misdiagnosed as a variety of tumors, including ependymoma that potentially harbors LOH 22q. Our data indicate that cyclin D1 is a target of hSNF5/INI1in primary tumors.     

Expression of P0 glycoprotein in CNS glia: effects of overexpression in N20.1 cells

To examine effects of expression of the PNS myelin P0 glycoprotein in glial cells of CNS lineage, we transfected murine N20.1 glial cells with a rat P0 cDNA. A stably transfected cell line expressing high levels of P0 message showed P0 immunostaining, along with changes in morphology. Polymerase chain reaction (PCR) identified the predicted rat P0 sequence in the transfected N20.1 cells and further revealed low levels of mouse P0 message in the nontransfected cells and in primary mouse astrocytes. This is the first evidence of endogenous expression of message for P0 glycoprotein in CNS glia. Quantitative RT-PCR confirmed the expression of rat P0 mRNA in the transfected N20.1 cells, at levels about 400 times greater than murine P0 in nontransfected cells. A 27-kD band was detected in the transfected cells by Western blot with P0 antibody, but not in mock-transfected or nontransfected N20.1 cells. Immunocytochemistry following permeabilization showed intracellular vesicular localization of P0 in the cytoplasm and perinuclear rings in transfected cells, with a similar pattern but much lower levels in nontransfected cells. Faint surface staining for P0 protein without permeabilization was seen only on the transfected cells. A few transfected cells with membrane sheets stained more intensely for surface P0. Quantitative RT-PCR was used to determine if P0 overexpression altered expression of other myelin-related genes compared with glial fibrillary acidic protein (GFAP); the ratios of myelin basic protein (MBP)/GFAP and proteolipid protein (PLP)/GFAP were increased 2- to 3-fold in the P0-transfected cells. We conclude that P0 overexpression alters N20.1 gene expression and cell morphology, and shifts the cells from astroglial to oligodendroglial phenotype.     

Correlation between encoded protein overexpression and copy number of the HER2 gene with survival in non-small cell lung cancer

The HER2 oncogene, which encodes the tyrosine kinase receptor, is commonly overexpressed in several types of cancer. Treatment using a humanized monoclonal antibody bound to HER2 product is becoming standard therapy for advanced breast cancer. Overexpression occurs in approximately 30% of non-small cell lung cancers (NSCLCs) and has been associated with poor prognosis. However, the frequency of a genetic aberration in the HER2 gene in lung cancer and the association between gene amplification and prognosis are poorly defined. To clarify these relationships, we simultaneously analyzed protein overexpression by immunohistochemistry (IHC) and determined the gene copy number by FISH in 50 surgical specimens of NSCLC. A low-grade increase in the copy number (3 to 8 copies) of the HER2 gene was detected in 44% of tumors. Most represented polysomy of chromosome 17. Protein overexpression was observed in 26%. Overexpression was detected in adenocarcinoma more frequently than in squamous cell carcinoma. No significant correlation was observed between copy number increase and overexpression. Neither gene copy number increase nor overexpression correlated with survival. We conclude that the significance of HER2 status in NSCLC is different from that in breast cancer because high-grade amplification occurs rarely.     

Defining a role for c-Myc in breast tumorigenesis

The proto-oncogene c-myc is commonly amplified and overexpressed in human breast tumors, and the tumorigenic potential of c-myc overexpression in mammary tissue has been confirmed by both in vitro and in vivo models of breast cancer. However, the mechanisms by which Myc promotes tumorigenesis are not well understood. Recent evidence indicates that Myc can promote cell proliferation as well as cell death via apoptosis. These studies provide new insight and impetus in defining a role for c-Myc in breast tumorigenesis and may point toward novel targets for breast cancer therapy.     

Promoter demethylation in MMTV/N-rasN transgenic mice required for transgene expression and tumorigenesis

We studied demethylation within the transgene promoter in transgenic mice carrying the N-ras proto-onco-gene driven by the mouse mammary tumor long terminal repeat (MMTV/N-ras^N) and the relationship of demethy!ation to transgene overexpression and tumorigenesis. Demethylation at Fspl or Clal sites correlated with age of the animal and transgene expression in nontumorous mammary gland. Demethylation preceded expression in this tissue. In lymphomas and mammary tumors, the promoter Fspl and Clal sites were significantly more demethylated than in nontumorous control tissues. The Aval, Cfol, and Hpall sites were also found to be undermethylated in older animals and showed differences between tumor and control tissues. Two additional sites (Eagl and Narl) remained fully methylated in all tissues. In contrast with normal tissue, demethylation at the Fspl and Clal sites and expression were not correlated in tumor tissue. An increase in expression in normal tissue initially occurred and was correlated with the level of promoter demethylation; this increase was followed by a further increment in transgene expression when tumors developed. Thus, promoter demethylation leading to transgene overexpression was associated with long-latency tumorigenesis in MMTV/N-ras^N transgenic mice. Demethylation of proto-oncogene promoters may therefore be a mechanism of carcinogenesis that requires further investigation in human tumors.     

Absence of p53 gene mutation and infrequent overexpression of p53 protein in hepatoblastoma

Ten cases of hepatoblastoma were studies for overexpression of p53 protein by immunohistochemistry and for possible p53 gene mutation by single strand conformation polymorphism (SSCP) analysis and direct DNA sequencing of the polymerase chain reaction products. Only one case of the macrotrabecular type at stage IV showed overexpression of p53 protein. No DNA mobility shift was found in any of the cases studies by SSCP analysis. DNA sequencing performed on the case showing overexpression of p53 protein revealed no mutation within exons 5 to 8. The associated adrenal cortical carcinoma of the same case also showed overexpression of p53 protein, but no mutation of the p53 gene. These results indicate that mutation of the p53 gene is infrequent in hepatoblastoma. This observation supports the view that mutation of the p53 gene is not as important in the oncogenesis of childhood neoplasms as in adult cancers.     

Cell cycle-related variation and tissue-restricted expression of human cyclin D1 protein

Recent evidence from genetic studies suggests that abnormalities of some of the members of the cyclin superfamily may be intimately associated with tumourigenesis, most likely through deregulation of the cell cycle control. In an attempt to elucidate the potential role of cyclin D1 (a gene located within the 11q13 amplicon and a candidate BCL-1, PRAD-1 oncogene) in the pathogenesis of human neoplasias, we have developed and characterized a novel monoclonal antibody specifically recognizing cyclin D1 protein in various assays including immunohistochemistry on frozen and paraffin sections. Using the DCS-6 antibody as a tool, we now show a characteristic cell cycle-dependent variation of the cyclin D1 protein in human cultured cells and report on the first immunohistochemical study of this G1 cyclin in a range of normal human tissues and breast carcinomas. Analysis of normal tissues revealed generally low levels of cyclin D1 protein, mainly restricted to the proliferative zones of some epithelial tissues, and the lack of its expression in several human tissues including lymph nodes, spleen, and tonsils. In contrast, pronounced overexpression/nuclear accumulation of cyclin D1 was found in 37 per cent of cases in a series of 35 primary ductal carcinomas of the breast. We conclude that the DCS-6 antibody provides a potentially useful tool for the establishment of simple methods suitable for verifying any diagnostic and/or prognostic value of this novel marker on large series of histological specimens and opens the way for biochemical, immunocytochemical, and immunohistochemical studies of the role played by cyclin D1 aberrations in human oncogenesis.     

p53过度表达与结直肠癌淋巴结转移、微转移关系的研究

 目的　探讨结直肠癌病例中p5 3过度表达与淋巴结转移、微转移间关系。方法　收集 1989～2 0 0 1年根治性手术切除 ,有完整检查资料的结直肠癌标本 74例经溶脂法检查淋巴结共 35 0 3枚 ,均经4 μm间断连续切片 ,HE染色及免疫组织化法 (CK18+EMA)检查淋巴结转移和微转移。肿块经免疫组化法 (DO - 0 7)检查 p5 3过度表达。 结果　检出转移淋巴结 2 2 3枚 ,微转移淋巴结 38枚。全组中 31例有 p5 3过度表达。结肠癌中 p5 3过度表达与淋巴结转移、微转移无关 (r =0 .19,P =0 .2 0 ;r =- 0 .2 3,P=0 .13)。直肠癌中p5 3过度表达与淋巴结转移、微转移无关 (r =0 .0 2 ,P =0 .90 ;r =0 .13,P =0 .32 )。结论　p5 3过度表达与结直肠癌淋巴结转移、微转移程度无明显相关性。     

Correlation between mutations in p53 gene and protein expression in human lymphomas

A discordance between p53 protein overexpression and the presence of mutations in the gene has been observed in many types of tumors, including human lymphomas. To probe this finding, we have studied a large series of 94 lymphomas of different pathologic types and histologic differentiation. Analyzing exons 5–9, we have found mutations in the p53 gene in 7 of 94 cases distributed in different subtypes: 4/12 (33%) high-grade B-cell non-Hodgkin's lymphomas (B-NHLs), in 1 of 5 (20%) high-grade mucosa-associated lymphomas (MALT), in 1 of 22 (4.5%) anaplastic large cell lymphoma (ALCL), and in 1 of 24 (4%) T-cell NHLs. Immunostaining with anti-p53 antibody DO-7 was possible in 87 lymphomas, and overexpression of p53 protein was observed in 16 cases (18%). A discrepancy between the results of SSCP and immunostaining was detected in 18 tumor samples. Two cases with mutations in the gene showed no altered protein expression and 16 cases overexpressed p53 protein had no point mutations. In these cases, the possibility that mutations occur outside the exons studied has been tested and the entire coding sequence analyzed. Only one case showed a mutation in exon 10, and we found two cases carrying a polymorphism in exon 4 and in intron 10. We conclude that mutations in p53 occur mainly in high-grade B-cell NHLs. Although not limited to a specific subtype of lymphoma, they may be rare in Hodgkin's disease and in low-grade lymphomas. The discrepancies between overexpression and presence of mutations suggest (1) the existence of another mechanism to stabilize the p53 protein, and (2) that the immunohistochemistry cannot be used to predict mutations in the gene. Am. J. Hematol. 55:1-8, 1997. © 1997 Wiley-Liss, Inc.     

MTA1表达与鼻咽癌浸润转移的关系

目的:研究肿瘤转移相关基因Mta1(metastasis-associated gene 1)表达与鼻咽癌浸润转移的关系.方法:应用逆转录聚合酶链反应(RT-PCR)技术,对43例鼻咽癌组织和20例正常鼻咽部组织,检测了Mta1 mRNA的表达.结果:43例鼻咽癌组织的Mta1 mRNA平均表达水平(2.36±0.87)明显高于鼻咽部正常组织(0.87±0.45)(P＜0.01);鼻咽癌组织中Mta1 mRNA高表达率与临床分期、T分期、N分期关系密切;43例鼻咽癌有19例Mta1 mRNA高表达,高表达率为44.2%.结论:Mta1基因在鼻咽癌组织中的表达明显高于正常组织,其高表达与鼻咽癌浸润转移关系密切.