人胰腺癌中MDM2癌基因的表达和对野生型p53的拮抗作用

为研究ＭＤＭ２癌基因和ｐ５３在胰腺癌中的相互关系和作用，我们用基因重组、基因转移和分子杂交技术探讨了ＭＤＭ２在胰腺癌细胞系中的表达和扩增情况，并研究了ＭＤＭ２对野生型（ｗｔ）ｐ５３的拮抗作用。结果显示，５株人胰腺癌细胞系均有ＭＤＭ２基因不同程度的表达。ＭＤＭ２表达载体转染转化的胰腺癌细胞系ＰＣ－２／ｓ－ｗｔｐ５３细胞（已转染有野生型ｐ５３的胰腺癌细胞）后，ＭＤＭ２可以部分地解除野生型ｐ５３对胰腺癌细胞生长的抑制作用，证实ＭＤＭ２基因对野生型ｐ５３的功能有拮抗作用。     

横纹肌肉瘤中MDM2及p53基因表达的原位杂交检测

目的观察ＭＤＭ２、ｐ５３癌基因在横纹肌肉瘤（ＲＭＳ）发病中的作用及其与临床病理、预后间的关系。方法对确诊并有随访的３１例ＲＭＳ标本,用原位杂交技术进行ＭＤＭ２、ｐ５３定位观察。结果发现ＭＤＭ２及ｐ５３癌基因表达阳性率分别为７７４％（２４／３１例）和６６７％（２１／３１例）,其阳性率与年龄、性别和ＲＭＳ组织类型无明显关联（Ｐ＞０．０５）,但阳性率及其强度与ＲＭＳ分化程度（Ⅰ级与Ⅲ级）,转移与否及存活率差异有显著性（Ｐ＜０．０５）。结论ＭＤＭ２、ｐ５３阳性检出率有助于判断ＲＭＳ恶性程度及预测肿瘤的转移和预后。     

骨肿瘤MDM2和p53基因的改变

目的从基因水平研究ＭＤＭ２和ｐ５３基因在骨肿瘤中的表达，探讨其在骨肿瘤发生发展中的作用。方法用地高辛标记原位杂交技术研究了３８例骨肿瘤（骨肉瘤１２例，软骨肉瘤１０例，骨巨细胞瘤１４例，软骨母细胞瘤２例）ＭＤＭ２和ｐ５３的表达情况，并分析两种基因表达之间的相互关系。结果ＭＤＭ２在骨肉瘤、软骨肉瘤和骨巨细胞瘤中的阳性率分别为４１．７％、５０．０％和３５．７％。ｐ５３的阳性率分别为５８．３％、４０．０％和２１．４％。２例软骨母细胞瘤ＭＤＭ２和ｐ５３均为阳性。ＭＤＭ２与ｐ５３基因表达呈显著正相关（Ｐ＜０．００５）。结论ＭＤＭ２和ｐ５３基因改变是骨肿瘤的一种常见现象，可能参与骨肿瘤的发生与发展。     

p-MAPK、cyclinD1和p53蛋白在大肠癌中的表达及其相关性

目的：检测ｐ－ＭＡＰＫ、ｃｙｃｌｉｎＤ１和ｐ５３蛋白在大肠癌中的表达，并探讨其相关性。方法：免疫组织化学Ｓ－Ｐ法。结果：１４０例大肠癌中ｐ－ＭＡＰＫ、ｃｙｃｌｉｎＤ１和ｐ５３蛋白的阳性表达率分别为９２．９％（１３０／１４０）、８７．１％（１２２／１４０）和６６．４％（９３／１４０）；５０例相应癌旁肠粘膜中阳性表达率分别为４．０％（２／５０）、６．０％（３／５０）和２．０％（１／５０）。三者阳性表达     

大肠腺癌p53基因cDNA突变与突变型p53基因蛋白表达的相互关系和意义

目的：探讨大肠腺癌突变型ｐ５３基因蛋白表达与ｐ５３基因ｃＤＮＡ突变间的相互关系和意义。方法：对１００例新鲜大肠腺癌组织采用ＰＡｂ２４０单克隆抗体免疫组织化学染色（ＬＳＡＢ法）检测突变型ｐ５３基因蛋白表达、ＲＴ－ＰＣＲ－ＳＳＣＰ检测ｐ５３基因ｃＤＮＡ突变，并比较它们间相互关系。结果：１００例大肠腺癌中，７６例ＰＡｂ２４０单抗阳性（７６％），５１例ｐ５３基因ｃＤＮＡ突变阳性（５１％）；ＰＡｂ２４０单抗反应与ｐ５３基因ｃＤＮＡ突变比较，两者皆阳性４９％，两者中一者阳性２９％，两者皆阴性２２％（Ｐ＜０．０００１）。结论：ｐ５３基因ｃＤＮＡ突变与其基因蛋白产物结构改变高度吻合，大肠腺癌ｐ５３基因ｍＲＮＡ（ｃＤＮＡ）突变是参与和影响突变型ｐ５３基因蛋白结构改变和生物学行为的主要因素     

鼻咽癌,宫颈癌和肺癌中p53基因突变和表达的对比研究

目的对比研究ｐ５３在鼻咽癌、宫颈癌、肺癌中的表达及其与ｐ５３基因突变的关系，为进一步研究ｐ５３在癌变过程中的作用奠定基础。方法应用免疫组化结合ＰＣＲ－ＳＳＣＰ银染技术检测２４例鼻咽癌、９例宫颈癌、１０例肺癌中ｐ５３基因的表达与突变。结果２３／２４例鼻咽癌、６／９例宫颈癌、９／１０例肺癌有ｐ５３的过表达，在两种发病与ＤＮＡ致瘤病毒有关的鼻咽癌、宫颈癌中未能检测到ｐ５３基因突变；而发病与化学致癌物有关的肺癌中有５／１０例检测到有ｐ５３基因突变，其中１例发生在外显子８，４例发生在外显子５。这５例均有ｐ５３的过表达。结论（１）鼻咽癌、宫颈癌、肺癌中存在着ｐ５３蛋白的过表达。（２）肺癌中ｐ５３的过表达大部分与ｐ５３基因突变有关，而鼻咽癌与宫颈癌中ｐ５３过表达与ｐ５３基因突变关系不大，其与致瘤性ＤＮＡ病毒的作用是否有关有待进一步研究。     

c─erbB─2、ras p21及p53在大肠癌表达的免疫组化研究

对４８例大肠癌连续切片进行ｃ─ｅｒｂＢ─２、ｒａｓｐ２１及ｐ５３３种癌相关基因产物的免疫组化ＡＢＣ法及ＡＰＡＡＰ－ＩＧＳＳ双重标记研究，结果表明：（１）癌相关基因ｃ─ｅｒｂＢ─２、ｒａｓｐ２１和ｐ５３阳性表达率在癌组织分别为５０．２％（２５／４８）、４３．８％（２１／４８）和５４．２％（２６／４８）；癌旁粘膜分别为２５％（１２／４８）、１８．８（９／４８）和２０．８％（１０／４８）；而在＂正常＂结肠粘膜则分别为１２．５％（６／４８）、６．３％（３／４８）和０．（２）有２种以上癌相关基因产物同时表达者在癌组织中有２６例（５４．２％）．在癌旁粘膜有５例（１０．５％），而在正常粘膜则未见有２种癌相关基因同时表达，（３）癌组织中ｃ─ｅｒｂＢ─２＋ｐ２１、ｃ─ｅｒｂＢ─２＋ｐ５３、ｐ２１＋ｐ５３及ｃ─ｅｒｂＢ─２＋ｐ２１＋ｐ５３同时表达者分别为２．１％、１２．５％、１６．７％及２２．９％，（４）ＡＰＡＡＰ－ＩＧＳＳ双重免疫标记结果表明，单一癌细胞可同时表达２种癌基因产物。上述结果提示：ｃ─ｅｒｂＢ─２、ｒａｓｐ２１及ｐ５３可能与大肠病的发生有关，三者在大肠病的发生中可能起协同作用。     

肺癌中MTS1/p16和p53基因产物的表达与细胞增殖的关系

目的：研究肺癌中ＭＴＳ１／ｐ１６和ｐ５３基因产物的表达与细胞增殖的关系。方法：应用Ｓ－Ｐ免疫组织化学方法研究６２例肺癌组织中ｐ１６蛋白和ｐ５３蛋白的表达情况，并进行增殖细胞核抗原（ＰＣＮＡ）检测，计算细胞增殖指数（ｐｒｏｌｉｆｅｒａｔｉｏｎｉｎｄｅｘ，ＰＩ）。结果：６２例肺癌组织中ｐ１６蛋白和ｐ５３蛋白阳性率分别为５８．１％和５９．７％。腺癌ｐ１６蛋白的阳性率明显高于小细胞癌（Ｐ＜０．０５）；淋巴结转移阳性组ｐ１６蛋白的表达显著低于阴性组（Ｐ＜０．０５）；ＰＩ分级为Ⅱ级的ｐ１６蛋白表达显著高于Ⅳ级（Ｐ＜０．０５）。不同组织类型肺癌中ｐ５３蛋白的表达未见明显差异，淋巴结转移阳性组ｐ５３蛋白的表达高于阴性组（Ｐ＜０．０１）；不同ＰＩ分级中ｐ５３蛋白的表达，Ⅳ级明显高于Ⅰ级（Ｐ＜０．０５）和Ⅱ级（Ｐ＜０．０５），Ⅲ级明显高于Ⅰ级（Ｐ＜０．０５）和Ⅱ级（Ｐ＜０．０１）。ｐ１６蛋白低表达和ｐ５３蛋白过表达之间未见明显相关。结论：ｐ１６蛋白低表达和ｐ５３蛋白过表达均有促进肺癌细胞增殖的作用，ｐ１６蛋白的表达与肺癌的细胞分化有关，ｐ５３蛋白过表达对肺癌细胞的转移起重要作用。抑癌基因ｐ５３对ＭＴＳ１／ｐ１６基因无明显调控作     

免疫组化和PCR—SSCP检测p53基因异常的一致性探讨

目的：探讨免疫组化检测ｐ５３蛋白表达和ＰＣＲ－ＳＳＣＰ检测ｐ５３基因突变的一致性。方法：单克隆抗体ＤＯ－７检测８５例非小细胞肺癌（ＮＳＣＬＣ）的ｐ５３蛋白表达，ＰＣＲ－ＳＳＣＰ检测其中３１例腺癌的ｐ５３基因突变。结果：８５例ＮＳＣＬＣ中ｐ５３蛋白表达阳性率为６８％（５８／８５），３１例腺癌中ｐ５３蛋白表达阳性率为６１％（１９／３１），１４例（４６％）出现ｐ５３基因５～８外显子突变，ｐ５３蛋白免疫组化和ＰＣＲ－ＳＳＣＰ检测ｐ５３基因突变无显著相关（χ２＝０．１，Ｐ＝０．７６），其一致率为５２％。结论：ｐ５３蛋白表达并不能很好地反映ｐ５３基因突变。     

横纹肌肉瘤p16基因及其蛋白表达的研究

目的探讨ｐ１６基因及其蛋白表达与横纹肌肉瘤发生发展的关系。方法应用ＰＣＲＳＳＣＰ分析及免疫组化方法检测了４４例人横纹肌肉瘤（ＲＭＳ）中ｐ１６基因状态和４７例ＲＭＳ中ｐ１６蛋白表达情况。结果２例有ｐ１６基因纯合性缺失,６例突变。ｐ１６基因异常的８例ＲＭＳ中６例伴广泛浸润、复发或转移。ｐ１６蛋白表达阴性率为３８．３％（１８／４７例）。ｐ１６基因异常的８例中有７例ｐ１６蛋白阴性,１例为阳性。另有１１例ｐ１６蛋白阴性而未查见基因异常。在腺泡状横纹肌肉瘤中,ｐ１６基因异常发生率及ｐ１６蛋白表达缺失率均高于胚胎性横纹肌肉瘤。ｐ１６蛋白阴性的ＲＭＳ更易伴发广泛浸润、复发或转移。结论ｐ１６基因异常及ｐ１６蛋白表达缺失可能与部分横纹肌肉瘤的发生发展有关     

Utility of fluorescence in situ hybridization to detect MDM2 amplification in liposarcomas and their morphological mimics

The atypical lipomatous tumor (ALT)/well-differentiated liposarcoma (WDLS) and the de-differentiated liposarcoma (DDLS) represent the most common category of liposarcomas. ALT/WDLSs and DDLSs are often difficult to distinguish from other tumors with similar morphological characteristics. In this study, we investigated whether the detection of amplified or overexpressed murine double-minute 2 (MDM2) can be a useful diagnostic ancillary aid. We used fluorescent in situ hybridization (FISH) and immunohistochemistry (IHC) to detect MDM2 amplification and protein overexpression, respectively, in 49 WDLSs, 5 DDLSs, 23 myxoid liposarcomas, 25 benign lipomatous tumors, and 75 spindle and pleomorphic sarcomas. MDM2 amplification was detected in 48 of 49 WDLSs, 5 of 5 DDLSs, 2 of 9 malignant peripheral nerve sheath tumors, and 2 of 10 myxofibrosarcomas. We did not detect MDM2 amplification in any of the benign lipomatous tumors. FISH-mediated detection of MDM2 amplification was the most valuable diagnostic aid for ALT/WDLS, as determined by using the Fisher exact test to compare two different diagnoses of 19 biopsies. On the contrary, unequivocal nuclear overexpression of MDM2 was found in only 10 of 50 ALT/WDLSs. The sensitivity and specificity of MDM2 amplification in distinguishing a DDLS from spindle and pleomorphic sarcomas were 100% and 95%, respectively, while those of MDM2 overexpression were 100% and 87%, respectively. In conclusion, our results indicate that FISH-mediated detection of MDM2 amplification is the most useful adjunct in the diagnosis of both ALT/WDLS and DDLS. However, IHC-mediated detection of MDM2 protein is useful only for the diagnosis of DDLS.     

Molecular abnormalities of p53, MDM2, and H-ras in synovial sarcoma.

Forty-nine cases of synovial sarcoma were evaluated for mutation of the p53 gene, amplification of the MDM2 gene and mutation of the H-ras gene, and for the relation of these factors to overall survival and clinicopathologic parameters. All investigations were carried out on formalin-fixed paraffin-embedded materials. Furthermore, we evaluated the expression of p53 protein, MDM2, and p21(WAF1/CIP1) immunohistochemically in these cases, together with an assessment of proliferative activities using monoclonal antibody MIB-1. Nine of the 49 cases (18.4%) had p53 gene alteration detected by polymerase chain reaction-single-strand conformation polymorphism (PCR-SSCP) and direct sequencing. Eleven cases (24%) showed nuclear accumulation of p53 protein in more than 10% of the tumor cells. Among them, only three cases contained gene mutations. There was no correlation between p53 nuclear accumulation and p53 gene alteration. MDM2 gene amplification, as shown by differential PCR, was observed in 19 out of 47 cases (40%). Nineteen out of 49 cases (38.8%) showed immunoreactivity for MDM2. MDM2 gene amplification and the expression of MDM2 protein showed a significant positive relationship (P = 0.0004). Moreover, MDM2 immunoreaction was significantly correlated with nuclear accumulation of p53 protein (P = 0.023). Positive immunoreaction for p21(WAF1/CIP1) was observed in 21 out of 48 cases (43.8%). p21(WAF1/CIP1) expression was correlated with p53 protein expression. H-ras gene mutations were seen in only three cases (6.1%). All mutations were in codon 12 (one GGC-to-AGC [Gly-to-Ser] mutation and two GGC-to-GAC [Gly-to-Ap] mutations). The gene alteration of p53, MDM2, and H-ras did not affect the patients' prognosis. Although the cases with positive immunoreaction for p53 tended to have a worse prognosis, the difference was not statistically significant (P = 0.13). No correlation was observed between MIB-1 LI and the immunohistochemical expression of p53, MDM2, and p21(WAF1/CIP1) or the mutation status of p53 and H-ras. On the other hand, high MIB-1 LI (more than 10) significantly correlated with poor prognosis (P < 0.0001). Our results suggest that p53 gene mutation does not appear to be a major prognostic factor and H-ras mutations are infrequent in synovial sarcoma.     

MDM2 and CDK4 gene amplification in Ewing's sarcoma

Amplification of the MDM2 gene, which maps to chromosome band 12q13 and encodes a p53-binding protein, may result in functional inactivation of p53 and has been observed in various bone and soft tissue sarcomas. Published studies have included few cases of Ewing's sarcoma (ES) or peripheral neuroectodermal tumour (PNET), a tumour group in which alterations of the p53 pathway have so far not been extensively studied. We examined two ES cell lines, RD-ES and SK-ES-1, and 30 specimens from 27 patients (24 ES, 6 PNET; 19 primary, 4 local recurrence, 7 metastasis) for MDM2 gene amplification by Southern blot analysis. All 30 clinical specimens had been confirmed to contain sufficient ES/PNET DNA by the demonstration of a rearrangement of the t(11;22)-associated EWS gene using an EWS cDNA probe on the same blots. MDM2 gene amplification was detected in 3 of 30 specimens (10 per cent), including two ES and one PNET, but in neither of the cell lines. The three cases with amplification were morphologically typical primary tumours. Two of the three cases also showed co-amplification of the CDK4 gene, which endoces a cyclin-dependent kinase and also maps to band 12q13. Clinically, all three cases had metastatic disease at diagnosis, compared with only 1 of 15 MDM2-negative cases where the primary tumour was studied. The difference was statistically significant (P=0.005), suggesting an association of MDM2 amplification with advanced stage. Further accural and multivariate analysis of ES/PNET cases with MDM2 gene amplification will be necessary to confirm the clinical significance of these findings. The results suggest that the prevalence of MDM2 gene amplification in ES/PNET is comparable to other sarcomas, and implicate dysfunction of the p53 pathway in a subset of ES/PNET. The biological significance of CDK4 co-amplification remains to be determined.     

DISTINCT mdm2/p53 EXPRESSION PATTERNS IN LIPOSARCOMA SUBGROUPS: IMPLICATIONS FOR DIFFERENT PATHOGENETIC MECHANISMS

Recent findings have indicated that TP53 inactivation in sarcomas may result from mutation and/or deletion of the TP53 gene or, alternatively, from binding to the MDM2 gene products. To investigate further a possible role of the two genes in sarcomas, 24 large and deep-seated lipomas and 74 liposarcomas of various subtypes were analysed for mdm2 and p53 overexpression by immunocytochemistry. Nineteen cases of the same series were also molecularly analysed for both MDM2 gene amplification and TP53 mutations, and a further ten cases for non-random chromosomal abnormalities. In the retroperitoneal well-differentiated–dedifferentiated (WD–DD) group, 15/16 WD and 8/8 DD liposarcomas displayed the mdm2+/p53+ phenotype, consistent with MDM2 gene amplification in the absence of TP53 mutations. In the non-retroperitoneal WD–DD group, 5/11 WD liposarcomas also retained the mdm2+/p53+ phenotype whereas all DD liposarcomas showed an immunophenotype and, when assessed, a genotype consistent with mutant TP53. Null mdm2 immunophenotype, coupled with evidence of a specific chromosome translocation t(12;16), was constantly observed in both the usual and the cellular subtypes of myxoid liposarcoma, three cases of which also showed TP53 alterations at the genetic or protein level. Neither mdm2 nor p53 overexpression was observed in the lipomas. The results show the existence of three main pathogenetically distinct groups of liposarcoma. The first retroperitoneal WD–DD group, which represents a novel class of tumours within a single histological category of sarcoma, where MDM2-mediated inactivation of p53 could be related to the pathogenetic mechanism. The second is the non-retroperitoneal WD–DD group, where the TP53 mutations appear to correlate with the dedifferentiation process. The third is the myxoid group, which is characterized by its own unique cytogenetic profile and never shows any involvement of TP53 or MDM2 genes. As for diagnostic significance, the absence of mdm2 and p53 reactivity in lipomas seems to represent a useful marker for differential diagnosis from lipoma-like WD liposarcomas. © 1997 by John Wiley & Sons, Ltd.     

Altered expression and molecular abnormalities of cell-cycle-regulatory proteins in rhabdomyosarcoma.

Rhabdomyosarcoma is the most commonly occurring soft-tissue sarcoma in children. Some reports have discussed the altered expression and molecular abnormalities of cell-cycle-regulatory proteins in rhabdomyosarcoma; however, variable frequencies of occurrence have been noted. In the current study, among 72 cases of rhabdomyosarcoma, the authors evaluated for the expression of p53, MDM2, p16, p21/WAF1, p27, cyclin D1, cyclin E, pRb and E2F-1 protein immunohistochemically and assessed for proliferative activities using MIB-1. We also analyzed the mutation of the p53 gene in 45 cases, the amplification of the MDM2 gene in 18 cases and the mutation of the H-ras gene in 29 cases, using formalin-fixed paraffin-embedded materials. Furthermore, we assessed the correlation between clinicopathologic factors and the results of both immunohistochemical and molecular analyses. Alveolar type affected older patients, and it had a significantly higher mitotic rate compared with the embryonal type (P=0.0226). p53 overexpression was detected in 22 (30.6%) of 72 cases, and 10 (22.2%) of 45 cases had p53 gene abnormalities. As for MDM2, its overexpression was found in nine (12.5%) of 72 cases, and three (16.7%) of 18 cases showed MDM2 amplification. A statistically significant association was observed between immunoreaction for MDM2 and p53 overexpression (P=0.0002), and p53 and MDM2 overexpression was significantly correlated with high MIB-1 labeling indices. E2F-1 labeling indices showed a significantly higher score in alveolar type compared with that seen in embryonal type (P=0.0334), but MIB-1 did not. In conclusion, our study suggests that p53 overexpression may be related to tumor progression because tumors with p53 overexpression have a high proliferative activity in the current study. Alveolar type had a significantly higher both mitotic rate and E2F-1 labeling indices when compared with the embryonal type. The current study is the first report of the correlation of E2F-1 with alveolar rhabdomyosarcoma.     

Overexpression of p21WAF1/CIP1 and MDM2 characterizes serous borderline ovarian tumors

Ovarian epithelial tumors are classically divided into benign, malignant, and borderline or of low malignant potential. It is controversial whether this last group of tumors should be considered benign or malignant. Expression of cell cycle markers has recently been linked to tumor behavior and response to treatment. It has been shown that one of the pathways through which the p53 gene controls the cell cycle is by transactivating p21WAF1/CIP1, a cyclin-dependent kinase (cdk) inhibitor. By inhibiting cdks, p21WAF1/CIP1 blocks the G-1 to S-phase transition in the cell cycle. p53 can be regulated by MDM2 (murine double minute-2) through direct inactivation or promotion of its cytoplasmic degradation. In an attempt to investigate the cell cycle checkpoint mechanisms of these tumors, we studied the expression of p53, Ki-67, MDM2, and p21WAF1/CIP1 by immunohistochemistry. We analyzed the expression of these proteins in 19 cystadenomas (8 serous and 11 mucinous), 40 borderline tumors (31 serous and 9 mucinous), and 18 serous carcinomas of the ovary. p21WAF1/CIP1 was expressed in 7 of 19 (37%) benign cystadenomas, 32 of 40 (80%) borderline tumors (93.5% of serous and 33% of mucinous), and in 9 of 18 (50%) serous carcinomas. Ki-67 was only weakly expressed in 8 of 19 (42%) benign cystadenomas, all borderline tumors showed Ki-67 staining in less than 50% of the cells, and 55% of serous carcinomas stained in more than 50% of tumor cells. p53 was absent in all but 1 of the cystadenomas, was expressed in 9 of 40 (22.5%) borderline tumors (25.8% of serous and 11% of mucinous), and in 10 of 18 (55%) carcinomas. All 11 implants of serous borderline tumors expressed p21WAF1/CIP1. Most serous borderline tumors expressed higher levels of MDM2 compared with the benign cystadenomas and carcinomas. Four of the serous borderline implants (40%) expressed MDM2. Coexpression of p21WAF1/CIP1 and MDM2 characterizes serous borderline tumors of the ovary and their implants, which suggests that these cell cycle control proteins are important in these tumors and may be related to tumor progression. Low expression of p53 protein in serous borderline tumors might be in part mediated by MDM2. This suggests that the p53 pathway is intact in most of these tumors, in contrast with carcinomas, in which high expression of p53 has been related to mutations of this gene.     

Analysis of glioma cell lines for amplification and overexpression of MDM2

Recently, amplification of the gene encoding a p53 binding protein, MDM2, was determined in 8% of the cases constituting a large series of glioblastomas. Here we have utilized Southern blot analysis to examine 30 cell lines established from such tumors, and our investigation has revealed large increases in MDM2 gene dosage in two cases, one of which showed coamplification of the CDK4 gene that resides in close proximity to MDM2 in chromosomal region 12q13–14. Northern analysis demonstrated overexpression of MDM2 mRNA in the two cell lines with gene amplification, and overexpression of MDM2 protein was evident in each of these by immunohistochemical and Western blot analysis. Analysis of TP53 cDNAs revealed normal TP53 sequences in the cell lines with MDM2 amplification; these results are consistent with those of previous studies suggesting that MDM2 amplification occurs only in tumors expressing wild-type p53. In total, these data suggest that MDM2 amplification in glioblastoma cell lines occurs at a frequency (6.7%) comparable to that determined in primary tumors; occurs in cell lines expressing wild-type p53; and can involve the coamplification of additional genes.     

Overexpression of p53 protein and MDM2 in papillary carcinomas of the thyroid: Correlations with clinicopathologic features

Expression of p53 protein and MDM2 was evaluated in paraffin-embedded tissue from 78 patients with papillary carcinomas of the thyroid (PCT), in order to elucidate the relationship between them and their correlations with some clinicopathologic features implicated in tumor progression. These proteins were expressed in nuclei of tumor cells, but not in non-tumor cells. Staining was defined as positive when 10% or more of tumor cells expressed these proteins. The number of cases positive for p53 protein was 21/78 (27%), and that positive for MDM2 was 26/78 (33%). Co-overexpression of p53 protein and MDM2 was observed in 12/78 cases (15%). A significant positive relationship was found between them (P < 0.01); p53-positive cases tended to be also positive for MDM2 and vice versa. Statistical analysis revealed that overexpression of p53 protein significantly correlated with large tumor size (P = 0.0271) and the presence of capsular invasion (P = 0.04). There were significant positive correlations between tumor size and intrathyroidal invasion and between tumor size and capsular invasion in PCT, suggesting that p53 protein overexpression is associated only with tumor progression (tumor size). However, we could not find any significant correlations between MDM2 expression and clinicopathologic features. Our findings suggest that overexpression of p53 protein and MDM2 in papillary carcinoma of the thyroid is associated with the progression of the tumors, and that p53 may be a marker of the progression of PCT.     

P53/MDM2 overexpression in metastatic endometrial cancer: correlation with clinicopathological features and patient outcome

Several studies have reported that p53/mdm2 distortions play a pivotal role in the development and progression of various human malignancies. However, the number of reports having evaluated simultaneously the components of the P53-pathway alterations in advanced-stage human endometrial carcinomas (EC) is low. In this study, we examined the expression of P53/MDM2 proteins in primary and metastatic ECs, and analyzed the clinicopathological characteristics as well as the survival outcome of patients in relation to P53/MDM2 overexpression. The study group comprised 36 patients with advanced EC, whose primary and metastatic tumor slides were sufficient for analysis. Immunohistochemical assessment was made by applying anti-human P53 and MDM2 antibodies and a highly sensitive EnVision⁺/HPR visualization system. Nuclear P53 overexpression was seen in 11 (31%) primary ECs and 12 (33%) metastatic tumors. There was a significant correlation between P53 overexpression (in primary cancers and metastatic tumors) and MDM2 overexpression in metastatic tumors. Nuclear MDM2 overexpression was noted in 42% (15/36) of primary carcinomas and in 47% (17/36) of metastatic tumors. A significant association existed between MDM2 overexpression and histological grading (G1 + G2 versus G3, P = 0.043). P53/MDM2 overexpression occurred simultaneously in 7 out of 36 (19%) primary ECs and in 9 out of 36 (25%) metastatic lesions. Concomitant overexpression of these proteins was reported in 7 out of 36 (19%) cases and tended to be higher in tumors showing VSI compared to neoplasms lacking vascular space invasion (P = 0.051). P53 overexpression, either in primary ECs (P < 0.0001) or metastatic lesions (P < 0.0001), was significantly associated with poor survival in univariate analysis. Moreover, the log-rank test demonstrated that simultaneous P53/MDM2 overexpression was also correlated with decreased length of survival. There was no correlation between MDM2 overexpression and patient survival. Multivariate Cox regression analysis revealed that only P53 overexpression is an independent predictor of survival. In conclusion, our data support the view that patients with P53 overexpression are significantly associated with an unfavorable outcome, whereas MDM2 overexpression is not related to decreased survival length in women operated on for advanced-stage EC.     

High-resolution genomic profiling of adenomas and carcinomas of the salivary glands reveals amplification, rearrangement, and fusion of HMGA2

Carcinoma ex pleomorphic adenoma (Ca-ex-PA) is an epithelial malignancy developing within a benign salivary gland pleomorphic adenoma (PA). Here we have used genome-wide, high-resolution array-CGH, and fluorescence in situ hybridization to identify genes amplified in double min chromosomes and homogeneously staining regions in PA and Ca-ex-PA and to identify additional genomic imbalances characteristic of these tumor types. Ten of the 16 tumors analyzed showed amplification/gain of a 30-kb minimal common region, consisting of the 5'-part of HMGA2 (encoding the three DNA-binding domains). Coamplification of MDM2 was found in nine tumors. Five tumors had cryptic HMGA2-WIF1 gene fusions with amplification of the fusion oncogene in four tumors. Expression analysis of eight amplified candidate genes in 12q revealed that tumors with amplification/rearrangement of HMGA2 and MDM2 had significantly higher expression levels when compared with tumors without amplification. Analysis of individual HMGA2 exons showed that the expression of exons 3-5 were substantially reduced when compared with exons 1-2 in 9 of 10 tumors with HMGA2 activation, indicating that gene fusions and rearrangements of HMGA2 are common in tumors with amplification. In addition, recurrent amplifications/gains of 1q11-q32.1, 2p16.1-p12, 8q12.1, 8q22-24.1, and 20, and losses of 1p21.3-p21.1, 5q23.2-q31.2, 8p, 10q21.3, and 15q11.2 were identified. Collectively, our results identify HMGA2 and MDM2 as amplification targets in PA and Ca-ex-PA and suggest that amplification of 12q genes (in particular MDM2), deletions of 5q23.2-q31.2, gains of 8q12.1 (PLAG1) and 8q22.1-q24.1 (MYC), and amplification of ERBB2 may be of importance for malignant transformation of benign PA.