Single-strand conformational polymorphism (SSCP)-detected p53 gene mutations are a less sensitive marker of malignancy in pleural fluids than p53 immunostaining

p53 immunostaining has been advocated as a marker of malignancy in pleural biopsies and serous fluids. The object of this study was to compare the sensitivity and specificity of p53 immunostaining for the detection of malignant cells in pleural fluids with a technique designed to detect p53 gene mutations in exons 5, 6, 7 and 8 by SSCP and nucleotide sequencing. Five out of eight pleural fluids containing adenocarcinoma showed p53 immunostaining and two of these also showed polymorphisms on SSCP and a mutation on sequencing. None of the 10 benign pleural fluids showed immunostaining for p53 or polymorphisms on SSCP. We believe that the poor sensitivity of p53 gene mutation by SSCP is mainly due to DNA from the background reactive cells 'swamping' the mutant DNA. We do not advocate its use as a diagnostic aid.     

Diagnostic associations of p53 immunostaining in fine needle aspiration cytology of the breast

Duplicate cytospin preparations were made from 46 symptomatic breast fine needle aspirates. One of each pair was assigned to benign or malignant categories by one experienced observer as part of the 'triple approach'patient assessment. the other was immunostained with DO7, a monoclonal antibody to recombinant p53 protein, and rated by another observer as positive or negative for nuclear staining, unaware of the cytodiagnosis. Positive controls included carcinomas known to have mutant p53, while negative controls were of the reagent substitution type. of the 26 aspirates with a benign cytodiagnosis (verified by the triple approach), 23 were p53 protein-negative and three positive. of the 20 with a malignant cytodiagnosis (histologically confirmed), six were p53 protein-negative and 14 positive (exact P <0.0001). As a diagnostic test this would give 70% sensitivity and 88% specificity.     

Role of the newer p53 family proteins in malignancy

The most recently identified members of the p53 family, p63 and p73, share certain structural and functional similarities with p53. Both p63 and p73 can bind to canonical p53-DNA-binding sites, transactivate the promoters of known p53 target genes and induce apoptosis. Despite these similarities there are many important differences. In contrast to p53, p63 and p73 give rise to multiple distinct protein isoforms that have different functional properties. Upstream signaling pathways involved in the activation of p63 and p73 differ from those involved in p53 activation. Only a subset of the DNA damaging agents that induce p53 can induce p73. Cellular and viral oncoproteins can discriminate between p53 and the newer family members. In addition, the levels of p63 and p73 are affected by certain states of cellular differentiation. Finally, it is becoming clear that the newest members of the p53 family are not classical tumor suppressor genes. In contrast to the high prevalence of p53 mutations in human cancers, p63 and p73 mutations are rare. Indeed, levels of p73 increase during malignant progression. In addition, unlike p53-/- mice, mice lacking p63 and p73 do not develop tumors, but instead have significant developmental abnormalities. Mutations in p63 have also been detected in humans with the ectodermal dysplastic syndrome EEC. Further studies are required to determine whether qualitative or quantitative differences in the expression of p63 and p73 isoforms are important in the development of human cancers.     

Establishment and characterization of a human cell line derived from a uterine papillary serous carcinoma with wild-type p53 function

Uterine papillary serous carcinoma is an uncommon histologic subtype of endometrial cancer that behaves aggressively and has a poor prognosis. We successfully established a uterine papillary serous carcinoma cell line. The population-doubling time was approximately 16 h. Although loss of p53 function is considered critical for the molecular pathogenesis of uterine papillary serous carcinoma, p53 was not only mutated but functionally active in this cell line. This newly established cell line should be useful for investigating the characteristics of uterine papillary serous carcinoma.     

VEGF和p53蛋白在宫颈癌中的表达

目的:探讨血管内皮生长因子VEGF和p53蛋白在宫颈癌及正常宫颈组织中的表达及意义。方法:采用免疫组化SP法检测VEGF和p53蛋白在55例宫颈癌、15对经放化疗前后的宫颈癌组织、18例正常宫颈组织中的表达情况。结果:VEGF和p53蛋白在宫颈癌中强阳性表达、正常宫颈组织中少量表达,两者之间有显著性差异(P<0.05);VEGF和p53的表达与宫颈癌临床分期显著相关(P<0.05),与分化程度显著相关(P<0.05),与放化疗前后相关(P<0.05);VEGF与p53蛋白在宫颈癌组织中阳性表达呈正相关(P<0.01)。结论:VEGF和p53蛋白在宫颈癌的演进中,尤其在宫颈癌的浸润和转移方面,很可能起着相互促进的作用。因此,联合检测VEGF和p53蛋白的表达可提高宫颈癌的转移及预测临床诊治的准确性。     

p63 is a suppressor of tumorigenesis and metastasis interacting with mutant p53

p53 mutations, occurring in two-thirds of all human cancers, confer a gain of function phenotype, including the ability to form metastasis, the determining feature in the prognosis of most human cancer. This effect seems mediated at least partially by its ability to physically interact with p63, thus affecting a cell invasion pathway, and accordingly, p63 is deregulated in human cancers. In addition, p63, as an 'epithelial organizer', directly impinges on epidermal mesenchimal transition, stemness, senescence, cell death and cell cycle arrest, all determinant in cancer, and thus p63 affects chemosensitivity and chemoresistance. This demonstrates an important role for p63 in cancer development and its progression, and the aim of this review is to set this new evidence that links p63 to metastasis within the context of the long conserved other functions of p63.     

Determination of sensitive positions to mutations in human p53 protein

Over last several years, we demonstrated that the mutations are more likely to occur at randomly unpredictable amino acid pairs in a protein. We therefore can in principle predict the amino acid pairs sensitive to the future mutations in a protein. However, we still need to predict the positions at which the sensitive amino acid pairs are located in a protein. In this study, we use a probabilistic approach to analyze the effect of 191 mutations in human p53 protein and can approximately estimate the sensitive positions to mutations in human p53 protein.     

Increased expression of SIRT2 is a novel marker of cellular senescence and is dependent on wild type p53 status

Sirtuins (SIRT) belonging to the NAD+ dependent histone deacetylase III class of enzymes have emerged as master regulators of metabolism and longevity. However, their role in prevention of organismal aging and cellular senescence still remains controversial. In the present study, we now report upregulation of SIRT2 as a specific feature associated with stress induced premature senescence but not with either quiescence or cell death. Additionally, increase in SIRT2 expression was noted in different types of senescent conditions such as replicative and oncogene induced senescence using multiple cell lines. Induction of SIRT2 expression during senescence was dependent on p53 status as depletion of p53 by shRNA prevented its accumulation. Chromatin immunoprecipitation revealed the presence of p53 binding sites on the SIRT2 promoter suggesting its regulation by p53, which was also corroborated by the SEAP reporter assay. Overexpression or knockdown of SIRT2 had no effect on stress induced premature senescence, thereby indicating that SIRT2 increase is not a cause of senescence; rather it is an effect linked to senescence-associated changes. Overall, our results suggest SIRT2 as a promising marker of cellular senescence at least in cells with wild type p53 status.     

Pharmacological targeting of p53 through RITA is an effective antitumoral strategy for malignant pleural mesothelioma

Malignant mesothelioma, a very aggressive tumor associated to asbestos exposure, is expected to increase in incidence, and unfortunately, no curative modality exists. Reactivation of p53 is a new attractive antitumoral strategy. p53 is rarely mutated in mesothelioma, but it is inactivated in most tumors by the lack of p14^ARF. Here, we evaluated the feasibility of this approach in pleural mesothelioma by testing RITA and nutlin-3, two molecules able to restore p53 function through a different mechanism, on a panel of mesothelioma cell lines representing the epithelioid (NCI-H28, NCI-H2452, IST-MES 2), biphasic (MSTO-211H), and sarcomatoid (NCI-H2052) histotypes compared with the normal mesothelial HMC-hTERT. RITA triggered robust caspase-dependent apoptosis specifically in epithelioid and biphasic mesothelioma cell lines, both through wild-type and mutant p53, concomitant to p21 downregulation. Conversely, nutlin-3 induced a p21-dependent growth arrest, rather than apoptosis, and was slightly toxic on HMC-hTERT.

Interestingly, we identified a previously undetected point mutation of p53 (p.Arg249Ser) in IST-MES 2, and showed that RITA is also able to reactivate this p53 mutant protein and its apoptotic function. RITA reduced tumor growth in a MSTO-211H-derived xenograft model of mesothelioma and synergized with cisplatin, which is the mainstay of treatment for this tumor.

Our data indicate that reactivation of p53 and concomitant p21 downregulation effectively induce cell death in mesothelioma, a tumor characterized by a high intrinsic resistance to apoptosis. Altogether, our findings provide the preclinical framework supporting the use of p53-reactivating agents alone, or in combination regimens, to improve the outcome of patients with mesothelioma.     

抑癌基因p53与肿瘤研究的最新进展

p53基因是迄今为止已发现的与人类肿瘤发生相关性最高的抑癌基因,其主要生物学功能是通过调控DNA修复、细胞周期停滞和诱导细胞凋亡,维持基因组和细胞稳定,抑制肿瘤生长;肿瘤血管再生、微小RNA（microRNA,miRNA）及肿瘤干细胞是近几年来肿瘤发生机理研究领域的热点,本文综述了p53基因在肿瘤血管再生、miRNA、肿瘤干细胞中作用的最新研究进展及其在肿瘤治疗中的应用。     

鼻咽癌组织p53相关蛋白的纯化

鼻咽癌是南方常见的恶性肿瘤,ＮＰＣ与ＥＢ病毒关系十分密切。通过免疫亲和层析的方法了解ＮＰＣ肿瘤标本中ｐ５３与病毒或细胞蛋白间潜在的相互作用。这种相互作用可能引起Ｐ５３在ＮＰＣ组织中累积。建立单克隆抗体ｐＡｂ１８０１,ｐＡｂ２４０１免疫亲和层析柱,从ＮＰＣ转移淋巴结分离ｐ５３结合蛋白。     

Selenium activates p53 and p38 pathways and induces caspase-independent cell death in cervical cancer cells

The mechanisms of sodium selenite-induced cell death in cervical carcinoma cells were studied during 24 h of exposure in the HeLa Hep-2 cell line. Selenite at the employed concentrations of 5 and 50 μmol/L produced time- and dose-dependent suppression of DNA synthesis and induced DNA damage which resulted in phosphorylation of histone H2A.X. These effects were influenced by pretreatment of cells with the SOD/catalase mimetic MnTMPyP or glutathione-depleting buthionine sulfoximine, suggesting the significant role of selenite-generated oxidative stress. Following the DNA damage, selenite activated p53-dependent pathway as evidenced by the appearance of phosphorylated p53 and accumulation of p21 in the treated cells. Concomitantly, selenite activated p38 pathway but its effect on JNK was very weak. p53- and p38-dependent signaling led to the accumulation of Bax protein, which was preventable by specific inhibitors of p38 (SB 203580) and p53 (Pifithrin-α). Mitochondria in selenite-treated cells changed their dynamics (shape and localization) and released AIF and Smac/Diablo, which initiated caspase-independent apoptosis as confirmed by the caspase-3 activity assay and the low effect of caspase inhibitors z-DEVD-fmk and z-VAD-fmk on cell death. We conclude that selenite induces caspase-independent apoptosis in cervical carcinoma cells mostly by oxidative stress-mediated activation of p53 and p38 pathways, but other selenite-mediated effects, in particular mitochondria-specific ones, are also involved.     

The diagnostic and molecular characteristics of malignant mesothelioma and ovarian/peritoneal serous carcinoma

B. Davidson The diagnostic and molecular characteristics of malignant mesothelioma and ovarian/peritoneal serous carcinoma Malignant mesothelioma and ovarian/peritoneal serous carcinoma are two of the most common tumours affecting the serosal cavities. Unlike other malignant tumours diagnosed at this anatomical site, such as lung and breast carcinoma, malignant mesothelioma and serous carcinoma share a common histogenesis, may be difficult to differentiate morphologically, and co‐express many of the diagnostic markers used by cytopathologists in effusion diagnosis. Selected markers have nevertheless shown sufficient sensitivity and specificity to differentiate serous carcinoma from malignant mesothelioma effectively. Recently, our group applied high throughput technology to the identification of new markers that may aid in differentiating these two cancer types and validated several of these markers in follow‐up studies. This review will present current data regarding the diagnostic and biological aspects of malignant mesothelioma and ovarian/peritoneal serous carcinoma.     

宫颈癌中p53表达和病毒感染的研究

应用PCR对121例宫颈脱落细胞和组织分别检测HPV-16／18和HSV-2DNA。同时应用免疫组化S-P法检测76例宫颈组织中P53过度表达。结果发现,宫颈癌组织中HPV-16／18和HSV-2阳性率分别为61．3％和32．3％,慢性宫颈炎组HPV-16／18和HSV-2阳性率分别为22．5％和20．0％,与正常宫颈组比较均有显著性差异。宫颈癌中HPV-16／18和HSV-2混合感染率为16．1％。p53过度表达率在慢性宫颈炎、宫颈上皮内瘤变(CIN)、宫颈癌组织中呈梯度递增。另外,宫颈癌组织中P53过度表达与HPV-16／18、HSV-2的感染无相关性。提示：宫颈癌的发生与HPV-16／18关系密切,HSV-2可能与HPV-16／18协同作用导致宫颈癌的发生。宫颈组织中p53过度表达率与宫颈癌的进程有关,这在宫颈癌的防治方面有一定的临床意义。