The role of the p53 molecule in cancer therapies with radiation and/or hyperthermia.

In recent years, cancer-related genes have been analyzed as predictive indicators for cancer therapies. Among those genes, the gene product of a tumor suppressor gene p53 plays an important role in cancer therapy, because the p53 molecule induces cell-cycle arrest, apoptosis and depression of DNA repair after cancer therapies such as radiation, hyperthermia and anti-cancer agents. An abnormality of the p53 gene might introduce low efficiency in their cancer therapies. Mutations of p53 are observed at a high frequency in human tumors, and are recognized in about half of all malignant tumors in human. In the both systems of a human cell culture and their transplanted tumor, the sensitivities to radiation, heat and anti-cancer agents were observed in wild-type p53 cells, but not in mutated or deleted p53 cells. In this review, we discuss the p53 activation signaling pathways through the modification of p53 molecules such as phosphorylation after radiation and/or hyperthermia treatments.

Aberrant Cell Cycle Regulation in Cervical Carcinoma

Carcinoma of the uterine cervix is one of the most common malignancies among women worldwide. Human papillomaviruses (HPV) have been identified as the major etiological factor in cervical carcinogenesis. However, the time lag between HPV infection and the diagnosis of cancer indicates that multiple steps, as well as multiple factors, may be necessary for the development of cervical cancer. The development and progression of cervical carcinoma have been shown to be dependent on various genetic and epigenetic events, especially alterations in the cell cycle checkpoint machinery. In mammalian cells, control of the cell cycle is regulated by the activity of cyclin-dependent kinases (CDKs) and their essential activating coenzymes, the cyclins. Generally, CDKs, cyclins, and CDK inhibitors function within several pathways, including the p16INK4A-cyclin D1-CDK4/6-pRb-E2F, p21WAF1-p27KIP1-cyclinE-CDK2, and p14ARF-MDM2-p53 pathways. The results from several studies showed aberrant regulation of several cell cycle proteins, such as cyclin D, cyclin E, p16 INK4A, p21WAF1, and p27KIP1, as characteristic features of HPV- infected and HPV E6/E7 oncogene-expressing cervical carcinomas and their precursors. These data suggested further that interactions of viral proteins with host cellular proteins, particularly cell cycle proteins, are involved in the activation or repression of cell cycle progression in cervical carcinogenesis.

The potential roles of p53 tumor suppressor in nucleotide excision repair (NER) and base excision repair (BER)

The p53 tumor suppressor has long been envisaged to preserve genetic stability by the induction of cell cycle checkpoints and apoptosis. More recently, p53 has been implicated to play roles in DNA repair responses to genotoxic stresses. UV-damage and the damage caused by certain chemotherapeutics including cisplatin and nitrogen mustards are known to be repaired by the nucleotide excision repair (NER) pathway which is reportedly regulated by p53 and its downstream genes. There are evidences to suggest that the base excision repair (BER) induced by the base-damaging agent methyl methanesulfonate (MMS) is partially deficient in cells lacking functional p53. This result suggests that the activity of BER might be also dependent on the p53 status. In this review, we discuss the possibilities that p53 regulates BER as well as NER; these are one of the most significant potentials of p53 tumor suppressor for repairing the vast majority of DNA damages that is incurred from various environmental stresses.

Apoptosis: future directions in cancer therapy.

Cancer as a multifactorial disease results in gain of immortality due to defective apoptosis. The primary mode of cell death by apoptosis induced by various modes of treatment often fail in vivo. The in vitro environment is less complex while the in vivo environment is influenced by various external regulatory signals besides the existence of multiple, parallel and independent apoptotic pathways. Further, specific preference for an apoptotic pathway in a certain cell type would significantly alter the apoptotic responses. Identification of defects in preferred pathways and choosing alternative and potentially inducible pathways would help in deciding on apoptosis-based treatment protocols. Mechanisms involved in the execution of apoptosis may also not be unique to apoptotic pathways since similar events, possibly with strict control, do occur during mitosis. Further evaluation may yield new dimensions to apoptosis and apoptosis-based therapy.

Proteína p53 em nefroblastomas / P53 protein in nephroblastomas

A imunoexpressão do antígeno p53 foi estudada em 35 pacientes com idade média de 38 +/- 29 meses. Todos foram tratados com nefrectomia e quimioterapia, em 17 se associou também a radioterapia. O tempo médio de seguimento foi de 69 +/- 66 meses. Em 5 deles a histologia era defavorável. Em 10 casos (28,5 por cento) a marcação foi positiva, com ordem crescente de positividade para epitélio, blastema e estroma. Cinco pacientes faleceram da doença, nenhum deles com histologia desfavorável. Não se encontrou relação significante entre o padrão imunohistoquímico e os parêmetros seguintes: sobrevida dos pacientes, estádio e grau do tumor.

P53 regulation and function in normal cells and tumors

Mechanisms to protect organisms from the consequences of DNA damage include the tumor suppressor p53 pathway. p53 protein binds specifically to a DNA consensus sequence to induce growth inhibitory genes or nonspecifically to damaged sites leading to DNA repair or apoptosis. While p53 protein is susceptible to post-translational modifications and binding to other proteins, few of the modifications or associations have been demonstrated in the context of the cell. We used a novel, sensitive DNA binding assay to examine p53 proteins in lysates prepared from cells responding to DNA damage. Non-transformed progenitor keratinocytes exhibited rapid and sustained induction of activated p53 protein binding to the consensus sequence, correlated with sustained induction of a downstream target gene, the cyclin-dependent kinase inhibitor p21. Binding to a mismatched DNA probe was transient and correlated with total p53 protein in the lysate, suggesting that most or all of the endogenous p53 proteins induced after damage were capable of mismatched DNA binding. Squamous cell carcinoma (SCC) derivates showed defects in induction p53 protein after DNA damage and disproportional losses in DNA binding, compared to total p53 protein steady state levels, along with increased NDN2/p53 association. Maximum induction of endogenous p53 protein binding to the mismatched probe correlated with transcription-independent induction of apoptosis. We suggest a model in which activated forms of p53 carry out transcription-dependent functions in growth arrest and DNA repair which may vary by cell type, while most or all wild type forms of p53 are capable of binding to damaged DNA. p53 protein loss of binding to damaged DNA causes failure of cells to trigger apoptosis and may lead to resistance to chemotherapy. Implications for new directions in therapeutics include functional molecular profiling of individual patient tumors for p53 DNA binding to predict treatment response better than mutational analysis alone and targeted activation of p53 DNA damage binding in tumor cells for increasing their sensitivity to chemotherapy.

Potentials and limitations of adenovirus-p53 gene therapy for brain tumors

We investigated the antineoplastic potentials of recombinant adenovirus containing wild-type p53 cDNA (Ad5CMV-p53) for malignant gliomas. In four human glioma cell lines (U-251 and LG expressing endogenous mutant p53, and U-87 and EFC-2 expressing wild-type p53) and two rat glioma cell lines (9L and C6, each expressing mutant and wild-type p53), gene transfer efficiency determined by X-gal staining and Western blotting was varied (10-99% at 10-500 multiplicity of infection, MOI). Growth inhibitory effect was drastic (>90% at 100 MOI) in U-251 cells and only moderate or minimal in other cell lines harboring wild-type p53 or low gene transfer efficiency. Ex vivo transduction of U-251 cells with Ad5CMV-p53 suppressed the in vivo tumorigenicity of the cells. Histopathologic examination for Ad5CMV-p53 toxicity to rat brains showed inflammatory reactions in half of the tested brains at 10(8) MOI. U-251 cells were inoculated intracerebrally in nude mice and injected Ad5CMV-p53 into the tumor, in which neither the tumor suppression nor the survival benefit was observed. In conclusion, heterogeneity of the cellular subpopulations of malignant glioma in p53 status, variable and insufficient gene delivery to tumor, and adenoviral toxicity to brain at higher doses may be limiting factors to be solved in developing adenovirus-p53 gene therapy for malignant gliomas.

Potentials and limitations of adenovirus-p53 gene therapy for brain tumors

We investigated the antineoplastic potentials of recombinant adenovirus containing wild-type p53 cDNA (Ad5CMV-p53) for malignant gliomas. In four human glioma cell lines (U-251 and LG expressing endogenous mutant p53, and U-87 and EFC-2 expressing wild-type p53) and two rat glioma cell lines (9L and C6, each expressing mutant and wild-type p53), gene transfer efficiency determined by X-gal staining and Western blotting was varied (10-99% at 10-500 multiplicity of infection, MOI). Growth inhibitory effect was drastic (>90% at 100 MOI) in U-251 cells and only moderate or minimal in other cell lines harboring wild-type p53 or low gene transfer efficiency. Ex vivo transduction of U-251 cells with Ad5CMV-p53 suppressed the in vivo tumorigenicity of the cells. Histopathologic examination for Ad5CMV-p53 toxicity to rat brains showed inflammatory reactions in half of the tested brains at 10(8) MOI. U-251 cells were inoculated intracerebrally in nude mice and injected Ad5CMV-p53 into the tumor, in which neither the tumor suppression nor the survival benefit was observed. In conclusion, heterogeneity of the cellular subpopulations of malignant glioma in p53 status, variable and insufficient gene delivery to tumor, and adenoviral toxicity to brain at higher doses may be limiting factors to be solved in developing adenovirus-p53 gene therapy for malignant gliomas.

Induction of apoptosis in colon cancer cells by nonsteroidal anti-inflammatory drugs

Epidemiological studies have demonstrated that nonsteroidal anti-inflammatory drugs (NSAIDs) decrease the incidence of colon cancer. In addition, NSAIDs reduce the number and size of polyps in patients with familial adenomatous polyposis. The mechanisms of the anti-neoplastic effect of NSAIDs are still far from complete understanding, but one possible mechanism is the induction of apoptosis. Several lines of evidence suggest that NSAIDs-induced apoptosis in colon cancer cells are mediated through the cyclooxygenase (COX)-independent pathway. In this study we explored the mechanism of NSAIDs-induced apoptosis in the colon cancer cell line, HT-29. We confirmed that NSAIDs induce apoptosis in HT-29 cells irrespective of their COX-selectivity. Indomethacin enhanced the expression of p21waf-1 in HT-29 cells. However the expression of apoptosis-related genes such as Fas, bcl-2 and bax was not affected by indomethacin. Intra- and extra-cellular calcium chelators, protein tyrosine kinase (PTK) inhibitor, protein kinase A (PKA) inhibitor and protein kinase C (PKC) inhibitors did not influence indomethacin-induced apoptosis in HT-29 cells. We concluded that NSAIDs-induced apoptosis in colon cancer cells may be independent from signals transducted through [Ca++]i, PTK, PKA, PKC or the expression of apoptosis-related genes. In contrast, our results demonstrating the induction of p21waf-1 transcription by NSAIDs suggest the possible association of NSAIDs-induced apoptosis and cell-cycle control in colon cancer cells.

La proteína p53 y los oncogenes de papilomavirus humanos en la carcinogénesis del cuello uterino / P53 and human papillomavirus in the carcinogenesis of the uterine cervix

El genoma de algunos papilomavirus humanos (HPV) se encuentra presente en la mayoría de los tumores de cervix. Los productos de los oncogenes virales E6 y E7 inactivan a los productos de los genes supresores p53 y Rb. La proteína p53 controla la progresión de las células del reposo al crecimiento. Los daños al genoma celular elevan los niveles de p53 e inhiben esta progresión. Esto permite la reparación del DNA antes de su duplicación. Las células que pierden este control replican su DNA dañado, lo cual genera inestabilidad genómica. La proteína E6 de los HPV de alto riesgo interacciona con p53 e induce su degración. En las lesiones del cervix, la expresión de los oncogenes E6 y E7 podrían contribuir a la inestabilidad genómicca celular, y permitiría acumular alteraciones que las llevaran al desarrollo tumoral

Análisis inmunohistoquímico de p53 y c-erbB-2 en el carcinoma de mama / Immunohistochemical analysis of p53 and c-erbB-2 in breast cancer

We have studied by immunohistochemistry the nuclear expression of p53 and the finding of oncoprotein c-erbB-2 in a series of 85 breast carcinomas. The tissue was fixed in buffered formalin and embedded in paraffin. The primary antisera were obtained from Biogenex (USA), Bp53-12, monoclonal, used in a dilution of 1:100 for p53 and from Triton Diagnostics (USA), pAB-1, polyclonal, used in a dilution of 1:200 for c-erbB2. The detection system was the Vectastain Elite kit obtained from Vector Laboratories (USA). The results were compared with several morphological features of the tumors such as size and type of the tumor, extension of the intraductal component, nuclear grade, axillary lymph node metastasis and estrogen receptor data as well as with total and disease free survival. The oncoprotein p53 was detected in the nuclei in 25 (29.4 percent) of our cases (Fig 1). Its presence was correlated with tumor size (p < 0.01), high nuclear grade (p < 0.0001) and estrogen receptor negative tumors (p < 0.0001). There was an inverse relationship between p53 expression and 5 year disease free survival (p < 0.005). In 21 (24,7 percent) of the cases we found amplification of c-erbB-2. The staining pattern was membranous (Fig. 2). It was correlated significantly with the ductal type of invasive carcinoma (p < 0.05), an associated extensive intraductal component (p < 0.001), estrogen receptor negative tumors (p < 0.0001) and shortening of disease free survival in the patients with positive axillary lymph nodes (p < 0.005). In 9 cases we found staining for both markers without statistically significant relationship with the other features studied. We conclude that the presence of these genetic alterations demonstrated by immunohistochemistry were, in our series, unfavourable prognostic factors in invasive breast cancer.

Carcinoma temprano con epitelio atípico de vesícula biliar (Reporte de un caso con expresión aumentada del P53) / Early gallbladder carcinoma with atypical epitheelium

El gen supresor de tumor P53 se cree que juega un rol importante en la progresión de los tumores malignos a través de la mutación y la expresión aumentada. La inmunohistoquímica de la proteina P53 y el Ki-67 fueron realizados en un carcinoma temprano con epitelio atípico de vesícula biliar. La expresión aumentada de la proteina P53 fue encontrado en el área del adenocarcinoma con una distribución difusa de las células positivas (index:76+/-24 por ciento), y en las áreas de epitelio atípico con focos de células positivas, de distribución difusa (Index:30+/-14 por ciento), mientras el index del Ki-67 fue 6+/- en el área del adenocarcinoma y 6 +/- en el área del epitelio atípico en cada área correspondiente a la medida del P53. Esto sugiere que la expresión aumentada del P53 ocurre en carcinomas tempranos y/o epitelio atípico, al menos en algunos casos y que esto es un evento temprano en el desarrollo del carcinoma de vesícula biliar además de que el epitelio pueda ser parte del carcinoma.

Malignidad primaria del hígado

The most common liver tumor in USA and most of the western world is metastasic. Primary hepatic tumors can derive from hepatocytes (hepatocarcinoma), biliary ducts (cholangiocarcinoma) and Kupffer's cells (hemangiosarcoma). The most common primary liver tumor in the world is the hepatocarcinoma, this is due to the high prevalence of the lesion in many areas of Africa and Asia, although it is unusual in the western hemisphere. Risk factors for hepatocarcinoma include: HBsAg carrier state, hepatitis C infection, cirrhosis of the liver, alcoholic liver disease, aflatoxin exposure, thorium dioxide, pesticides, anabolic steroids and hemochromatosis. The regenerative nodule can be considered as a preneoplastic lesion. Several tumoral markers have been found helpful in the diagnosis of this tumor: the classic one AFP is present only in 43 percent of the cases, HBsAg in studies in Taiwan in 58 percent of non neoplastic hepatocytes, AAT in 70 percent of the patients. These marquers are expressed in the liver cell in several forms: granulofibrilar occupying most of the cytoplasm and diffuse granular mainly in malignant hepatocytes. Immunoperoxidase stain shows a thick expression of the granular antigen in regenerating cells: this is true for AFP, AAT and HBsAg. The morphology of the tumor is multicentric in cirrhotic livers or focal in normal ones. Characteristically has a notorius tendency to invade the venous system and generally can appear as: a) nodular, b) massive, c) diffuse. The most common histologic pattern is the trabecular-pseudoglandular. The OMS classification of this lesion described 12 different types. In general the prognosis is very poor with the exception of the fibrolamelar type seen in young people that frequently can be resected...