Mutated P53 Cells Research Articles

PARP-1 Expression Influences Cancer Stem Cell Phenotype in Colorectal Cancer Depending on p53

Poly(ADP-ribose) polymerase-1 (PARP-1) is a protein involved in multiple physiological processes. Elevated PARP-1 expression has been found in several tumours, being associated with stemness and tumorigenesis. In colorectal cancer (CRC), some controversy among studies has been described. In this study, we analysed the expression of PARP-1 and cancer stem cell (CSC) markers in CRC patients with different p53 status. In addition, we used an in vitro model to evaluate the influence of PARP-1 in CSC phenotype regarding p53. In CRC patients, PARP-1 expression correlated with the differentiation grade, but this association was only maintained for tumours harbouring wild-type p53. Additionally, in those tumours, PARP-1 and CSC markers were positively correlated. In mutated p53 tumours, no associations were found, but PARP-1 was an independent factor for survival. According to our in vitro model, PARP-1 regulates CSC phenotype depending on p53 status. PARP-1 overexpression in a wild type p53 context increases CSC markers and sphere forming ability. By contrast, those features were reduced in mutated p53 cells. These results could implicate that patients with elevated PARP-1 expression and wild type p53 could benefit from PARP-1 inhibition therapies, meanwhile it could have adverse effects for those carrying mutated p53 tumours.

An Observed Effect of p53 Status on the Bystander Response to Radiation-Induced Cellular Photon Emission.

In this study, we investigated the potential influence of p53 on ultraviolet (UV) signal generation and response of bystander cells to the UV signals generated by beta-irradiated cells. Five cell lines of various p53 status (HaCaT, mutated; SW48, wild-type; HT29, mutated; HCT116+/+, wild-type; HCT116-/-, null) were irradiated with beta particles from tritium. Signal generation (photon emission at 340 ± 5 nm) was quantified from irradiated cells using a photomultiplier tube. Bystander response (clonogenic survival) was assessed by placing reporter cell flasks directly superior to irradiated signal-emitting cells. All cell lines emitted significant quantities of UV after tritium exposure. The magnitudes of HaCaT and HT29 photon emission at 340 nm were similar to each other while they were significantly different from the stronger signals emitted from SW48, HCT116+/+ and HCT116-/- cells. In regard to the bystander responses, HaCaT, HCT116+/+ and SW48 cells demonstrated significant reductions in survival as a result of exposure to emission signals. HCT116-/- and HT29 cells did not exhibit any changes in survival and thus were considered to be lacking the mechanisms or functions required to elicit a response. The survival response was found not to correlate with the observed signal strength for all experimental permutations; this may be attributed to varying emission spectra from cell line to cell line or differences in response sensitivity. Overall, these results suggest that the UV-mediated bystander response is influenced by the p53 status of the cell line. Wild-type p53 cells (HCT116+/+ and SW48) demonstrated significant responses to UV signals whereas the p53-null cell line (HCT116-/-) lacked any response. The two mutated p53 cell lines exhibited contrasting responses, which may be explained by unique modulation of functions by different point mutations. The reduced response (cell death) exhibited by p53-mutated cells compared to p53 wild-type cells suggests a possible role of the assessed p53 mutations in radiation-induced cancer susceptibility and reduced efficacy of radiation-directed therapy.

Dose Response of Soft X-Ray-Induced Bystander Cell Killing Affected byp53Status

A radiation-induced bystander response, which is generally defined as a cellular response that is induced in nonirradiated cells that received bystander signals from directly irradiated cells within an irradiated cell population. In our earlier X-ray microbeam studies, bystander cell killing in normal human fibroblasts had a parabolic relationship to the irradiation dose. To elucidate the role of p53 in the bystander cell killing, the effects were assessed using human non-small cell lung cancer cells expressing wild-type or temperature-sensitive mutated p53. The surviving fraction of bystander wild-type p53 cells showed a parabolic relationship to the irradiation dose; survival was steeply reduced up to 0.45 Gy, recovered toward to 2 Gy, and remained at control levels up to 5 Gy. In contrast, in the mutated p53 cells at a nonpermissive temperature, the surviving fraction was steeply reduced up to 1 Gy and remained at the reduced level up to 5 Gy. When the mutated p53 cells were incubated at a permissive temperature, the decrease in the surviving fraction at 2 Gy was suppressed. The wild-type p53 cells were not only restrained in releasing bystander signals at 2 Gy, but were also resistant to the signals released by the mutated p53 cells. These results suggest that the X-ray-induced bystander cell killing depends on both the irradiation dose and the p53 status of the targeted cells and the bystander cells.

Expression of p53-regulated genes in human cultured lymphoblastoid TSCE5 and WTK1 cell lines after spaceflight in a frozen state

The 53 kDa tumor suppressor protein p53 is generally thought to contribute to the genetic stability of cells and to protect cells from DNA damage through the activity of p53-centered signal transduction pathways. To clarify the effect of space radiation on the expression of p53-dependent regulated genes, gene expression profiles were compared between two human cultured lymphoblastoid cell lines: one line (TSCE5) has a wild-type p53 gene status, and the other line (WTK1) has a mutated p53 gene status. Frozen human lymphoblastoid cells were stored in a freezer in the International Space Station (ISS) for 133 days. Gene expression was analyzed using DNA chips after culturing the space samples for 6 h on the ground after their return from space. Ground control samples were also cultured for 6 h after being stored in a frozen state on the ground for the same time period that the frozen cells were in space. p53-Dependent gene expression was calculated from the ratio of the gene expression values in wild-type p53 cells and in mutated p53 cells. The expression of 50 p53-dependent genes was up-regulated, and the expression of 94 p53-dependent genes was down-regulated after spaceflight. These expression data identified genes which could be useful in advancing studies in basic space radiation biology. The biological meaning of these results is discussed from the aspect of gene functions in the up- and down-regulated genes after exposure to low doses of space radiation.

Abstract 2026: MicroRNA-214 is a negative regulator of p53

Abstract The p53 transcription factor mediates multiple cellular functions including the response to genotoxic stress, differentiation, cell cycle and apoptosis. Since p53 inactivation is a critical event in tumor development, the regulation of p53 and its pathway is of prime importance in the understanding of tumorigenesis. While a number of microRNAs have been shown to be regulated by p53, microRNA regulation of p53 is still not well documented. Here we demonstrate that miR-214, a putative oncogenic miRNA, regulates p53 protein but not mRNA level by direct targeting 3′UTR of p53. Ectopic expression of miR-214 represses p53 and enhances the resistance to radiation- or chemotherapeutic agent-induced cell cycle arrest and apoptosis in wild-type but not mutated p53 cells. Introduction of p53 cDNA lacking 3′UTR largely abrogates miR-214-induced cell growth and survival. Depletion of endogenous miR-214 elevates protein level of p53 and induces apoptosis in human cancer cell lines expressing wild-type p53. These findings indicate that miR-214 is an important posttranscriptional regulator of p53 and exerts its oncogenic activity through targeting p53. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2026.

Protein microarray analysis of apoptosis-related protein expression following heat shock in human tongue squamous cell carcinomas containing different p53 phenotypes

Purpose: Hyperthermia is useful in the treatment of human head and neck cancers, because it is relatively easy to regulate temperatures when compared to tumors located in deep organs. In this study, attention was focused on p53 as a possible predictive indicator for the efficacy of hyperthermic cancer therapy.Methods: Two kinds of cell lines were used. These were derived from a human squamous cell carcinoma (SAS) and had identical genetic backgrounds except for their p53 gene status. It was previously reported that the heat sensitivity and frequency of apoptosis in wild-type p53 cells (SAS/neo) were clearly elevated when compared with mutated p53 cells (SAS/mp53). In order to study the expression of apoptosis related proteins after heat treatment, protein microarray analysis was used.Results: The expression of apoptosis inhibitory proteins such as Bcl-2, Bcl-xL, NF-κB, COX2, STAT3, IL-6, and IKKα/1 was seen to increase after heat treatment in SAS/mp53 cells, but not in SAS/neo cells.Conclusion: The result of these observations indicates that apoptosis inhibitory proteins (such as Bcl-2, Bcl-xL, IL-6, etc.) were highly induced in SAS/mp53 cells after heat treatment when compared to control SAS/neo cells.

The Role of p53 Molecule in Radiation and Hyperthermic Therapies

In recent years, cancer-related genes have been analyzed at the molecular level as predictive indicators for cancer therapy. Among those genes, the tumor suppressor gene p53 is worthy of notice in cancer therapy, because the p53 molecule prevents the malignant degeneration of non-cancer cells by regulating cell-cycle arrest, apoptosis, and DNA repair. An abnormality of the p53 gene introduces a genetic instability and increases the incidence of carcinogenesis and teratogenesis. Therefore, p53 is called a guardian of the genome. Mutations of p53 are observed at a high frequency in human tumors, and are recognized in about half of all malignant tumors in human head and neck cancers. We previously reported that radio- and heat-sensitivities of human cultured tongue squamous cell carcinoma cells are p53-dependent, and are closely correlated with the induction of apoptosis. In a human cell culture system, the interactive hyperthermic enhancement of radiosensitivity was observed in wild-type p53 cells, but not in mutated p53 cells. In a transplanted tumor system, the combination therapies of radiation and hyperthermia induced efficient tumor growth depression and apoptosis in the wild-type p53 tumors. In this review, we discuss the p53 activation signaling pathways through the modification of p53 molecules, such as phosphorylation after radiation and hyperthermia treatments.

Resistance of colon cancer cells to long-term 5-fluorouracil exposure is correlated to the relative level of Bcl-2 and Bcl-X(L) in addition to Bax and p53 status.

Defects in apoptosis have been implicated in chemoresistance of colon cancer cells. We report here the ability to resist to 5-fluorouracil-induced apoptosis of 8 colon cancer cell lines differing in p53 and bax status: p53(-/0)bax(+/+) for TC7, SW480, HT-29; p53(+/+)bax(-/-) for LS174T, LoVo; p53(+/+) bax(+/-) for HCT116; p53(+/+) or p53(+/0)bax(+/+) for LS513 or HCT-EB, respectively. To approximate to the in vivo therapy, the cell lines were exposed to a long-term treatment of 5-FU. The analysis of proteins implicated in the apoptotic pathway has shown that the independent analysis of p53 or bax status was not sufficient to predict the extent of drug-resistance of all cell lines. In p53(+/+) cell lines but not in p53(-/0) cell lines, a low level of the pro-apoptotic Bax protein was correlated with a greater resistance of cells to 5-FU. In addition, we found that high levels of anti-apoptotic Bcl-2 and Bcl-x(L) proteins combined with a low level of Bax were correlated to high 5-FU resistance of wild-type p53 cell lines. The same correlation was obtained for 2 out of 3 mutated p53 cell lines. In conclusion, the relative levels of Bcl-2, Bcl-x(L) and Bax may altogether contribute to determine the resistance of a majority of colon tumor cells to long-term 5-FU treatment, whatever their p53 status.

Cytotoxic effects of topotecan combined with various active G2/M-phase anticancer drugs in human tumor-derived cell lines.

Topotecan (TPT) is a DNA-Topoisomerase I poison that exhibits antitumor activity. TPT, like other DNA-damaging agents, arrests or delays cell cycle progression during S- and G2-phase in a wide variety of tumor-derived cell lines. Particularly, the G2-arrest gives time for the cell to repair its DNA lesions prior to starting a new cell cycle. Based on these observations, we assessed the interaction between TPT and G2/M-active agents in p53-mutated cell lines of diverse origin in order to achieve cell toxicity. Two short-term sequential schedules were administered (TPT --> G2/M-active drug at the interval of greatest TPT-induced G2/M-phase cell arrest, and G2/M-active drug --> TPT), in three human tumor-derived cell lines with proven sensitivity to the following drugs: Bleomycin in HEp-2 (squamous larynx carcinoma); Docetaxel in SKBr-3 (breast adenocarcinoma); Etoposide in NCI-H23 (non-small-cell lung cancer). Our results show that: 1) Sequential TPT --> G2/M-active drugs are synergistic when administration overlapped the maximum percentage of TPT-induced G2/M-phase cell arrest interval in all three mutated p53 cell lines; 2) the reverse sequential schedule (G2/M-active drug --> TPT) was antagonistic, and being only additive for Etoposide --> TPT association. In conclusion, our findings further support the potential cytotoxic role of TPT in combination with other active drugs when the correct schedule of administration is applied. In addition, they provide a rationale for new applications in clinical trials using short-term sequential TPT --> G2/M-active drugs.