p53 siRNA - a therapeutic tool with significant implication in modulation of apoptosis and angiogenic pathways (original) (raw)
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Suppression of p53R2 gene expression with specific siRNA sensitizes HepG2 cells to doxorubicin
Gene, 2017
p53R2 is a p53-inducible protein that contributes to DNA repair by providing dNTPs in response to DNA damage. The roles of p53R2 in cancer cells and malignancies still remain controversial. Herein, we examined the effects of p53R2 silencing on HepG2 human hepatocellular carcinoma (HHC) cell line (wild-type p53) viability, apoptosis and cell cycle arrest in the presence and absence of doxorubicin. Cell transfection was performed using a liposomal approach. Gene silencing was determined by quantitative real-time PCR and western blot analysis. To evaluate the cell growth rate after transfection, trypan blue dye exclusion assay was employed. The cytotoxicity of the doxorubicin and p53R2 siRNA as single agents or in combination against HepG2 cell was analyzed by MTT assay and the drug combination effects was evaluated by calculating the combination index. The effects of treatments on different stages of cell cycle were analyzed by flow cytometry using propidium iodide (PI) and induction ...
Cancer Research, 2006
p53 is degraded in cervical cancer cells by the human papillomavirus E6 and can be stabilized with short interfering RNA (siRNA) molecules targeting E6 mRNA. In this in vitro study, we show that E6 siRNA-induced p53 activation is transient in HeLa cervical cancer cells despite continuous suppression of E6 mRNA; activation can be sustained if the endogenous p53 antagonists COP1, MDM2, Pirh2, and c-Jun-NH 2 -kinase are also targeted by siRNAs or by inhibiting the nuclear export of p53 with leptomycin B. The direct targeting of any one of these four cellular p53 antagonists had no effect on p53 activity when E6 was intact, but inhibited the fading off of E6 siRNA-induced p53 activation in nonstress conditions. The effect was additive when multiple cellular antagonists were concomitantly inhibited, indicating that all these proteins degrade p53 when E6 is inactivated. The antiproliferative effect induced by E6 silencing was enhanced when the endogenous p53 antagonists were additionally targeted. In conclusion, if human papillomavirus E6 is inhibited under nonstress conditions, the subsequent p53 activation is quickly reversed by the endogenous p53 degenerative machinery. The present results indicate that several cellular p53 antagonists must be inhibited for sustained p53 activity if E6 siRNA therapy is attempted and if no combined genotoxic therapy is applied. (Cancer Res 2006; 66(24): 11817-24) Requests for reprints: Sakari Hietanen,
Augmentation of antitumor effects of p53 Gene therapy by combination with HDAC inhibitor
Cancer Biology & Therapy, 2005
We have previously shown that the HDAC inhibitors (HDACI) activate the p53 molecule through acetylation of 320 and 373 lysine residues, upregulate PIG3 and NOXA and induce apoptosis in cancer cells expressing wild and pseudo-wild type p53 genes (Terui T, et al. Cancer Res 2003; 63:8948-54). It has also been reported that expression of the Coxackie adenovirus receptor and subsequent transfection efficiency of the adenovirus in cancer cells were enhanced by HDACI treatment. In this study, we extended these observations to explore the combination effect of adenoviral vector carrying wild type p53 (Ad-p53) gene therapy with a HDACI, sodium butyrate (SB), on xenografted human gastric cancer cells (KATO-III) and hepatocellular carcinoma cells (HuH7) in nude mice. We first confirmed an increased expression of Coxackie adenovirus receptors with an associated increment of transgene (X-gal) expression by SB treatment in KATO-III cells. We then injected Ad-p53 into subcutaneous tumors of KATO-III and HuH7 combined with intraperitoneal administration of SB and found a significantly higher growth suppressive effect than single treatments of each. Even a complete regression of tumors was observed in three of five mice treated with this combination while with single treatment no tumor regression was observed. Tumors treated with the combination showed higher numbers of TUNEL positive cells than those treated with a single modality. Moreover, necrotic changes were more evident in tumors treated with the combination than separately, a compatible finding to the observation that vascularity revealed by CD34 staining was poorer in tumors treated with the combination than those treated with p53 gene or SB alone. This was further supported by the finding that BAI-1 (brain specific angiogenesis inihibitor-1), an inhibitor of vascularization, was induced by SB treatment in KATO-III and HuH7 cells transfected with Ad-p53. Thus SB was shown to be an efficient potentiator of p53 gene therapy for cancer.
Biological research, 2014
Vascular endothelial growth factor (VEGF) is involved in the growth of new blood vessels that feed tumors and kinesin spindle protein (KSP) plays a critical role in mitosis involving in cell proliferation. Simultaneous silencing of VEGF and KSP, an attractive and viable approach in cancer, leads on restricting cancer progression. The purpose of this study is to examine the therapeutic potential of dual gene targeted siRNA cocktail on human hepatocellular carcinoma Hep3B cells. The predesigned siRNAs could inhibit VEGF and KSP at mRNA level. siRNA cocktail showed a further downregulation on KSP mRNA and protein levels compared to KSP-siRNA or VEGF-siRNA, but not on VEGF expression. It also exhibited greater suppression on cell proliferation as well as cell migration or invasion capabilities and induction of apoptosis in Hep3B cells than single siRNA simultaneously. This could be explained by the significant downregulation of Cyclin D1, Bcl-2 and Survivin. However, no sigificant diffe...
Cell Cycle, 2008
Mutant p53 proteins are thought to have acquired a "gain of function" (GOF) activity that mainly contributes to tumor aggressiveness. Previously we reported that constitutive downregulation of mutant p53 by RNA interference reduces the tumorigenicity of cancer cells in an animal model; however, effects of adaptation to long-term mutant p53 inhibition could not be excluded. To address this point, mimicking more physiological conditions, we now describe the establishment of a lentiviral-based system for conditional interference with mutant p53 expression. In vivo studies assessed the efficacy of conditional RNA interference in inhibiting gain of function activity of mutant p53 proteins by reducing tumor growth ability. Moreover by using this system, microarray data were validated in vitro and in vivo and putative mutant p53 target genes that may contribute to its gain of function effects in cancer were identified. Results are confirmatory that depletion of mutant p53 protein impacts on tumor malignancy and validated the inducible lentiviral-based system as an efficient tool to study the gain of function activity of human tumor derived p53 mutants.
Inhibition of tumor angiogenesis by p53: a new role for the guardian of the genome
Journal of Molecular Medicine, 2007
The p53 tumor suppressor protein has long been recognized as the central factor protecting humans from cancer. It has been famously dubbed "the guardian of the genome" due to its ability to respond to genotoxic stress, such as DNA damage and other stress signals, and to protect the genome by inducing a variety of biological responses including DNA repair, cell cycle arrest, and apoptosis. However, the tumor suppressive effects of p53 go far beyond its roles in mediating these three processes. There is growing evidence that p53 also exerts its effects on multiple aspects of tumor formation, including suppression of metastasis and, as summarized in this review, inhibition of new blood vessel development (angiogenesis). The p53 protein has been shown to limit angiogenesis by at least three mechanisms:
Control of cell migration: a tumour suppressor function for p53
Biology of The Cell, 2006
Much remains to be learned about how cancer cells acquire the property of migration, a prerequisite for invasiveness and metastasis. Loss of p53 functions is assumed to be a crucial step in the development of many types of cancers, leading to dysregulation of cell cycle checkpoint controls and apoptosis. However, emerging evidence shows that the contribution of the tumour suppressor p53 to the control of tumorigenesis is not restricted to its wellknown anti-proliferative activities, but is extended to other stages of cancer development, i.e. the modulation of cell migration. This interesting alternative function has been proposed in light of the effect of p53 on specific features of migrating cells, including cell spreading, establishment of cell polarization and the production of protrusions. The effects of p53 on cell motility are largely mediated through the regulation of Rho signalling, thereby controlling actin cytoskeletal organization. These recent studies connect the regulation of proliferation to the control of cell migration and define a new concept of p53 function as a tumour suppressor gene, suggesting that p53 might be involved in tumour invasion and metastasis. This review focuses on emerging data concerning the properties of p53 that contribute to its atypical role in the regulation of cell migration.
2018
BACKGROUND: Angiogenesis is vital for development of normal tissue and wound healing; but it play an important role in development of some diseases such as different types of cancer. Vascular endothelial growth factor (VEGF) and VEGF receptors (VEGFR) are two important key factors in this process. Previous studies have shown that downregulation of VEGFR1 inhibits cell proliferation, migration, and vascular permeability of endothelial cells. So, blocking VEGF and VEGFR1 have been considered as a target to prevent the growth of tumors. METHODS: In this study, VEGFR1 gene expression was suppressed in AGS and SNU5 cancer cells using RNA interference (RNAi) technology. Down-regulation of VEGFR1 was assessed at mRNA and protein levels using realtime polymerase chain reaction (PCR), and western blot methods. Moreover, the viability and apoptosis of these cells were analyzed using MTT and flow cytometry techniques. RESULTS: VEGFR1 expression was significantly down-regulated both in mRNA and...
p53 Inhibits Angiogenesis by Inducing the Production of Arresten
Cancer Research, 2012
Several types of collagen contain cryptic antiangiogenic noncollagenous domains that are released upon proteolysis of extracellular matrix (ECM). Among those is Arresten, a collagen-derived antiangiogenic factor (CDAF) that is processed from a1 collagen IV. However, the conditions under which Arresten is released from collagen IV in vivo or whether the protein functions in tumor suppressor pathways remain unknown. Here, we show that p53 induces the expression of a1 collagen IV and release of Arresten-containing fragments from the ECM. Comparison of the transcriptional activation of COL4A1 with other CDAF-containing genes revealed that COL4A1 is a major antiangiogenic gene induced by p53 in human adenocarinoma cells. p53 directly activated transcription of the COL4A1 gene by binding to an enhancer region 26 kbp downstream of its 3 0 end. p53 also stabilized the expression of full-length a1 collagen IV by upregulation of a(II) prolyl-hydroxylase and increased the release of Arresten in the ECM through a matrix metalloproteinase (MMP)-dependent mechanism. The resulting upregulation of a1 collagen IV and production of Arresten by the tumor cells significantly inhibited angiogenesis and limited tumor growth in vivo. Furthermore, we show that immunostaining of Arresten correlated with p53 status in human prostate cancer specimens. Our findings, therefore, link the production of Arresten to the p53 tumor suppressor pathway and show a novel mechanism through which p53 can inhibit angiogenesis. Cancer Res; 72(5);
P53: Its Role in Carcinogenesis And Also A Novel Target for Anti-Cancer Therapy
IOSR Journal of Dental and Medical Sciences
p53 is a nuclear binding phosphoprotein, which regulates the gene expressions and controls several key genes involved in gene regulation. It facilitates DNA repair. If the damage is beyond repair, p53 triggers apoptosis of the cell. p53 has been shown to regulate apoptosis in both transcription dependent and independent manner. Various types of genotoxic and non genotoxic stresses can lead to p53 activation like radiations, mutagens like Aflotoxins, benzopyrines, alkylating agents etc. and also the agents which damage mitotic spindle, cause ribonucleotide depletion, hypoxia, heat stroke, exposure to nitric acid etc. These damages are either repaired or apoptosis is triggered. p53 pathways play a critical role in prevention of carcinogenesis. It may turn oncogenic when there is loss of function, dominant negative activity and oncogenic activity of mutant p53. It plays a critical role in oncogenesis by several mechanisms, so targeting p53 is a favorite modality for developing a new approach in cancer therapeutics. The approach includes gene therapy to restore p53 function, inhibition of p53-Mdm interaction, restoration of mutant p53 to wild p53, targeting p53 family proteins, eliminating mutant p53 and p53 based vaccine. It is very interesting to study p53 molecule in this perspective because, this unique molecule, has the property to trigger oncogenesis as well as it can be targeted therapeutically to treat cancers.