• Korean Journal of Head & Neck Oncology
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• v.22 no.1
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• pp.8-14
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• 2006

Objectives: The p53 tumor suppressor gene has a key role in cellular control mechanisms involving apoptosis and DNA repair, leading to the G1 arrest following DNA damage. Its mutation is one of the most frequent alterations in human cancers. Ki-67 is identified in replicating cells of both benign and malignant lesions, so it can be the predictor of proliferative activity. The aim of this study is to evaluate the expression of p53 and Ki-67 in salivary gland tumors. Materials and Methods: Immunohistochemical analysis was used to detect expression of p53 and Ki-67 in paraffin-embedded samples from 31 benign and 27 malignant salivary gland tumors. Results were analyzed between benign and malignant tumors and compared with the clinical parameters such as stage and recurrence in malignant tumors. Results: p53 overexpression was detected in 19.6% of benign tumors and 40.7% of malignant tumors, but there was no statistical significance. p53 was significantly expressed in Warthin's tumor(45.5%) compared with pleomorphic adenoma(5.9%). Only 5.9% of pleomorphic adenoma were positive for p53, while 60% of carcinoma ex pleomorphic adenoma were positive for p53. Ki-67 was expressed in 3.2% of benign tumors and 51.9% of malignant tumors, which showed significant higher expression in malignant tumors. In malignant tumors, p53 and Ki-67 expressions bore no correlation to stage and recurrence. Conclusion: p53 overexpression is not associated with the progression of malignant tumors, and Ki-67 overexpression can be used as biologic indicator of malignant salivary gland tumors.

• Journal of Life Science
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• v.15 no.5 s.72
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• pp.726-733
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• 2005

Histone deacetylase (HDAC) inhibitors target key steps of tumor development. They inhibit proliferation, induce differentiation and/or apoptotic cell death, and exhibit potent antimetastatic and antiangiogenic properties in cancer cells in vitro and in vivo. Although they are emerging as a promising new treatment strategy in malignancy, how they exert their effect on human non-small cell lung cancer cells is as yet unclear. The present study was undertaken to investiate the underlying mechanism of a HDAC inhibitor trichostatin A (TSA)-induced growth arrest and its effect on the cell cycle control gene products in a human lung carcinoma cell line A549. TSA treaoent induced the growth inhibition and morphological changes in a concentration-dependent manner. Treatment of A549 cells with TSA resulted in a concentration-dependent increased G1 (under 100 ng/ml) and/or G2/M (200 ng/ml) cell population of the cell cycle as determined by flow cytometry Moreover, 200 ng/ml TSA treatment significantly induced the population of sub-G1 cells (23.0 fold of control). This anti-proliferative effect of TSA was accompanied by a marked inhibition of cyclins, positive regulators of cell cycle progression, and cyclin-dependent kinases (Cdks) expression and concomitant induction of tumor suppressor p53 and Cdk inhibitors such as p21 and p27 Although further studies are needed, these findings provide important insights into the possible molecular mechanisms of the anti-cancer activity of TSA in human lung carcinoma cells.

• Applied Microscopy
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• v.32 no.4
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• pp.411-419
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• 2002

Metallothionein (MT) is a family of ubiquitous, low molecular weight ($6,000{\sim}7,000D$), cysteine-rich ($30{\sim}35%$) inducible protein with a high affinity to metal ions and has no aromatic amino acids and histidine. Some of the known functions of MT include detoxification of heavy metals and alkylating agents and neutralization of free radicals. Also, this protein has been reported to involve in tumor pathophysiology and therapy resistance. MT expression may affect a number of cellular processes including gene expression, apoptosis, proliferation and differentiation. Many reports on the physiological and biochemical properties of MT have been published, but ultrastructural reports on the localization of MT in human gastric cancer tissues are extremely rare. The present study was undertaken to examine the ultrastructural features and the localization of MT within the gastric adenocarcinoma. Ultrastructures of gastric cancer cells were characterized by the high nuclear cytoplasmic ratio, the interdigitation between cells, the irregular nucleus containing much heterochromatin and the wide distribution of free ribosomes in the cytoplasm. Immunohistochemical reaction for MT was prominent in the gastric adenocarcinoma. And the immunogold labellings were more prominent within the nucleus than the cytoplasm. Particularly, immunogold particles were numerously seen at nulcleolus or nucleolar associated heterochromatin. These results suggest that MT expression by gastric cancer cells is associated with cell proliferative activity and is possibly synthesized in the cytoplasm, and then the protein is transported into the nucleus to participate in any transcriptional steps.

• Biomedical Science Letters
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• v.12 no.2
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• pp.81-89
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• 2006

The ischemia-responsive 94 gene (irp94) encoding a 94 kDa endoplasmic reticulum resident protein was investigated its molecular properties associated with unfoled protein responses. First, the expression of irp94 mRNA was tested after the reperfusion of the transient forebrain ischemia induction at the central nervous system in three Mongolian gerbils. Second, irp94 expression in PC12 cells, which are derived from transplantable rat pheochromocytoma cultured in the DMEM media, was tested at transcriptional and translational levels. The half life of irp94 mRNA was also determined In PC12 cells. Last, the changes of irp94 mRNA expression were investigated by the addition of various ER stress inducible chemicals (A23187, BFA, tunicamycin, DTT and $H_2O_2$) and proteasome inhibitors, and heat shock. High level expression of irp94 mRNA was detected after 3 hours reperfusion in the both sites of the cerebral cortex and hippocampus of the gerbil brain. The main regulation of irp94 mRNA expression in PC 12 cells was determined at the transcriptional level. The half life of irp94 mRNA in PC12 cells was approximately 5 hours after the initial translation. The remarkable expression of irp94 mRNA was detected by the treatment of tunicamycin, which blocks glycosylation of newly synthesized polypeptides, and $H_2O_2$, which induces apoptosis. When PC12 cells were treated with the cytosol proteasome inhibitors such as ALLN (N-acetyl-leucyl-norleucinal) and MG 132 (methylguanidine), irp94 mRNA expression was increased. These results indicate that expression of irp94 was induced by ER stress including oxidation condition and glycosylation blocking in proteins. Expression of irp94 was increased when the cells were chased after heat shock, suggesting that irp94 may be involved in recovery rather than protection against ER stresses. In addition, irp94 expression was remarkably increased when cytosol proteasomes were inhibited by ALLN and MG 132, suggesting that irp94 plays an important role for maintaining the ERAD (endoplasmic reticulum associated degradation) function.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.10
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• pp.4353-4358
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• 2014

Background: PI3/AKT and NF-kB signaling pathways are constitutively active in acute myeloid leukemia and cross-talk between the two has been shown in various cancers. However, their role in acute myeloid leukemia has not been completely explored. We therefore used cell penetrating inhibitor peptides to define the contributions of AKT and NF-kB to survival and multi drug resistance (MDR) in HL-60 cells. Materials and Methods: Inhibition of AKT and NF-kB activity by AKT inhibitor peptide and NBD inhibitor peptide, respectively, resulted in decreased expression of mRNA for the MDR1 gene as assessed by real time PCR. In addition, treatment of HL-60 cells with AKT and NBD inhibitor peptides led to inhibition of cell viability and induction of apoptosis in a dose dependent manner as detected by flow cytometer. Results: Finally, co-treatment of HL-60 cells with sub-optimal doses of AKT and NBD inhibitor peptides led to synergistic apoptotic responses in AML cells. Conclusions: These data support a strong biological link between NF-kB and PI3-kinase/AKT pathways in the modulation of antiapoptotic and multi drug resistant effects in AML cells. Synergistic targeting of these pathways using NF-kB and PI3-kinase/AK inhibitor peptides may have a therapeutic potential for AML and possibly other malignancies with constitutive activation of these pathways.

• THE JOURNAL OF KOREAN ORIENTAL ONCOLOGY
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• v.6 no.1
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• pp.67-79
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• 2000

Objective : Hedyotis diffusa has been used as an anticancer agent for several decades in oriental medicine. We test whether the methanol extract of the herb affects transcriptional activation factors including $NF-{\kappa}B$ and AP-1. Methods : 1. HL-60 cells were treated with various concentrations(from 200 to $50{\mu}g/ml$) of methanol extract and $H_2O$ extract($200{\mu}g/ml$)of hedyotis diffusa, After 48h later, the cells were tested for viability by MTT assay. 2. The HL-60 cells were treated with $200{\mu}g/ml$ of methanol extract for the indicated periods. First. Nuclear extracts were isolated and incubated with oligonucleotide probe of $NF-{\kappa}B$ and AP-1. Second. Nuclear extracts were isolated and reacted with p50, p65. c-rel pan-Jun, c-Jun, JunB. JunD antibody on ice for 30min. Finally The cell lysates were prepared and analyzed by western blotting using anti-Fas, anti-FasL and anti-p53 antibody. Results : 1. The methanol extract decreases the viability of human lymphoid origin leukemia HL-60 cells in a dose-dependent manner. 2. $NF-{\kappa}B$ is rapidly activated by the addition of the methanol extract, reaches a peak at 30min and gradually returns to resting level. We confirm that $NF-{\kappa}B$ is a heterodimer mainly composed of p65 subunit with c-Rel. 3. Transcriptional activation of AP-1 is detected at 30min and reaches a maximum at 1hr after stimulation of the cells with the methanol extract. AP-1 is mainly composed with Jur-D and partially Jug-B proteins. 4. the methanol extract of Hedyotis diffusa induces the expression of Fas, Fas ligand and p53 proteins of HL-60 cells in a time dependent fashion. Conclusions : These results suggest that the methanol extract of Hedyotis diffusa exerts anticancer effects to induce the death of human leukomic HL-60 cells via activation of trascriptional factors such as $NF-{\kappa}B$ and AP-1, increase in expression of Fas mediated signalling proteins, and induction of tumor suppressor gene. p53.

• Asian Pacific Journal of Cancer Prevention
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• v.16 no.9
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• pp.3723-3727
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• 2015

The p53 tumor suppressor protein is a principal mediator of growth arrest, senescence, and apoptosis in response to a broad array of cellular damage. p53 is a substrate for the ubiquitin-proteasome system, however, the ubiquitin-conjugating enzymes (E2s) involved in p53 ubiquitination have not been well studied. UBE2Q1 is a novel E2 ubiquitin conjugating enzyme gene. Here, we investigated the effect of UBE2Q1 overexpression on the level of p53 in the MDA-MB-468 breast cancer cell line as well as the interaction between UBE2Q1 and p53. By using a lipofection method, the p53 mutated breast cancer cell line, MDA-MB-468, was transfected with the vector pCMV6-AN-GFP, containing UBE2Q1 ORF. Western blot analysis was employed to verify the overexpression of UBE2Q1 in MDA-MB-468 cells and to evaluate the expression level of p53 before and after cell transfection. Immunoprecipitation and GST pull-down protocols were used to investigate the binding of UBE2Q1 to p53. We established MDA-MB-468 cells that transiently expressed a GFP fusion proteins containing UBE2Q1 (GFP-UBE2Q1). Western blot analysis revealed that levels of p53 were markedly lower in UBE2Q1 transfected MDA-MB-468 cells as compared with control MDA-MB-468 cells. Both in vivo and in vitro data showed that UBE2Q1 co-precipitated with p53 protein. Our data for the first time showed that overexpression of UBE2Q1can lead to the repression of p53 in MDA-MB-468 cells. This repression of p53 may be due to its UBE2Q1 mediated ubiquitination and subsequent proteasome degradation, a process that may involve direct interaction of UBE2Q1with p53.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.8
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• pp.4067-4073
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• 2012

Obesity is an established risk factor for colorectal cancer. Pioglitazone is a peroxisome proliferator activated receptor$receptor{\gamma}$ ($PPAR{\gamma}$) agonist that induces differentiation in adipocytes and induces growth arrest and/or apoptosis in vitro in several cancer cell lines. In the present study, we investigated the effect of pioglitazone on the development of azoxymethane-induced colon aberrant crypt foci (ACF) in KK-$A^{\mathcal{Y}}$ obesity and diabetes model mice, and tried to clarify mechanisms by which the $PPAR{\gamma}$ ligand inhibits ACF development. Administration of 800 ppm pioglitazone reduced the number of colon ACF/mouse to 30% of those in untreated mice and improved hypertrophic changes of adipocytes in KK-$A^{\mathcal{Y}}$ mice with significant reduction of serum triglyceride and insulin levels. Moreover, mRNA levels of adipocytokines, such as leptin, monocyte chemoattractant protein-1 and plasminogen activator inhibitor-1, in the visceral fat were decreased. PCNA immunohistochemistry revealed that pioglitazone treatment suppressed cell proliferation in the colorectal epithelium with elevation of p27 and p53 gene expression. These results suggest that pioglitazone prevented obesity-associated colon carcinogenesis through improvement of dysregulated adipocytokine levels and high serum levels of triglyceride and insulin, and increase of p27 and p53 mRNA levels in the colorectal mucosa. These data indicate that pioglitazone warrants attention as a potential chemopreventive agent against obesity-associated colorectal cancer.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.11
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• pp.6605-6611
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• 2013

Background: WEE1 is a G2/M checkpoint regulator protein. Various studies have indicated that WEE1 could be a good target for cancer therapy. The main aim of this study was to asssess the tumor suppressive potential of WEE1 silencing in two different breast cancer cell lines, MCF7 which carries the wild-type p53 and MDA-MB468 which contains a mutant type. Materials and Methods: After WEE1 knockdown with specific shRNAs downstream effects on cell viability and cell cycle progression were determined using MTT and flow cytometry analyses, respectively. Real-time PCR and Western blotting were conducted to assess the effect of WEE1 inhibition on the expression of apoptotic (p53) and anti-apoptotic (Bcl2) factors and also a growth marker (VEGF). Results: The results showed that WEE1 inhibition could cause a significant decrease in the viability of both MCF7 and MDA-MB-468 breast cancer cell lines by more than 50%. Interestingly, DNA content assays showed a significant increase in apoptotic cells following WEE1 silencing. WEE1 inhibition also induced upregulation of the apoptotic marker, p53, in breast cancer cells. A significant decrease in the expression of VEGF and Bcl-2 was observed following WEE1 inhibition in both cell lines. Conclusions: In concordance with previous studies, our data showed that WEE1 inhibition could induce G2 arrest abrogation and consequent cell death in breast cancer cells. Moreover, in this study, the observed interactions between the pro- and anti-apoptotic proteins and decrease in the angiogenesis marker expression confirm the susceptibility to apoptosis and validate the tumor suppressive effect of WEE1 inhibition in breast cancer cells. Interestingly, the levels of the sensitivity to WEE1 silencing in breast cancer cells, MCF7 and MDA-MB468, seem to be in concordance with the level of p53 expression.

• Journal of Gastric Cancer
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• v.16 no.4
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• pp.221-229
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• 2016

Purpose: Dysregulated microRNAs (miRNAs) can contribute to cancer development by leading to abnormal proliferation of cells, apoptosis, and differentiation. Although several miRNAs that are related to gastric cancer have been identified, the reported results have been inconsistent. The aim of this study was to determine miRNA expression profiles and validate miRNAs up- and down-regulated in gastric cancer. Materials and Methods: We evaluated 34 primary gastric cancer tissues and paired adjacent nontumorous gastric tissues. Total RNA was extracted, and low-molecular-weight RNAs (<200 nucleotides) were isolated for further analysis. Two pairs of tissues were processed for GeneChip microarray analysis, and the identified up- and down-regulated miRNAs were validated by real-time quantitative polymerase chain reaction (qPCR). Results: In the set of differentially expressed miRNAs, 5 were overexpressed by more than 2 fold, and 5 were reduced by 2 fold or less in gastric cancer tissues compared with normal gastric tissues. Four of these miRNAs (miR-196b-5p, miR-375, miR-483-5p, and miR-486-5p) were then validated by qPCR, and the relative expression levels of 2 miRNAs (miR-196b-5p and miR-375) were significantly different between cancer and normal tissues. Conclusions: Our results revealed that the expression of miR-196b-5p and miR-375 significantly correlates with gastric cancer. These miRNAs could therefore serve as diagnostic biomarkers of gastric cancer.