Showing papers in "Oncology Reports in 2011"
TL;DR: By developing HDAC isoforms as potential biomarkers for breast cancer metastasis, the present study can be extended to developing therapies for Breast cancer invasion.
Abstract: Histone deacetylases (HDACs) are associated with the development and progression of cancer, but it is not known which of the HDAC isoforms play important roles in breast cancer metastasis. This study identified the specific HDAC isoforms that are necessary for invasion and/or migration in human breast cancer cell lines. MDA-MB-231 cells were significantly more invasive and expressed higher levels of matrix metalloproteinase-9 (MMP-9) compared to MCF-7 cells. We compared the expression of HDAC isoforms between MCF-7 and MDA-MB-231 cells and found greater expression of HDAC4, 6 and 8 in MDA-MB-231 cells by RT-PCR and Western blot analyses. In addition, apicidin, a histone deacetylase inhibitor, was shown to attenuate the invasion, migration and MMP-9 expression in MDA-MB-231 cells. Using specific siRNAs directed against HDAC1, 4, 6 and 8, we show that inhibition of HDAC1, 6 and 8, but not HDAC4, are responsible for invasion and MMP-9 expression in MDA-MB-231 cells. We analyzed the invasiveness of MCF-7 cells overexpressing HDAC1, 4, 6 or 8 and found that overexpression of HDAC1, 6 or 8 increased invasion and MMP-9 expression. By developing HDAC isoforms as potential biomarkers for breast cancer metastasis, the present study can be extended to developing therapies for breast cancer invasion.
162 citations
TL;DR: It is found that exosomes might impair the cytotoxic activity of PBMCs when DCs are present and FasL and TRAIL were present in the exosome suspension and addition of an anti-FasL antibody may decrease the percentage of apoptosis of DCs andPBMCs.
Abstract: This study was performed to identify the origin of the ascites-derived exosomes from patients with ovarian cancer and to observe the effect of exosomes on anti-tumor immunity. Exosomes were isolated from the ascites of patients with ovarian epithelial cancer by ultracentrifugation plus density gradient centrifugation. The origin of exosomes was identified by immunoelectronmicroscopy (IEM). The growth curve of the tumor cell line SKOV3 cultured with or without exosomes was analyzed. The apoptosis of autogeneic tumor cells (ATCs) and SKOV3 cells affected by exosomes was measured by flow cytometry (FCM) and light phase contrast microscopy. The cytotoxic effect of the peripheral blood mononuclear cells (PBMCs) stimulated by exosomes and/or dendritic cells (DCs) on ovarian cancer cells was measured using a CCK-8 assay. The levels of IFN-γ released by PBMCs stimulated by exosomes and/or DCs were measured by ELISA. The apoptosis of PBMCs and DCs affected by exosomes was measured by FCM and light microscopy. Whether the mature process of DCs was affected by exosomes was studied by FCM. The ratio of CD4+ T cell and CD8+ T cell were measured by FCM. FasL and TRAIL molecules on exosomes were detected by western blot analysis. The human FasL antagonistic antibody was used to block the apoptosis of DCs and PBMCs induced by exosomes. The receptors of TRAIL DR4 and DR5 on PBMCs and DCs were detected by FCM. In 41 patients examined, we isolated exosomes from the ascites of 35 patients. We detected TCR, CD20, HLA-DR, B7-2, HER2/neu, CA125 and Histone H2A on exosomes. We found that exosomes might impair the cytotoxic activity of PBMCs when DCs are present. We found that exosomes had no effect on the growth and apoptosis of SKOV3 cells. However, exosomes may induce apoptosis of precursors, mature DCs and PBMCs. We found that FasL and TRAIL were present in the exosome suspension and addition of an anti-FasL antibody may decrease the percentage of apoptosis of DCs and PBMCs. We conclude that exosomes exist in ascites of 85.4% of patients with ovarian cancer. Moreover, these exosomes may be of multi-origin. Exosomes had no effect on the growth and apoptosis of tumor cells but impaired the cytotoxic activity of PBMCs in the presence of DCs. Exosomes also may induce apoptosis of the precursors of DCs, DCs and PBMCs. FasL and TRAIL on exosomes may partly account for the apoptosis of cells of the immune system.
162 citations
TL;DR: In this article, the SPONGE method was applied to knock down let-7d in OECM-1 and two primary OSCC cell types, and the results showed that knockdown promoted epithelial-mesenchymal transition (EMT) traits and migratory/invasive capabilities in OSCC cells.
Abstract: Oral squamous cell carcinoma (OSCC) is a prevalent cancer worldwide. Let-7 family has been shown to function as a tumor suppressor through regulating multiple oncogenic signaling. Recent study reported that combined underexpression of miR-205 and let-7d showed negative correlation with the survival prognosis of head and neck cancer patients. However, the let-7d-involved mechanism in regulating OSCC is still unclear. In this study, we first demonstrated that let-7d expression was significantly decreased while Twist and Snail expression was increased in OSCC cancer cell lines and primary cultures as compared to normal human oral keratinocyte cells. To further investigate the role of let-7d in OSCC, we applied the SPONGE method to knock down let-7d in OECM-1 and two primary OSCC cell types. The results showed that knockdown of let-7d promote epithelial-mesenchymal transition (EMT) traits and migratory/invasive capabilities in OSCC cells. Furthermore, down-expression of let-7d significantly activated Twist and Snail expressions and chemo-resistant abilities of OSCC cells. Notably, overexpression of let-7d effectively reversed the EMT phenotype, blocked migratory/invasive abilities, and further increased the chemosensitivity in oral cancer tumor initiating ALDH1+ cells. In sum, these results show that let-7d negatively modulates EMT expression and also plays a role in regulating chemo-resistant ability in oral cancer.
142 citations
TL;DR: The results suggest that the increased MET copy number measured by qPCR plays an important role in determining prognosis in gastric cancer patients, however, the predictive role of MET amplification for treatment response should be further explored in upcoming clinical trials.
Abstract: Identification of critical genes which play pivotal roles in controlling tumor growth and survival will establish the basis for developing therapeutic targets. With the aim of establishing personalized medicine for treatment of solid tumors, we focused on MET amplification in gastric cancer patients, given the extreme sensitivity to c-Met inhibitor in MET amplified gastric cancer cell lines. We tested MET amplification and activation of c-Met in various gastric cancer cell lines and tissue samples from 482 gastric cancer patients who underwent curative surgery. Gastric cancer cell lines with MET amplification by quantitative real-time PCR (qPCR) and FISH predicted sensitivity to PHA-665,752, a selective c-Met kinase inhibitor. Of the 472 patients who had DNA sample available for qPCR analysis, 100 patients (21.2%) had a MET copy number greater than 4.0 copies and demonstrated poorer survival following curative surgery with statistical significance (5-year OS; 50.0 vs. 59.1%; MET amplification (+) vs. MET amplification (-); P = 0.0134). These results suggest that the increased MET copy number measured by qPCR plays an important role in determining prognosis in gastric cancer patients. However, the predictive role of MET amplification for treatment response should be further explored in upcoming clinical trials.
140 citations
TL;DR: The study showed that miR-25 was highly expressed both in clinical ovarian cancer samples and cell lines and that it was a potential therapeutic target for ovarian cancer intervention and directly regulates apoptosis by targeting Bim in ovarian cancer.
Abstract: MicroRNAs (miRNAs) are emerging as a class of small regulatory RNAs whose alterations are implicated in the initiation and progression of human cancers. Our study showed that miR-25 was highly expressed both in clinical ovarian cancer samples and cell lines. Down-regulation of miR-25 in ovarian cancer cells induced apoptosis whereas overexpression of miR-25 enhanced cell proliferation. The effects of miR-25 abrogation were partly mediated by the intrinsic apoptosis pathway. Many pro-apoptotic proteins such as Bim, Bax and caspase-3 were up-regulated after transfection. Furthermore, luciferase assays demonstrated that Bim was the direct target of miR-25. Introducing Bim cDNA without 3'UTR abrogated miR-25-induced cell survival. Finally, there was an inverse relationship between Bim and miR-25 expression in ovarian cancer tissues. Taken together, these data indicate that miR-25 directly regulates apoptosis by targeting Bim in ovarian cancer and that miR-25 could be a potential therapeutic target for ovarian cancer intervention.
131 citations
TL;DR: The expression level of IL- 17 mRNA in gastric tumors was associated with the depth of the tumors, lymph-vascular invasion and lymph node involvement, suggesting that IL-17 obviously was related to tumor progression.
Abstract: Recently, a subset of IL-17 producing T cells distinct from Th1 or Th2 cells has been described as key players in inflammation and autoimmune diseases as well as cancer development. In this study, we investigated the expression level of IL-17 and T helper 17 (Th17)-related cytokines in gastric cancer tissues and assessed the association of their expression with angiogenesis and their clinicopathological parameters. Tumor and adjacent normal tissues were obtained from 82 patients with gastric cancer. IL-17, IL-21 and IL-23 mRNA expression levels were quantified by real-time RT-PCR. Th17 infiltration, microvessel density and neutrophil infiltration in tumor tissues were examined by immunohistochemistry and double immunofluorescence histochemistry. Expression of IL-17, IL-21 and IL-23 mRNA was found to be significantly up-regulated in tumor tissues compared with adjacent normal tissues. The expression level of IL-17 mRNA strongly and positively correlated with that of IL-21 mRNA in tumor tissue. The number of vascular endothelial cells and infiltrating neutrophils was significantly larger in tumors expressing a high level of IL-17 mRNA than in tumors expressing a low level of IL-17 mRNA. In tumor tissues most CD4+ cells were stained with anti-IL-17 antibody. The expression level of IL-17 mRNA in gastric tumors was associated with the depth of the tumors, lymph-vascular invasion and lymph node involvement, suggesting that IL-17 obviously was related to tumor progression. IL-17 and IL-21, which regulates IL-17, would be potential therapeutic targets for the treatment of gastric cancer.
129 citations
TL;DR: It is demonstrated that quercetin induced FasL-related apoptosis by transactivation through activation of c-jun/AP-1 and promotion of histone H3 acetylation in HL-60 cells.
Abstract: Quercetin, a naturally occurring flavonoid abundant in fruits and vegetables, has been demonstrated as a multipotent bioflavonoid with great potential for the prevention and treatment of cancer. Apoptosis is thought to be an important response to most chemotherapeutic agents in leukemia cells. However, the underlying mechanism of induction of apoptosis by quercetin involving epigenetic regulation is poorly understood. In the present study, by evaluation of fragmentation of DNA, poly (ADP-ribose) polymerase (PARP) and procaspases, we found that quercetin was able to induce apoptosis of human leukemia HL-60 cells in a dose-dependent manner. Quercetin triggered the extrinsic apoptosis pathway through activation of caspase-8 and induction of Bid cleavage, Bax conformation change and cytochrome c release. Furthermore, quercetin induced Fas ligand (FasL) expression involving activation of the extracellular signal-regulated kinase (ERK) and Jun N-terminus kinase (JNK) signaling pathways. In addition to activation of c-Jun, quercetin increased histone H3 acetylation which resulted in the promotion of the expression of FasL. Quercetin exhibited potential for the activation of histone acetyltransferase (HAT) and the inhibition of histone deacetyltransferase (HADC), both of which contributed to histone acetylation. However, only the activation effect on HAT was associated with the ERK and JNK pathway. These results demonstrated that quercetin induced FasL-related apoptosis by transactivation through activation of c-jun/AP-1 and promotion of histone H3 acetylation in HL-60 cells.
118 citations
TL;DR: In this study, using real-time PCR and Western blotting, it is found that Nampt was overexpressed at the mRNA and protein levels, respectively, in established gastric cancer cells and human Gastric cancer tissues, and may be a new therapeutic target for gastric cancers.
Abstract: Nicotinamide phosphoribosyltransferase (Nampt), an enzyme involved in the NAD⁺ salvage pathway, is over-expressed and important in the carcinogenesis in several types of cancers. The expression of Nampt and its role in gastric cancer remain largely unknown. In this study, using real-time PCR and Western blotting we found that Nampt was overexpressed at the mRNA and protein levels, respectively, in established gastric cancer cells and human gastric cancer tissues. The specific Nampt inhibitor FK866 repressed gastric cancer cell proliferation, as assessed by MTT assay. Using transwell and soft agar clonogenic assays, we also found that FK866 suppressed gastric cancer cell migration and anchorage-independent growth, respectively. These inhibitory effects of FK866 were accompanied by significantly decreased expression of VEGF, MMP2, MMP9 and NF-κB. As determined by MTT assay and flow cytometry, FK866 also increased the chemo-sensitivity of gastric cancer cells to fluorouracil by greater inhibition of cell proliferation and the induction of apoptosis. Our findings indicate that Nampt may be a new therapeutic target for gastric cancer.
117 citations
TL;DR: MiR-152 and miR-148a may be involved in the carcinogenesis of ovarian cancer through deregulation of cell proliferation and down-regulated in ovarian cancer cell lines.
Abstract: microRNAs (miRs) are endogenous small non-coding RNAs that are aberrantly expressed in various carcinomas. miR-152 and miR-148a have not been comprehensively investigated in ovarian cancer. Thus, the aim of this study was to identify the role of miR-152 and miR-148a in epithelial ovarian cancer. Total RNA was extracted from tissues of 78 patients with epithelial ovarian cancer, 17 normal ovarian epithelium tissues and two ovarian cancer cell lines. Using quantitative real-time PCR (qRT-PCR) followed by the 2-ΔΔCT method for calculating the results, we found that the expression levels of miR-152 were significantly decreased in ovarian cancer tissues compared to normal ovarian epithelium tissues (p<0.05). However, although the expression of miR-148a was also decreased in 65% of patients, no statistically significant difference in expression was found. A strong correlation was found between the expression of miR-152 and miR-148a (p<0.001, Pearson's correlation). The relationship between miR-152 or miR-148a expression levels in ovarian cancer and clinicopathological features, response to therapy and short-term survival was analyzed and the results showed that no correlation existed. In addition, we found that both miR-152 and miR-148a were down-regulated in ovarian cancer cell lines. After miR-152 or miR-148a mimics were transfected into ovarian cancer cell lines, the MTT cell proliferation assay showed that cell proliferation was significantly inhibited. Taken together, miR-152 and miR-148a may be involved in the carcinogenesis of ovarian cancer through deregulation of cell proliferation. They may be novel biomarkers for early detection or therapeutic targets of ovarian cancer.
114 citations
TL;DR: It is suggested that miR-320a may be a novel therapeutic candidate for the treatment of colorectal cancer and useful for identifying CRC patients that are at an elevated risk for developing liver metastasis.
Abstract: MicroRNAs (miRNAs) have been implicated in regulating diverse cellular pathways. Although there is emerging evidence that various miRNAs function as oncogenes or tumor suppressors in colorectal cancer (CRC), the role of miRNAs in mediating liver metastasis remains unexplored. The expression profile of miRNAs in liver metastasis and primary CRC tissues was analyzed by miRNA microarrays and verified by real-time polymerase chain reaction (PCR). In 62 CRC patients, the expression levels of miR-320a were determined by real-time PCR, and the effects on migration and invasion of miR-320a were determined using a transwell assay. miR-320a target genes were confirmed by luciferase assay, real-time PCR and western blot analysis. A set of miRNAs was found to be dysregulated in the liver metastasis tissues compared to matched primary CRC tissues, and the expression levels of miR-320a were significantly decreased in the liver metastasis tissues examined. miR-320a was correlated with tumor progression in CRC. miR-320a was downregulated in liver metastatic colon cancer cells and inhibited liver metastatic colon cancer cell migration and invasion. miR-320a directly binds to the 3'UTR of neuropilin 1 (NRP-1), a protein that functions as a co-receptor of vascular epithelial growth factor. miR-320a downregulated the expression of NRP-1 at both the mRNA and protein levels. These data demonstrated that miR-320a may be useful for identifying CRC patients that are at an elevated risk for developing liver metastasis. Our findings suggest that miR-320a may be a novel therapeutic candidate for the treatment of colorectal cancer.
110 citations
TL;DR: Results indicate that irradiation induces an alteration to a malignant phenotype consistent with epithelial-mesenchymal transition (EMT) in colorectal cancer cells.
Abstract: Radiotherapy remains a major approach to adjuvant therapy for patients with advanced rectal cancer. Nevertheless, the effects of radiation on malignant processes have yet to be clarified. The aim of this study was to assess the biological effects of radiation on colorectal cancer (CRC) cells with special reference to epithelial-mesenchymal transition (EMT), a key developmental program often activated during cancer invasion and metastasis. We investigated the effect of radiation on two colorectal cancer cell lines, CaR1 and DLD1, assessing cell morphology, motility, migration and invasive ability. Expression of molecules associated with EMT was determined using RT-PCR, Western blotting, and immunofluorescence staining in control and irradiated cells. We also used real-time RT-PCR to examine the expression of molecules associated with EMT before and after chemoradiotherapy. Thus, we studied 26 rectal cancer patients who received preoperative chemoradiotherapy followed by radical surgery. In addition, we examined the relationship between disease recurrence and the expression of a number of proteins. Irradiation caused CRC cells to undergo phenotypic changes characteristic of EMT: spindle-cell shape, loss of polarity, intercellular separation and pseudopodia formation. Irradiation enhanced cell migration and invasiveness. In irradiated CRC cells, molecular changes consistent with EMT were observed. In clinical samples, we observed molecular changes consistent with EMT, and those changes were significantly enhanced in patients with recurring disease. These results indicate that irradiation induces an alteration to a malignant phenotype consistent with EMT in colorectal cancer cells.
TL;DR: Analysis of the relations between clinicopathological features and expression of the eight circadian genes in cancer tissue showed that high expression ofThe Bmal1 gene and lowexpression of the Per1 gene correlated with liver metastasis.
Abstract: Circadian rhythms are daily oscillations in various biological processes, generated by the feedback loops of eight core circadian genes: Period1 (Per1), Period2 (Per2), Period3 (Per3), Cryptochrome1 (Cry1), Cryptochrome2 (Cry2), Clock, Bmal1 and Casein Kinase I e (CKIe). Recent studies have suggested that circadian genes participate in the growth and development of various cancers. This study examined the relations of circadian gene expression to clinicopathological factors and outcomes in patients with colorectal cancer. We studied surgical specimens of cancer tissue and adjacent normal mucosa obtained from 202 patients with untreated colorectal cancer. The relative expression levels of the circadian genes in the specimens were measured by quantitative real-time, reverse-transcription polymerase chain reaction. Expression of the Clock gene and the CKIe gene in cancer tissue were significantly higher compared to that in adjacent normal mucosa. Expression of the Per1 and Per3 genes in cancer tissue was significantly lower compared to that in adjacent normal mucosa. Analysis of the relations between clinicopathological features and expression of the eight circadian genes in cancer tissue showed that high expression of the Bmal1 gene and low expression of the Per1 gene correlated with liver metastasis. On analysis of the relations between outcomes and gene expression, high expression of the Per2 gene was associated with significantly better outcomes than low expression of the Per2 gene. Overexpression of the Bmal1 gene and reduced expression of the Per1 gene may thus be useful predictors of liver metastasis. Moreover, reduced expression of the Per2 gene may be a predictor of outcomes in patients with colorectal cancer.
TL;DR: The data demonstrate that miR-122 plays an important role in HBV-related hepatocarcinogenesis by targeting NDRG3, a member of the N-myc downstream-regulated gene (NDRG) family, which represents key diagnostic markers and potential therapeutic targets for HBV -related HCC.
Abstract: microRNAs (miRNAs) are short, non-coding RNAs with post-transcriptional regulatory functions that participate in diverse biological pathways. miR-122, a liver-specific miRNA, has been found to be down-regulated in hepatocellular carcinoma (HCC) and HCC-derived cell lines. In this study, miR-122 was down-regulated in the hepatitis B virus (HBV)-related HCC cell line HepG2.2.15 compared to HepG2. NDRG3, a member of the N-myc downstream-regulated gene (NDRG) family, was up-regulated in HepG2.2.15 and was identified as a target gene of miR-122. An inverse correlation between the expression of miR-122 and the NDRG3 protein was noted in HBV-related HCC specimens. The transfection of the miR-122 expression vector into the HepG2.2.15 cell line repressed the transcription and expression of NDRG3, which subsequently reversed the malignant phenotype of the cells. The replication of HBV, expression of viral antigens and proliferation of cells were significantly inhibited by restoration of miR-122. The data demonstrate that miR-122 plays an important role in HBV-related hepatocarcinogenesis by targeting NDRG3. Thus, miR-122 and NDRG3 represent key diagnostic markers and potential therapeutic targets for HBV-related HCC.
TL;DR: It is suggested that GA might modulate through blocking the p38, JNK, PKC and PI3K/AKT signaling pathways and reducing the NF-κB protein level, resulting in the inhibition of MMP-2 and -9 of PC-3 human prostate cancer cells.
Abstract: epidemiological studies have demonstrated that a natural diet or consumption of fruits or vegetables can decrease the risk of cancer development. cancer cells can migrate to and invade other organs or tissues that cause more difficulty to treat them and this also results in the need for treatments targeting multiple cellular pathways. Gallic acid (GA) has been demonstrated to possess multiple biological activities including anticancer function. However, no report exist on GA inhibited invasion and migration of human prostate cancer cells. We investigated the effects of migration and invasion in GA-treated pc -3 human prostate cancer cells with a series of in vitro exper - iments. Boyden chamber transwell assay was used to examine the migration and invasion of pc -3 cells. Western blotting, real- time PCR and gelatin zymography were used for determining the protein levels, gene expression and enzyme activities of matrix metalloproteinase-2 (MMP-2) and -9 in vitro. r esults indicated that GA inhibited the invasion and migration of PC-3 cells and these effects are dose-dependent. GA inhibited the protein levels of MMP-2 and -9, son of sevenless homolog 1 (SOS1), growth factor receptor-bound protein 2 (GRB2), protein kinase C (PKC) and nuclear factor-κ B (NF-κB) p65, c-Jun N-terminal kinase (JNK), extracellular signal-regulated kinase 1/2 (ERK1/2), p38, p-AKT (Thr308) and p-AKT (Ser473), but it promoted the levels of phosphatidylinositol 3-kinase (PI3K) and AKT in PC-3 cells. GA also reduced the enzyme activities of MMP-2 and -9 in the examined cells. Moreover, the down- regulation of focal adhesion kinase (FAK) and Ras homolog gene family, member A (Rho A) mRNA expression levels, and up-regulation of the tissue inhibitor of metalloproteinase-1 (TIMP1) gene levels occurred in GA-treated PC-3 cells after 24 h treatment. Based on these observations, we suggest that GA might modulate through blocking the p38, JNK, PKC and PI3K/AKT signaling pathways and reducing the NF-κB protein level, resulting in the inhibition of MMP-2 and -9 of PC-3 human prostate cancer cells.
TL;DR: Evidence is provided that miR-146a can directly target SMAD4, and a role in the development of gastric cancer may be suggested by modulating cell proliferation and apoptosis, which is one of the most common malignant diseases worldwide.
Abstract: MicroRNAs (miRNAs) have emerged as important gene regulators and are recognized as oncogenes or tumor suppressor genes in carcinogenesis. Gastric cancer is one of the most common malignant diseases worldwide. Our previous studies have revealed that miR-146a is upregulated in gastric epithelial cells infected with Helicobacter pylori (H. pylori) and in mucosal tissues from H. pylori-positive patients. However, the role of miR-146a in gastric cancer is largely unknown. In the current study, we showed that miR-146a was upregulated in 20 gastric cancer tissues compared with matched non-tumor adjacent tissues by quantitative RT-PCR. Furthermore, ectopic expression of miR-146a could improve cell proliferation in vitro by using Cell Counting kit 8 (CCK-8). We also found that miR-146a inhibited apoptosis of gastric cancer cells by flow cytometry (FCM) and Caspase-Glo® 3/7 assay. Using target prediction algorithms, luciferase reporter assay and Western blot assay, SMAD family member 4 (SMAD4) was identified as a target gene of miR-146a in gastric cancer. Moreover, an inverse correlation was observed between the expression of SMAD4 mRNA and miR-146a in gastric cancer tissues (R=-0.731, P=0.039, Pearson's correlation). Taken together, our results provide important evidence that miR-146a can directly target SMAD4, and suggest that miR-146a may play a role in the development of gastric cancer by modulating cell proliferation and apoptosis. miR-146a could serve as a potential biomarker and therapeutic target against gastric cancer.
TL;DR: This study indicates that downregulated miR-221/222 can sensitize glioma cells to TMZ by regulating apoptosis independently of p53, and shows that low expression of miR/222 sensitized gliomas cells to temozolomide (TMZ).
Abstract: A previous study showed that miR-221/222 can regulate cell apoptosis. p53 is a well known tumor suppressor which can influence the chemosensitivity of glioma cells. However, the effect of miR-221/222 in gliomas with different p53 status is unknown. Here, we demostrate that knockdown of miR-221/222 increases apoptosis in human gliomas of different p53 types (U251 cells, p53 mutant-type; LN308 cells, p53 null-type; and U87 cells, p53 wild-type). Furthermore, the effect of miR-221/22 caused no change of p53 expression in the glioma cells studied. In addition, when a specific siRNA against p53 was employed in U87 cells, no attenuation of apoptosis was found after knockdown of miR-221/222. Importantly, we found that As-miR-221/222-treated cells increased expression of Bax, cytochrome c, Apaf-1 and cleaved-caspase-3. Our results showed that low expression of miR-221/222 sensitized glioma cells to temozolomide (TMZ); in addition, ectopic expression of PUMA by pcDNA-PUMA had a similar effect. Taken together, our study indicates that downregulated miR-221/222 can sensitize glioma cells to TMZ by regulating apoptosis independently of p53 status.
TL;DR: Whether curcumin can reverse chemoresistance by downregulating NF-κB in human gastric cancer cells is investigated and it is found thatCurcumin potentiates the antitumor effects of chemotherapeutics in Gastric cancer by suppressing NF-σκB and NF-γB-regulated anti-apoptotic genes.
Abstract: Gastric cancer remains one of the major health problems worldwide. Chemotherapy is an important therapeutic modality for gastric cancer, but the success rate of this treatment is limited because of chemoresistance. The ubiquitously expressed transcription factor NF-κB has been suggested to be associated with chemoresistance of gastric cancer. Agents that can either enhance the effects of chemotherapeutics or overcome chemoresistance to chemotherapeutics are needed for the treatment of gastric cancer. Curcumin, a component of turmeric, is one such agent that has been shown to suppress NF-κB and increase the efficacy of chemotherapy. In this study, we investigated whether curcumin can reverse chemoresistance by downregulating NF-κB in human gastric cancer cells. SGC-7901 human gastric cancer cells was treated with chemotherapeutics (etoposide and doxorubicin) or by combined application of curcumin and chemotherapeutics. The viability of SGC-7901 cells was measured by MTT assay. Apoptosis of SGC-7901 cells was detected using the TUNEL and Annexin V/PI methods. The protein levels of NF-κB were analyzed by immunocytochemical staining. EMSA was used to confirm the increased nuclear translocation of RelA. The protein levels of p-IκBα, Bcl-2 and Bcl-xL were analyzed by Western blotting. The chemotherapeutics (etoposide and doxorubicin) suppressed the growth of SGC-7901 cells, in a time-dose-dependent manner. Use of curcumin in addition to these agents can suppress cell growth further (inhibitory rate: doxorubicin vs. doxorubicin + curcumin, 33% vs. 45%, p<0.05; etoposide vs. etoposide + curcumin, 35% vs. 48%, p<0.05). Furthermore, chemotherapeutics induced apoptosis of SGC-7901 cells and activated NF-κB. The combination of curcumin and chemotherapeutics induced apoptosis of SGC-7901 cells further, attenuated the activation of NF-κB, and reduced expression of the NF-κB-regulated anti-apoptotic gene products Bcl-2 and Bcl-xL. Curcumin potentiates the antitumor effects of chemotherapeutics in gastric cancer by suppressing NF-κB and NF-κB-regulated anti-apoptotic genes.
TL;DR: Results indicate that OA suppresses the M2 polarization of macrophages and tumor cell proliferation by inhibiting STAT3 activation, and may be a potentially new agent that can be used for the prevention and treatment of various malignant tumors, including glioma.
Abstract: Tumor-associated macrophages (TAMs) polarized to the M2 phenotype promote tumor cell proliferation and are associated with a poor prognosis in patients with high grade glioma. We previously revealed that corosolic acid, a triterpenoid compound, inhibits the M2 polarization of human monocyte-derived macrophages (HMDM). In the present study, we examined whether oleanolic acid (OA), a triterpenoid compound whose structure is similar to corosolic acid, also shows inhibitory effects on M2 polarization in HMDM. OA significantly inhibited the expression of CD163, one of the phenotype markers of M2 macrophages, as well as suppressed the secretion of IL-10, one of the anti-inflammatory cytokines preferentially produced by M2 macrophages, thus suggesting that OA suppresses the M2 polarization of macrophages. Furthermore, OA inhibited the proliferation of U373 human glioblastoma cells, and the activation of signal transducer and activator of transcription-3 (STAT3) in both human macrophages and glioblastoma cells. These results indicate that OA suppresses the M2 polarization of macrophages and tumor cell proliferation by inhibiting STAT3 activation. Therefore, OA may be a potentially new agent that can be used for the prevention and treatment of various malignant tumors, including glioma.
TL;DR: Evaluated EGFR gene mutations in squamous cell carcinoma and its clinicopathological features demonstrate that EGFR mutational analysis should be performed not only in adenocarcinoma, but also in SCC to allow accurate diagnosis and treatment.
Abstract: Epidermal growth factor receptor (EGFR) gene mutations have been reported to be clinically significant in non-small cell lung cancer (NSCLC). However, because most previous studies focused only on adenocarcinomas, EGFR mutations in other histotypes are poorly investigated. We evaluated the frequency of EGFR gene mutations in squamous cell carcinoma (SCC) and its clinicopathological features. In total, 89 frozen tumor specimens that had been first diagnosed as SCCs, were examined for EGFR mutations in exons 19 and 21 using direct sequencing, PNA-enriched sequencing and SmartAmp2. Additionally, pathological investigation, including immunostaining for p63 and TTF-1, alcian blue staining and EGFR mutation-specific immunohistochemistry in mutation-positive samples was also performed. The frequency of EGFR mutations was 5.6% (5/89); all mutations were deletions in EGFR exon 19. Immunohistological investigation of these samples revealed that two of five were positive for p63 and TTF-1 staining, and showed production of mucin, as evidenced by alcian blue staining. Consequently, three of the samples were considered to be true SCC at final pathological diagnosis, while the remaining two samples were revised to adenosquamous carcinoma and adenocarcinoma. The final frequency of the EGFR mutations in true SCC was 3.4% (3/87). In conclusion, EGFR mutations were found in a small, but significant, number of SCC tumor samples and thus EGFR mutational analysis was useful in the accurate diagnosis of SCC. Our data demonstrate that EGFR mutational analysis should be performed not only in adenocarcinoma, but also in SCC to allow accurate diagnosis and treatment.
TL;DR: It is revealed that aberrantly expressed miRNAs play important roles in NPC tumorigenesis and may serve as potential targets for novel therapeutic strategies in the future.
Abstract: Nasopharyngeal carcinoma (NPC) is posing a serious health problem worldwide. The association between its pathogenesis and microRNAs (miRNA) has not been elucidated. In this study, miRNA expression profiling was performed to screen the miRNA expression changes in 8 NPC tissues and 4 normal nasopharyngeal tissues. Thirty-four miRNAs were identified to be differentially expressed; of these, one miRNA (miR-18a) was overexpressed and 33 miRNAs (miR-34b, miR-34c, let-7 family, etc.) were underexpressed in NPC tissues compared to the normal samples. Validation was performed by real-time quantitative PCR for two altered miRNAs (miR-34b and let-7g) and one non-differentially expressed miRNA (miR-30c). Unsupervised hierarchical clustering analysis showed that the aberrant miRNAs were correlated with the clinical stage of NPC patients. In addition to several biological pathways that are well characterised in NPC and which were significantly targeted by the underexpressed miRNAs, two novel pathways, nervous system development and sensory perception of sound, were identified to be strongly associated with NPC development. Furthermore, a c-Myc centered miRNA regulatory network was inferred in NPC. Our study reveals that aberrantly expressed miRNAs play important roles in NPC tumorigenesis and may serve as potential targets for novel therapeutic strategies in the future.
TL;DR: Cell proliferation and chemoresistance to doxorubicin were promoted by overexpression of miR-21 in T24 cells and may usher in new therapeutic strategies in treating this disease.
Abstract: Transitional cell carcinomas (TCCs) of the urinary bladder are common malignancies with a high recurrence rate Since microRNA-21 (miR-21) may contribute to tumorigenesis and chemoresistance in many cancer types, we aimed to investigate its efficacy in TCCs The expression of miR-21 and its target PTEN was determined by real-time qRT-PCR and western blotting, respectively in tumor tissues as well as adjacent non-tumor mucosa The effect of miR-21 on cell proliferation and chemosensitivity to doxorubicin were measured using the MTT method Cell apoptosis induced by doxorubicin was investigated using flow cytometry in the T24 cell line BCL-2, AKT and pAKT were detected by western blotting for analysis of potential mechanisms miR-21 was significantly up-regulated in tumor tissues while PTEN was expressed in lower levels compared to non-tumor tissues A negative correlation between expression of miR-21 and PTEN was established in vivo Cell proliferation and chemoresistance to doxorubicin were promoted by overexpression of miR-21 in T24 cells BCL-2 up-regulation could be achieved by miR-21 overexpression, which prevented T24 cells from apoptosis induced by doxorubicin Furthermore, the miR-21 induced BCL-2 up-regulation could be cancelled by the PI3K inhibitor LY294002 These data verified the oncogenic role of miR-21 in TCCs and may usher in new therapeutic strategies in treating this disease
TL;DR: Cisplatin or 5-fluorouracil alter miRNA expression in esophageal cancer cells, and IPA suggests that these miRNAs may target molecular pathways involved in cell survival after chemotherapy.
Abstract: Neoadjuvant chemotherapy is often used in the treatment of advanced esophageal cancer. In this study, we determined the impact of chemotherapy on microRNA (miRNA) expression in esophageal cancer cells, and whether identified changes might have biological relevance. Two esophageal carcinoma cell lines (one adenocarcinoma and one squamous cell carcinoma) were treated with cisplatin or 5-fluorouracil for 24 or 72 h. RNA was extracted from cells following 24-h treatment, and used for microarray studies. Promising miRNA candidates were selected for RT-PCR validation. Target prediction using TargetScan, combined with bioinformatic analysis (Ingenuity Pathway Analysis, IPA), was performed to evaluate the implications of the altered miRNA expression. Thirteen miRNAs (miR-199a-5p, miR-302f, miR-320a, miR-342-3p, miR-425, miR-455-3p, miR-486-3p, miR-519c-5p, miR-548d-5p, miR-617, miR-758, miR-766, miR-1286) were deregulated after 24- and/or 72-h treatment in both cell lines, and most miRNAs presented similar expression changes after short- or long-term exposure. IPA revealed that the major networks which incorporate the predicted targets, include functions such as 'Cell death', 'Cell cycle', 'Cellular growth and proliferation', 'DNA replication, recombination, and repair' and 'Drug metabolism'. Cisplatin or 5-fluorouracil alter miRNA expression in esophageal cancer cells. IPA suggests that these miRNAs may target molecular pathways involved in cell survival after chemotherapy.
TL;DR: Nestin plays important roles in cell growth, migration, invasion and adhesion to extra-cellular matrices in glioma cells and may serve as a novel candidate for molecular-targeted therapy for gliomas, including glioblastomas.
Abstract: Nestin, a class VI intermediate filament protein, was originally described as a neuronal stem cell marker during central nervous system development. Nestin is expressed in gliomas, and its expression levels are higher in gliomas with high WHO histopathological classification grades than in those with low grades. In the present study, we examined whether nestin regulates the biological activities of human glioma cells. Immunohistochemically, the nestin expression patterns in 10 human glioblastoma patients were examined. The expression levels of nestin in A172, a human high-grade glioma cell line, and KG-1-C, a human low-grade glioma cell line, were examined using real-time PCR, Western blot and immunofluorescence analyses. An expression vector carrying a short hairpin RNA targeting nestin was stably transfected into A172 (Sh) cells. The effects of decreased expression levels of nestin in Sh cells on cell growth, migration, invasion, adhesion to extracellular matrices and fibrillar actin expression on three-dimensional culture plates were examined. The nestin expression vector was transiently transfected into KG-1-C (Nes) cells, and the effects of the nestin overexpression on cell growth and migration were examined. Nestin was expressed in the cytoplasm of the glioblastoma cells in all cases examined. Sh cells showed marked decreases in the expression levels of nestin mRNA and protein, and the growth rate of Sh cells was lower than that of sham (Sc) cells. In contrast, the adhesion activity of Sh cells to types I and IV collagens, fibronectin and laminin was higher than that of Sc cells. Fibrillar actin was clearly detected at the periphery of colonies of Sh cells at the attachment sites on three-dimensional culture plates. The migration and invasion of Sh cells were markedly inhibited compared with those of Sc cells. In contrast, the levels of nestin expression markedly increased in the Nes cells, which were transiently transfected with the nestin expression vector. The growth rate and motility of Nes cells were higher than those of the mock cells. In conclusion, nestin plays important roles in cell growth, migration, invasion and adhesion to extra-cellular matrices in glioma cells. Nestin may serve as a novel candidate for molecular-targeted therapy for gliomas, including glioblastomas.
TL;DR: The highly metastatic breast cancer cell line MDA-MB-231 exhibited the strongest binding to MAL and the highest expression levels of α2,3-sialic acid residues among the selected cell lines, depending on mRNA expression levels
Abstract: Aberrant sialylation is closely associated with the malignant phenotype of cancer cells and metastatic potential. However, the precise nature of the molecules in breast cancers has not been unveiled. In this study, we investigated the expression levels of α2,3-sialic acid residues of 50 primary tumor cases, 50 pair-matched lymph node metastasis tumor samples and in the MDA-MB-231, T-47D and MCF-7 breast cancer cell lines with different metastatic potential. The expression of α2,3-sialic acid residues was analyzed by histochemistry, cytochemistry and flow cytometry with Maackia amurensis lectin (MAL). The invasion and migration abilities of cells were examined using cell adhesion and transwell in vitro assays. Pair-matched lymph node metastasis tumor samples exhibited higher levels of expression of α2,3-sialic acid residues compared to that of primary tumors (P=0.0432). Furthermore, of 38 tumors cases in T1/T2 stages, 31 (81.58%) had weak staining for MAL, which specifically binds to α2,3-sialic acid residues, whereas of 12 tumor cases in T3/T4 stages, only 1 (8.33%) had weak reactions for MAL. The highly metastatic breast cancer cell line MDA-MB-231 exhibited the strongest binding to MAL and the highest expression levels of α2,3-sialic acid residues among the selected cell lines, depending on mRNA expression levels of α2,3-sialyltransferase gene. The adhesion, invasion and migration activities confirmed that MDA-MB-231 exhibited the greater cell adhesion to, migration toward and invasion to Matrigel. Taken together, the high expression of α2,3-sialic acid residues in breast cancer was associated with metastatic potential. This property may be important for developing new therapeutic approaches for breast cancer.
TL;DR: The findings suggest that ANGPTL4 is one of the factors involved in the progression of human colorectal cancer, especially venous invasion and distant metastasis.
Abstract: There is strong evidence that the angiopoietin family is involved in the regulation of tumour progression. Recently, it has been reported that angiopoietin-like 4 (ANGPTL4) expression in cancer cells promotes the metastatic process by increasing vascular permeability. The present study was conducted to examine ANGPTL4 expression and its association with clinicopathological factors and prognosis in human colorectal cancers. We examined 144 cases of surgically-resected human colorectal adenocarcinomas by immunohistochemistry, RT-PCR and Western blot analysis. Also, overall survival was investigated. Among 144 cases of adenocarcinoma, 95 cases (66.0%) showed positive staining in the cytoplasm of the carcinoma cells for ANGPTL4. Histologically, well, moderately, poorly differentiated adenocarcinoma or mucinous carcinoma showed 55.2, 79.3, 61.5 or 44.4% expression of ANGPTL4, respectively. The expression of ANGPTL4 was correlated with the depth of tumour invasion (p<0.0005), Vienna classification (category 3-5)(p<0.00005), venous invasion (p<0.0005) and Duke's classification (p<0.005). However, ANGPTL4 expression was not correlated with overall survival. However, all patients (100%) with distant metastasis showed immunopositivity for ANGPTL4. The mRNA and the protein expression of ANGPTL4 were shown in four resected samples and cultured cell lines by RT-PCR or western blot analysis. These findings suggest that ANGPTL4 is one of the factors involved in the progression of human colorectal cancer, especially venous invasion and distant metastasis.
TL;DR: SIRT1 silencing was found to induce cell growth arrest in HCC cells, suggesting an association of SIRT1 expression with HCC development and that Sirt1 plays a role in cancer cell growth.
Abstract: Silent mating type information regulation 2 homolog 1 (SIRT1) is a multifaceted, nicotinamide adenine dinucleotide-dependent protein deacetylase with involvement in a wide variety of cellular processes ranging from cancer to aging. Expression of SIRT1 was evaluated in 90 cases of he p atocellular carcinoma (HCC) and five HCC cell lines. The relationship between the mutation status of p53 and expression of SIRT1 was also investigated in 10 fresh HCC tissues. Synthetic small interfering RNA was used to silence SIRT1 gene expression by RNA interference (RNAi), and cell growth and cell cycle progression were assessed. Expression of SIRT1 was significantly elevated in the HCC tissues when compared to that of non-tumor tissues (p<0.001). Overexpression of SIRT1 and p53 was observed in 56% (50 of 90) and in 30% (27 of 90) of the HCCs, respectively. Expression of SIRT1 showed significant correlation with gender (p=0.023), serum AFP levels (p=0.030), viral infection (p=0.005) and p53 expression (p<0.021). Western blot analysis found no correlation between p53 mutation and expression levels of SIRT1. SIRT1 silencing was found to induce cell growth arrest in HCC cells. These results suggest an association of SIRT1 expression with HCC development and that SIRT1 plays a role in cancer cell growth.
TL;DR: A miRNA expression profile was identified by miRCURY LNA Array and aberrantly expressed miRNAs may offer new clues to tumorigenesis of gastric cancers.
Abstract: miRNAs are small 19 to 22 nucleotide sequences of RNA that negatively regulate gene expression. miRNA expression profiles may become useful biomarkers for diagnostics, prognosis and prediction of response to treat, and it could be a powerful tool for cancer prevention and therapeutics. Several miRNA expression profiles of miRNAs in gastric cancer have been reported, but these studies screened only few miRNAs and samples used in experiments include several different subtypes of gastric cancers, which decrease the sensitivity to identify new aberrant miRNAs. In this study, a miRNA expression profile was identified by miRCURY LNA Array (v.14.0) between intestinal-type gastric cancers and normal tissues. Forty miRNA precursors were up-regulated and thirty-six miRNA were down-regulated in intestinal-type gastric cancers (p<0.01). Sixteen new miRNAs were found in intestinal-type gastric cancers. Seventeen new miRNAs were found in intestinal-type gastric cancers. miR-145, miR-27a, miR-494 are differently expressed between intestinal-type and diffuse-type gastric cancers. miR-32, miR-182 and miR-143 dysregulated expression levels are related with different pathological stages of intestinal-type gastric cancers (p<0.01). Taken together, aberrantly expressed miRNAs may offer new clues to tumorigenesis of gastric cancers. miR-32, miR-182 and miR-143 may be potential diagnostic biomarkers for intestinal-type gastric cancers.
TL;DR: The results indicated that miRNAs are not only involved in carcinogenesis of colorectum, but may also participate in the progression such as with liver metastases in human Colorectal cancers.
Abstract: At present, a full understanding of the mechanisms by which colorectal cancer (CRC) distant metastases form is still beyond our reach because of the intricate regulation of gene expression. MicroRNAs (miRNAs) are shown to be involved in various human diseases including cancers through negative regulation of target gene expression at the post-transcriptional level. However, there are only a few studies on the roles of miRNA aberrations in liver metastasis of human colorectal cancer. To identify miRNA expression patterns associated with liver metastasis in human colorectal cancer, the miRNA expression profiles of colorectal cancer tissues with liver metastasis and their non-metastatic counterparts were studied using microRNA microarrays and further confirmed by quantitative RT-PCR. We show that 28 miRNAs are differentially expressed in the colorectal carcinomas with liver metastasis compared to the non-metastatic counterparts. Of these, 4 miRNAs including miR-150*, miR-125b-2*, miR-1179 and miR-139-3p were up-regulated in colorectal cancers with liver metastasis while the others were down-regulated. The target genes of selected deregulated miRNAs were predicted through bioinformatic techniques with two functional analyses, gene ontology and KEGG analysis, which showed that categories of high enrichment GOs and specific pathways targeted by dysregulated miRNAs were involved in liver metastasis during human colorectum carcinogenesis. Our results indicated that miRNAs are not only involved in carcinogenesis of colorectum, but may also participate in the progression such as with liver metastases in human colorectal cancers.
TL;DR: Conclusively, regulation of proliferation in nuclear ER-positive endometrial cancer cells is mediated by both ER-Notch signaling and GPR30-PI3K/AKT signaling, whereas only the latter pathway is involved in the regulation of growth in nuclear EMTs.
Abstract: To elucidate the mechanisms of nuclear estrogen receptor (ER)-mediated and G protein-coupled receptor 30 (GPR30)-mediated signaling in the regulation of proliferation in ER-positive and ER-negative endometrial cancer cells, two human endometrial carcinoma cell lines, Ishikawa (ER-positive) and KLE (ER-negative), were used. PCR and Western blot analyses were used to determine the effects of estrogen stimulation on the activation of Notch and GPR30-PI3K/AKT signaling. Cell growth was investigated using MTT assays. Overexpression of ER in ER-negative cells was achieved by plasmid transfection and was used to investigate the effects on cellular growth and Notch signaling. GPR30-mediated signaling was evaluated using siRNA interference. Estrogen stimulated cell proliferation in both cell lines, it activated Notch signaling in ER-positive Ishikawa cells, but not in ER-negative KLE cells. Blockade of this signaling by a Notch inhibitor resulted in partial arrest of estrogen-induced cell proliferation in Ishikawa cells. Overexpression of ER in KLE cells restored estrogen-enhanced Notch signaling and further promoted cell growth. GPR30, as a new G-protein-coupled estrogen receptor, was detected in both cell lines, but was stronger in ER-negative KLE cells. Depletion of GPR30 in KLE cells abolished estrogen-induced PI3K/AKT signaling activation and resulted in inhibition of cell proliferation. Conclusively, regulation of proliferation in nuclear ER-positive endometrial cancer cells is mediated by both ER-Notch signaling and GPR30-PI3K/AKT signaling, whereas only the latter pathway is involved in the regulation of growth in nuclear ER-negative endometrial cancer cells.
TL;DR: The results suggest that Ang-(1-7) may have therapeutic potential against advanced lung carcinoma and inactivation of the PI3K/Akt, P38 and JNK signal pathways is demonstrated.
Abstract: The local renin-angiotensin system (RAS) is one of the crucial components in the tumor microenvironment. Recent evidence suggests that the local RAS plays an important role in tumor metabolism, survival, angiogenesis and invasion processes. Angiotensin-(1-7) [Ang-(1-7)] is an endogenous peptide of the RAS with vasodilator and anti-proliferative properties. Previous studies have demonstrated that Ang-(1-7) inhibits both the growth of human lung cancer cells in vitro and tumor angiogenesis in vivo through activation of the MAS receptor. This study investigated the anti-metastatic effect of Ang-(1-7) in A549 human lung adenocarcinoma cells in vitro. We found that Ang-(1-7) reduced the cell migratory and invasive abilities by reducing the expression and activity of MMP-2 and MMP-9. Furthermore, we demonstrated that the anti-migration and anti-invasion effect of Ang-(1-7) was mediated through inactivation of the PI3K/Akt, P38 and JNK signal pathways. Our results suggest that Ang-(1-7) may have therapeutic potential against advanced lung carcinoma as a new agent.