Anti-Survivin (N111) Antibody (A25779)

Aberrant promoter region hypermethylation of upstream transcription factors may be responsible for silencing entire anti-neoplastic gene networks. In this study, we explored whether transcription factor coding gene, caudal-related homeobox 2 (CDX2), is silenced by promoter hypermethylation in lung cancer, and examined its potential tumor-suppressive functions. Semi-quantitative RT-PCR showed that four of six lung cancer cell lines exhibited no or weak CDX2 expression. Expression of CDX2 was correlated to CDX2 promoter region methylation status, as determined by methylation-specific PCR (MSP) and bisulfite sequencing. Restoration of CDX2 expression was induced by treatment with demethylating drug 5-aza-2'-deoxycytidine (5-AZA) in lung cancer cell lines. Methylation of CDX2 was common in human primary lung cancer (61 of 110 tumors, 55.45%), but no methylation was found in normal lung tissues. Re-expression of CDX2 suppressed lung cancer cell proliferation and blocked cells in G1 phase. β-catenin/TCF activity and downstream genes expression were inhibited by re-expression of CDX2, and increased by depletion of CDX2. In conclusion, CDX2 is frequently methylated in lung cancer, and expression of CDX2 is regulated by promoter region hypermethylation. CDX2 may serve as a tumor suppressor in lung cancer and inhibits lung cancer cell proliferation by suppressing Wnt signaling.

The present study evaluated whether the magnetic nanoparticles of Fe(3)O(4) (MNPs-Fe(3)O(4)) could enhance the activity of artesunate (ART), and to explore its potential mechanisms. Cytotoxicity of the copolymer of ART with MNPs-Fe(3)O(4) on K562 cells was detected by MTT assay and the apoptosis rate of K562 cells was measured by flow cytometry. Protein expression levels of bcl-2, bax, bcl-rambo, caspase-3, and survivin in K562 cells were measured by Western blot. After being incubated with the copolymer of ART with MNPs-Fe(3)O(4) for 48 hours, the growth inhibition rate of K562 cells was significantly increased compared with that of K562 cells treated with ART alone (P < 0.05), and the apoptosis rate of K562 cells was increased significantly compared with that of K562 cells treated with ART alone, suggesting that MNPs-Fe(3)O(4) can enhance the activity of ART. Interestingly, the copolymer-induced cell death was attenuated by caspase inhibitor Z-VAD-FMK. Our results also showed that treatment with the copolymer of MNPs-Fe(3)O(4) and ART increased the expression of bcl-2, bax, bcl-rambo, and caspase-3 proteins, and decreased the expression of survivin protein in K562 cells compared with ART treatment alone. These results suggest that MNPs-Fe(3)O(4) can enhance ART-induced apoptosis, which may be related to the upregulation of bcl-rambo and downregulation of survivin.

BACKGROUND:

Pancreatic cancer is a leading cause of cancer-related deaths in the world with a 5-year survival rate of less than 6%. Currently, there is no successful therapeutic strategy for advanced pancreatic cancer, and new effective strategies are urgently needed. Recently, an arginine deprivation agent, arginine deiminase, was found to inhibit the growth of some tumor cells (i.e., hepatocellular carcinoma, melanoma, and lung cancer) deficient in argininosuccinate synthetase (ASS), an enzyme used to synthesize arginine. The purpose of this study was to evaluate the therapeutic efficacy of arginine deiminase in combination with gemcitabine, the first line chemotherapeutic drug for patients with pancreatic cancer, and to identify the mechanisms associated with its anticancer effects.

METHODS:

In this study, we first analyzed the expression levels of ASS in pancreatic cancer cell lines and tumor tissues using immunohistochemistry and RT-PCR. We further tested the effects of the combination regimen of arginine deiminase with gemcitabine on pancreatic cancer cell lines in vitro and in vivo.

RESULTS:

Clinical investigation showed that pancreatic cancers with reduced ASS expression were associated with higher survivin expression and more lymph node metastasis and local invasion. Treatment of ASS-deficient PANC-1 cells with arginine deiminase decreased their proliferation in a dose- and time-dependent manner. Furthermore, arginine deiminase potentiated the antitumor effects of gemcitabine on PANC-1 cells via multiple mechanisms including induction of cell cycle arrest in the S phase, upregulation of the expression of caspase-3 and 9, and inhibition of activation of the NF-κB survival pathway by blocking NF-κB p65 signaling via suppressing the nuclear translocation and phosphorylation (serine 536) of NF-κB p65 in vitro. Moreover, arginine deiminase can enhance antitumor activity of gemcitabine-based chemotherapy in the mouse xenograft model.

CONCLUSIONS:

Our results suggest that arginine deprivation by arginine deiminase, in combination with gemcitabine, may offer a novel effective treatment strategy for patients with pancreatic cancer and potentially improve the outcome of patients with pancreatic cancer.

PURPOSE:

This study was conducted to investigate the small double-stranded RNA (dsRNA) mediated anti-tumor effects of p21WAF1/ClP1 (p21) transcriptional activation in vitro in the human glioma SHG-44 cell line.

MATERIALS AND METHODS:

Human glioma SHG-44 cells were transfected with dsRNA using LipofectAMINE 2000 transfection reagent. Real-time PCR and Western blot analysis were conducted to detect p21 and survivin mRNA and protein levels, respectively. Cell proliferation was examined by MTT assay. Cell cycle distribution and apoptosis were detected by flow-cytometric analysis.

RESULTS:

We found that dsRNA targeting p21 promoter (dsP21) significantly induced the expression of p21 at transcription and protein levels, and reduced the expression of survivin. AS well, dsP21 transcription significantly inhibited human glioma SHG-44 cell proliferation. Analysis of cell cycle distribution revealed that dsP21 transfection increased accumulation of cells in the G0/G1 phase and reduced accumulation of cells in the S phase. Further analysis revealed that dsP21 transcription led to an increase in both early and late stages of apoptosis in human glioma SHG-44 cells.

CONCLUSION:

In the present study, P21 activation by RNA-induced gene activation (RNAa) induced anti-tumor activity in vitro in a human glioma SHG-44 cell line. The results suggested that RNAa could be used for human glioma treatment by targeted activation of tumor suppressor genes.

BACKGROUND:

Colorectal cancer remains one of the leading causes of cancer death worldwide. Traditional Chinese Medicine (TCM) has played a positive role in colorectal cancer treatment. There is a great need to establish effective herbal formula for colorectal cancer treatment. Based on TCM principles and clinical practices, we have established an eight herbs composed formula for colorectal cancer treatment, which is Teng-Long-Bu-Zhong-Tang (TLBZT). We have demonstrated the anticancer effects of TLBZT against colorectal carcinoma in vitro. In present study, we evaluated the anticancer potential of TLBZT, used alone or in combination with low dose of 5-Fluorouracil (5-Fu), in CT26 colon carcinoma in vivo.

METHODS:

CT26 colon carcinoma was established in BALB/c mice and treated with TLBZT, 5-Fu, or TLBZT plus 5-Fu. The tumor volumes were observed. Apoptosis was detected by TUNEL assay. Caspases activities were detected by colorimetric assay. Cell senescence was indentified by senescence β-galactosidase staining. Gene expression and angiogenesis was observed by immunohistochemistry or western blot.

RESULTS:

TLBZT significantly inhibited CT26 colon carcinoma growth. TLBZT elicited apoptosis in CT26 colon carcinoma, accompanied by Caspase-3, 8, and 9 activation and PARP cleavage, and downregulation of XIAP and Survivin. TLBZT also induced cell senescence in CT26 colon carcinoma, with concomitant upregulation of p16 and p21 and downregulation of RB phosphorylation. In addition, angiogenesis and VEGF expression in CT26 colon carcinoma was significantly inhibited by TLBZT treatment. Furthermore, TLBZT significantly enhanced anticancer effects of 5-Fu in CT26 colon carcinoma.

CONCLUSIONS:

TLBZT exhibited significantly anticancer effect, and enhanced the effects of 5-Fu in CT26 colon carcinoma, which may correlate with induction of apoptosis and cell senescence, and angiogenesis inhibition. The present study provides new insight into TCM approaches for colon cancer treatment that are worth of further study.

Effective therapeutics against triple negative breast cancer (TNBC), which has no standard-of-care therapy, needs to be developed urgently. Here we demonstrated a strategy of integrating indocyanine green (ICG), paclitaxel (PTX), and survivin siRNA into one thermosensitive poly(2-(2-methoxyethoxy)ethyl methacrylate-co-oligo(ethylene glycol) methacrylate)-co-2-(dimethylamino)ethyl methacrylate-b-poly(D,L-lactide-co-glycolide) (P (MEO2MA-co-OEGMA-co-DMAEMA)-b-PLGA) nanoparticle (NP-IPS) for triple-punch strategy against TNBC. The NP-IPS significantly enhanced the stability of ICG. Controlled release of the PTX in tumor regions was triggered by the hyperthermia produced by laser irradiated ICG. The NP-IPS exhibited remarkable antitumor efficacy (almost complete ablation of the tumor xenografts) due to the combinational effects of chemotherapy, photothermal therapy, and gene therapy with low drug dose (ICG, 0.32 μmol/kg; PTX, 0.54 μmol/kg; siRNA, 1.5 mg/kg) and minimal side effects. Taken together, our current study demonstrates a nanoplatform for triple-therapy, which reveals a promising strategy for TNBC treatment.

MicroRNA-31 (miR-31) plays important roles in colon cancer development. However, the underlying mechanism is still not clear. We have explored the functions of miR-31 on proliferation of colon cancer cells as well as the underlying mechanism. E2F2 was identified as a direct target of miR-31. miR-31 regulated the proliferation of colon cancer cells by targeting E2F2. Moreover, in the present study, E2F2 acted as a tumor suppressor in colon cancer by repressing the expression of survivin and regulating the expression of CCNA2, C-MYC, MCM4 and CDK2. A possible mechanism for the function of miR-31 on colon cancer proliferation is presented and indicates that miR-31 might become a target for anti-cancer drug design.

Poor viability of transplanted bone marrow mesenchymal stem cells (BMSCs) is well‑known, but developing methods for enhancing the viability of BMSCs requires further investigation. The aim of the present study was to elucidate the effects of infrasound on the proliferation and apoptosis of BMSCs, and to determine the association between survivin expression levels and infrasound on BMSCs. Primary BMSCs were derived from Sprague Dawley rats. The BMSCs, used at passage three, were divided into groups that received infrasound for 10, 30, 60, 90 or 120 min, and control groups, which were exposed to the air for the same durations. Infrasound was found to promote proliferation and inhibit apoptosis in BMSCs. The results indicated that 60 min was the most suitable duration for applied infrasound treatment to BMSCs. The protein and mRNA expression levels of survivin in BMSCs from the two treatment groups that received 60 min infrasound or air, were examined by immunofluorescence and quantitative polymerase chain reaction. Significant differences in survivin expression levels were identified between the two groups, as infrasound enhanced the expression levels of survivin. In conclusion, infrasound promoted proliferation and inhibited apoptosis in BMSCs, and one mechanisms responsible for the protective effects may be the increased expression levels of survivin.

MicroRNAs play important roles in carcinogenesis by negatively regulating the expression of target genes. Here we explore the biological function of miR-155 and the underlying mechanism in colorectal carcinoma. We validate, for the first time, that E2F2 is a direct target of miR-155 using western blot and a luciferase reporter assay and that miR-155 regulates the proliferation and cell cycle of colorectal carcinoma cells by targeting E2F2 using siRNA technology. We also found, for the first, time that E2F2 acts as a tumor suppressor in colorectal carcinoma. Overall, miR-155 plays an important role in colorectal carcinoma tumorigenesis by negative regulation of its targets including E2F2 and may be a potential therapeutic target for colorectal carcinoma treatment.

Ran, a member of the Ras GTPase family, has important roles in nucleocytoplasmic transport. Herein, we detected Ran expression in pancreatic cancer and explored its potential role on tumour progression. Overexpressed Ran in pancreatic cancer tissues was found highly correlated with the histological grade. Downregulation of Ran led to significant suppression of cell proliferation, cell cycle arrest at the G1/S phase and induction of apoptosis. In vivo studies also validated that result. Further studies revealed that those effects were at least partly mediated by the downregulation of Cyclin A, Cyclin D1, Cyclin E, CDK2, CDK4, phospho-Rb and Survivin proteins and up regulation of cleaved Caspase-3.

Histone deacetylase (HDAC) inhibitors have recently emerged as a new class of anti-cancer agents. Trichostatin A (TSA), a classical HDAC inhibitor, has been demonstrated to induce cell cycle arrest, promote cell apoptosis, and inhibit metastasis. However, the molecular mechanism underlying TSA function has not been fully elucidated. In the current study, we found that TSA treatment induced altered expression of cell cycle-associated genes in HCT116 cells by RT-PCR array. Among the 84 genes related to cell cycle control, 34 genes were significantly altered by TSA treatment, with 7 genes upregulated and 27 genes downregulated. Interestingly, gene expression of minichromosome maintenance protein-2 (MCM-2) was significantly downregulated by TSA treatment. This was confirmed by quantitative RT-PCR and Western blotting. Moreover, silencing of MCM-2 by siRNA led to cell cycle arrest and apoptosis in HCT116 cells. In addition, TSA caused an increase of phosphorylated JNK, which was involved in downregulation of MCM-2. Together, our results suggest that MCM-2 is a noval therapeutic target of TSA in colon cancer cells.

T-cell acute lymphoblastic leukemias (T-ALLs) are clonal lymphoid malignancies with a poor prognosis, and still a lack of effective treatment. Here we examined the interactions between the mammalian target of rapamycin (mTOR) inhibitor rapamycin and idarubicin (IDA) in a series of human T-ALL cell lines Molt-4, Jurkat, CCRF-CEM and CEM/C1. Co-exposure of cells to rapamycin and IDA synergistically induced T-ALL cell growth inhibition and apoptosis mediated by caspase activation via the intrinsic mitochondrial pathway and extrinsic pathway. Combined treatment with rapamycin and IDA down-regulated Bcl-2 and Mcl-1, and inhibited the activation of phosphoinositide 3-kinase (PI3K)/mTOR and extracellular signal-related kinase (ERK). They also played synergistic pro-apoptotic roles in the drug-resistant microenvironment simulated by mesenchymal stem cells (MSCs) as a feeder layer. In addition, MSCs protected T-ALL cells from IDA cytotoxicity by up-regulating ERK phosphorylation, while rapamycin efficiently reversed this protective effect. Taken together, we confirm the synergistic antitumor effects of rapamycin and IDA, and provide an insight into the potential future clinical applications of combined rapamycin-IDA regimens for treating T-cell malignancies.

Hepatocellular carcinoma remains one of the most prevalent malignancies worldwide. Curcuma aromatica and Polygonum cuspidatum are one of the commonly used paired-herbs for liver cancer treatment. Curcumin and resveratrol are the major anticancer constituents of Curcuma aromatica and Polygonum cuspidatum, respectively. Curcumin and resveratrol have been found to exhibit a synergistic anticancer effect in colon cancer. However, the combined effect of curcumin and resveratrol against hepatocellular carcinoma remains unknown. In the present study, we evaluated the combined effects of curcumin and resveratrol in hepatocellular carcinoma Hepa1-6 cells. The results showed that curcumin and resveratrol significantly inhibited the proliferation of Hepa1-6 cells in a dose- and time-dependent manner. The combination treatment of curcumin and resveratrol elicited a synergistic antiproliferative effect in Hepa1-6 cells. The apoptosis of Hepa1-6 cells induced by the combination treatment with curcumin and resveratrol was accompanied by caspase-3, -8 and -9 activation, which was completely abrogated by a pan caspase inhibitor, Z-VAD-FMK. Combination of curcumin and resveratrol upregulated intracellular reactive oxygen species (ROS) levels in Hepa1-6 cells. The ROS scavenger, NAC, partially attenuated the apoptosis and caspase activation induced by the combination treatment of curcumin and resveratrol. In addition, the combination of curcumin and resveratrol downregulated XIAP and survivin expression. These data suggest that the combination treatment of curcumin and resveratrol is a promising novel anticancer strategy for liver cancer. The present study also provides new insights into the effective mechanism of paired-herbs in traditional Chinese medicine.

8-Methoxypsoralen (8-MOP), a naturally occurring compound, is a potent modulator of epidermal cell growth and differentiation in combination with ultraviolet light. However, there is little information on 8-MOP contribution to cell apoptosis alone. In the study, we evaluated 8-MOP, independently of its photoactivation, induced apoptosis in human hepatocellular carcinoma HepG2 cells. And we provide a molecular explanation linking 8-MOP to induce apoptosis. In HepG2 cells, treatment with 8-MOP induced the cell apoptosis in both dose-dependent and time-dependent manners. IC(50) values of 8-MOP were 8.775, 5.398 μM for 48 and 72 h, respectively. Further study showed that 8-MOP decreased the procaspase-3, procaspase-8, and procaspase-9, increased the ratio of Bax/Bcl-2 and decreased the survivin. Moreover, 8-MOP decreased differentiated embryonic chondrocyte gene1 (DEC1). Overexpression of DEC1 antagonized partially apoptosis induced by 8-MOP. And overexpression of DEC1 abolished the decrease of survivin and the activation of caspase-3 induced by 8-MOP partially. So, down regulation of DEC1 is involved in 8-MOP-induced apoptosis in HepG2 cells. Here, it is demonstrated that DEC1 possesses anti-apoptotic effects in 8-MOP-treated HepG2 cells. The findings provide more of a basis for 8-MOP as an anti-tumor agent in cancer therapy.

Notch receptor signaling pathway (NRSP) is increasingly linked to carcinogenesis. Non-small cell lung cancer (NSCLC) appears to actively utilize this conserved developmental pathway. The aims of this study are to determine whether or not Notch 1-4 are overexpressed in NSCLC tissues compared with normal lung tissues and whether inhibiting NRSP could induce caspase-dependent or caspase-independent apoptosis. Immunohistochemistry was used to evaluate the expression of Notch 1-4 in 101 NSCLC tissue samples and 30 normal lung tissue samples. DAPT was used to repress NRSP in SK-MES-1 cells. Apoptosis was determined by Annexin V and PI staining. Cleaved poly ADP-ribose polymerase (PARP) was measured by Western blot; X-linked inhibitor of apoptosis protein (XIAP) and Survivin were assessed by qRT-PCR and Western blot; the release of second mitochondria-derived activator of caspase (Smac) from mitochondria to cytoplasm was evaluated by Western blot; the subcellular locations of endonuclease G (Endo G) and apoptosis inducing factor (AIF) were observed by Western blot and indirect immunofluorescence analysis. (Mech Dev, 98, 2000, 95) Notch 1-4 are up-regulated in NSCLC tissues and Notch 1, 2 are positively correlated with lymph node metastasis, (Proc Natl Acad Sci U S A, 106, 2009, 22293) DAPT treatment could inhibit NRSP and induce apoptosis, with a marked increase in cleaved PARP, decreases in XIAP and Survivin proteins and concomitant release of Smac, EndoG, and AIF from mitochondria, indicating that inhibiting NRSP by DAPT triggers caspase-dependent and caspase-independent apoptosis.

Cancerous inhibitor of protein phosphatase 2A (CIP2A) and survivin are aberrantly expressed in a wide range of human cancers, including lung tumors. In order to assess the expressions of these two proteins in Chinese non-small-cell lung cancer (NSCLC) patients and determine their correlation with prognosis, NSCLC tissues and adjacent non-cancerous normal lung tissues were collected from 97 patients undergoing surgical treatment and evaluated by immunohistochemistry staining. CIP2A or survivin immunoreactivity was detected in significantly more NSCLC tissues than in adjacent non-cancerous lung tissues (P < 0.05). Moreover, CIP2A expression in NSCLC correlated with TNM stage, while survivin expression correlated with TNM stage and lymph node metastasis. Kaplan-Meier survival analysis showed that the overall survival times in patients expressing either CIP2A or survivin protein in NSCLC were shorter. COX regression analysis indicated that expression of CIP2A protein was an independent prognostic factor for NSCLC patients (HR = 3.631, P = 0.015). Therefore, CIP2A expression in Chinese NSCLC patients may be a useful biomarker of biological malignancy.

AIM:

To examine epigenetic changes and the function of HOXA11 in human gastric cancer (GC).

MATERIALS & METHODS:

Seven GC cell lines, five cases of normal gastric mucosa and 112 cases primary GC samples were used in this study.

RESULTS:

Expression of HOXA11 and lack of promoter region methylation were found in NCI-N87, MKN45, BGC823 and HGC27 cells. Loss of expression and complete methylation were found in AGS gastric cancer cells. Reduced expression and partial methylation were found in MGC803 and SGC7901 cells. Restoration of HOXA11 expression was induced by 5-aza-2'-deoxycytidine. HOXA11 was methylated in 81.25% (91/112) of primary GCs. The presence of methylation was associated with male gender, tumor size, tumor differentiation and lymph node metastasis (all p < 0.05). Restoration of HOXA11 expression reduced cell proliferation, invasion, migration and induced apoptosis and G2/M phase arrest. HOXA11 was found to inhibit Wnt signaling by upregulating NKD1 expression.

CONCLUSION:

Epigenetic silencing of HOXA11 promotes GC proliferation, migration and invasion through activation of Wnt signaling.