Aberrant DNA Methylation in Human Cancers

DNA methylation, one of the best-characterized epigenetic modifications, plays essential roles in diseases, including human cancers. In recent years, our understanding on DNA methylation with human cancers has made significant progress, which was facilitated by stunning development in the analysis of the human methylome of multiple cancer types. In this review, recent developments in the characterization of aberrant DNA methylation involved in human cancers development were discussed with special emphasis on the mechanisms of aberrant DNA methylation in human cancers. We also summarize the recent treatment strategy for human cancers with de-methylation drugs.

Carcinogen-Macromolecular Adducts As Biomarkers in Human Cancer Risk Assessment

Substantial data have been generated during the past 5 years in both experimental systems and human populations which shed light on the potential role of carcinogen-macromolecular adducts in human cancer risk assessment. The use of DNA and protein adducts is based on the fundamental concept in chemical carcinogenesis that most genotoxins are metabolized to electrophilic 'ultimate' carcinogens that are capable of forming covalent adducts with cellular macromolecules. This report examines the relative usefulness and limitations of using DNA and protein adducts and related techniques for assessing human exposure to genotoxic carcinogens. Data discussed in this report clearly demonstrate that these biomarkers not only allow early detection of potential cancer hazard in humans, but they can also significantly increase the power of conventional cancer epidemiological studies in determining true causal relationships. In addition, such biomarkers can improve extrapolation of cancer risks from laboratory animals to humans or from one human population to another.

Studies on hematoporphyrin-photosensitized effects on human cancer cells in vitro——TEM and SEM observation

The ECa109 esophageal cancer epithelial cell line cells was treated with hematoporphyrin one hour and followed by irrediation with 20W black light at a distance of two cm for ten minutes.The cancer cells were examined by TEM and SEM after treatment for one,three and five days.By exposure to hematoporphyrin derivative(HpD)and photoirradiation,the proportion of dark-cells with high electron density were decreased and the proportion of empty cells with low electron density was increased.The latter showed distruction of cancer cells.

Met as a target in human cancer

Inappropriate Met-hepatocyte growth factor/scatter factor （HGF/SF） signaling is a hallmark of most types of solid tumors in humans and often correlates with poor prognosis. There are several mechanisms by which Met signaling is deregulated including Met and/or HGF/SF overexpression, mutation, autocrine signaling, and ligand-independent activation. Because Met is an attractive therapeutic target for a multitude of cancers, extensive research towards finding Met and HGF/SF inhibitors is ongoing.？ In parallel with these efforts, several mouse models have been developed in our lab that will be valuable for preclinical testing of Met-targeted therapeutics.

World’s First Myoblast Treatment of Human Cancer Found Safe and Efficacious

Evolution of placental mammals over the past 160 million years witnesses the relative sparing of muscles from cancer attacks. In 1) nude mice with human gastrointestinal or lung tumors, and 2) human subjects with liver, lung or gastrointestinal tumors, intra-tumor implantation of allogeneic human myoblasts induced cancer apoptosis, inhibiting metastasis and tumor growth. We postulate four mechanisms of cancer apoptosis: a) myoblasts releasing tumor necrosis factor-α (TNF-α);b) deprivation of nutrients and oxygen;c) local inflammatory and immunologic attacks;and d) prevention from metastasis. These basic and clinical studies demonstrated preliminary safety and efficacy of intra-tumor myoblast implantation in the development of prevention and treatment for cancer, now the number one disease killer of mankind.

Unraveling links between aging,circadian rhythm and cancer:Insights from evidence-based analysis

Aging and circadian rhythms have been connected for decades,but their molecular interaction has remained unknown,especially for cancers.In this situation,we summarized the current research actuality and problems in this field using the bibliometric analysis.Publications in the PubMed and Web of Science databases were retrieved.Overall,there is a rising trend in the publication volume regarding aging and circadian rhythms in the field of cancer.Researchers from USA,Germany,Italy,China and England have greater studies than others.Top three publication institutions are University of California System,UDICE-French Research Universities and University of Texas System.Current research hotspots include oxidative stress,breast cancer,melatonin,cell cycle,calorie restriction,prostate cancer and NF-κB.In conclusion,results generated by bibliometric analysis indicate that many approaches involve in the complex interactions between aging and circadian rhythm in cancer.These established and emerging research directions guide our exploration of the regulatory mechanisms of aging and circadian rhythms in cancer and provide a reference for developing new research avenues.

Tyrosine kinase inhibitors and human epidermal growth factor receptor-2 positive breast cancer

The body of evidence investigating human epidermal growth factor receptor-2(HER2)directed therapy in patients with breast cancer(BC)has been growing within the last decade.Recently,the use of tyrosine kinase inhibitors(TKIs)has been of particular interest in the treatment of human malignancies.This literature commentary is intended to highlight the most recent findings associated with the widely-studied TKI agents and their clinical significance in improving the outcomes of HER2 positive BC.

Mutation analysis of large tumor suppressor genes LATS1 and LATS2 supports a tumor suppressor role in human cancer

In recent years, human cancer genome projects provide unprecedented opportunities for the discovery of cancer genes and signaling pathways that contribute to tumor development. While numerous gene mutations can be identified from each cancer genome, what these muta- tions mean for cancer is a challenging question to address, especially for those from less understood putative new cancer genes. As a powerful approach, in silico bioinformatics analysis could efficiently sort out mutations that are predicted to damage gene function. Such an analysis of human large tumor suppressor genes, LATS1 and LATS2, has been carried out and the results support a role of hLATS1/12 as negative growth regulators and tumor suppressors.

Parasporal Proteins from Bacillus thuringiensis and Their Cytotoxicity on Human Cancer Cells

Parasporins(PSs) represent a novel functional category of crystal proteins(Cry) produced by non-insecticidal Bacillus thuringiensis. A distinct feature for PSs is their specific cytotoxicity against human cancer cells from diverse origins, other than hemolytic or insecticidal activity. As structurally/functionally Cry proteins, parasporins are expressed as protoxins that require protease cleavage for activation. Currently,identified PSs is classified into 6 groups: PS1, PS2, PS3, PS4, PS5 and PS6, which are heterogeneous in cytotoxic spectrum and activity level. Some PSs have been explored for their mode of anticancer activities, reports mainly include pore formation induced by binding to putative receptors on cell membrane and apoptosis by intracellular Ca2+concentration. Further work should focus on the identification of new PS or PS homologs and better understanding of their anticancer mechanism before possible application in cancer therapy.

The role of TRPC6 in HGF-induced cell proliferation of human prostate cancer DU145 and PC3 cells

Hepatocyte growth factor （HGF） is a glycoprotein that induces prostate cancer cell proliferation, migration and invasion. The activation of transient receptor potential canonical 6 （TRPC6） channels is considered important in promoting prostate cancer cell proliferation. In this study, we assessed the role of endogenous TRPC6 channels in the HGF-induced cell proliferation of prostate cancer. Reverse transcription-PCR and Western blotting were used to investigate TRPC6 expression. Electrophysiological techniques （whole-cell patch clamp configuration） and Ca^2＋ imaging analysis were used to investigate the channel activity in cells. The effects of TRPC6 channels on cell cycle progression, cell apoptosis and cell growth were also examined. TRPC6 and c-MET were expressed in DU145 and PC3 cells. In addition, functional TRPC6 channels were present in DU145 and PC3 cells, and TRPC6 knockdown suppressed TRPC-Iike currents evoked by oleoyl-2-acetyl-sn-glycerol （OAG）. Inhibition of TRPC6 channels in DU145 and PC3 cells abolished OAG- and HGF-induced Ca^2＋ entry. Furthermore, inhibition of TRPC6 channels arrested DU145 and PC3 cells at the G2/M phase and suppressed HGF-induced cell proliferation. Collectively, our results indicate that TRPC6 has an important role in HGF-induced DU145 and PC3 cell proliferation.

Anti-cancer effect of ethylacetate fraction from Orostachys japonicus on HT-29 human colon cancer cells by induction of apoptosis through caspase-dependent signaling pathway

Objective: To investigate the anti-colon cancer effects of ethylacetate fraction from Orostachys japonicus(0. japonicus) on HT-29 cancer cells. Methods: The viability of HT-29 cells was assayed by the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2 H-tetrazolium(MTS) method. Apoptosis induction and cell cycle inhibition were confirmed by fluorescein isothiocyanate and propidium iodide staining using flow cytometry.Morphological changes in the nucleus were observed, using a fluorescence microscope with4',6-diamidino-2-phenylindole(DAPI) nuclear staining. The expression levels of the upstream and downstream proteins involved in the anti-cancer mechanism were confirmed by Western blotting. Results: After treating HT-29 cells with different concentrations of ethylacetate fraction from O. japonicus, the viability of cells decreased in a concentration-dependent manner,while apoptosis induction and apoptotic body formation increased. Cell cycle analysis showed that the arrest occurred at the sub-G_1 and S phase. Among the upstream and downstream proteins involved in anti-cancer activity, the level of B cell lymphoma-2 decreased, and the bcl-2-associated x protein increased. The level of pro-caspase-3, pro-caspase-8, and pro-caspase-9 decreased, while the level of cleaved-caspase-3, cleaved-caspase-8, and cleaved-caspase-9 increased. Moreover, the phosphorylation, that is, activation of extracellular signal regulated kinase 1/2, Jun-N-terminal kinase, and p38 increased. Conclusions: Combining the above results, it is thought that the survival of HT-29 cells is suppressed by ethylacetate fraction from0. japonicus through mitochondrial regulation-induced caspase cascade activation, induction of apoptosis and cell cycle arrest.

Anti-proliferation activity of terpenoids isolated from Euphorbia kansui in human cancer cells and their structure-activity relationship

Euphorbia kansui is a commonly used traditional Chinese medicine for the treatment of edema,pleural effusion,and asthma,etc.According to the previous researches,terpenoids in E.kansui possess various biological activities,e.g.,anti-virus,anti-allergy,antitumor effects.In this work,twenty five terpenoids were isolated from E.kansui,including thirteen ingenane-and eight jatrophane-type diterpenoids(with two new compounds,kansuinin P and Q)and four triterpenoids.Eighteen of them were analyzed by MTS assay for in vitro anticancer activity in five human cancer cell lines.Structure-activity relationship for 12 ingenane-type diterpenoids in colorectal cancer Colo205 cells were preliminary studied.Significant anti-proliferation activities were observed in human melanoma cells breast cancer MDA-MB-435 cells and Colo205 cells.More than half of the isolated ingenane-type diterpenoids showed inhibitory activities in MDA-MB-435 cells.Eight ingenane-and one jatrophane-type diterpenoids possessed much lower IC_(50) values in MDA-MB-435 cells than positive control staurosporine.Preliminary structure-activity relationship analysis showed that substituent on position 20 was important for the activity of ingenane-type diterpenoids in Colo205 cells and substituent on position 3 contributed more significant biological activity of the compounds than that on position 5 in both MDA-MB-435 and Colo205 cells.

Viral Etiology Relationship between Human Papillomavirus and Human Breast Cancer and Target of Gene Therapy

Objective To explore the viral etiology of human breast cancer to determine whether there are novel molecular targets for gene therapy of breast cancer and provide evidence for the research of gene therapy and vaccine development for breast cancer. Methods PCR was used to screen HPV16 and HPV18 oncogenes E6 and E7 in the SKBR3 cell line and in 76 paraffin embedded breast cancer tissue samples. RNA interference was used to knock down the expression of HPV18 E6 and E7 in SKBR3 cells, then the changes in the expression of cell-cycle related proteins, cell viability, colony formation, metastasis, and cell cycle progression were determined. Results HPV18 oncogenes E6 and E7 were amplified and sequenced from the SKBR3 cells. Of the patient samples, 6.58% and 23.68% were tested to be positive for HPV18 E6 and HPV18 E7. In the cell culture models, the knockdown of HPV18 E6 and E7 inhibited the proliferation, metastasis, and cell cycle progression of SKBR3 cell. The knockdown also clearly affected the expression levels of cell cycle related proteins. Conclusion HPV was a contributor to virus caused human breast cancer, suggesting that the oncogenes in HPV were potential targets for gene therapy of breast cancer.

Expression of cell cycle regulator p57^(kip2), cyclinE protein and proliferating cell nuclear antigen in human pancreatic cancer: An immunohistochemical study

AIM: To investigate the effects of p57kip2, cyclinE protein and proliferating cell nuclear antigen (PCNA) on occurrence and progression of human pancreatic cancer.METHODS: The expression of p57kip2, cyclinE protein and PCNA in tumor tissues and adjacent tissues from 32patients with pancreatic cancer was detected by SP immunohistochemical technique.RESULTS: The positive expression rate of p57kip2 protein in tumor tissues was 46.9%, which was lower than that in adjacent pancreatic tissues (x2 = 5.317, P＜0.05). P57kip2protein positive expression remarkably correlated with tumor cell differentiation (P＜0.05), but not with lymph node metastasis (P＞0.05). The positive expression rate of cyclinE protein in tumor tissues was 68.8%, which was higher than that in adjacent pancreatic tissues (x2 = 4.063,P＜0.05). CyclinE protein positive expression significantly correlated with tumor cell differentiation and lymph node metastasis (P＜0.05). The positive expression rate of PCNA in the tumor tissues was 71.9%, which was higher than that in adjacent pancreatic tissues (x2 = 5.189, P＜0.05).PCNA positive expression remarkably correlated with tumor cell differentiation and lymph node metastasis (P＜0.05).CONCLUSION: The decreased expression of p57kip2 and/or overexpression of cyclinE protein and PCNA may contribute to the occurrence and progression of pancreatic cancer.p57kip2, cyclinE protein, and PCNA play an important role in occurrence and progression of pancreatic cancer.

Influence of Osteopontin Short Hairpin RNA on the Proliferation and Invasion of Human Renal Cancer Cells

The influence of short hairpin RNA(shRNA)-mediated osteopontin(OPN)gene silencing on the proliferation and invasion of human renal cancer ACHN cells was investigated.Four types of OPN shRNA recombinant plasmids were constructed and RT-PCR assays were used to screen the most highly functional shRNA recombinant plasmids,which were transferred into the cultured ACHN cells by LipofectamineTM 2000.The cells transfected by shRNA expression vectors(ACHN/OPN)were visualized under an inverted microscope and screened...

Highly Efficient Labeling of Human Lung Cancer Cells Using Cationic Poly-L-lysine-Assisted Magnetic Iron Oxide Nanoparticles

Cell labeling with magnetic iron oxide nanoparticles(IONPs)is increasingly a routine approach in the cellbased cancer treatment.However,cell labeling with magnetic IONPs and their leading effects on the biological properties of human lung carcinoma cells remain scarcely reported.Therefore,in the present study the magnetic c-Fe2O3nanoparticles(MNPs)were firstly synthesized and surface-modified with cationic poly-L-lysine(PLL)to construct the PLL-MNPs,which were then used to magnetically label human A549 lung cancer cells.Cell viability and proliferation were evaluated with propidium iodide/fluorescein diacetate double staining and standard 3-(4,5-dimethylthiazol-2-diphenyl-tetrazolium)bromide assay,and the cytoskeleton was immunocytochemically stained.The cell cycle of the PLL-MNPlabeled A549 lung cancer cells was analyzed using flow cytometry.Apoptotic cells were fluorescently analyzed with nuclear-specific staining after the PLL-MNP labeling.The results showed that the constructed PLL-MNPs efficiently magnetically labeled A549 lung cancer cells and that,at low concentrations,labeling did not affect cellular viability,proliferation capability,cell cycle,and apoptosis.Furthermore,the cytoskeleton in the treated cells was detected intact in comparison with the untreated counterparts.However,the results also showed that at high concentration(400 lg m L-1),the PLL-MNPs would slightly impair cell viability,proliferation,cell cycle,and apoptosis and disrupt the cytoskeleton in the treated A549 lung cancer cells.Therefore,the present results indicated that the PLL-MNPs at adequate concentrations can be efficiently used for labeling A549 lung cancer cells and could be considered as a feasible approach for magnetic targeted anti-cancer drug/gene delivery,targeted diagnosis,and therapy in lung cancer treatment.

Radiosensitivity of human colon cancer cell enhanced by immunoliposomal docetaxel

AIM: To enhance the radiosensitivity of human colon cancer cells by docetaxel.METHODS: Immunoliposomal docetaxel was prepared by coupling monodonal antibody against carcinoembryonic antigen to cyanuric chloride at the PEG terminus of liposome. LoVo adenocarcinoma cell line was treated with immunoliposomal docetaxel or/and irradiation. MTT colorimetric assay was used to estimate cytotoxicity of immunoliposomal docetaxel and radiotoxicity. Cell cycle redistribution and apoptosis were determined with flow cytometry. Survivin expression in LoVo cells was verified by immunohistochemistry. D801 morphologic analysis system was used to semi-quantify immunohistochemical staining of survivin.RESULTS: Cytotoxicity was induced by immunoliposomal docetaxel alone in a dose-dependent manner. Immunoliposomal docetaxel yielded a cytotoxicity effect at a low dose of 2 nmol/L. With a single dose irradiation, the relative surviving fraction of LoVo cells showed a dosedependent response, but there were no significant changes as radiation delivered from 4 to 8 Gy. Compared with liposomal docetaxel or single dose irradiation,strongly radiopotentiating effects of immunoliposomal docetaxel on LoVo cells were observed. A low dose of immunoliposomal docetaxel could yield sufficient radiosensitivity. Immunoliposomal docetaxel were achieved both specificity of the conjugated antibody and drug radiosensitization. Combined with radiation,immunoliposomal docetaxel significantly increased the percentage of G2/M cells and induced apoptosis, but significantly decreased the percentage of cells in G2/G1 and S phase by comparison with liposomal docetaxel.Immunohistochemical analysis showed that the brown stained survivin was mainly in cytoplasm of LoVo cells.Semi-quantitative analysis of the survivin immunostaining showed that the expression of survivin in LoVo cells under irradiation with immunoliposomal docetaxel was significantly decreased.CONCLUSION: Immunoliposomal docetaxel is strongly effective for target radiosensitation in LoVo colon carcinoma cells, and may offer the potential to improve local radiotherapy.

Molecular Mechanism of Induction on Apoptosis of Human Esophageal Cancer HCE-4 Cells by Active Components from Astragalus membranaceus

[Objectives] To investigate the pharmacologic effects of active components from A. membranaceus on human esophageal cancer HCE-4 cells and its apoptosis mechanism. [Methods] The viabilities of HCE-4 cells were measured by MTT assay. The induction of active components from A. membranaceus on apoptosis of HCE-4 cells was detected by Annexin V-FITC/PI double staining. The apoptotic-related protein expression levels were determined by Western blotting. [Results] Formononetin and astragaloside IV suppressed the proliferation of HCE-4 cells in a dose-dependent manner. The Annexin V-FITC/PI double staining results showed that formononetin and astragaloside IV could induce HCE-4 cells apoptosis in a time-dependent manner. The Western blotting results showed that formononetin and astragaloside IV could significantly down-regulate p-AKT,pro-caspase-3,and increase cle-caspase-3 protein expression in HCE-4 cells. [Conclusions]Active components from A. membranaceus such as formononetin and astragaloside IV significantly inhibited the proliferation of human esophageal cancer HCE-4 cells by inducing mitochondrial dependent apoptosis via AKT signaling pathway.

Epigenetic modification regulates both expression of tumor-associated genes and cell cycle progressing in human colon cancer cell lines: Colo-320 and SW1116

The aim of this study is to assess the effects of DNA methylation and histone acetylation, alone or in combination, on the expression of several tumor-associated genes and cell cycle progression in two established human colon cancer cell lines: Colo-320 and SW1116. Treatments with 5-aza-2'-deoxycytidine (5-aza-dC) and trichostatin A, alone or in combination, were applied respectively. The methylation status of the CDKN2A promoter was determined by methylation-specific PCR, and the acetylated status of the histones associated with the p21wAF1 and CDKN2A genes was examined by chromatin immunoprecipitation. The expression of the CDKN2A, p21WAF1, p53, p73, APC, c-myc, c-Ki-ras and survivin genes was detected by real-time RT-PCR and RT-PCR. The cell cycle profile was established by flow cytometry.We found that along with the demethylation of the CDKN2A gene promoter in both cell lines induced by 5-aza-dC alone or in combination with TSA, the expression of both CDKN2A and APC genes increased. The treatment of TSA or sodium butyrate up-regulated the transcription of p21 WAF1 significantly by inducing the acetylation of histones H4 and H3, but failed to alter the acetylation level of CDKN2A-associated histones. No changes in transcription of p53, p73,c-myc, c-Ki-ras and survivin genes were observed. In addition, TSA or sodium butyrate was shown to arrest cells at the G1 phase. However, 5-aza-dC was not able to affect the cell cycle progression. In conclusion, regulation by epigenetic modification of the transcription of tumor-associated genes and the cell cycle progression in both human colon cancer cell lines Colo-320 and SW1116 is gene-specific.

Effects of Estradiol and Tamoxifen on Proliferation of Human Breast Cancer Cells and Human Endometrial Cells

The effects of estradiol and tamoxifen on the proliferation of estrogen receptor positive cells and the relationship between the tamoxifen tolerance and cell origin were investigated The tissues of human endometrium and breast cancer were randomly selected following dissection for primary cell culture After the breast cancer cells and endometrial cells were treated with 1×10 -8 mol/L estradiol and/or 1 ×10 -6 tamoxifen, 3H-labelled thymine nucleotide was used to trace the kinetics of cell proliferation There was no significant difference in the inhibition on the human endometrial cells between tamoxifen-treated group (6 3%) and control group (6 4%), but tamoxifen could significantly inhibit the proliferation of the human breast cancer cells (45 84%) as compared with control group (52 72%) Moreover, tamoxifen could significantly stimulate the proliferation of tamoxifen resistant breast cancer cells (9 64%) as compared with control group (6 32%) Estradiol could significantly stimulate the proliferation of all the three kinds of cells as compared with control group The combined use of estradiol and tamoxifen could inhibit the proliferation of the endometrial cells and breast cancer cells as compared with estradiol used alone, but on the tamoxifen resistant breast cancer cells, they could more significantly stimulate the proliferation than E2 It was concluded that E2 could stimulate the proliferation of these three kinds of cells However, the inhibitive effects of tamoxifen on the proliferation of these cells were dependent on the estradiol