AIM: To establish the DNA methylation patterns of the promoter CpG islands of 14 'drug-resistance' genes in hepatocellular carcinoma (HCC).METHODS: The methylation specific polymerase chain reaction in conjunc...AIM: To establish the DNA methylation patterns of the promoter CpG islands of 14 'drug-resistance' genes in hepatocellular carcinoma (HCC).METHODS: The methylation specific polymerase chain reaction in conjunction with sequencing verification was used to establish the methylation patterns of the 14 genes in the liver tissues of four healthy liver donors, as well as tumor and the paired non-cancerous tissues of 30 HCC patients.RESULTS: While 11 genes (ATP-binding cassette, sub-family G (WHITE), member 2(ABCG2), activating transcription factor (ATF2), beta-2-microglobulin (B2M), deoxycytidine kinase(DCK, occludin (OCLN, v-raf-1 murine leukemia viral oncogene homolog (RAF/), ralA binding protein 1 (RALBP1),splicing factor (45 kD) (SPF45), S-phase kinase-associated protein 2 (p45) (SKP2), tumor protein p53 (Li-Fraumeni syndrome) (TP53) and topoisomerase (DNA) II beta (TOP2B) maintained the unmethylated patterns, three genes displayed to various extents the hypermethylation state in tumor tissues in comparison with the normal counterparts. The catalase (CAT) was hypermethylated in tumor and the neighboring non-cancerous tissue of one case (3.3%). Both glutathione S-transferase pi (GSTp/) (80%, 24/30 in tumor and 56.7%,17/30 in the paired non-cancerous tissues) and cystic fibrosis transmembrane conductance regulator, ATP-binding cassette (sub-family C, member 7) (CF-FR) (77%, 23/30 in tumor and 50%, 15/30 in the paired non-cancerous tissues) genes were prevalently hypermethylated in HCC as well as their neighboring non-cancerous tissues. No significant difference in the hypermethylation occurrence was observed between the HCC and its neighboring non-cancerous tissues.CONCLUSION: Hypermethylation of promoter CpG islands of both CF-FR and GSTpigenes occurs prevalently in HCC,which may correlate with the low expression of these two genes at the mRNA level and has the profound etiological and clinical implications. It is likely to be specific to the early phase of HCC carcinogenesis.展开更多
Lung cancer is one of the leading causes of death with one of the lowest survival rates. However, a subset of lung cancer patients who are of Asian origin and carry somatic mutations in epidermal growth factor recepto...Lung cancer is one of the leading causes of death with one of the lowest survival rates. However, a subset of lung cancer patients who are of Asian origin and carry somatic mutations in epidermal growth factor receptor or EGFR have responded remarkable well to two tyrosine kinase inhibitors, gefitinib and erlotinib. While EGFR mutation profiles have been reported from Japan, South Korea, and Taiwan, there is no such report from mainland of China where the largest pool of patients reside. In this report, we identified ten somatic mutations from a total of 41 lung cancer patients in China. Among them, seven mutations were found in 17 adenocarcinomas. In contrast to previous reports, eight of these mutations are deletions in exon 19 and two of these deletions are homozygous. These results suggest that a large portion of Chinese adenocarcinoma patients could benefit from gefitinib or erlotinib. This unique mutation profile provides a rationale to develop the next generation of EGFR inhibitors more suitable for the Chinese population.展开更多
DNA methylation is the most intensively studied epigenetic phenomenon, disturbances of which result in changes ingene transcription, thus exerting drastic imparts onto biological behaviors of cancer. Both the global d...DNA methylation is the most intensively studied epigenetic phenomenon, disturbances of which result in changes ingene transcription, thus exerting drastic imparts onto biological behaviors of cancer. Both the global demethylation andthe local hypermethylation have been widely reported in all types of tumors, providing both challenges and opportunitiesfor a better understanding and eventually controlling of the malignance. However, we are still in the very early stage ofinformation accumulation concerning the tumor associated changes in DNA methylation pattern. A number of excellentrecent reviews have covered this issue in depth. Therefore, this review will summarize our recent data on DNA methy-lation profiling in cancers. Perspectives for the future direction in this dynamic and exciting field will also be given.展开更多
To determine the possible role of the epigenetic mechanisms in carcinogenesis of the hepatocellular carcinoma, we methylation-profiled the promoter CpG islands of twenty four genes both in HCC tumors and the neighbori...To determine the possible role of the epigenetic mechanisms in carcinogenesis of the hepatocellular carcinoma, we methylation-profiled the promoter CpG islands of twenty four genes both in HCC tumors and the neighboring non-cancerous tissues of twenty eight patients using the methylation-specific PCR (MSP) method in conjunction with the DNA sequencing. In comparison with the normal liver tissues from the healthy donors, it was found that while remained unmethylated the ABL, CAV, EPO, GATA3, LKB1, NEP, NFL, NIS and p27^(KIP1) genes, varying extents of the HCC specific hypermethylation were found associated with the ABO, AR, CSPG2, cyclin al, DBCCR1, GALR2, IRF7, MGMT, MT1A, MYOD1, OCT6, p57^(KIP2), p73, WT1 genes, and demethylation with the MAGEA1 gene, respectively. Judged by whether the hypermethylated occurred in HCC more frequently than in their neighboring normal tissues, the hypermethylation status of the AR, DBCCR1, IRF7, OCT6, and p73 genes was considered as the event specific to the late stage, while that the rest that lacked such a distinguished contrast, as the event specific to the early stage of HCC carcinogenesis. Among all the clinical pathological parameters tested for the association with, the hypermethylation of the cyclin al gene was more prevalent in the non-cirrhosis group (P=0.021) while the hypermethylated p16^(INK4a) gene was more common in the cirrhosis group (P=0.017). The concordant methylation behaviors of nineteen genes, including the four previously studied and their association with cirrhosis has been evaluated by the best subgroup selection method. The data presented in this report would enable us to shape our understanding of the mechanisms for the HCC specific loss of the epigenetic stability of the genome, as well as the strategy of developing the novel robust methylation based diagnostic and prognostic tools.展开更多
AIM:Through exploring the regulation of gene expression during hepatocarcinogenesis induced by aflatoxin B1 (AFB1),to find out the responsible genes for hepatocellular carcinoma (HCC) and to further understand the und...AIM:Through exploring the regulation of gene expression during hepatocarcinogenesis induced by aflatoxin B1 (AFB1),to find out the responsible genes for hepatocellular carcinoma (HCC) and to further understand the underlying molecular mechanism.METHODS:Tree shrews ( Tupaia belangeri chinensis)were treated with or without AFB1 for about 90 weeks. Liver biopsies were performed regularly during the animal experiment. Eight shares of total RNA were respectively isolated from 2 HCC tissues, 2 HCC-surrounding noncancerous liver tissues, 2 biopsied tissues at the early stage(30^th week) of the experiment from the same animals as above, 1 mixed sample of three liver tissues biopsied at the beginning (0^th week) of the experiment, and another i mixed sample of two liver tissues from the untreated control animals biopsied at the 90^th week of the experiment. The samples were then tested with the method of Atlas^TM cDNA microarray assay. The levels of gene expression in these tissues taken at different time points during hepatocarcinogenesis were compared.RESULTS:The profiles of differently expressed genes were quite different in different ways of comparison.At the same period of hepatocarcinogenesis, the genes in the same function group usually had the same tendency for up-or down-regulation. Among the checked 588 genes that were known to be related to human cancer, 89 genes (15.1%) were recognized as “important genes” because they showed frequent changes in different ways of comparison. The differentially expressed genes during hepatocarcinogenesis could be classified into four categories: genes up-regulated in HCC tissue, genes with similar expressing levels in both HCC and HCC-surrounding liver tissues which were higher than that in the tissues prior to the development of HCC,genes down-regulated in HCC tissue, and genes up-regulated prior to the development of HCC but down-regulated after the development of HCC.CONCLUSION: A considerable number of genes could change their expressing levels both in HCC and in HCC-surrounding non-cancerous liver tissues. A few modular genes were up-regulated only in HCC but not in surrounding liver tissues, while some apoptosis-related genes were down-regulated in HCC and up-regulated in surrounding liver tissues. To compare gene-expressing levels among the liver tissues taken at different time points during hepatocarcinogenesis may be helpful to locate the responsible gene (s) and understand the mechanism for AFB1 induced liver cancer.展开更多
AIM: To investigate the effect of cell adhesion molecule Pselectin monoclonal antibody (Mab) on metastasis and immune function of mice orthototopically implanted with human gastric cancer tissue.METHODS: SCID mice wer...AIM: To investigate the effect of cell adhesion molecule Pselectin monoclonal antibody (Mab) on metastasis and immune function of mice orthototopically implanted with human gastric cancer tissue.METHODS: SCID mice were implanted orthotopically with SGC-7901 human gastric carcinoma tissue. Starting from day 3 after operation, animals were given intravenously PBS or P-selectin Mab (100 μg/injection) (for both normal mice and tumor-implanted mice with tumors), twice weekly for 3weeks. Two animals in each group were sacrificed randomly at the 1st, 2nd, 4th week and 6th week. While T cell and B cell transformation indices were determined with the 3H TdR infiltration method, the NK cell activity was detected by the LDH release method.RESULTS: The metastatic rate in the P-selectin Mab treated group was lower than that in the PBS treated group (with tumors). The NK activity of normal mice increased over time.The immune functions (T, B cell function, NK activity) of the tumor group in the 6th week were significantly lower than those in the 4th week, but the change was attenuated by Pselectin Mab.CONCLUSION: P-selectin Mab could suppress the metastasis of gastric cancer with no adverse effect on host immune function.展开更多
Using a combination of hybridization of PAC to a cDNA library and RACE technique, we isolated a novel cDNA, designated as C17orf25 (Chromosome 17 open rea(ling frame 25, previously named it HC71A), from the deletion r...Using a combination of hybridization of PAC to a cDNA library and RACE technique, we isolated a novel cDNA, designated as C17orf25 (Chromosome 17 open rea(ling frame 25, previously named it HC71A), from the deletion region on chromosome 17p13.3. The cDNA encodes a protein of 313 amino acids with a calculated molecular mass of 34.8 kDa. C17orf25 is divided into 10 exons and 9 introns, spanning 23 kb of genomic DNA. Northern blot analysis showed that the mRNA expression of C17orf25 was decreased in hepatocellular carcinoma samples as compared to adjacent noncancerous liver tissues from the same patients. The transfection of C17or25 into the hepatocellular carcinoma cell SMMC7721 and overexpression could inhibit the cell growth. The above results indicate that C17orf25 is a novel human gene, and the cloning and preliminary characterization of C17orf25 is a prerequisite for further functional analysis of this novel gene in human hepatocellular carcinoma.展开更多
AIM: To construct a novel HBV antisense RNA delivery system targeting hapatocellular carcinoma and study its inhibitory effect in vitro and in vivo.METHODS: GE7,a 16-peptide specific to EGFR, and HA20,a homologue of N...AIM: To construct a novel HBV antisense RNA delivery system targeting hapatocellular carcinoma and study its inhibitory effect in vitro and in vivo.METHODS: GE7,a 16-peptide specific to EGFR, and HA20,a homologue of N-terminus of haemagglutinin of influenza viral envelope protein, were synthesized and conjugated with polylysin. The above conjugates were organized into the pEBAF-as-preS2, a hepatocarcinoma specific HBV antisense expression vector, to construct a novel HBV antisense RNA delivery system, named AFP-enhancing 4-element complex. Hepatocelluar carcinoma HepG2.2.15 cells was used to assay the in vitro inhibition of the complex on HBV. Expression of HBV antigen was assayed by ELISA. BALB/c nude mice bearing HepG2.2.15 cells were injected with AFP-enhancing 4-element complex. The expression of HBV antisense RNA was examined by RT-PCR and the size of tumor in nude mice were measured.RESULTS: The AFP-enhancing 4-element complex was constructed and DNA was completely trapped at the slot with no DNA migration when the ratio of polypeptide to plasmid was 1:1.The expression of HBsAg and HBeAg of HepG2.2.15 cells was greatly decreased after being transfected by AFP-enhancing 4-element complex. The inhibitory rates were 33.4 % and 58.5 % respectively. RTPCR showed HBV antisense RNA expressed specifically in liver tumor cells of tumor-bearing nude mice. After 4injections of AFP-enhancing 4-element complex containing 0.2 μg DNA, the diameter of the tumor was 0.995 cm±0.35,which was significantly smaller than that of the control groups (2.215 cm±0.25, P＜0.05).CONCLUSION: AFP-enhancing 4-element complex could deliver HBV antisense RNA targeting on hepatocarcinoma and inhibit both HBV and liver tumor cells in vitro and in vivo.展开更多
AIM:Bcl-2/adenovirus E1B 19 ku interacting protein 2-like (BNIPL-2) is a novel protein recently identified in our laboratory.BNIPL-2 is homologous to human BNIP-2,a potentially proapoptotic protein,and can interact wi...AIM:Bcl-2/adenovirus E1B 19 ku interacting protein 2-like (BNIPL-2) is a novel protein recently identified in our laboratory.BNIPL-2 is homologous to human BNIP-2,a potentially proapoptotic protein,and can interact with Bcl-2 and Cdc42GAP and promote apoptosis in BEL-7402 cells.Here we report the gene-expression profile regulated by BNIPL-2 in human hepatocarcinoma Hep3B cells and the analysis of its potential roles in cell apoptosis.METHODS:BNIPL-2 was overexpressed in Hip3B cells using tetracycline indecible or Tet-on system.Screened by Western blot,the cells with low background and high induction fold if BNIPL-2 were obtained.We performed Altlas human cDNA expression array hybridization on these cells and analyzed the data with Quantarray coftwate to identify BNIPL-2-regulated genes and their expression profile.RT-PCR was used to confirm the altered expression level of part of genes identified by the Atlas arrar hybridization.RESULTS:Fifteen of 588 genes spotted on the Atlas membrane showed altered expression levels in BNIPL-2-transfected Hep3B-Tet-on cells,in which 8 genes involved in cell apoptosis or growth inhibition were uregulated and 7 genes involved in cellular proliferation were down-regulated following overexpression of BNIPL-2.CONCLUSION:cDNA arrar is a powerful tool to explore gene expression profiles under inducible conditions.The data obtained usind the cDNA expression microarray techonlogy indicates that BNIPL-2 may play its roles in apoptosis through regulating the expression of gnees associated with cell apoptosis,growth inhibition and cell proliferation .展开更多
AIM: To find out key genes responsible for hepatocarc inogenesis and to further understand the underlying molecular mechanism through investigating the differential gene expression between human normal liver tissue an...AIM: To find out key genes responsible for hepatocarc inogenesis and to further understand the underlying molecular mechanism through investigating the differential gene expression between human normal liver tissue and hepatocellular carcinoma (HCC).METHODS: DNA microarray was prepared by spotting PCR products of 1 000 human genes including 445 novel genes, 540 known genes as well as 12 positive (housekeeping) and 3 negative controls (plant gene) onto treated glass slides. cDNA probes were prepared by labeling normal liver tissue mRNA and cancer liver tissue mRNA with Cy3-dUTP and Cy5-dUTP separately through reverse transcription. The arrays were hybridized against the cDNA probe and the fluorescent signals were scanned. The dataobtained from repeated experiments were analyzed. RESULTS: Among the 20 couple samples investigated (from cancerous liver tissue and normal liver tissue), 38 genes including 21 novel genes and 17 known genes exhibited different expressions. CONCLUSION: cDNA microarray technique is powerful to identify candidate target genes that may play important roles in human carcinogenesis. Further analysis of the obtained genes is helpful to understand the molecular changes in HCC progression and ultimately may lead to the identification of new targets for HCC diagnosis and intervention.展开更多
To understand the DNA-methylation mediated gene silencing mechanisms, we analyzed in cell culture of the promoter function of the MAGE-A1 gene, which is frequently demethylated and over-expressed in human hepatocellul...To understand the DNA-methylation mediated gene silencing mechanisms, we analyzed in cell culture of the promoter function of the MAGE-A1 gene, which is frequently demethylated and over-expressed in human hepatocellular carcinoma. We have established the correlation of the DNA methylation of the promoter CpG island with expression status of this gene in a panel of the established liver cancer cell lines. The crucial CpG dinucleotide(s) within the minimal promoter subjected to the control mediated by DNA methylation with profound biological functions was also delineated.Furthermore, a novel sequence-specific DNA-protein interaction at the -30 CpG dinucleotide upstream of the gene was found having a vital part to play in the DNA methylation mediated transcription silencing of the MAGE-A1 gene. Our results would not only provide new insights into the DNA methylation mediated mechanisms over transcription of the MAGE-A1 gene, but also pave the way for further defining the cross-talk among DNA methylation, histone modification and chromatin remodeling in detail.展开更多
The human RNA methyltransferase like 1 gene (RNMTL1) is one of thirteen newly discovered genes within a 116 Kb segment of the chromosome 17pl3.3 that suffers from a high frequent loss of heterozygosity in human hepato...The human RNA methyltransferase like 1 gene (RNMTL1) is one of thirteen newly discovered genes within a 116 Kb segment of the chromosome 17pl3.3 that suffers from a high frequent loss of heterozygosity in human hepatocellular carcinoma in China[1-5]. To understand the molecular mechanisms underlying transcription control of the RNMTL1 gene in human cancers, we decline using of the conventional approach where the cis-elements bound by the known transcription factors are primary targets, and carried out the systematic analyses to dissect the promoter structure and identify/characterize the key cis-elements that are responsible for its strong expression in cell. The molecular approaches applied included 1, the primer extension for mapping of the transcription starts; 2, the transient transfection/reporter assays on a large number of deletion and site-specific mutants of the promoter segment for defining the minimal promoter and the crucial elements within; and 3, the electrophoresis mobility shift assay with specific antibodies for reconfirming the nature of the transcription factors and their cognate cis-elements. We have shown that the interaction of an ATF/CREB element (-38 to -31) and its cognate transcription factors play a predominant role in the promoter activity of the RNMTL1 gene. The secondary DNA structures of the ATF/CREB element play a more vital role in the protein-DNA interaction. Finally, we reported a novel mechanism underlying the YY1 mediated transcription repression, namely, the ATF/CREB dependent transcription-repression by YY1 is executed in absence of its own sequence-specific binding.展开更多
CT120, a novel membrane-associated gene implicated in lung carcinogenesis, was previously identified from chromosome 17p13.3 locus, a hot mutation spot involved in human malignancies. In the present study, we further ...CT120, a novel membrane-associated gene implicated in lung carcinogenesis, was previously identified from chromosome 17p13.3 locus, a hot mutation spot involved in human malignancies. In the present study, we further determined that CT120 ectopic expression could promote cell proliferation activity of NIH3T3 cells using MTS assay, and monitored the downstream effects of CT120 in NIH3T3 cells with Atlas mouse cDNA expression arrays. Among 588 known genes, 133 genes were found to be upregulated or downregulated by CT120. Two major signaling pathways involved in cell proliferation, cell survival and anti-apoptosis were overexpressed and activated in response to CT120: One is the Raf/MEK/Erk signal cascades and the other is the PI3K/Akt signal cascades, suggesting that CT120 might contribute, at least in part, to the constitutively activation of Erk and Akt in human lung caner cells. In addition, some tumor metastasis associated genes cathepsin B, cathepsin D, cathepsin L, MMP-2/TIMP-2 were also upregulated by CT120, upon which CT120 might be involved in tumor invasiveness and metastasis. In addition, CT120 might play an important role in tumor progression through modulating the expression of some candidate 'Lung Tumor Progression' genes including B-Raf, Rab-2, BAX, BAG-1, YB-1, and Cdc42.展开更多
N-ros is one of the transforming genes in human hepatic cancer cells. It has been found that N-ras was overexpressed at the mENA and protein level in hepatoma cells. In order to explore the biological roles of N-ras i...N-ros is one of the transforming genes in human hepatic cancer cells. It has been found that N-ras was overexpressed at the mENA and protein level in hepatoma cells. In order to explore the biological roles of N-ras in human hepatic oaroinogenesis and the potential application in control of cancer cell growth, a pseudotype retrovirus containing antisense sequence of human N-ras was constructed and packaged. A recombinant rebrovirus vector containing antisense or sense sequences of N-ras oDNA was constructed by pZIP-NeoSV(X)1. The pseudotype virus was packaged and rescued by transfeotion and infection in PA317 and ψ2 helper cells. It has been demonstrated that the pseudotype retrovirus containing antisense N-ras sequence did inhibit the growth of human PLC/PEF/5 hepatoma cells accompanied with inhibition of p21 expression, while the retrovirus containing sense sequence had none. The pseudotype virus had no effect on human diploid fibroblasts.展开更多
We have identified IC53-2, a human homologue of the rat C53 gene from a human placenta cDNA library (GeneBank Accession No. AF217982). IC53-2 can bind to the CDK5 activator p35 by in vitro association assay. IC53-2 is...We have identified IC53-2, a human homologue of the rat C53 gene from a human placenta cDNA library (GeneBank Accession No. AF217982). IC53-2 can bind to the CDK5 activator p35 by in vitro association assay. IC53-2 is mapped to human chromosome 17q21.31. The IC53-2 transcript is highly expressed in kidney, liver, skeletal muscle and placenta. It is abundantly expressed in SMMC-7721, C-33A, 3AO, A431and MCF-7 cancer cell lines by RT-PCR assay. Stable transfection of IC53-2 cDNA into the hepatocellularcarcinoma SMMC-7721 cell remarkably stimulates its growth in vitro. The above results indicate thatIC53-2 is a novel human gene, which may be involved in the regulation of cell proliferation.展开更多
OBJECTIVE To examine the possible risk factors for stomach cancer among men and women in Shanghai, China. METHODS In urban Shanghai, in-person interviews were completed for 311 cases newly diagnosed with stomach cance...OBJECTIVE To examine the possible risk factors for stomach cancer among men and women in Shanghai, China. METHODS In urban Shanghai, in-person interviews were completed for 311 cases newly diagnosed with stomach cancer of ages 30-74. Data were collected from April 1999 to October 1999 and compared to 1579 population-based controls (controls in three cancer studies used together). Information on demographic variables, smoking, diet consumption and others was collected from all subjects. Unconditional logistic regression was used to adjust the possible factors. RESULTS Stomach cancer risk in men rose with smoking, eating hot and fried foods, chronic gastritis and a family history of gastric cancer among men; the risk among women was associated with the consumption of preserved,pickled and fried foods,heavy drinking, chronic gastritis, a history of ulcer disease and a family history of gastric cancer. A doseresponse relationship was found (trends test, P<0.01) among men smokers. In contrast, the consumption of fresh vegetables and fruits, beans (especially soybeans) and soy products, plant oil, and eggs and egg products, was inversely associated with stomach cancer risk. After adjustment of the potential confounding variables, these associations remained significant. CONCLUSIONS The present findings provide further evidence that smoking, eating salted foods (especially salted vegetables), oil-fried foods(including fried cereal, eggs, and peanuts), chronic gastritis, a family history of gastric cancer and so on increase the risk of stomach carcinoma in Shanghai. Fresh vegetables and fruits, beans and soybean products (even after adjusted for use of fresh vegetables and fruits), plant oil, and so on may have protective effects.展开更多
OBJECTIVE To investigate the relationship between BMI, WHR and biliary tract cancers (CBT).METHODS A population-based case-control study was conducted in urban Shanghai from June 1, 1997 to May 31,2001 involving inter...OBJECTIVE To investigate the relationship between BMI, WHR and biliary tract cancers (CBT).METHODS A population-based case-control study was conducted in urban Shanghai from June 1, 1997 to May 31,2001 involving interviews with 627 new cases of biliary tract cancers aged 35 to 74 years and 959 frequency-matched population controls by gender and age in five-year groups. All subjects were interviewed in person by trained interviewers using a structured questionnaire. An unconditional logistic regression was performed to calculate adjusted odds ratios (ORs) and 95% confidence intervals (CIs).RESULTS Obesity was associated with an increased risk of gallbladder cancer across adulthood at ages 20-29 and 30-39 in females. Compared with subjects in the lowest quartile of WHR, ORs for the highest quartile and P for trend for cancers of gallbladder and extrahepatic bile duct both reached significant levels among males and females.CONCLUSION Our observations in urban Shanghai suggested that obesity in early adult life may contribute to the risk of gallbladder cancer, and increased WHR may substantially elevated risk of cancers of the gallbladder and extrahepatic bile duct.展开更多
基金Supported by the National High Technology Research and Development Program of China (863 program),No.2002AA2Z3353 and the Science Foundation of Shanghai Municipal Government,No.02DJ14056
文摘AIM: To establish the DNA methylation patterns of the promoter CpG islands of 14 'drug-resistance' genes in hepatocellular carcinoma (HCC).METHODS: The methylation specific polymerase chain reaction in conjunction with sequencing verification was used to establish the methylation patterns of the 14 genes in the liver tissues of four healthy liver donors, as well as tumor and the paired non-cancerous tissues of 30 HCC patients.RESULTS: While 11 genes (ATP-binding cassette, sub-family G (WHITE), member 2(ABCG2), activating transcription factor (ATF2), beta-2-microglobulin (B2M), deoxycytidine kinase(DCK, occludin (OCLN, v-raf-1 murine leukemia viral oncogene homolog (RAF/), ralA binding protein 1 (RALBP1),splicing factor (45 kD) (SPF45), S-phase kinase-associated protein 2 (p45) (SKP2), tumor protein p53 (Li-Fraumeni syndrome) (TP53) and topoisomerase (DNA) II beta (TOP2B) maintained the unmethylated patterns, three genes displayed to various extents the hypermethylation state in tumor tissues in comparison with the normal counterparts. The catalase (CAT) was hypermethylated in tumor and the neighboring non-cancerous tissue of one case (3.3%). Both glutathione S-transferase pi (GSTp/) (80%, 24/30 in tumor and 56.7%,17/30 in the paired non-cancerous tissues) and cystic fibrosis transmembrane conductance regulator, ATP-binding cassette (sub-family C, member 7) (CF-FR) (77%, 23/30 in tumor and 50%, 15/30 in the paired non-cancerous tissues) genes were prevalently hypermethylated in HCC as well as their neighboring non-cancerous tissues. No significant difference in the hypermethylation occurrence was observed between the HCC and its neighboring non-cancerous tissues.CONCLUSION: Hypermethylation of promoter CpG islands of both CF-FR and GSTpigenes occurs prevalently in HCC,which may correlate with the low expression of these two genes at the mRNA level and has the profound etiological and clinical implications. It is likely to be specific to the early phase of HCC carcinogenesis.
基金supported by the GIBH funds provided by the Chinese Academy of Sciences,the City of Guangzhou and Guangdong Province
文摘Lung cancer is one of the leading causes of death with one of the lowest survival rates. However, a subset of lung cancer patients who are of Asian origin and carry somatic mutations in epidermal growth factor receptor or EGFR have responded remarkable well to two tyrosine kinase inhibitors, gefitinib and erlotinib. While EGFR mutation profiles have been reported from Japan, South Korea, and Taiwan, there is no such report from mainland of China where the largest pool of patients reside. In this report, we identified ten somatic mutations from a total of 41 lung cancer patients in China. Among them, seven mutations were found in 17 adenocarcinomas. In contrast to previous reports, eight of these mutations are deletions in exon 19 and two of these deletions are homozygous. These results suggest that a large portion of Chinese adenocarcinoma patients could benefit from gefitinib or erlotinib. This unique mutation profile provides a rationale to develop the next generation of EGFR inhibitors more suitable for the Chinese population.
基金The research performed in this lab is supported by Shanghai Science Foundation(NO.04DZ14006)National Natural Science Foundation(NO.30450001)+1 种基金Major State Basic Research Development program of China(NO.2004CB51 8804)the National High Technology Re-search and Development Program of China(NO.2002AA2Z3352).
文摘DNA methylation is the most intensively studied epigenetic phenomenon, disturbances of which result in changes ingene transcription, thus exerting drastic imparts onto biological behaviors of cancer. Both the global demethylation andthe local hypermethylation have been widely reported in all types of tumors, providing both challenges and opportunitiesfor a better understanding and eventually controlling of the malignance. However, we are still in the very early stage ofinformation accumulation concerning the tumor associated changes in DNA methylation pattern. A number of excellentrecent reviews have covered this issue in depth. Therefore, this review will summarize our recent data on DNA methy-lation profiling in cancers. Perspectives for the future direction in this dynamic and exciting field will also be given.
基金supported by the National High Technology Research and Development Program of China(863 Program)(2001AA217011,2002AA2Z3352)the Major State Basic Research Development Program of China (973 Program)(G1998051004)the Science Foundation of Shanghai Municipal Government(02DJ14056)to JingDe ZHU.
文摘To determine the possible role of the epigenetic mechanisms in carcinogenesis of the hepatocellular carcinoma, we methylation-profiled the promoter CpG islands of twenty four genes both in HCC tumors and the neighboring non-cancerous tissues of twenty eight patients using the methylation-specific PCR (MSP) method in conjunction with the DNA sequencing. In comparison with the normal liver tissues from the healthy donors, it was found that while remained unmethylated the ABL, CAV, EPO, GATA3, LKB1, NEP, NFL, NIS and p27^(KIP1) genes, varying extents of the HCC specific hypermethylation were found associated with the ABO, AR, CSPG2, cyclin al, DBCCR1, GALR2, IRF7, MGMT, MT1A, MYOD1, OCT6, p57^(KIP2), p73, WT1 genes, and demethylation with the MAGEA1 gene, respectively. Judged by whether the hypermethylated occurred in HCC more frequently than in their neighboring normal tissues, the hypermethylation status of the AR, DBCCR1, IRF7, OCT6, and p73 genes was considered as the event specific to the late stage, while that the rest that lacked such a distinguished contrast, as the event specific to the early stage of HCC carcinogenesis. Among all the clinical pathological parameters tested for the association with, the hypermethylation of the cyclin al gene was more prevalent in the non-cirrhosis group (P=0.021) while the hypermethylated p16^(INK4a) gene was more common in the cirrhosis group (P=0.017). The concordant methylation behaviors of nineteen genes, including the four previously studied and their association with cirrhosis has been evaluated by the best subgroup selection method. The data presented in this report would enable us to shape our understanding of the mechanisms for the HCC specific loss of the epigenetic stability of the genome, as well as the strategy of developing the novel robust methylation based diagnostic and prognostic tools.
基金Supported by National Natural Science Foundation of China,NO.39860072 and NO.39869001Natural Science Foundation of Guangxi Zhuang Autonomous Region,NO.9817137
文摘AIM:Through exploring the regulation of gene expression during hepatocarcinogenesis induced by aflatoxin B1 (AFB1),to find out the responsible genes for hepatocellular carcinoma (HCC) and to further understand the underlying molecular mechanism.METHODS:Tree shrews ( Tupaia belangeri chinensis)were treated with or without AFB1 for about 90 weeks. Liver biopsies were performed regularly during the animal experiment. Eight shares of total RNA were respectively isolated from 2 HCC tissues, 2 HCC-surrounding noncancerous liver tissues, 2 biopsied tissues at the early stage(30^th week) of the experiment from the same animals as above, 1 mixed sample of three liver tissues biopsied at the beginning (0^th week) of the experiment, and another i mixed sample of two liver tissues from the untreated control animals biopsied at the 90^th week of the experiment. The samples were then tested with the method of Atlas^TM cDNA microarray assay. The levels of gene expression in these tissues taken at different time points during hepatocarcinogenesis were compared.RESULTS:The profiles of differently expressed genes were quite different in different ways of comparison.At the same period of hepatocarcinogenesis, the genes in the same function group usually had the same tendency for up-or down-regulation. Among the checked 588 genes that were known to be related to human cancer, 89 genes (15.1%) were recognized as “important genes” because they showed frequent changes in different ways of comparison. The differentially expressed genes during hepatocarcinogenesis could be classified into four categories: genes up-regulated in HCC tissue, genes with similar expressing levels in both HCC and HCC-surrounding liver tissues which were higher than that in the tissues prior to the development of HCC,genes down-regulated in HCC tissue, and genes up-regulated prior to the development of HCC but down-regulated after the development of HCC.CONCLUSION: A considerable number of genes could change their expressing levels both in HCC and in HCC-surrounding non-cancerous liver tissues. A few modular genes were up-regulated only in HCC but not in surrounding liver tissues, while some apoptosis-related genes were down-regulated in HCC and up-regulated in surrounding liver tissues. To compare gene-expressing levels among the liver tissues taken at different time points during hepatocarcinogenesis may be helpful to locate the responsible gene (s) and understand the mechanism for AFB1 induced liver cancer.
基金the Youth Science Foundation of Shanghai Public Administration No.131984Y15
文摘AIM: To investigate the effect of cell adhesion molecule Pselectin monoclonal antibody (Mab) on metastasis and immune function of mice orthototopically implanted with human gastric cancer tissue.METHODS: SCID mice were implanted orthotopically with SGC-7901 human gastric carcinoma tissue. Starting from day 3 after operation, animals were given intravenously PBS or P-selectin Mab (100 μg/injection) (for both normal mice and tumor-implanted mice with tumors), twice weekly for 3weeks. Two animals in each group were sacrificed randomly at the 1st, 2nd, 4th week and 6th week. While T cell and B cell transformation indices were determined with the 3H TdR infiltration method, the NK cell activity was detected by the LDH release method.RESULTS: The metastatic rate in the P-selectin Mab treated group was lower than that in the PBS treated group (with tumors). The NK activity of normal mice increased over time.The immune functions (T, B cell function, NK activity) of the tumor group in the 6th week were significantly lower than those in the 4th week, but the change was attenuated by Pselectin Mab.CONCLUSION: P-selectin Mab could suppress the metastasis of gastric cancer with no adverse effect on host immune function.
基金the National 863High Technology Research and Development Pro-gram of China (Zl9-02--0l-0l) to Wan DF and theProject of Ch
文摘Using a combination of hybridization of PAC to a cDNA library and RACE technique, we isolated a novel cDNA, designated as C17orf25 (Chromosome 17 open rea(ling frame 25, previously named it HC71A), from the deletion region on chromosome 17p13.3. The cDNA encodes a protein of 313 amino acids with a calculated molecular mass of 34.8 kDa. C17orf25 is divided into 10 exons and 9 introns, spanning 23 kb of genomic DNA. Northern blot analysis showed that the mRNA expression of C17orf25 was decreased in hepatocellular carcinoma samples as compared to adjacent noncancerous liver tissues from the same patients. The transfection of C17or25 into the hepatocellular carcinoma cell SMMC7721 and overexpression could inhibit the cell growth. The above results indicate that C17orf25 is a novel human gene, and the cloning and preliminary characterization of C17orf25 is a prerequisite for further functional analysis of this novel gene in human hepatocellular carcinoma.
基金the National Natural Science Foundation Community,No.39970333
文摘AIM: To construct a novel HBV antisense RNA delivery system targeting hapatocellular carcinoma and study its inhibitory effect in vitro and in vivo.METHODS: GE7,a 16-peptide specific to EGFR, and HA20,a homologue of N-terminus of haemagglutinin of influenza viral envelope protein, were synthesized and conjugated with polylysin. The above conjugates were organized into the pEBAF-as-preS2, a hepatocarcinoma specific HBV antisense expression vector, to construct a novel HBV antisense RNA delivery system, named AFP-enhancing 4-element complex. Hepatocelluar carcinoma HepG2.2.15 cells was used to assay the in vitro inhibition of the complex on HBV. Expression of HBV antigen was assayed by ELISA. BALB/c nude mice bearing HepG2.2.15 cells were injected with AFP-enhancing 4-element complex. The expression of HBV antisense RNA was examined by RT-PCR and the size of tumor in nude mice were measured.RESULTS: The AFP-enhancing 4-element complex was constructed and DNA was completely trapped at the slot with no DNA migration when the ratio of polypeptide to plasmid was 1:1.The expression of HBsAg and HBeAg of HepG2.2.15 cells was greatly decreased after being transfected by AFP-enhancing 4-element complex. The inhibitory rates were 33.4 % and 58.5 % respectively. RTPCR showed HBV antisense RNA expressed specifically in liver tumor cells of tumor-bearing nude mice. After 4injections of AFP-enhancing 4-element complex containing 0.2 μg DNA, the diameter of the tumor was 0.995 cm±0.35,which was significantly smaller than that of the control groups (2.215 cm±0.25, P＜0.05).CONCLUSION: AFP-enhancing 4-element complex could deliver HBV antisense RNA targeting on hepatocarcinoma and inhibit both HBV and liver tumor cells in vitro and in vivo.
基金Supported by a grant from the National Key Basic Research Program(973)of China,No.2002CB5131130 and grants from the National 863High Technology Research and Development Program of China,No.2001AA221141,2002BA71 IA02,and 2001AA227121
文摘AIM:Bcl-2/adenovirus E1B 19 ku interacting protein 2-like (BNIPL-2) is a novel protein recently identified in our laboratory.BNIPL-2 is homologous to human BNIP-2,a potentially proapoptotic protein,and can interact with Bcl-2 and Cdc42GAP and promote apoptosis in BEL-7402 cells.Here we report the gene-expression profile regulated by BNIPL-2 in human hepatocarcinoma Hep3B cells and the analysis of its potential roles in cell apoptosis.METHODS:BNIPL-2 was overexpressed in Hip3B cells using tetracycline indecible or Tet-on system.Screened by Western blot,the cells with low background and high induction fold if BNIPL-2 were obtained.We performed Altlas human cDNA expression array hybridization on these cells and analyzed the data with Quantarray coftwate to identify BNIPL-2-regulated genes and their expression profile.RT-PCR was used to confirm the altered expression level of part of genes identified by the Atlas arrar hybridization.RESULTS:Fifteen of 588 genes spotted on the Atlas membrane showed altered expression levels in BNIPL-2-transfected Hep3B-Tet-on cells,in which 8 genes involved in cell apoptosis or growth inhibition were uregulated and 7 genes involved in cellular proliferation were down-regulated following overexpression of BNIPL-2.CONCLUSION:cDNA arrar is a powerful tool to explore gene expression profiles under inducible conditions.The data obtained usind the cDNA expression microarray techonlogy indicates that BNIPL-2 may play its roles in apoptosis through regulating the expression of gnees associated with cell apoptosis,growth inhibition and cell proliferation .
基金Supported by the Special Fund of Chinese Academy of Sciences, No. KSCX1-06
文摘AIM: To find out key genes responsible for hepatocarc inogenesis and to further understand the underlying molecular mechanism through investigating the differential gene expression between human normal liver tissue and hepatocellular carcinoma (HCC).METHODS: DNA microarray was prepared by spotting PCR products of 1 000 human genes including 445 novel genes, 540 known genes as well as 12 positive (housekeeping) and 3 negative controls (plant gene) onto treated glass slides. cDNA probes were prepared by labeling normal liver tissue mRNA and cancer liver tissue mRNA with Cy3-dUTP and Cy5-dUTP separately through reverse transcription. The arrays were hybridized against the cDNA probe and the fluorescent signals were scanned. The dataobtained from repeated experiments were analyzed. RESULTS: Among the 20 couple samples investigated (from cancerous liver tissue and normal liver tissue), 38 genes including 21 novel genes and 17 known genes exhibited different expressions. CONCLUSION: cDNA microarray technique is powerful to identify candidate target genes that may play important roles in human carcinogenesis. Further analysis of the obtained genes is helpful to understand the molecular changes in HCC progression and ultimately may lead to the identification of new targets for HCC diagnosis and intervention.
基金supported by the National High Technology Research and Development Program of China(863 Program)(2002AA2Z3352)the Major State Basic Research Development Program of China(973 Program)(G1998051004)the Science Foundation of Shanghai Municipal Government(02DJ14056)to Jing De ZHU.
文摘To understand the DNA-methylation mediated gene silencing mechanisms, we analyzed in cell culture of the promoter function of the MAGE-A1 gene, which is frequently demethylated and over-expressed in human hepatocellular carcinoma. We have established the correlation of the DNA methylation of the promoter CpG island with expression status of this gene in a panel of the established liver cancer cell lines. The crucial CpG dinucleotide(s) within the minimal promoter subjected to the control mediated by DNA methylation with profound biological functions was also delineated.Furthermore, a novel sequence-specific DNA-protein interaction at the -30 CpG dinucleotide upstream of the gene was found having a vital part to play in the DNA methylation mediated transcription silencing of the MAGE-A1 gene. Our results would not only provide new insights into the DNA methylation mediated mechanisms over transcription of the MAGE-A1 gene, but also pave the way for further defining the cross-talk among DNA methylation, histone modification and chromatin remodeling in detail.
基金This work is supported by the 973 projects of China (G1998051004) to Jingde Zhu and (G199805l200) to Dafang Wan, respectively.Thanks are due to Hongyu Zhang and other mem-bers in Jingde Zhu's lab for assistance and helps onnumerous occasions.
文摘The human RNA methyltransferase like 1 gene (RNMTL1) is one of thirteen newly discovered genes within a 116 Kb segment of the chromosome 17pl3.3 that suffers from a high frequent loss of heterozygosity in human hepatocellular carcinoma in China[1-5]. To understand the molecular mechanisms underlying transcription control of the RNMTL1 gene in human cancers, we decline using of the conventional approach where the cis-elements bound by the known transcription factors are primary targets, and carried out the systematic analyses to dissect the promoter structure and identify/characterize the key cis-elements that are responsible for its strong expression in cell. The molecular approaches applied included 1, the primer extension for mapping of the transcription starts; 2, the transient transfection/reporter assays on a large number of deletion and site-specific mutants of the promoter segment for defining the minimal promoter and the crucial elements within; and 3, the electrophoresis mobility shift assay with specific antibodies for reconfirming the nature of the transcription factors and their cognate cis-elements. We have shown that the interaction of an ATF/CREB element (-38 to -31) and its cognate transcription factors play a predominant role in the promoter activity of the RNMTL1 gene. The secondary DNA structures of the ATF/CREB element play a more vital role in the protein-DNA interaction. Finally, we reported a novel mechanism underlying the YY1 mediated transcription repression, namely, the ATF/CREB dependent transcription-repression by YY1 is executed in absence of its own sequence-specific binding.
文摘CT120, a novel membrane-associated gene implicated in lung carcinogenesis, was previously identified from chromosome 17p13.3 locus, a hot mutation spot involved in human malignancies. In the present study, we further determined that CT120 ectopic expression could promote cell proliferation activity of NIH3T3 cells using MTS assay, and monitored the downstream effects of CT120 in NIH3T3 cells with Atlas mouse cDNA expression arrays. Among 588 known genes, 133 genes were found to be upregulated or downregulated by CT120. Two major signaling pathways involved in cell proliferation, cell survival and anti-apoptosis were overexpressed and activated in response to CT120: One is the Raf/MEK/Erk signal cascades and the other is the PI3K/Akt signal cascades, suggesting that CT120 might contribute, at least in part, to the constitutively activation of Erk and Akt in human lung caner cells. In addition, some tumor metastasis associated genes cathepsin B, cathepsin D, cathepsin L, MMP-2/TIMP-2 were also upregulated by CT120, upon which CT120 might be involved in tumor invasiveness and metastasis. In addition, CT120 might play an important role in tumor progression through modulating the expression of some candidate 'Lung Tumor Progression' genes including B-Raf, Rab-2, BAX, BAG-1, YB-1, and Cdc42.
文摘N-ros is one of the transforming genes in human hepatic cancer cells. It has been found that N-ras was overexpressed at the mENA and protein level in hepatoma cells. In order to explore the biological roles of N-ras in human hepatic oaroinogenesis and the potential application in control of cancer cell growth, a pseudotype retrovirus containing antisense sequence of human N-ras was constructed and packaged. A recombinant rebrovirus vector containing antisense or sense sequences of N-ras oDNA was constructed by pZIP-NeoSV(X)1. The pseudotype virus was packaged and rescued by transfeotion and infection in PA317 and ψ2 helper cells. It has been demonstrated that the pseudotype retrovirus containing antisense N-ras sequence did inhibit the growth of human PLC/PEF/5 hepatoma cells accompanied with inhibition of p21 expression, while the retrovirus containing sense sequence had none. The pseudotype virus had no effect on human diploid fibroblasts.
基金supported by grants(G1998051004,and 2002CB513100)from the China State Key Basic Research Program(to DF WAN,and to WX QIN)grants(2001AA221141,2002BA711A02)from the National 863 High Technology Research and Development Program of China(to WX QIN).
文摘We have identified IC53-2, a human homologue of the rat C53 gene from a human placenta cDNA library (GeneBank Accession No. AF217982). IC53-2 can bind to the CDK5 activator p35 by in vitro association assay. IC53-2 is mapped to human chromosome 17q21.31. The IC53-2 transcript is highly expressed in kidney, liver, skeletal muscle and placenta. It is abundantly expressed in SMMC-7721, C-33A, 3AO, A431and MCF-7 cancer cell lines by RT-PCR assay. Stable transfection of IC53-2 cDNA into the hepatocellularcarcinoma SMMC-7721 cell remarkably stimulates its growth in vitro. The above results indicate thatIC53-2 is a novel human gene, which may be involved in the regulation of cell proliferation.
文摘OBJECTIVE To examine the possible risk factors for stomach cancer among men and women in Shanghai, China. METHODS In urban Shanghai, in-person interviews were completed for 311 cases newly diagnosed with stomach cancer of ages 30-74. Data were collected from April 1999 to October 1999 and compared to 1579 population-based controls (controls in three cancer studies used together). Information on demographic variables, smoking, diet consumption and others was collected from all subjects. Unconditional logistic regression was used to adjust the possible factors. RESULTS Stomach cancer risk in men rose with smoking, eating hot and fried foods, chronic gastritis and a family history of gastric cancer among men; the risk among women was associated with the consumption of preserved,pickled and fried foods,heavy drinking, chronic gastritis, a history of ulcer disease and a family history of gastric cancer. A doseresponse relationship was found (trends test, P<0.01) among men smokers. In contrast, the consumption of fresh vegetables and fruits, beans (especially soybeans) and soy products, plant oil, and eggs and egg products, was inversely associated with stomach cancer risk. After adjustment of the potential confounding variables, these associations remained significant. CONCLUSIONS The present findings provide further evidence that smoking, eating salted foods (especially salted vegetables), oil-fried foods(including fried cereal, eggs, and peanuts), chronic gastritis, a family history of gastric cancer and so on increase the risk of stomach carcinoma in Shanghai. Fresh vegetables and fruits, beans and soybean products (even after adjusted for use of fresh vegetables and fruits), plant oil, and so on may have protective effects.
文摘OBJECTIVE To investigate the relationship between BMI, WHR and biliary tract cancers (CBT).METHODS A population-based case-control study was conducted in urban Shanghai from June 1, 1997 to May 31,2001 involving interviews with 627 new cases of biliary tract cancers aged 35 to 74 years and 959 frequency-matched population controls by gender and age in five-year groups. All subjects were interviewed in person by trained interviewers using a structured questionnaire. An unconditional logistic regression was performed to calculate adjusted odds ratios (ORs) and 95% confidence intervals (CIs).RESULTS Obesity was associated with an increased risk of gallbladder cancer across adulthood at ages 20-29 and 30-39 in females. Compared with subjects in the lowest quartile of WHR, ORs for the highest quartile and P for trend for cancers of gallbladder and extrahepatic bile duct both reached significant levels among males and females.CONCLUSION Our observations in urban Shanghai suggested that obesity in early adult life may contribute to the risk of gallbladder cancer, and increased WHR may substantially elevated risk of cancers of the gallbladder and extrahepatic bile duct.
