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Viral kinetics of Enterovirus 71 in human habdomyosarcoma cells

World Journal of Gastroenterology 2011年第36期17卷 4135-4142页

作者： Jing Lu Li-Na Yi hsiang-fu kung Ming-Liang He Ya-Qing He Hong Zan Stanley Ho Center for Emerging Infectious Diseases and Li Ka Shing Institute of Health SciencesFaculty of MedicineThe Chinese University of Hong KongHong KongChina Shenzhen Center for the Disease Control and Prevention Shenzhen 518055Guangdong ProvinceChina Institute for Immunology University of California at IrvineCA 92697United States

AIM:To characterise the viral kinetics of enterovirus 71 (EV71).METHODS:In this study,human rhabdomyosarcoma (RD) cells were infected with EV71 at different multiplicity of infection (MOI).After infection,the cytopathic effect (CPE) was monitored and recorded using a phase contrast microscope associated with a CCD camera at different time points post viral infection (0,6,12,24 h post infection).Cell growth and viability were measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay in both EV71 infected and mock infected cells at each time point.EV71 replication kinet-ics in RD cells was determined by measuring the total intracellular viral RNA with real-time reverse-transcription polymerase chain reaction (qRT-PCR).Also,the intracellular and extracellular virion RNA was isolated and quantified at different time points to analyze the viral package and secretion.The expression of viral protein was determined by analyze the levels of viral structure protein VP1 with Western blotting.RESULTS:EV71 infection induced a significant CPE as early as 6 h post infection (p.i.) in both RD cells infected with high ratio of virus (MOI 10) and low ratio of virus (MOI 1).In EV71 infected cells,the cell growth was inhibited and the number of viable cells was rapidly decreased in the later phase of infection.EV71 virions were uncoated immediately after entry.The intracellular viral RNA began to increase at as early as 3 h p.***.and the exponential increase was found between 3 h to 6 h p.***.in both infected groups.For viral structure protein synthesis,results from western-blot showed that intracellular viral protein VP1 could not be detected until 6 h p.***.in the cells infected at either MOI 1 or MOI 10;and reached the peak at 9 h p.***.in the cells infected with EV71 at both MOI 1 and MOI 10.Simultaneously,the viral package and secretion were also actively processed as the virus underwent rapid replication.The viral package kinetics was comparable for both MOI 1 and MOI 10 infected groups.It was observed that

关键词：细胞病变效应肠病毒动力学人类逆转录聚合酶链反应病毒结构蛋白免疫印迹分析病毒感染

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Cancer Immunotherapy Targeting the Telomerase Reverse Transcriptase

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Cellular & Molecular Immunology 2006年第1期3卷 1-9页

作者： Longfei Huo Janice WS Tang Junjian Huang Peitang Huang Cuifen Huang hsiang-fu kung Marie C. Lin Open Laboratory of Chemical Biology of the Institute of Molecular Technology. Department of Chemistry The University of Hong Kong Pokfulam Hong Kong China Li Ka Shing Institute of Health Sciences and Centre for EmergingInfectious Diseases The Chinese University of Hong Kong Shatin HongKong China Beijing Institute of Biotechnology Beijing China Open Laboratory of ChemicalBiology of the Institute of Molecular Technology for Drug Discovery Department of Chemistry The University of Hong Kong Hong KongChina

The human telomerase reverse transcriptase （hTERT） is expressed in more than 85% of tumor cells but is usually not found in normal cells, which makes hTERT as an ideal tumor-associate antigen （TAA） to develop potential vaccine specifically destroying cancers without impairing normal tissues in human cancer immunotherapy. Here are reviewed the fundamental advances of studies on immunogenicity of hTERT or its peptides and the early clinical trials using the hTERT vaccine approach in the last decades.

关键词： cancer telomerase reverse transcriptase immunotherapy lymphocyte

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High Expression of p300 in Human Breast Cancer Correlates with Tumor Recurrence and Predicts Adverse Prognosis

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Chinese Journal of Cancer Research 2011年第3期23卷 201-207页

作者： Xiang-sheng Xiao Mu-yan Cai Jie-wei Chen Xin-yuan Guan hsiang-fu kung Yi-xin Zeng Dan Xie State Key Laboratory of Oncology in South China Guangzhou 510060China Department of Breast Oncology Guangzhou 510060China Department of Pathology Sun Yat-sen University Cancer CenterGuangzhou 510060China The Chinese University of Hong Kong Hong KongChina

Objective：Transcriptional coactivator p300 has been shown to play a variety of roles in the transcription process and mutation of p300 has been found in certain types of human cancers.However,the expression dynamics of p300 in breast cancer （BC） and its effect on BC patients＇ prognosis are poorly understood.Methods：In the present study,the methods of tissue microarray and immunohistochemistry （IHC） were used to investigate the protein expression of p300 in BCs.Receiver operating characteristic （ROC） curve analysis,Spearman＇s rank correlation,Kaplan-Meier plots and Cox proportional hazards regression model were utilized to analyze the data.Results：Based on the ROC curve analysis,the cutoff value for p300 high expression was defined when the H score for p300 was more than 105.High expression of p300 could be observed in 105/193 （54.4%） of BCs,in 6/25 （24.0%） of non-malignant breast tissues,respectively （P=0.004）.further correlation analysis showed that high expression of p300 was positively correlated with higher histological grade,advanced clinical stage and tumor recurrence （P0.05）.In univariate survival analysis,a significant association between high expression of p300 and shortened patients＇ survival and poor progression-free survival was found （P0.05）.Importantly,p300 expression was evaluated as an independent prognostic factor in multivariate analysis （P0.05）.Conclusion：Our findings provide a basis for the concept that high expression of p300 in BC may be important in the acquisition of a recurrence phenotype,suggesting that p300 high expression,as examined by IHC,is an independent biomarker for poor prognosis of patients with BC.

关键词： Breast cancer p300 Tumor recurrence Prognosis

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A new polymorphism in the GRP78 is not associated with HBV invasion

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World Journal of Gastroenterology 2009年第39期15卷 4958-4961页

作者： Xiao Zhu Yi Wang Tao Tao Dong-Pei Li Fei-Fei Lan Wei Zhu Dan Xie hsiang-fu kung Institute of Oncology Affiliated Tumor Hospital of Guangzhou Medical College Guangzhou 510095 Guangdong Province China State Key Laboratory of Oncology in South China CancerCenter Sun Yat-sen University Guangzhou 510060 GuangdongProvince China Department of Digestive Diseases Zibo Central Hospital Zibo 255000 Shandong Province China Zhongshan School of Medicine Sun Yat-sen University Guangzhou 510080 Guangdong Province China Laboratory of Integrated Biosciences School of Life Sciences Sun Yat-sen University Guangzhou 510275 Guangdong Province China

AIM:To examine the association between -86 bp(T>A) in the glucose-regulated protein 78 gene(GRP78) and hepatitis B virus(HBV) invasion.METHODS:DNA was genotyped for the single-nucleotide polymorphism by polymerase chain reaction followed by sequencing in a sample of 382 unrelated HBV carriers and a total of 350 sex-and age-matched healthy controls.Serological markers for HBV infection were determined by enzyme-linked immunosorbent as-say kits or clinical chemistry testing.RESULTS:The distributions of allelotype and genotype in cases were not significantly different from those in controls.In addition,our fi ndings suggested that neither alanine aminotransferase/hepatitis B e antigen nor HBV-DNA were associated with the allele/genotype variation in HBV infected individuals.CONCLUSION:-86 bp T>A polymorphism in GRP78 gene is not related to the clinical risk and acute exacerbation of HBV invasion.

关键词：基因多态性病毒入侵调节蛋白 HBV 乙型肝炎病毒酶联免疫检测试剂盒病毒DNA 基因分型

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Transactivation of the TIEG1 confers growth inhibition of transforming growth factor-β-susceptible hepatocellular carcinoma cells

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World Journal of Gastroenterology 2012年第17期18卷 2035-2042页

作者： Lei Jiang Yiu-Kay Lai Jin-Fang Zhang Chu-Yan Chan Gang Lu Marie CM Lin Ming-Liang He Ji-Cheng Li hsiang-fu kung Laboratory of Internal Medicine The First AffiliatedHospital of Wenzhou Medical College Wenzhou 325000 Zhejiang Province China Department of Life Science National Tsing HuaUniversity Hsinchu 30013 Taiwan Stanley Ho Centre for Emerging Infectious Diseases The Chinese University of Hong Kong Shatin 999077 HongKong China Brain Tumor Centre and Division ofNeurosurgery Department of Surgery The Chinese University ofHong Kong Shatin 999077 Hong Kong China Institute of Cell Biology Zhejiang UniversityHangzhou 310058 Zhejiang Province China

AIM:To investigate the role of transforming growth factor(TGF)-β-inducible early gene 1(TIEG1) in TGF-β-induced growth inhibition in hepatocellular carcinoma(HCC) cells.METHODS:Human hepatocyte and HCC cell lines with varied susceptibilities to TGF-β1 were tested by methylthiazoletetrazolium(MTT) assay.The expression changes of Smad2,Smad3,Smad4,Smad7,TIEG1 and TIEG2 gene following treatment with TGF-β1 in a TGF-β-sensitive hepatocyte cell line(MIHA),a TGF-β-sensitive hepatoma cell line(Hep3B) and two TGF-β-insensitive hepatoma cell lines(HepG2 and Bel7404) were examined.SiRNA targeting TIEG1 was transfected into Hep3B cells and the sensitivity of cells to TGF-β1 was examined.Overexpression of TIEG1 was induced by lentiviral-mediated transduction in TGF-β1-resistant hepatoma cell lines(Bel7404 and HepG2).MTT assay and 4',6-Diamidino-2-phenylindole staining were used to identify cell viability and apoptosis,respectively.The expression level of stathmin was measured by reverse transcriptase polymerase chain reaction and Western-blotting analysis,and stathmin promoter activity by TIEG1 was monitored by a luciferase reporter gene system.RESULTS:TIEG1 was significantly upregulated by TGF-β1 in the TGF-β1-sensitive HCC cell line,Hep3B,but not in the resistant cell lines.The suppression of TIEG1 by siRNAs decreased the sensitivity of Hep3B cells to TGF-β1,whereas the overexpression of TIEG1 mediated growth inhibition and apoptosis in TGF-β1-resistant HCC cell lines,which resembled those of TGF-β1-sensitive HCC cells treated with TGF-β1.Our data further suggested that stathmin was a direct target of TIEG1,as stathmin was signif icantly downregulated by TIEG1 overexpression,and stathmin promoter activity was inhibited by TIEG1 in a dose-dependent manner.CONCLUSION:Our data suggest that transactivation of TIEG1 conferred growth inhibition of TGF-β-susceptible human HCC cells.

关键词：逆转录聚合酶链反应转化生长因子细胞生长敏感性肝癌 Western印迹荧光素酶报告基因肝细胞株

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Suppressing tumor growth of nasopharyngeal carcinoma by hTERTC27 polypeptide delivered through adeno-associated virus plus adenovirus vector cocktail

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Chinese Journal of Cancer 2012年第12期31卷 588-597页

作者： Xiong Liu Xiang-Ping Li Ying Peng Samuel S.Ng Hong Yao Zi-Feng Wang Xiao-Mei Wang hsiang-fu kung Marie C.M. Lin Department of Otolaryngology Nanfang Hospital Nanfang Medical UniversityGuangzhou Guangdong 510515 P. R. China Department of Neurology The Sun Yat-sen Memorial Hospital Sun Yat-sen UniversityGuangzhou Guangdong 510120 P. R. China Brain Tumor Center Divison of Neurosurgery Department of Surgery The Chinese University of Hong KongShatin Hong Kong China Center for Emerging Infectious Diseases The Chinese University of Hong Kong ShatinHong Kong P. R. China Medical College Shenzhen UniversityShenzhen Guangdong 518061 P. R. China

Nasopharyngeal carcinoma(NPC) is a metastatic carcinoma that is highly prevalent in Southeast Asia. Our laboratory has previously demonstrated that the C-terminal 27-kDa polypeptide of human telomerase reverse transcriptase (hTERTC27) inhibits the growth and tumorigenicity of human glioblastoma and melanoma cells. In this study, we investigated the antitumor effect of hTERTC27 in human C666-1 NPC cells xenografted in a nude mouse model. A cocktail of vectors comprising recombinant adeno-associated virus (rAAV) and recombinant adenovirus (rAdv) that each carry hTERTC27 (rAAV-hTERTC27 and rAdv- hTERTC27; the cocktail was abbreviated to rAAV/rAdv-hTERTC27) was more effective than either rAAV- hTERTC27 or rAdv-hTERTC27 alone in inhibiting the growth of C666-1 NPC xenografts. furthermore, we established three tumors on each mouse and injected rAAV/rAdv-hTERTC27 into one tumor per mouse. Although hTERTC27 expression could only be detected in the injected tumors, reduced tumor growth was observed in the injected tumor as well as the uninjected tumors, demonstrating that the vector cocktail could provoke an antitumor effect on distant, metastasized tumors. further studies showed the observed antitumor effects included inducing necrosis and apoptosis and reducing microvessel density. Together, our data suggest that the rAAV/rAdv-hTERTC27 cocktail can potently inhibit NPC tumor growth in both local and metastasized tumors and should be further developed as a novel gene therapy strategy for NPC.

关键词：重组腺相关病毒抑制肿瘤生长抑制腺病毒载体鸡尾酒鼻咽癌多肽人端粒酶逆转录酶

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A glycolysis-based ten-gene signature correlates with the clinical outcome, molecular subtype and IDH1 mutation in glioblastoma

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Journal of Genetics and Genomics 2017年第11期44卷 519-530页

作者： Cong Chen Yu Shi Yong Li Zhi-Cheng He Kai Zhou Xiao-Ning Zhang Kai-Di Yang Jin-Rong Wu hsiang-fu kung Yi-Fang Ping Xiu-Wu Bian Institute of Pathology and Southwest Cancer Center Southwest Hospital Third Military Medical University Key Laboratory of Tumor Immunopathology of Ministry of Education of China Third Military Medical University

Reprogrammed metabolism is a hallmark of cancer. Glioblastoma（GBM） tumor cells predominantly utilize aerobic glycolysis for the biogenesis of energy and intermediate nutrients. However, in GBM, the clinical significance of glycolysis and its underlying relations with the molecular features such as IDH1 mutation and subtype have not been elucidated yet. Herein, based on glioma datasets including TCGA（The Cancer Genome Atlas）, REMBRANDT（Repository for Molecular Brain Neoplasia Data） and GSE16011 we established a glycolytic gene expression signature score（GGESS） by incorporating ten glycolytic genes. Then we performed survival analyses and investigated the correlations between GGESS and IDH1 mutation as well as the molecular subtypes in GBMs. The results showed that GGESS independently predicted unfavorable prognosis and poor response to chemotherapy of GBM patients. Notably, GGESS was high in GBMs of mesenchymal subtype but low in IDH1-mutant GBMs. furthermore, we found that the promoter regions of tumor-promoting glycolytic genes were hypermethylated in IDH1-mutant GBMs.Finally, we found that high GGESS also predicted poor prognosis and poor response to chemotherapy when investigating IDH1-wild type GBM patients only. Collectively, glycolysis represented by GGESS predicts unfavorable clinical outcome of GBM patients and is closely associated with mesenchymal subtype and IDH1 mutation in GBMs.

关键词： Glycolysis Glioblastoma Prognosis IDH1 Molecular subtype

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Cancer gene therapy targeting angiogenesis:An updated review

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World Journal of Gastroenterology 2006年第43期12卷 6941-6948页

作者： Ching-Chiu Liu Zan Shen hsiang-fu kung Marie CM Lin Institute of MolecularTechnology for Drug Discovery and Synthesis Department ofChemistry The University of Hong Kong Pokfulam Hong KongChina Centre for Emerging Infectious Diseases TheChinese University of Hong Kong Shatin Hong Kong China

Since the relationship between angiogenesis and tumor growth was established by Folkman in 1971, scientists have made efforts exploring the possibilities in treating cancer by targeting angiogenesis. Inhibition of angiogenesis growth factors and administration of angiogenesis inhibitors are the basics of anti- angiogenesis therapy. Transfer of anti-angiogenesis genes has received attention recently not only because of the advancement of recombinant vectors, but also because of the localized and sustained expression of therapeutic gene product inside the tumor after gene transfer. This review provides the up-to-date information about the strategies and the vectors studied in the field of anti-angiogenesis cancer gene therapy.

关键词：肿瘤生长临床基因治疗血管生成

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Pluronic L-81 ameliorates diabetic symptoms in db/db mice through transcriptional regulation of microsomal triglyceride transfer protein

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World Journal of Gastroenterology 2009年第24期15卷 2987-2994页

作者： Wo-Shing Au Li-Wei Lu Sidney Tam Otis King Hung Ko Billy KC Chow Ming-Liang He Samuel S Ng Chung-Man Yeung Ching-Chiu Liu hsiang-fu kung Marie C Lin Institute of Molecular Technology for Drug Discovery and Synthesis Department of Chemistry The University of Hong Kong Hong Kong China Department of Pathology The University of Hong Kong Hong Kong China Division of Clinical Biochemistry Queen Mary Hospital Hong Kong China School of Biological Sciences The University of Hong Kong China Stanley Ho Center for Emerging Infectious Diseases The Chinese University of Hong Kong Shatin Hong Kong China

AIM: To test whether oral L-81 treatment could im-prove the condition of mice with diabetes and to investigate how L-81 regulates microsomal triglyceride transfer protein (MTP) activity in the liver. METHODS: Genetically diabetic (db/db) mice were fed on chow supplemented with or without L-81 for 4 wk. The body weight, plasma glucose level, plasma lipid profile, and adipocyte volume of the db/db mice were assessed after treatment. Toxicity of L-81 was also evaluated. To understand the molecular mecha-nism, HepG2 cells were treated with L-81 and the effects on apolipoprotein B (apoB) secretion and mRNA level of the MTP gene were assessed.RESULTS: Treatment of db/db mice with L-81 sig-nificantly reduced and nearly normalized their body weight, hyperphagia and polydipsia. L-81 also markedly decreased the fasting plasma glucose level, improved glucose tolerance, and attenuated the elevated levels of plasma cholesterol and triglyceride. At the effective dosage, little toxicity was observed. Treatment of HepG2 cells with L-81 not only inhibited apoB secretion, but also signif icantly decreased the mRNA level of the MTP gene. Similar to the action of insulin, L-81 exerted its effect on the MTP promoter. CONCLUSION: L-81 represents a promising candidate in the development of a selective insulin-mimetic mol-ecule and an anti-diabetic agent.

关键词：酯转运蛋白抗糖尿病甘油三酯微粒体小鼠 db 转录调控基因表达水平

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Artificial cold exposure induced stroke in renovascular hypertensive rats and its association with cold-inducible RNA binding protein mRNA expression in brain tissue and blood pressure

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Neural Regeneration Research 2007年第8期2卷 455-460页

作者： Xiaoaena Shi Jianwen Lin Ying Peng Lally L.K. Chan hsiang fu kung Marie C. Lin Ruxun Huang Department of Neurology Second Affiliated Hospital Guangzhou University of Traditional Chinese Medicine Guangzhou 51012~Guangdong Province China Department of Neurology First Affiliated Hospital of Sun Yat-sen University Guangzhou 510008 Guangdong Province China Department of Chemistry Open Laboratory of Chemical Biology Institute of Molecular Technology for Drug Discovery and SynthesisUniversity of Hong Kong Hong Kong Department of Neurology Second Affiliated Hospital of Sun Yat-sen University Guangzhou 510008 Guangdong Province China Li Ka Shing Institute of Health Sciences and Centre for Emerging Infectious Diseases. the Chinese Universityof Hong Kong Hong Kong

BACKGROUND： High incidence of stroke at interchange period of autumn and winter was demonstrated by epidemiological survey, and the specific causes should be further investigated. OBJECTIVE： To investigate the influence of artificial cold exposure on the incidence of stroke in renovascular hypertensive rats （RHR）, and analyze the association with blood pressure and cold-inducible RNA binding protein （CIRP） mRNA expression in brain tissue. DESIGN： A completely randomized grouping design, a randomized control animal trial. SETTINGS： Lab of Neurology, the First Affiliated Hospital of Sun Yat-sen University; Department of Chemistry, Open laboratory of Chemical Biology, Institute of Molecular Technology for Drug Discovery and Synthesis, University of Hong Kong. MATERIALS： Male SD rats （n=460）, weighing 80 - 100 g were obtained from Guangdong Province Health Animal Unit. A modified RXZ-300A intelligent artificial climate cabinet （Ningbo Jiangnan Instrument Co. ,Ltd., China）. METHODS： The experiment were processed in the Lab of Neurology, the First Affiliated Hospital of Sun Yat-sen University and the Open Laboratory of Chemical Biology, Institute of Molecular Technology for Drug Discovery and Synthesis, University of Hong Kong from October 2004 to November 2005. Rats （n = 400） were operated to establish 2-kidney 2-clip RHR model as described previously. The sham-operated rats （n =60） served as normotensive controls. Eight weeks later, 300 of RHR were randomly selected according to their systolic blood pressure （SBP） and divided into 3 sub-groups （n =100 per group）： mild hypertensive group （SBP of 160 - 200 mm Hg）, moderate hypertensive group （SBP of 200 - 220 mm Hg） and severe hypertensive group （SBP 〉 220 mm Hg）. Each group was further divided into two groups （n =50） under ACE and non-ACE. Normal sham-operated SD rats （n =60）, SBP 〈 140 mm Hg, were randomly divided into two groups： Sham-operated control group （n =30） under ACE and non-ACE. To establish the ACE and non-ACE treatment, rats were housed individually in ar

关键词： artificial cold exposure hypertension renovascular hypertensive rats （RHR） stroke cold-inducible RNA binding protein

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