To explore the regulation of eIF4E, we screened the protein interacting with eIF4E from human cDNA library by using yeast two-hybrid system. Several clones interacting with eIF4E were identified. One of them was homologous with HUWE1 (HECT, UBA and WWE domain containing 1, also named as ARF-BP1, HECTH9 or HUWE1). Cell co-immunoprecipitation showed that eIF4E could bind to HUWE1 in mammalian cells. We also found that HUWE1 bearing the HECT domain is necessary for its association with eIF4E.

AIM: Pancreatic and duodenal homeobox gene 1(pdx-1) is a crucial transcription factor in pancreatic islet development and differentiation. This study was conducted to evaluate whether pdx-1 delivered by adeno-associated virus (AAV) could induce liver cells to differentiate into insulin-producing cells in diabetic rats and thus provide more information for cell replacement therapy for diabetes. METHODS: Recombinant AAV vector was employed to deliver pdx-1to STZ-induced diabetic rats via portal vein (4×1011). Blood glucose and body weight were monitored. Gene expression of pdx-1 and insulin were determined by RT-PCR and immunocytotochemistry (ICC) at the 6th week after the injection. RESULTS: AAV-pdx-1 group showed obvious gene expression of pdx-1 and insulin by RT-PCR analysis and the presence of more insulin-positive cells by ICC. Hyperglycemia was partially ameliorated and body weight was also increased in AAV-pdx-1 treated diabetic rats, though still significantly different from those in the non-diabetic group. CONCLUSION: The data indicate that AAV-pdx-1 can induce more rat liver cells into insulin-producing cells in vivo, thereby ameliorate hyperglycemia. Further experiments are needed to explore which subpopulation of liver cells responds to this development shift and the mechanism of this development shift induced by pdx-1 in order to improve the differentiation efficiency.

Human gene therapy needs to express exogenous DNA at the targeting cells,producing a practical and efficient therapeutic dosage at an approp-riate time(quantitative pharmacology)with a safe man-ner.Recombinant adeno-associated virus(rAAV)Vec-tom possess a number of properties and recent progress in rAAV production made it rapidly become the reagent of choice for therapeutic gene tmasfer.Over 60 clinical trials of gene therapy based on rAAV have been carried out.The dose response reaction between rAAV vectors and gene expression activity or clinical outcome is one of major aspects of these trials.Most studies showed that vector genomes(vg)and gene expression had a concentration-dependent relationship during a certain scope.However,gene expr~sion Can be afffected by viral serotypes,tissue tropisms,cell targeting,drug regulation,injection route,age and sex,etc.Thus,these aspects should be carefully comidered by scienti-sts,pharmacologisis and physicians during animal ex-periments or clinical trails.KEY WORDS gene therapy;viral vector;dose-re-sponse;quantitative pharmacology;clinical thempy

Equipping tumor-targeting carrier with cell-penetrating peptide with high transduction efficacy has become a trend. According to the fusion modes and tumor-targeting mechanisms, cell-penetrating peptides can be divided into five categories: first, self-targeting cell-penetrating peptides; second, fusion carriers made of targeting ligands and cell-penetrating peptides; third, cell-penetrating peptide-modified nano-carrier; fourth, tumor-microenvironment-targeting cell-penetrating peptides; fifth, other special cell-penetrating peptides. Fusion carriers, which can not be effectively released into the cytoplasm after transducted into target cells, can be further modified to increase their efficacy. In all, cell-penetrating peptide introduced into tumor-targeting carrier should provide a new era for anti-tumor pharmacy research and cancer therapy.

How to reduce immune response is an unprecedented challenge for rAAV gene medicine. Recent studies suggested that lowering dosage of the vector used could reduce immune response caused by rAAV gene medicine. Nevertheless, it would also decrease the transgene expression, leading to failure of gene treatment. It is therefore important to take appropriate steps to maintain high gene expression level and pharmacodynamic, while the dosage of rAAV used is reduced. Here, steps to enhancing gene therapy, such as optimization of the administration, reconstruction of the viral vector and selection of the promoter, are discussed in order to achieve maximum outcome.

Cell adhesion mediated by cell adhesion molecules (CAMs) constitutes essential life phenomenon. In inflammation, immunity, infection, thrombosis, tumor metastasis and wound healing, cell adhesion comes into being the basic physiological and pathological process. Intervening with cell adhesion has been the important therapeutic and prophylactic strategies for diseases. Accumulated evidence has indicated that plant polysaccharides especially those exacted from Chinese traditional and herbal drugs displayed various pharmacological effects such as anti-inflammation, anti-cancer, anti-infection, immunomodulation, cardiovascular protective effects and so on. In this paper, the research progress of plant polysaccharides on cell adhesion is reviewed.

Hepatits B virus( HBV) infection is a global epidemic which seriously harms the public health. In spite of the great progress in hepatits B prevention and treatment, there is few ef-fective medicine. Research findings show that liver damage and degree of liver failure are sophisticatedly related to the interaction between HBV and the immune response of host. All these make it important to know the replication mechanism and the contrac-tion process, in order to lay a preliminary solid foundation for studying HBV drug targets and making a new ant-virus strategy. This article aims to summarize HBV viral replication process, while focusing on the latest research findings about drug targets, to find a new kind of anti-HBV drugs, and to explore the under-lying mechanism of effective drugs.

Objective:To construct K-ras-targeted siRNAs (K-ras siRNA) and to investigate the inhibitory effects of K-ras siRNAs on the growth and migration of lung cancer A549 cells (containing mutant K-ras gene) and NCI-H446 cells (containing wild-type K-ras gene). Methods: Four K-ras siRNAs (K-ras siRNA1～K-ras siRNA3 targeting wild-type K-ras and K-ras siRNA4 targeting mutant K-ras) were designed and artificially synthesized; they were used to transfect A549 cells and NCI-H446 cells. The expressions of Ras mRNA and protein were examined by RT-PCR and Western blot-ting. The inhibitory effects of K-ras siRNAs on the proliferations of A549 and NCI-H446 cells were determined by MTT assay. The effects of K-ras siRNAs on the cell migration and apoptosis were observed by Transwell assay and Hoechst 33258 staining, respectively. Results: Mutant K-ras-targeted siRNA (K-ras siRNA4) specifically inhibited the K-ras ex-pression but had no influence on H-ras and N-ras expression in A549 cells. K-ras siRNA4 inhibited the proliferation of A549 cells but did not inhibit that of NCI-H446 cells, which contained wild type K-ras gene. K-ras siRNA4 also induced apoptosis and inhibited migration of A549 cells. Conclusion: Mutant K-ras-targeted siRNA4 can inhibit the proliferation, migration and induce apoptosis of A549 cells. It may be a potential and personalized drug for the treatment against lung cancer containing mutant K-ras gene.

Gene medicine based on recombinant adeno-associated virus (rAAV) vector has rapidly become the prior-choose reagent for gene therapy, since it had been shown that the rAAV was able to stably express many genes in vivo without detectable side-effect. However, recent findings of CTL immune responses to AAV capsid in a clinical trial highlighted a new issue regarding safety that previously was not identified in animal studies. Obviously it is so important to understand the interaction of rAAV with the immune system in details for the safety and success of rAAV gene medicine. In this review we evaluate several current hypotheses aiming to explain the cellular immunotoxicity, also analysis the current findings including the presentation kinetics of the capsid antigen and the activation of CTL. Focusing on the key steps of the immune response several solutions are proposed, including immunosuppression, optimization of vector and improvement of purity, in order to insure clinical safety and efficacy of rAAV.

Purpose To investigate human soluble TRAIL(sTRAIL)protein expression and purification and its potential anti-tumor activity on hepatocellular carcinoma(HepG2).Methods Soluble TRAIL gene ligated with expression vector pPIC9 was transfected into GS115(his4)and the recombination strain expressing sTRAIL was screened by MD plate.The effects of different media,methanol inducement period,methanol concentration,and pH were investigated and optimized using shaking flask.The anti-tumor activity of sTRAIL with HepG2 cells Was analyzed after purification.Results The highest expression of sTRAIL was obtained at pH 6.0,1% methanol in BMMY medium,with the concentration of(58.7±2.4)mg/L at 48 h.Recombinant sTRAIL protein could induce HepG2 cells apoptosis and inhibit HepG2 cells proliferation effectively.Conclusion The optimized condition of human sTRAIL expression and purification Was developed and the obtained recombinant sTRAIL protein may be a promising therapeutic agent for hepatocellular carcinoma.