• Journal of Pharmaceutical Investigation
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• v.34 no.5
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• pp.351-362
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• 2004

The basic concept underlying gene therapy is that human diseases may be treated by the transfer of genetics material into specific cells of a patient in order to correct or supplement defective genes responsible for disease development. There are several systems that can be used to transfer foreign genetic material into the human body. Both viral and non-viral vectors are developed and evaluated for delivering therapeutic genes. Viral vectors are biological systems derived from naturally evolved viruses capable of transferring their genetics materials into host cells. However, the limitations associated with viral vectors, in terms of their safety, particularly immunogenecity, and their limited capacity of transgenic materials, have encouraged researchers to increasingly focus on non-viral vectors as an alternative to viral vectors. Although non-viral vectors are less efficient than viral ones, they have the advantages of safety, simplicity of preparation and high gene encapsulation capability. This article reviews the most recent studies highlighting the advantages and the limitation of gene delivery systems focused on non-viral systems compared to viral systems.

• Journal of Pharmaceutical Investigation
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• v.30 no.1
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• pp.1-12
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• 2000

Gene therapy is a method for the treatment of diseases with introducing the gene-engineered materials into a patient with gene-deficiency disease (e.g. cystic fibrosis) or cancer to produce a therapeutic protein in a patient's cells. Successful gene therapy requires establishing both gene expression systems and delivery systems. Viral and non-viral vectors have been used for gene delivery. Viral vectors have a high transfection efficiency, but are limited in relations to issues of safety, toxicity and immunogenecity. Non-viral vectors are easy to prepare and relatively safe. However, non-viral vectors have a low transfection efficiency. Cationic liposomes are the most available among non-viral vectors. Cationic liposomes have been used to transfect cells both in vitro and in vivo experiments. Besides, several formulations containing cationic lipid are being used in clinical trials in cases of cystic fibrosis or cancer. A crucial subject to the further development of gene delivery vectors will be a long-term gene expression with following characteristics; protecting and deliverying DNA efficiently, non-toxic and non-immunogenic, and easy to produce in large scale.

• Archives of Pharmacal Research
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• v.24 no.1
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• pp.1-15
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• 2001

Gene therapy has emerged as a new concept of therapeutic strategies to treat diseases which do not respond to the conventional therapies. The principle of gene therapy is to Introduce genetic materials into patient cells to produce therapeutic proteins in these cells. Gene therapy is now at the stage where a number of clinical trials have been carried out to patients with gene-deficiency disease or cancer. Genetic materials for gene therapy are generally composed of gene expression system and gene delivery system. For the clinical application of gene therapy in a way which conventional drugs are used, researches have been focused on the design of gene delivery system which can offer high transfection efficiency with minimal toxicity. Currently, viral delivery systems generally provide higher transfection efficiency compared with non-viral delivery systems while non-viral delivery systems are less toxic, less immunogenic and manufacturable in large scale compared with viral systems. Recently, novel strategies towards the design of new non-viral delivery system, combination of viral and non-viral delivery systems and targeted delivery system have been extensively studied. The continued effort in this area will lead us to develop gene medicine as "gene as a drug" in the near future.

• Biomedical Science Letters
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• v.9 no.2
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• pp.67-74
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• 2003

Viral and non-viral vectors have been used in the delivery of genetic materials into animal cells and tissues, with each approach having pros and cons. Non-viral vectors have many useful merits such as easy preparation, low immunity and size tolerance of a transgene when compared to those of viral vectors. Delivery specificity may be achieved by complex formation between receptor ligands and a non-viral vector. In the present study, non-viral vector systems are investigated in an effort to find a practical delivery means for gene therapy, Receptor-ligand interaction between transferrin-receptor and transferrin was utilized for efficient gene transfer into cancer cells. A plasmid vector, pcDNA3 (LacZ) was ligated with a small duplexed oligo fragment in which a Biotin- VN$^{TM}$ phosphoramidite was placed in the middle of the oligo. The plasmid vector labeled by biotin was then conjugated with biotin-labeled transferrin via streptavidin. This trimeric conjugates were delivered to a hepatoma cell line, HepG2. The delivery efficiency of the trimeric conjugate was 2-fold higher than that of cationic liposomes used for transfection of a plasmid vector. These results demonstrate that a plasmid vector can be efficiently transferred into cells by forming a trimeric complex of plasmid vector-linker-ligand.

• BMB Reports
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• v.30 no.4
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• pp.269-273
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• 1997

Heptocvte-specific expression induced by Hepatitis B virus (HBV) enhancer 2-core gene promoter was examined in various hepatocyte and non-hepatocyte cell lines. using non-viral and retroviral vector systems in which chloramphenicol acetyltransferase (CAT) is used as a reporter. The non-viral plasmid containing the HBV enhancer 2-core promoter exhibited 22 and 66% of CAT activities in hepatoma cell lines. HepG2 and Hep3B, respectively when compared with CAT activity expressed by CMV promoter. The CAT activities, however. were found to be marginal in other tested hepatoma cell lines as well as mouse primary hepatocytes and non-hepatocytes. The HBV enhancer 2 located upstream the CMV promoter did not affect the CMV promoter activity nor provided hepatocyte-specific expression. Transfection of retroviral plasmid DNA containing the HBV enhancer 2-core promoter as an internal promoter exhibited high and specific CAT expression in HepG2 and Hep3B cell lines but the activity value was 5 to 10 fold lower than the non-viral plasmid with identical promoter. These results suggest that the usage of HBV enhancer 2-core promoter for liver specific expression is limited to certain vectors and hepatocyte cell lines.

• Molecules and Cells
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• v.37 no.2
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• pp.140-148
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• 2014

We identified four basic amino acid residues as nuclear localization signals (NLS) in the C-terminal domain of the prototype foamy viral (PFV) integrase (IN) protein that were essential for viral replication. We constructed seven point mutants in the C-terminal domain by changing the lysine and arginine at residues 305, 308, 313, 315, 318, 324, and 329 to threonine or proline, respectively, to identify residues conferring NLS activity. Our results showed that mutation of these residues had no effect on expression assembly, release of viral particles, or in vitro recombinant IN enzymatic activity. However, mutations at residues 305 (R ${\rightarrow}$ T), 313(R ${\rightarrow}$ T), 315(R ${\rightarrow}$ P), and 329(R ${\rightarrow}$ T) lead to the production of defective viral particles with loss of infectivity, whereas non-defective mutations at residues 308(R ${\rightarrow}$ T), 318(K ${\rightarrow}$ T), and 324(K ${\rightarrow}$ T) did not show any adverse effects on subsequent production or release of viral particles. Sub-cellular fractionation and immunostaining for viral protein PFV-IN and PFV-Gag localization revealed predominant cytoplasmic localization of PFV-IN in defective mutants, whereas cytoplasmic and nuclear localization of PFV-IN was observed in wild type and non-defective mutants. However sub-cellular localization of PFV-Gag resulted in predominant nuclear localization and less presence in the cytoplasm of the wild type and non-defective mutants. But defective mutants showed only nuclear localization of Gag. Therefore, we postulate that four basic arginine residues at 305, 313, 315 and 329 confer the karyoplilic properties of PFV-IN and are essential for successful viral integration and replication.

• Proceedings of the PSK Conference
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• 2000.10a
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• pp.105-107
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• 2000

• Asian Pacific Journal of Cancer Prevention
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• v.16 no.3
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• pp.1037-1040
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• 2015

Hepatitis C is a blood-borne infectious disease of liver, caused by a small enveloped, positive-single stranded RNA virus, called the hepatitis C virus (HCV). HCV belongs to the Flaviviridae family and has 6 genotypes and more than 100 subtypes. It is estimated that 185 million people are infected with HCV worldwide and 5% of these are in Pakistan. The study was designed to evaluate different genotypes of HCV circulating in District Mardan and to know about the behavior of these genotypes to different anti-viral regimes. In this study 3,800 patients were exposed to interferon alfa-2a plus Ribavirin treatment for 6-months and subjected to real-time PCR to check the viral response. Among these 3,677 (97%) patients showed no detectable HCV RNA while 123 (3%) patients (non-responders) remained positive for HCV RNA. Genotypes of their analyzed showed that most of them belonged to the 3a genotype. Non-responders (123) and relapsed (5) patients were subjected to PEG-interferon and Ribavirin therapy for next 6 months, which resulted into elimination of HCV RNA from 110 patients. The genotypes of the persisting resistant samples to anti-viral treatment were 3b, 2a, 1a and 1b. Furthermore, viral RNA from 6 patients remained un-typed while 4 patients showed mixed infections. HCV was found more resistant to antiviral therapy in females as compared to mals. The age group 36-45 in both females and males was found most affected by infection. In general 3a is the most prevalent genotype circulating in district Mardan and the best anti-viral therapy is PEG-interferon plus Ribavirin but it is common practice that due to the high cost patients receive interferon alfa-2a plus Ribavirin with consequent resistance in 3% patients given this treatment regime.

• Journal of Microbiology and Biotechnology
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• v.30 no.9
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• pp.1273-1281
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• 2020

Due to the broad host suitability of viral vectors and their high gene delivery capacity, many researchers are focusing on viral vector-mediated gene therapy. Among the retroviruses, foamy viruses have been considered potential gene therapy vectors because of their non-pathogenicity. To date, the prototype foamy virus is the only retrovirus that has a high-resolution structure of intasomes, nucleoprotein complexes formed by integrase, and viral DNA. The integration of viral DNA into the host chromosome is an essential step for viral vector development. This process is mediated by virally encoded integrase, which catalyzes unique chemical reactions. Additionally, recent studies on foamy virus integrase elucidated the catalytic functions of its three distinct domains and their effect on viral pathogenicity. This review focuses on recent advancements in biochemical, structural, and functional studies of foamy virus integrase for gene therapy vector research.

• BMB Reports
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• v.45 no.4
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• pp.213-220
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• 2012

Viruses have evolved to manipulate the host cell machinery for virus propagation, in part by interfering with the host cellular signaling network. Molecular studies of individual pathways have uncovered many viral host-protein targets; however, it is difficult to predict how viral perturbations will affect the signaling network as a whole. Systems biology approaches rely on multivariate, context-dependent measurements and computational analysis to elucidate how viral infection alters host cell signaling at a network level. Here we describe recent advances in systems analyses of signaling networks in both viral and non-viral biological contexts. These approaches have the potential to uncover virus- mediated changes to host signaling networks, suggest new therapeutic strategies, and assess how cell-to-cell variability affects host responses to infection. We argue that systems approaches will both improve understanding of how individual virus-host protein interactions fit into the progression of viral pathogenesis and help to identify novel therapeutic targets.