• Journal of Microbiology and Biotechnology
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• v.34 no.2
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• pp.280-288
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• 2024

The fusogenic membrane glycoprotein (FMG) derived from the human endogenous retrovirus-W (HERV-W) exhibits fusogenic properties, making it a promising candidate for cancer gene therapy. When cells are transfected with HERV-W FMG, they can fuse with neighboring cells expressing the receptor, resulting in the formation of syncytia. These syncytia eventually undergo cell death within a few days. In addition, it has been observed that an HERV-W env mutant, which is truncated after amino acid 483, displays increased fusogenicity compared to the wild-type HERV-W env. In this study, we observed syncytium formation upon transfection of HeLa and TE671 human cancer cells with plasmids containing the HERV-W 483 gene. To explore the potential of a semi-replication-competent retroviral (s-RCR) vector encoding HERV-W 483 for FMG-mediated cancer gene therapy, we developed two replication-defective retroviral vectors: a gag-pol vector encoding HERV-W 483 (MoMLV-HERV-W 483) and an env vector encoding VSV-G (pCLXSN-VSV-G-EGFP). When MoMLV-HERV-W 483 and pCLXSN-VSV-G-EGFP were co-transfected into HEK293T cells to produce the s-RCR vector, gradual syncytium formation was observed. However, the titers of the s-RCR virus remained consistently low. To enhance gene transfer efficiency, we constructed an RCR vector encoding HERV-W 483 (MoMLV-10A1-HERV-W 483), which demonstrated replication ability in HEK293T cells. Infection of A549 and HT1080 human cancer cell lines with this RCR vector induced syncytium formation and subsequent cell death. Consequently, both the s-RCR vector and RCR encoding HERV-W 483 hold promise as valuable tools for cancer gene therapy.

• Journal of Animal Science and Technology
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• v.46 no.3
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• pp.307-314
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• 2004

Xenotransplantation of porcine organs has the potential to overcome the severe. shortage of human tissues and organs available for human transplantation. The swine represents an ideal source of such organs because of their plentiful supply and their numerous anatomical and physiological similarities to the human. However, this procedure also carries with a number of safety issues relating to the zoonotic infections. Porcine endogenous retrovinJses(PERVs), \Wich are germ line transmitted and persist without symptoms in the pigs, are most concerning zoonotic viroses. In order to analyze the prevalence of PERV in domestic pigs, four kinds of pigs'(Landrace, Berkshire, Yorkshire, and Duroc) genomic DNA were isolated from their hair follicles. PCR analysis was carried out for detection of PERVs using subgroup A/B/C and E pol sequence primers. All pigs (20 heads) tested had high copy number of PERVs within genomes. Subgroup A/B/C and E pol gene sequences from 20 isolates were determined by direct sequencing. Sequence analysis showed pol sequences are highly conserved among intra- and inter-subspecies(99.l and 98.8%, respectively). As a first report of PERV prevalence in Korea pigs, our data would be the basic concepts of PERV transmission study in xenotransplantation.

• Asian-Australasian Journal of Animal Sciences
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• v.27 no.11
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• pp.1644-1651
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• 2014

Transgenic animals have become important tools for the production of therapeutic proteins in the domestic animal. Production efficiencies of transgenic animals by conventional methods as microinjection and retrovirus vector methods are low, and the foreign gene expression levels are also low because of their random integration in the host genome. In this study, we investigated the homologous recombination on the porcine ${\beta}$-casein gene locus using a knock-in vector for the ${\beta}$-casein gene locus. We developed the knock-in vector on the porcine ${\beta}$-casein gene locus and isolated knock-in fibroblast for nuclear transfer. The knock-in vector consisted of the neomycin resistance gene (neo) as a positive selectable marker gene, diphtheria toxin-A gene as negative selection marker, and 5' arm and 3' arm from the porcine ${\beta}$-casein gene. The secretion of enhanced green fluorescent protein (EGFP) was more easily detected in the cell culture media than it was by western blot analysis of cell extract of the HC11 mouse mammary epithelial cells transfected with EGFP knock-in vector. These results indicated that a knock-in system using ${\beta}$-casein gene induced high expression of transgene by the gene regulatory sequence of endogenous ${\beta}$-casein gene. These fibroblasts may be used to produce transgenic pigs for the production of therapeutic proteins via the mammary glands.

• Tuberculosis and Respiratory Diseases
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• v.41 no.2
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• pp.87-96
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• 1994

Background : Since tumor necrosis factor was discovered in 1975, TNF has been well known about its cytotoxic effect on tumor cells in vivo and in vitro. According to the recent improvement of molecular biological techinques, it is possible that exogenous TNF gene is transferred to tumor cells and is expressed in theirs. By virtue of TNF gene transfer, we have expected that TNF expressed in TNF-gene-transferred tumor cells would kill tumor cells in vivo without systemic side effect. The expected mechanisms in which antitumor effects of TNF expressed in TNF-gene-transferred tumor cells are working would be as followings. In the first mechanism, TNF expressed in TNF-gene-transferred tumor cells would kill tumor cells around(like homicide). In the second mechanism, TNF expressed in TNF-gene-transferred tumor cells would kill themselves(like suicide). In the third mechanism, TNF expressed in TNF-gene-transferred tumor cells would recruit immune effector cells and kill tumor cells indirectly. In the last mechanism, TNF expressed in TNF-gene-transferred tumor cells would augment cytokine such as interferon-$\gamma$ to kill tumor cells. Among these four mechanisms of antitumor effect, only the second mechanism has not been established yet. Therefore, to elucidate the second mechanism, We performed this study. Method : We transferred TNF-$\alpha$ gene to NCI-H2058, a human mesothelioma cell line and WEHI164, a murine fibrosarcoma cell line by using retroviral vector(pLT12SNTNF). And, We determined by using MTT assay whether TNF expressed in TNF-gene-transferred tumor cell lines would kill themselves like suicide or not. Then, if TNF-gene-transferred tumor cell lines would not suicide themselves, I would know more about the TNF sensitivity of TNF-gene-transferred tumor cell lines to exogenous TNF also by MTT assay. Result : NCI-H2058 and WEHI164 which were sensitive to TNF, became far less sensitive to endogenous and exogenous TNF after being transferred TNF-$\alpha$ gene to. Conclusion : TNF-gene-transfer to NCI-H2058 and WEHI164 gave them resistance to TNF.