• Journal of Pharmaceutical Investigation
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• 제31권3호
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• pp.185-190
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• 2001

Human papillomavirus (HPV) infection is known to cause cervical cancers. Human papillomavirus-like particles (VLP) have been studied as preventive vaccines of cervical cancers. To develop VLP as a therapeutic gene carrier, we studied the method to encapsulate cytokine genes in virus-like particles. HPV type 16 capsid L1 genes were amplified by polymerase chain reaction and cloned into T vector. L1 gene was then inserted into baculovirus transfer vector. The clone of baculovirus encoding L1 gene was isolated and used to express L1 protein in Sf 21 insect cells. VLP were purified by CsCl density gradient and ultracentrifugation. VLP were disassembled to capsomer units by treatment of a reducing agent. Given that interleukin-2 (IL-2) genes have been used in anticancer gene therapy and as a molecular adjuvant, IL-2 cytokine plasmids were chosen as a model gene. IL-2 plasmids were incubated with the disassembled capsomer suspension. To reassemble the particles, the mixture of capsomers and cytokine plasmids was dialyzed. The disassembly and reassembly of VLP were confirmed by transmission electron microscopy. The entrapment of cytokine plasmids in reassembled VLP was tested by the stability of plasmids against DNase I. After treatment of reassembled virus-like particles with DNase I, discrete IL-2 DNA band was observed. Our results indicate that IL-2 cytokine plasmid (3.5 kb size) can be encapsulated in the virus-like particles, suggesting the potential of VLP as a gene delivery system. Moreover, VLP containing the adjuvant cytokine plasmids might function as more effective subunit vaccines.

• IMMUNE NETWORK
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• 제9권5호
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• pp.158-168
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• 2009

Tumor therapy using cytokines has been developed for last two decades. Several recombinant cytokines and tumor cell vaccines produced by cytokine gene transfer have been in clinical trials, but several side effects hamper routine clinical applications. Many cytokines are originally expressed as membrane-bound form and then processed to secretory form exerting paracrine effects. Though functional differences of these two types of cytokines are elusive yet, the membrane-bound form of cytokine may exert its effects on restricted target cells as a juxtacrine, which are in physical contacts. With the efforts to improve antitumor activities of cytokines in cancer patients, developing new strategies to alleviate life-threatening side effects became an inevitable goal of tumor immunologists. Among these, tumor cell vaccines expressing cytokines as membrane-bound form on tumor cell surface have been developed by genetic engineering techniques with the hope of selective stimulation of the target cells that are in cell-to-cell contacts. In this review, recent progress of tumor cell vaccines expressing membrane-bound form of cytokines will be discussed.

• The Korean Journal of Pain
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• 제22권1호
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• pp.1-15
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• 2009

Neuropathic pain is often refractory to intervention because of the complex etiology and an incomplete understanding of the mechanisms behind this type of pain. Glial cells, specifically microglia and astrocytes, are powerful modulators of pain and new targets of drug development for neuropathic pain. Glial activation could be the driving force behind chronic pain, maintaining the noxious signal transmission even after the original injury has healed. Glia express chemokine, purinergic, toll-like, glutaminergic and other receptors that enable them to respond to neural signals, and they can modulate neuronal synaptic function and neuronal excitability. Nerve injury upregulates multiple receptors in spinal microglia and astrocytes. Microglia influence neuronal communication by producing inflammatory products at the synapse, as do astrocytes because they completely encapsulate synapses and are in close contact with neuronal somas through gap junctions. Glia are the main source of inflammatory mediators in the central nervous system. New therapeutic strategies for neuropathic pain are emerging such as targeting the glial cells, novel pharmacologic approaches and gene therapy. Drugs targeting microglia and astrocytes, cytokine production, and neural structures including dorsal root ganglion are now under study, as is gene therapy. Isoform-specific inhibition will minimize the side effects produced by blocking all glia with a general inhibitor. Enhancing the anti-inflammatory cytokines could prove more beneficial than administering proinflammatory cytokine antagonists that block glial activation systemically. Research on therapeutic gene transfer to the central nervous system is underway, although obstacles prevent immediate clinical application.

• Tuberculosis and Respiratory Diseases
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• 제51권2호
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• pp.135-146
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• 2001

배 경 : 암세포는 정상 세포와 다르므로 면역 기전에 의해 제거되어야 함에도 불구하고 암이 발생하는 것은 암세포가 면역 감시 체계를 회피하기 때문이다. 약제감수성 증강 유전자인 herpes simplex thymidine kinase(HSTK) 유전자를 이용하여 암세포를 파괴하여 종양 특이항원이 더 잘 유리되도록 하고 사이토카인 유전자를 이용하여 면역세포를 유도하여 이 장애를 극복할 수 있는지 보고자 이 실험을 하였다. 방 법 : Lewis 폐암 세포주(LLC)에 adenovirus를 이용하여 HSTK를 형질도입하고 이에 의해 ganciclovir에 대한 LLC의 감수성을 증강시키는지를 관찰하고 mixed population assay를 이용하여 bystander 효과를 관찰하였다. Ad-HSTK, Ad-IL-2, Ad-GMCSF의 형질도입이 LLC의 종양 형성 능력에 영향을 미치는지 관찰하였다. 또한 그러한 형질 도입이 기존의 종양에 대한 항암 효과를 가져오는지 관찰하였다. 항암 효과의 기전을 확인하기 위해 쥐의 비장을 관찰하였다. 결 과 : Ad-HSTK의 형질도입은 ganciclovir에 대한 LLC의 감수성을 현저하게 증강시켰다. Ad-HSTK, Ad-IL-2, Ad-GM-CSF를 형질도입한 LLC를 쥐에 주사하고 ganciclovir로 처리하였을 때 종양 형성 능력이 감소하였다. Ad-HSTK, Ad-IL-2, Ad-GM-CSF를 형질도입한 LLC를 종양백신으로 사용하였을때 종양 성장이 어느 정도 저해되는 것을 관찰하였다. 특히 HSTK와 GM-CSF를 복합 형질도입했을 때 더 강력한 항암효과가 일어나는 것을 관찰하였다. 그러나 HSTK와 IL-2을 복합 형질도입했을 때는 각각을 단독으로 형질도입했을 때보다 항암효과가 상승되지 않았다. HSTK와 GM-CSF의 복합 형질도입한 LLC를 종양백신으로 사용하였을 때 비장의 수상세포 침윤이 현저히 증가하였다. 결 론 : HSTK와 GM-CSF의 복합 형질도입으로 만든 종양백신은 수상세포를 활성화시키므로써 항암면역기능을 유의하게 증강시킬 수 있었다.

• 대한두경부종양학회지
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• 제19권1호
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• pp.25-33
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• 2003

Background and Objectives: The Herpes Simplex type 2 Defective Infectious Single Cycle virus (DISC virus) is attenuated virus originally produced as viral vaccines but are also efficient gene transfer vehicle. The main goals of this study were to examine the efficiencies of the gene transfer using DISC vectors for various head and neck squamous cell carcinoma cell lines and to evaluate the efficacy of vaccination with DISC virus carrying a immunomodulatory genes (GM-CSF) as cancer therapy in a organotopic oral cavity squamous cell cancer model. Materials and Methods : We determinated the gene transfer efficiency of DISC virus by x-gal stain method and proved gene and protein expression of DISC-GMCSF transfected SCCVII cells by RT-PCR and ELISA method. Also we evaluated the ex vivo vaccination effects of SCCVII/GMCSF (DISC-GMCSF transfected SCCVII vaccine) vaccine on preventing the recurrence of micro-residual tumor. After the vaccination of SCCVII/GMCSF, specific cytotoxic T-cell responses was evaluated by CTL assay. Results: At an MOI of 10 DISC virus showed 64-88% of transfection rates in various head and neck squamous cancer cell lines. SCCVII cells transduced by DISC virus vector (MOI=10) carrying the GM-CSF gene, produced 4.5 nanogram quantities of GM-CSF per $10^6$ cells. In vivo vaccination using tumor cells transduced ex vivo with DISC-GMCSF resulted in better protection rate against subsequent tumor recurrence in organotopic oral cavity cancer model. Although tumor free survival rate was not statistically significantly increased in vaccination group (p=0.078), tumor specific cytotocic T-cell responses were significantly increased in SCCVII/GMCSF vaccination group. Conclusion: These data demonstrate that; 1) The DISC virus vector is capable of efficient gene transfer to various head and neck squamous cancer cell lines, 2) GM-CSF secreting genetically modified tumor vaccine (SCCVII/GMCSF) efficiently protected against tumor recurrence in organotopic micro-residual oral cavity cancer model and produced tumor specific cytotoxic T-cell response. DISC virus-mediated, cytokine gene transfer may prove to be useful as a clinical therapy for head and neck cancers.

• Journal of Microbiology and Biotechnology
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• 제27권10호
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• pp.1892-1895
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• 2017

IK can downregulate interferon-gamma-induced major histocompatibility complex (MHC) class II expression through the MHC class II transactivator, which suggests that IK can inhibit the interactions between immune cells. We delivered adeno-associated virus serotype 2 (AAV2) encoding the genes for truncated IK (tIK) or green fluorescent protein (GFP) to DBA1/J mice via intravenous injection. Seven weeks after injection, collagen-induced arthritis was induced in the AAV2-treated mice. AAV2-tIK injection reduced the severity of arthritis and the percentage of pathogenic Th17 cells compared with AAV2-GFP injection. These results suggest a novel gene therapy strategy for treatment of inflammatory arthritis.

• 환경생물
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• 제23권2호
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• pp.152-156
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• 2005

Ionizing radiation is a well- known therapy factor for human carcinoma cells. Genotoxic stress mediates cell cycle control, transcription and cellular signaling. In this work, we have used a microarray hybridization approach to characterize the cell type-specific transcriptional response of human carcinoma MCF-7 and HeLa cell line to $\gamma-radiation$, such as 4Gy 4hr. We found that exposure to $\gamma-ray$ alters by at least a $log_2$ factor of 1.0 the expression of known genes. Of the 27 genes affected by irradiation, 11 are down- regulated in MCF-7 cells and 2 genes induced by radiation,15 are repressed in HeLa cells. Many genes were involved in known damage- response pathways for cell cycling, transcription factor and cellular signaling response. However, in MCF-7 cells, we observed gene expression pattern in chromatin, apoptosis, stress, differentiation, cytokine, metabolism, ribosome and calcium. In HeLa cells, it showed clearly the expression changes in adhesion and migration, lysosome, brain, genome instability and translation. These insights reveal new therapy directions for studying the human carcinoma cell response to radiation.

• Journal of Microbiology and Biotechnology
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• 제16권12호
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• pp.1919-1927
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• 2006

Tumor cells genetically engineered to secrete cytokines are effective in tumor therapy, but various unexpected side effects are observed, which may result from the bulk activation of various bystander cells. In this study, we tested tumor vaccines expressing various membrane-bound forms of IL-2 (mbIL-2) on MethA fibrosarcoma cells to focus antitumor immune responses to CTL. Chimeric forms of IL-2 with whole CD4, deletion forms of CD4, and TNF were expressed on the tumor cell surface, respectively. Tumor clones expressing mbIL-2 or secretory form of IL-2 were able to support the cell growth of CTLL-2, an IL-2-dependent T cell line, and the proliferation of spleen cells from 2C TCR transgenic mice that are responsive to the $p2Ca/L^d$ MHC class I complex. Expression of mbIL-2 on tumor cells reduced the tumorigenicity of tumor cells, and the mice that once rejected the live IL-2/TNF tumor clone acquired systemic immunity against wild-type MethA cells. The IL-2/TNF clone was inferior to other clones in tumor formation, and superior in the stimulation of the CD8+ T cell population in vitro. These results suggest that the IL-2/TNF clone is the best tumor vaccine, and may stimulate CD8+ T cells by direct priming. Expression of IL-2/TNF on tumor cells may serve as an effective gene therapy method to ameliorate the side effects encountered in the recombinant cytokine therapy and the conventional cytokine gene therapy using the secretory form of IL-2.

• Asian Pacific Journal of Cancer Prevention
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• 제13권11호
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• pp.5353-5361
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• 2012

Interleukin-18 (IL-18) is an immune-stimulatory cytokine with antitumor activity in preclinical models. It plays pivotal roles in linking inflammatory immune responses and tumor progression and is a useful candidate in gene therapy of lymphoma or lymphoid leukemia. A phase I study of recombinant human IL-18 (rhIL-18) in patients with advanced cancer concluded that rhIL-18 can be safely given in biologically active doses to patients with advanced cancer. Some viruses can induce the secretion of IL-18 for immune evasion. The individual cytokine activity might be potentiated or inhibited by combinations of cytokines. Here we focus on combinational effects of cytokines with IL-18 in cancer progression. IL-18 is an important non-invasive marker suspected of contributing to metastasis. Serum IL-18 may a useful biological marker as independent prognostic factor of survival. In this review we cover roles of IL-18 in immune evasion, metastasis and angiogenesis, applications for chemotherapy and prognostic or diagnostic significance.

• 동의생리병리학회지
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• 제18권2호
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• pp.468-473
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• 2004

Psoriasis is a chronic inflammatory disease of the skin characterized by epidermal hyperplasia, dermal angiogenesis, infiltration of activated T cells, and increased cytokine levels, and affects 1-3% of the world-wide population. Although many immunological and clinical reports indicate a role for the immune system in the pathogenesis of psoriasis, puzzling questions about psoriasis remain unsolved. During the several decade, immunosuppressor and PUVA treatment are ubiquitously used to psoriasis therapy. But recently, to promote terminal differentiation of keratinocytes, block either NK-Tcell or T-cell activation, and interrupting the angiogenic switch represent another therapeutic opportunity in psoriasis. To keep face with immunological therapy, the needs of newly designed prescription on the psoriasis treatments were demanded. With the object of understand the psoriasis from an orient medical point of view, patients were administrated the GY during several weeks. We investigated the changes of gene expression in involved and uninvolved skin samples during the oriental remedy. Microarray data showed several important results. First, Gene expression profiling is similar to each patient. Second, precursor proteins that organize cornified envelops are decreased at the end of remedy. But genes which related to apoptosis, G-protein signalling, and lipid metabolism are increased. Third, 68.5% of clustering genes localized on the psoriasis susceptibility locus. In our results indicated that GY influence on the keratinocytes hyperproliferation by regulating the gene, which located on the psoriasis susceptibility locus.