• Asian Pacific Journal of Cancer Prevention
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• 제16권14호
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• pp.5687-5690
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• 2015

Background: Factors predictive of survival have been identified in Western patients with metastatic clear cell renal cell carcinoma (mCCRCC) treated with sunitinib. Less is known, however, about factors predictive of survival in Japanese patients. This study evaluated factors prognostic of survival in Japanese patients with mCCRCC treated with first-line sunitinib. Materials and Methods: This retrospective study evaluated 46 consecutive Japanese mCCRCC patients treated with sunitinib as first line therapy. Clinical and biochemical markers associated with progression-free survival (PFS) were analyzed, with prognostic factors selected by uniand multivariate Cox regression analyses. Results: Univariate analysis showed that factors significantly associated with poor PFS included Memorial Sloan-Kettering Cancer Center poor risk scores, International Metastatic RCC Database Consortium poor risk and high (>0.5 mg/dl) serum C-reactive protein (CRP) concentrations (p<0.001 each). Multivariate analysis showed that high serum CRP was independently associated with poorer PFS (p=0.040). Six month disease control rate (complete response, partial response and stable disease) in response to sunitinib was significantly higher in patients with normal (${\leq}0.5mg/dl$) than elevated baseline CRP (p<0.001). Conclusions: CRP is a significant independent predictor of PFS for Japanese patients with mCCRCC treated with first-line sunitinib. Pretreatment CRP concentration may be a useful biomarker predicting response to sunitinib treatment.

• Tuberculosis and Respiratory Diseases
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• 제76권1호
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• pp.1-7
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• 2014

Maintenance therapy has emerged as a novel therapeutic paradigm for advanced non-small-cell lung cancer (NSCLC). Maintenance therapy that aims to sustain a clinically favorable state after first-line chemotherapy has two strategies. Switch maintenance therapy entails switching to a new and non-cross-resistant agent in an alternating or sequential manner, on completion of first-line chemotherapy. Continuous maintenance therapy keeps ongoing administration of a component of the current regimen after four to six cycles of chemotherapy, if there is a stable disease, or better response. Both maintenance therapies can be continued, until disease progression. The potential evidence regarding maintenance therapy includes providing the opportunity to receive additional treatment, through sustaining tumor shrinkage, and delayed emergence of tumor-related symptom. Thus far, debates over the parameters used to predict the effectiveness of maintenance therapy, financial burden, and uncertainty of improving the quality of life exist. Despite many debates, maintenance therapy, which is currently recommended, has been disclosed to be beneficial.

• 한국어류학회지
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• 제20권1호
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• pp.1-6
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• 2008

제브라피쉬 배아로부터 확보한 세포주로부터 외부형태와 세포 크기에 따라 3종류의 동형세포주를 확립하였다. 활발하게 증식하는 안정된 세포주 및 이들로부터 확립된 동형세포주의 세포특성은 변하지 않고 지속적으로 유지되었으며, 안정된 세포주로부터 총 18개의 콜로니를 확보하여 배양한 다음 3종류의 동형세포주를 선별하여 세포 특성을 분석하였다. 대부분의 동형세포주는 약 80% 정도 정상적인 염색체(2N=50)를 가지고 있었으며 FACs 분석과 일치하였다. 배아로부터 확립된 동형세포주에 항체테스트 결과, vimentin에서 양성을 보이는 결과로 볼 때 확립된 세포주는 분화된 섬유세포임이 확인되었다. 이러한 결과는, 확립된 동형세포주를 이용한 유전자조작과 어류복제에의 활용도를 높일 수 있음을 시사한다.

• 대한약학회:학술대회논문집
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• 대한약학회 2002년도 Proceedings of the Convention of the Pharmaceutical Society of Korea Vol.2
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• pp.324.1-324.1
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• 2002

Human 5-reductase type II(5AR2) is an important target for the treatment of benign prostatic hyperplasia. In this study we describe the establishment of cell line which stably expressed 3X FLAG tagged human 5AR2. We used this cell line as a cell based assay tool and source for 5AR2 enzyme. First a plasmid (3XFLAGpCMVl0-5AR2) for the expression of 5AR2 was constructed by the use of the vector 3XFLAGpCMV10 and transfected into the HEK 293. By selection with G418 sulfate. ten HEK 293 single cell clones were obtained of which three stably exhibited high 5AR2 activity. (omitted)

• Molecules and Cells
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• 제42권11호
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• pp.783-793
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• 2019

When endoplasmic reticulum (ER) functions are perturbed, the ER induces several signaling pathways called unfolded protein response to reestablish ER homeostasis through three ER transmembrane proteins: inositol-requiring enzyme 1 (IRE1), PKR-like ER kinase (PERK), and activating transcription factor 6 (ATF6). Although it is important to measure the activity of ATF6 that can indicate the status of the ER, no specific cell-based reporter assay is currently available. Here, we report a new cell-based method for monitoring ER stress based on the cleavage of $ATF6{\alpha}$ by sequential actions of proteases at the Golgi apparatus during ER stress. A new expressing vector was constructed by using fusion gene of GAL4 DNA binding domain (GAL4DBD) and activation domain derived from herpes simplex virus VP16 protein (VP16AD) followed by a human $ATF6{\alpha}$ N-terminal deletion variant. During ER stress, the GAL4DBD-VP16AD(GV)-$hATF6{\alpha}$ deletion variant was cleaved to liberate active transcription activator encompassing GV-$hATF6{\alpha}$ fragment which could translocate into the nucleus. The translocated GV-$hATF6{\alpha}$ fragment strongly induced the expression of firefly luciferase in HeLa Luciferase Reporter cell line containing a stably integrated 5X GAL4 site-luciferase gene. The established double stable reporter cell line HLR-GV-$hATF6{\alpha}$(333) represents an innovative tool to investigate regulated intramembrane proteolysis of $ATF6{\alpha}$. It can substitute active pATF6(N) binding motif-based reporter cell lines.

• Asian Pacific Journal of Cancer Prevention
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• 제14권9호
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• pp.5391-5396
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• 2013

Aim: To investigate the role of Golgi phosphoprotein 3 (GOLPH3) in tumour growth and metastasis of esophageal squamous cancer. Methods: A lentiviral shRNA-vector was utilized to stably knockdown GOLPH3 in Eca-109 esophageal squamous cancer cells. mRNA transcription and protein expression of GOLPH3 were examined by real-time quantitative PCR and Western blotting, respectively. Cell proliferation activity was assessed by MTT assay and invasion and migration potentials by matrigel invasion and transwell motility assays. Results: Stable knockdown in the GOLPH3 cell line was established. PD-A gene expression was significantly suppressed by lentivirus-mediated RNAi, which resulted in reducing the capacity for cell proliferation, migration, invasion and adhesion in vitro. In vivo, GOLPH3 depletion resulted in inhibition of tumour growth, with stable decrease in the expression of GOLPH3 in tumor xenografts. Conclusions: Our findings suggest that lentivirus mediated silencing of the GOLPH3 gene has a significant anti-tumour effect on esophageal squamous cancer in vitro and in vivo. In addition, the results indicate that GOLPH3 might be an effective molecular target for gene therapy in esophageal squamous cancer.

• Asian-Australasian Journal of Animal Sciences
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• 제20권7호
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• pp.1015-1022
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• 2007

The spermatogonial stem cell (SSCs) is unique in that it is the only cell in the adult male that can contribute genes to a subsequent generation. Permanent modification of the germ cell line may be realized if stem cells could be cultured, transfected with unique genes, and then transplanted into recipient testes. We developed a culture system that supported long-term viability of SSCs. We used a retrovirus vector (pMSCV including ${\beta}$-galactosidase) to stably transfect spermatogonia following long-term culture using the system developed. Expression of the reporter gene ${\beta}$-galactosidase controlled by the retroviral vector was stable in long-term cultured SSCs. We confirmed the retroviral-mediated ${\beta}$-galactsidase gene could be expressed in germ cells in recipient mice following SSCs transplantation.

• KSBB Journal
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• 제32권3호
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• pp.193-198
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• 2017

Sialylation is important in producing therapeutic proteins such as antibody, cytokine and fusion protein. Thus, enhancement of sialylation is usually performed in CHO cell cultures. ${\alpha}2,6$-Sialyltransferase (ST), which plays a key role in the attachment of ${\alpha}2,6-sialic$ acid, is present in human cells but not in Chinese hamster ovary (CHO) cells. Overexpression of ${\alpha}2,6-ST$ can be used for enhancing the degree of sialylation and achieving human-like glycosylation. In this study, we constructed CHO cells producing human cytotoxic T-lymphocyte antigen4-immunoglobulin (hCTLA4-Ig) as well as ${\alpha}2,6-ST$. Transfected CHO cells were selected using G418 and stable cell line was established. Profiles of viable cell density and hCTLA4-Ig titer in an overexpressed cell line were similar to those of a wild-type cell line. It was confirmed that the total amount of sialic acid was increased and ${\alpha}2,6-sialic$ acid was attached to the terminal residues of N-glycan of hCTLA4-Ig by ESI-LC-MS. Compared to 100% of ${\alpha}2,3-sialic$ acid in wild type cells, 70.9% of total sialylated N-glycans were composed of ${\alpha}2,6-sialic$ acid in transfected cells. In conclusion, overexpression of ${\alpha}2,6-ST$ in CHO cells led to the increase of both the amount of total sialylated N-glycan and the content of ${\alpha}2,6-sialic$ acid, which is more resemble to human-like structure of glycosylation.

• 약학회지
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• 제39권2호
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• pp.161-168
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• 1995

Brain dopamine systems play a central role in the control of movement, hormone release, and many complex behavior. The action of dopamine at its synapse is terminated predominately by high affinity reuptake into presynaptic terminals by dopamine transporter (DAT). The dopamine transporter(DAT) is membrane protein localized to dopamine-containing nerve terminals and closely related with cocaine abuse, Parkinsonism, and schizophrenia. In present study, the recombinant plasmid pRc/CMV-DAT, constructed by subcloning of a cDNA encoding a bovine DAT into eukaryotic expression vector pRc/CMV, was stably transfected into CV-1 cells(monkey kidney cell line). The DAT activities in the cell lines selected by Geneticin$^{R}$ were determined by measuring the uptake of $[^3H]$-dopamine. The transfected cell lines showed 30-50 fold higher activities than untransfected CV-1 cell line, and this result implies that DAT is well expressed and localized in transfected cells. The transfected cells accumulated $[^3H]$-dopamine in a dose-dependent manner with a $K_{m}$ of 991.6nM. Even though high doses of norepinephrine, epinephrine, serotonin, and choline neurotransmitters inhibited the uptake of $[^3H]$-dopamine, DAT in transfected cell line was proven to be much more specific to dopamine. The psychotropic drugs such as GBR12909, CFT, normifensine, clomipramine, desipramine, and imipramine inhibited significantly the dopamine uptake in tissue culture cells stably transfected with DAT cDNA. Radioactive in situ hybridization was done to map the cellular localization of DAT mRNA-containing cells in the adult rat central nervous system. The strong hybridization signals were detected only in the substantia nigra pars compacta and ventral tegmental area. The restricted anatomical localization of DAT mRNA-containing cells confirms the DAT as a presynaptic marker of dopamine-containing cells in the rat brain.

• IMMUNE NETWORK
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• 제8권4호
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• pp.130-136
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• 2008

Background: Human cytomegalovirus UL18, a MHC class I homologue, has been considered a natural killer (NK) cell decoy. It ligates LIR-1/ILT2 (CD85j), an NK inhibitory receptor, to prevent lysis of infected target cells. However, precise role of UL18 to NK cell cytotoxicity is yet elusive. Difficulty in clarifying the function of UL18 lies in complication in detecting UL18 mainly due to low level expression of UL18 on the surface and gradual loss of its expression. Methods: To overcome this hurdle, cDNA of cytoplasmic tail-less UL18 was constructed and expressed in swine endothelial cell (SEC). The expression level and its stability in the cell surface were monitored with FACS analysis. Results: Surface expression of UL18 is up-regulated by removing cytoplasmic tail portion from UL18F (a full sequence of UL18). SECs transfected with a cDNA of UL18CY (a cytoplasmic tail-less UL18) stably expressed UL18 molecule on the surface without gradual loss of its expression during 6 week continuous cultures. In the NK cytotoxicity assay, UL18 functions either inhibiting or activating NK cell cytotoxicity according to the source of NK cells. We found that there is individual susceptibility in determining whether the engagement of NK cell and UL18 results in overall inhibiting or activating NK cell cytotoxicity. Conclusion: In this study, we found that cytoplasmic tail is closely related to the regulatory function for controlling surface expression of UL18. Furthermore, by constructing stable cell line in which UL18 expression is up-regulated and stable, we provided a useful tool to clarify exact functions of UL18 on various immune cells having ILT2 receptor.