• Macromolecular Research
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• 제13권4호
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• pp.293-296
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• 2005

Transferrin ($T_{f}$) has been used as a targeting ligand for delivering liposome/interleukin-12 (IL-12) pDNA complexes to cancer cells mostly due to the greater number of transferrin receptors ($T_{f}R$) found on tumor cells than on normal cells. $T_{f}$ was conjugated to liposomes via the reaction of MPB-PE with thiol groups of $T_{f}$ introduced by a heterobifunctional cross-linking agent, N-succinimidyl-3-(2-pyridyldithio)propionate (SPDP). Four days after C26 inoculation when the tumor volume reached ${\sim}100mm^{3}$, tumor-bearing Balb/c mice were injected intravenously with $T_{f}-liposome/IL-12 pDNA$complexes twice a week for 3 weeks. Significant suppression of tumor growth was achieved in the group treated with the $T_{f}-liposome/IL-12 pDNA$ complexes, with a dose of $10{\mu}g$ of IL-12 pDNA showing the highest suppression effect among the tested doses. Similar results were obtained when the therapy was initiated one day after tumor inoculation, although in this case $30{\mu}g$ IL-12 pDNA/$T_{f}-liposome$ complexes showed a significant suppression of tumor growth between 19 and 23 days after tumor inoculation. This result indicates that the transferrin receptor-targeted liposomal system is an efficient delivery agent of therapeutic genes, such as IL-12, in mice and that its potential clinical use warrants further research investigation.

• Asian Pacific Journal of Cancer Prevention
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• 제15권6호
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• pp.2447-2451
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• 2014

Human papillomavirus (HPV) is the primary etiologic agent of cervical cancer. Consideration of safety and non human leukocyte antigen restriction, protein vaccine has become the most likely form of HPV therapeutic vaccine, although none have so far been reported as effective. Since tumor cells consistently express the two proteins E6 and E7, most therapeutic vaccines target one or both of them. In this study, we fabricated DC vaccines by transducing replication-defective recombinant adenoviruses expressing E6/E7 fusion gene of HPV-16, to investigate the lethal effects of specific cytotoxic T lymphocytes (CTL) against CaSki cells in vitro. Mouse immature dendritic cells (DC) were generated from bone marrow, and transfected with pAd-E6/E7 to prepare a DC vaccine and to induce specific CTL. The surface expression of CD40, CD68, MHC II and CD11c was assessed by flow cytometry (FCM), and the lethal effects of CTL against CaSki cells were determined by DAPI, FCM and CCK-8 methods. Immature mouse DC was successfully transfected by pAd-E6/E7 in vitro, and the transfecting efficiency was 40%-50%. A DC vaccine was successfully prepared and was used to induce specific CTL. Experimental results showed that the percentage of apoptosis and killing rate of CaSki cells were significantly increased by coculturing with the specific CTL (p <0.05). These results illustrated that a DC vaccine modified by HPV-16 E6/E7 gene can induce apoptosis of CaSki cells by inducing CTL, which may be used as a new strategy for biological treatment of cervical cancer.

• Asian Pacific Journal of Cancer Prevention
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• 제15권16호
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• pp.6495-6497
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• 2014

Rapidly increasing number of outstanding developments in the field of TRAIL mediated signaling have revolutionized our current information about inducing and maximizing TRAIL mediated apoptosis in resistant cancer cells. Data obtained with high-throughput technologies have provided finer resolution of tumor biology and now it is known that a complex structure containing malignant cells strictly coupled with a large variety of surrounding cells constitutes the tumor stroma. Utility of mesenchymal stem cells (MSCs) as cellular vehicles has added new layers of information. There is sufficient experimental evidence substantiating efficient gene deliveries into MSCs by retroviral, lentiviral and adenoviral vectors. Moreover, there is a paradigm shift in molecular oncology and recent high impact research has shown controlled expression of TRAIL in cancer cells on insertion of complementary sequences for frequently downregulated miRNAs. In this review we have attempted to provide an overview of utility of TRAIL engineered MSCs for effective killing of tumor and potential of using miRNA response elements as rheostat like switch to control expression of TRAIL in cancer cells.

• Journal of Microbiology
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• 제41권2호
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• pp.63-72
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• 2003

Chronic inflammatory bowel disease (IBD) such as Crohn's disease or ulcerative colitis, affects around 2 in every 1000 individuals in western countries and its incidence, particularly amongst children, is increasing. IBD shows extreme morbidity with impact on all aspects of quality of life. If left untreated, IBD can lead to death. Conventional treatment of IBD involves powerful immunosuppressive chemotherapies and surgical intervention. Long-term anti-inflammatory medication is required and so patients are often subject to a spectrum of unpleasant side effects. Interleukin-10 (IL-10) is a cytokine that acts to suppress inflammation. When however administered by injection, the high levels of IL-10 that are distributed throughout the body also lead to side effects. Lactococcus lactis can be genetically engineered to secrete biologically active cytokines. When applied to the mucosa, these L. lactis can actively deliver such cytokines. By use of this principle we developed a new therapeutic approach for IBD. Administration of L. lactis that secretes murine IL-10 cures and prevents IBD in mice. The use of the engineered L. lactis gets around the problem of delivering IL-10, allowing dramatic reduction of the effective dose. A sincere concern exists about the possible dangers of uncontrolled, deliberate release of genetically modified microorganisms, such as could occur following application in healthcare. We engaged in the establishment of adequate means for biological growth control of engineered L. lactis by targeted gene exchange between thyA and hIL-10.

• IMMUNE NETWORK
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• 제6권4호
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• pp.192-198
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• 2006

Background: Investigating strategy to enhance efficiency of gene transfer via adenovirus is critical to sustain gene expression in targeted cells or tissues to regulate immune responses. However, the use of adenovirus as a gene delivery method has been limited by the native tropism of the virus. In this study, the critical parameter is to improve the efficient binding of viral particles to the plasma membrane prior to cellular uptake. Methods: Human immunodeficiency virus (HIV-1) trans-acting activator of transcription (TAT), a protein transduction domain, was fused to the ectodomain of the coxsackie-adenovirus receptor (CAR). The CAR-TAT protein was produced from a Drosophila Schneider 2 cells (S2) transfected with CAR-TAT genes. The function of CARTAT was analyzed the efficiency of adenoviral gene transfer by flow cytometry, and then immunizing AdVGFP with CAR-TAT was transduced on dendritic cells (DCs). Results: S2 transfectants secreting CAR-TAT fusion protein has been stable over a period of 6 months and its expression was verified by western blot. Addition of CAR-TAT induced higher transduction efficiency for AdVGFP at every MOI tested. When mice were vaccinated with DC of which adenoviral transduction was mediated by CAR-TAT, the number of IFN-${\gamma}$ secreting T-cells was increased as compared with those DCs transduced without CAR-TAT. Conclusion: Our data provide evidence that CAR-TAT fusion protein enhances adenoviral transduction and immunogenecity of transgenes on DCs and may influence on the development of adenoviral-mediated anti-tumor immunotherapy.

• 대한핵의학회지
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• 제38권3호
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• pp.253-258
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• 2004

목적: 약물이나 유전자 전달에 이용되는 생체적합성이 높은 키토산의 간세포 지향성을 위해서 갈락토스를 수식하는 방법이 널리 이용되어지고 있다. 이번 연구에서는 갈락토스 수식 키토산의 간세포지향성 획득을 평가하는데 있어서 핵의학 영상법의 유용성에 대해 알아보고자 하였다. 대상 및 방법: 키토산에 NHS와 EDC를 이용하여 30 mol%의 lactobionic acid를 결합시켜 갈락토스 수식 키토산을 제조하였다. 얻어진 갈락토스 수식 키토산을 투석 시킨 후 동결 건조하여 얻어 낸 다음 키토산의 기본구조인 glucoseamine에 methyl기를 첨가시키는 반응을 하여 최종화합물을 합성하였다. GMC의 세포내 독성은 MTT assay를 통하여 확인하였다. 제조된 GMC에 $SnCl_2{\cdot}2H_2O$를 이용하여 $^{99m}Tc$을 표지하였다. 표지 후 안정성은 아세톤과 생리식염수를 이용하여 1시간까지 확인하였다. $^{99m}Tc$-GMC와 $^{99m}Tc$-MC 55.5 MBq (1.5 mCi)를 토끼의 외이정맥으로 주사 후, 감마카메라를 이용하여 10분, 30분, 60분, 90분 간격으로 전면 영상을 얻었다. 결과: 키토산에 lactobionic acid 30 mol%를 반응시켜 7.4 mol%의 galactose group이 키토산에 결합한 것을 확인하였다. 갈락토스 수식 키토산을 메틸화한 결과는 tri, di, mono가 각각 8.8%, 46%, 35.2%인 것을 확인하였다. MTT assay결과를 통해 GMC의 세포에 미치는 독성은 거의 없는 것을 확인할 수 있었다. 표지 효율은 메틸화시키지 않은 $^{99m}Tc$-GC가 아세톤과 생리식염수에서 각각 88%, 72%를 보인 반면에 $^{99m}Tc$-GMC는 96%정도를 보여 보다 높은 표지효율을 가지는 것을 확인하였다. $^{99m}Tc$-MC를 주사후 얻은 토끼 영상에서 키토산은 일반적으로 대부분 신장을 통해 배출되며, 간과 비장, 골격계에는 매우 적은 분포를 보이는데 비해. 갈락토스가 수식된 $^{99m}Tc$-GMC에서는 분포에 변화가 생겨 간, 신장, 그리고 방광에서 높은 방사능이 관찰되는 소견을 보였다. 결론: 핵의학 영상법은 갈락토스 리간드 수식 키토산의 간세포 지향성 여부를 평가하기 위한 생체내 평가법으로 이용될 수 있을 것으로 사료되었다.

• Childhood Kidney Diseases
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• 제24권1호
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• pp.1-13
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• 2020

Immunoglobulin (Ig)A vasculitis nephritis (IgAVN), also referred to as Henoch-Schönlein purpura nephritis, is a relatively benign disease in children. However, two 24-year European cohort studies have reported high sustained rates of hypertension, severe proteinuria, and renal dysfunction in patients with IgAVN. Notably, the incidence and exacerbation rates of proteinuria, hypertension, and renal dysfunction during pregnancy were high even in women who recovered from IgAVN before pregnancy. Patients with IgAVN need lifelong care. Trials have been performed to investigate early biomarkers and genes associated with poor prognosis to identify high-risk patients in whom IgAVN may progress to severe renal disease. Urinary IgA/cr, IgM/cr levels, and HLAB35 and angiotensinogen gene expression were shown to be predictors of progression of IgAVN to severe renal dysfunction. The 2019 Single Hub and Access point for paediatric Rheumatology in Europe (SHARE) initiative group published guidelines for pediatric IgAVN, following the Kidney Disease: Improving Global Outcomes (KDIGO) guidelines established in 2012. Compared with the KDIGO guidelines, the SHARE guidelines recommend earlier corticosteroid administration in cases of mild proteinuria (>0.5 g/d). Clinical trials of targeted budesonide delivery to the distal ileum, monoclonal antibody targeting C5, eculizumab and anti-CD20 monoclonal antibody administration, among others are currently underway in patients with IgA nephropathy. It is expected that newer therapeutic agents would become available for IgAVN in the near future. This review summarizes IgAVN with emphasis on recently published literature, including possible preventive strategies, predictive biomarkers for progression of IgAVN, and various treatments.

• Molecules and Cells
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• 제41권2호
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• pp.119-126
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• 2018

microRNA (miR)-612 shows anticancer activity in several types of cancers, yet its function in melanoma is still unclear. This study was undertaken to investigate the expression of miR-612 and its biological relevance in melanoma cell growth, invasion, and tumorigenesis. The expression and prognostic significance of miR-612 in melanoma were examined. The effects of miR-612 overexpression on cell proliferation, colony formation, tumorigenesis, and invasion were determined. Rescue experiments were conducted to identify the functional target gene(s) of miR-612. miR-612 was significantly downregulated in melanoma tissues compared to adjacent normal tissues. Low miR-612 expression was significantly associated with melanoma thickness, lymph node metastasis, and shorter overall, and disease-free survival of patients. Overexpression of miR-612 significantly decreased cell proliferation, colony formation, and invasion of SK-MEL-28 and A375 melanoma cells. In vivo tumorigenic studies confirmed that miR-612 overexpression retarded the growth of A375 xenograft tumors, which was coupled with a decline in the percentage of Ki-67-positive proliferating cells. Mechanistically, miR-612 targeted Espin in melanoma cells. Overexpression of Espin counteracted the suppressive effects of miR-612 on melanoma cell proliferation, invasion, and tumorigenesis. A significant inverse correlation (r = -0.376, P = 0.018) was observed between miR-612 and Espin protein expression in melanoma tissues. In addition, overexpression of miR-612 and knockdown of Espin significantly increased the sensitivity of melanoma cells to doxorubicin. Collectively, miR-612 suppresses the aggressive phenotype of melanoma cells through downregulation of Espin. Delivery of miR-612 may represent a novel therapeutic strategy against melanoma.

• Ultrasonography
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• 제32권1호
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• pp.59-65
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• 2013

목적: 본 연구는 초음파 조영제를 직접 합성하고 그 크기 및 소멸 시간 등의 특성을 분석하며, 기존의 초음파 조영제와의 영상을 비교하여 앞으로 초음파 조영제를 매개로 한 영상 유도 하 약물 및 유전자 치료제의 전달 시스템 구축의 기본을 갖추고자 한다. 대상 및 방법: 초음파 조영제는 21마이크로 몰의 DPPC(1, 2-Dihexadecanoyl-sn-glycero-3-phosphocholine, $C_{40}H_{80}NO_8P$), 9 마이크로 몰의 콜레스테롤, 1.9 마이크로 몰의 DCP(Dihexadecylphosphate, $[CH_3(CH_2)_{15}O]_2P(O)OH$) 및 클로로포름 및 SF6 기체를 이용하여 합성하였다. 약 12-24시간의 동결건조 후에 초음파 조영제는 SF6 기체와 초음파 처리(sonication) 법을 이용하여 합성하였고 그 초음파 조영제의 크기는 압출기를 이용하여 일정한 크기로 조정하였다. 합성된 초음파 조영제는 동적 광산란 측정법(dynamic light scattering measurement)을 이용하여 그 크기를 분석하였다. 한편 합성한 초음파 조영제의 소멸 양상을 평가하기 위하여 광학 현미경을 이용하여 그 숫자를 합성 직후, 12시간, 24시간, 36시간, 48시간, 60시간, 72시간 및 84시간 후에 평가하였다. 초음파 조영제로서의 효과를 평가하기 위하여 모형을 만들어 현재 임상적으로 사용되고 있는 초음파 조영제와 그 에코를 비교 분석하였다. 결과: 초음파 조영제는 동결건조-초음파 처리 방법을 통하여 성공적으로 합성 할 수 있었다. 합성된 초음파 조영제는 154.2 nm의 정점 (peak)에서 가장 많이 분포하였으며 합성 후 24시간부터는 입자의 숫자들이 감소하는 양상을 보였다. 소노뷰와 합성된 초음파 조영제의 에코밝기는 유의한 차이를 보이지 않았으나 소노뷰와 생리식염수 및 합성된 초음파 조영제와 생리식염수의 에코 밝기는 통계적으로 유의한 차이를 보였다 (p < 0.01). 결론: 본 연구를 통하여 초음파 조영제를 직접 합성할 수 있는 방법을 습득할 수 있었고 그 크기 및 특성을 분석할 수 있었다.