• Asian Pacific Journal of Cancer Prevention
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• 제14권9호
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• pp.4983-4988
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• 2013

Objectives: To establish a taxol-resistant cell line of human ovarian carcinoma (A2780/Taxol) and investigate its biological features. Methods: The drug-resistant cell line (A2780/Taxol) was established by continuous stepwise selection with increasing concentrations of Taxol. Cell morphology was assessed by microscopy and growth curves were generated with in vitro and in vivo tumor xenograft models. With rhodamine123 (Rh123) assays, cell cycle distribution and the apoptotic rate were analyzed by flow cytometry (FCM). Drug resistance-related and signal associated proteins, including P-gp, MRPs, caveolin-1, PKC-${\alpha}$, Akt, ERK1/2, were detected by Western blotting. Results: A2780/Taxol cells were established with stable resistance to taxol. The drug resistance index (RI) was 430.7. Cross-resistance to other drugs was also shown, but there was no significant change to radioresistance. Compared with parental cells, A2780/Taxol cells were significantly heteromorphous, with a significant delay in population doubling time and reduced uptake of Rh123 (p<0.01). In vivo, tumor take by A2780 cells was 80%, and tumor volume increased gradually. In contrast, with A2780/Taxol cells in xenograft models there was no tumor development. FCM analysis revealed that A2780/Taxol cells had a higher percentage of G0/G1 and lower S phase, but no changes of G2 phase and the apoptosis rate. Expression of P-gp, MRP1, MRP2, BCRP, LRP, caveolin-1, PKC-${\alpha}$, Phospho-ERK1/2 and Phospho-JNK protein was significantly up-regulated, while Akt and p38 MARK protein expression was not changed in A2780/Taxol cells. Conclusion: The A2780/Taxol cell line is an ideal model to investigate the mechanism of muti-drug resistance related to overexpression of drug-resistance associated proteins and activation of the PKC-${\alpha}/ERK$ (JNK) signaling pathway.

• Asian Pacific Journal of Cancer Prevention
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• 제13권7호
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• pp.3471-3476
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• 2012

Background and Objective: Histone deacetylase (HDAC) inhibitors represent a promising class of potential anticancer agents for treatment of human malignancies. In this study, we investigated the effect of trichostatin A (TSA), one such HDAC inhibitor, in combination with docetaxel (TXT), a cytotoxic chemotherapy agent or erlotinib, a novel molecular target therapy drug, on lung cancer A549 cells. Methods: A549 cells were treated with TXT, erlotinib alone or in combination with TSA, respectively. Cell viability, apoptosis, and cell cycle distribution were evaluated using MTT (3- (4, 5-dimethylthiazol-2-yl) -2, 5-diphenyltetrazolium bromide) assay, Hochst33258 staining and flow cytometry. Moreover, immunofluorescent staining and Western blot analysis were employed to examine alterations of ${\alpha}$-tubulin, heat shock protein 90 (hsp90), epidermal growth factor receptor (EGFR), and caspase-3 in response to the different exogenous stimuli. Results: Compared with single-agent treatment, co-treatment of A549 cells with TSA/TXT or TSA/erlotinib synergistically inhibited cell proliferation, induced apoptosis, and caused cell cycle delay at the $G_2/M$ transition. Treatment with TSA/TXT or TSA/erlotinib led to a significant increase of cleaved caspase-3 expression, also resulting in elevated acetylation of ${\alpha}$-tubulin or hsp90 and decreased expression of EGFR, which was negatively associated with the level of acetylated hsp90. Conclusions: Synergistic anti-tumor effects are observed between TXT or erlotinib and TSA on lung cancer cells. Such combinations may provide a more effective strategy for treating human lung cancer.

• Journal of Radiation Protection and Research
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• 제28권3호
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• pp.233-237
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• 2003

It has been known that ATM plays a central role in response of cells to ionizing radiation by enhancing DNA repair. We have investigated the feasibility of increasing radiosensitivity of tumor cells with the use of ATM inhibitors such as caffeine, pentoxifylline and wortmannin. Human colorectal cancer RKO.C cells and RKO-ATM cells (RKO cells overexpressing ATM) were used in the present study. The clonogenic cell survival in vitro indicated that RKO-ATM cells were markdely radioresistant than RKO.C cells. Treatment with 3 mM of caffeine significantly increased the radiosensitivity of cells, particulary the RKO-ATM cells, so that the radiosensitivity of RKO.C cells and RKO-ATM cells were almost similar. The radiation induced G2/M arrest in RKO-ATM cells was noticeably longer than that in RKO.C cells and caffeine treatment significantly reduced the length of the radiation induced G2/M arrest in both RKO.C and RKO-ATM cells. Pentoxifylline and wortmannin were also less effective than caffeine to radiosensitize RKO.C or RKO-ATM cells. However, wortmannin was more effective than caffeine against human lung adenocarcinoma A549 cells indicating the efficacy of ATM inhibitor to increase radiosensitivity is cell line dependent. For in vivo study, RKO.C cells were injected s.c. into the hind-leg of BALB/C-nuslc nude mice, and allowed to grow to 130mm3 tumor. The mice were i.p. injected with caffeine solution or saline and the tumors irradiated with 10 Gy of X-rays. The radiation induced growth delay was markedly increased by 1-2 mg/g of caffeine. It was concluded that caffeine increases radiosensitivity of tumor cells by inhibiting ATM kinase function, thereby inhibiting DNA repair, that occurs during the G2/M arrest after radiation.

• Asian Pacific Journal of Cancer Prevention
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• 제16권2호
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• pp.627-633
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• 2015

Background: To study the effect of parecoxib, a novel cyclooxygenase-2 selective inhibitor, on the radiation response of colorectal cancer (CRC) cells and its underlying mechanisms. Materials and Methods: Both in vitro colony formation and apoptosis assays as well as in vivo mouse xenograft experiments were used to explore the radiosensitizing effects of parecoxib in human HCT116 and HT29 CRC cells. Results: Parecoxib sensitized CRC cells to radiation in vitro with a sensitivity enhancement ratio of 1.32 for HCT116 cells and 1.15 for HT29 cells at a surviving fraction of 0.37. This effect was partially attributable to enhanced apoptosis induction by parecoxib combined with radiation, as illustrated using an in vitro apoptosis assays. Parecoxib augmented the tumor response of HCT116 xenografts to radiation, achieving growth delay more than 20 days and an enhancement factor of 1.53. In accordance with the in vitro results, parecoxib combined with radiation resulted in less proliferation and more apoptosis in tumors than radiation alone. Radiation monotherapy decreased microvessel density (MVD) and microvessel intensity (MVI), but increased the hypoxia level in xenografts. Parecoxib did not affect MVD, but it increased MVI and attenuated hypoxia. Conclusions: Parecoxib can effectively enhance radiation sensitivity in CRC cells through direct effects on tumor cells and indirect effects on tumor vasculature.

• Radiation Oncology Journal
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• 제18권4호
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• pp.329-336
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• 2000

목적 : 마우스 간암에서 방사선과 각종 항암제와의 복합요법을 시행하여, 방사선 감수성을 증가시킬 수 있는 약물을 탐색하고자 하였다. 방법 : C3H/HeJ마우스에 마우스 간암인 HCa-1을 이식하고, 평균 직경 8 mm에 이르렀을 때, 방사선 조사(25 Gy), 항암 약물(5-Fu, 150 mg/kg; adriamycln, 8 mg/kg; paclltaxel, 40 mg/kg; gemcltablne, 50 mg/kg), 또는 방사선과 항암 약물의 복합 치료를 시행하였다 치료에 대한 종양 반응은 종양 성장 지연과 항진 요인으로 분석하였다. 항진 효과를 보인 약물에 대하여 그 기전 연구는 조직 절편에서 apoptotic 수준을 평가하고, 또한 조절물질의 발현을 분석하였다. p53, Bcl-2, Bax, Bcl-XL, Bcl-XS, p21$^{WAF1/CIP1}$의 발현 분석은 westeblotting으로 하였다. 결과 : Gemcltabine 만이 방사선 감수성을 증가시키는 것으로 나타났다(항진요인:1.6). Gemcltabine과 방사선의 복합 치료는 apoptosis 유도에서는 부가적 수준만을 보였다. 조절울질의 발현 양상은 방사선 단독에 비하여 방사선과 gemcitabine의 병용시 p21$^{WAF1/CIP1}$의 증가가 유의하게 관찰되었다. 결론 : Gemcitabiue은 마우스 간암에서 방사선 감수성을 증가시키는 것으로 나타났다. 이를 조절하는 요소로서 p21$^{WAF1/CIP1}$ 이 관여할 것으로 생각 된다.

• Radiation Oncology Journal
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• 제25권4호
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• pp.233-241
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• 2007

목적: EGFR, HER2 과발현 인간 유방암 세포를 이용한 종양이식 마우스에서 EGFR/HER2 복합 Tyrosine Kinase 억제제인 GW572016이 방사선반응성에 미치는 영향을 알아보고 종양조직의 EGFR/HER2수용체 억제효과 및 EGFR down stream signal pathway 단백인 ERK 1/2, PI3k/Akt 억제효과를 알아보고자 하였다. 대상 및 방법: SUM 102와 SUM 149 EGFR 과발현 세포와 SUM 185, SUM 225 HER2 과발현 세포를 우측 옆구리 피하에 접종하여 종양이식마우스를 만들었다. 이식마우스는 2군으로 나누어 한 군은 GW572016에 의한 EGFR/HER2 수용체 억제와 down stream signal 단백의 활성 변화를 Immunoprecipitation과 Western blot의 방법을 사용하여 관찰하였고 다른 한군은 GW572016에 의한 방사선감수성 변화를 알아보기 위해 1) 대조군, 2) GW572016 단독군, 3) 방사선단독군, 4) GW572016+방사선병용투여군으로 나누어 종양성장을 비교 관찰하였다. GW572016에 의해서 SUM 149, SUM 185이식종양에서 EGFR및 HER2 수용체의 활성이 억제되었으며 특히 SUM 185, HER2 과발현 이식종양에서는 ERK 1/2 down stream 단백의 활성도 억제되었다 SUM 225 HEH2 과발현 이식종양에서는 이전의 in vitro실험에서와 달리 GW572016에 의해 HER2수용체의 활성변화가 없었으나 ERK 1/2, Akt의 활성은 모두 억제되었다. GW572016에 의해 SUM 149과 SUM 185에서 종양성장억제효과가 관찰되었고 특히 SUM 149에서는 GW572016과 방사선치료병용군에서 종양성장억제효과가 좀더 뚜렷하여 방사선감수성을 증가시키는 것으로 생각되었다. 결 론: GW572016은 EGFR 혹은 HER2 과발현 유방암세포에서 EGFR/HER2 수용체 억제와 down stream signal 단백의 활성을 억제시켰으며 SUM 149에서는 방사선감수성을 증가시키는 것으로 생각된다. 향후 EGFR을 표적으로 하는 억제제치료에서 EGFR 수용체억제뿐 아니라 down stream 단백의 활성억제 여부가 방사선 감수성 및 저항성의 극복과 관련이 있으리라는 근거를 설명할 수 있으며 향후 좀더 깊이 있는 연구가 필요하다.

• Biomolecules & Therapeutics
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• 제27권3호
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• pp.283-289
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• 2019

Brain aging induces neuropsychological changes, such as decreased memory capacity, language ability, and attention; and is also associated with neurodegenerative diseases. However, most of the studies on brain aging are focused on neurons, while senescence in astrocytes has received less attention. Astrocytes constitute the majority of cell types in the brain and perform various functions in the brain such as supporting brain structures, regulating blood-brain barrier permeability, transmitter uptake and regulation, and immunity modulation. Recent studies have shown that SIRT1 and SIRT2 play certain roles in cellular senescence in peripheral systems. Both SIRT1 and SIRT2 inhibitors delay tumor growth in vivo without significant general toxicity. In this study, we investigated the role of tenovin-1, an inhibitor of SIRT1 and SIRT2, on rat primary astrocytes where we observed senescence and other functional changes. Cellular senescence usually is characterized by irreversible cell cycle arrest and induces senescence- associated ${\beta}$-galactosidase (SA-${\beta}$-gal) activity. Tenovin-1-treated astrocytes showed increased SA-${\beta}$-gal-positive cell number, senescence-associated secretory phenotypes, including IL-6 and IL-$1{\beta}$, and cell cycle-related proteins like phospho-histone H3 and CDK2. Along with the molecular changes, tenovin-1 impaired the wound-healing activity of cultured primary astrocytes. These data suggest that tenovin-1 can induce cellular senescence in astrocytes possibly by inhibiting SIRT1 and SIRT2, which may play particular roles in brain aging and neurodegenerative conditions.

• Physical Therapy Rehabilitation Science
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• 제8권1호
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• pp.22-31
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• 2019

Objective: For knee osteoarthritis (OA), there is a demand for alternative modalities in order to delay surgery and to avoid the side effects of medications. This study compared the effects of applying seaweed pack and mudpack for the treatment of knee OA. Design: Randomized controlled trial. Methods: Twenty-five patients with knee OA who satisfied the criteria were included. The patients were divided into two groups according to the treatment method: mudpack (n=12) and seaweed pack (n=13). The two groups were treated for 20 minutes, twice a day for five continuous days at the Ocean Healing Center at Wando Island, South Korea. Participants were assessed by clinical scores (Western Ontario and McMaster Universities Osteoarthritis Index, Hospital for Special Surgery Knee Score, Knee injury and Osteoarthritis Outcome Score and 36-Item Short Form Health Survey) and lab results (erythrocyte sedimentation rate, C-reactive protein, insulin-like growth factor-1 [IGF-1], tumor necrosis $factor-{\alpha}$ [$TNF-{\alpha}$]) during the follow-up period. Results: For the most part, clinical scores improved after therapy and maintained improvements for four weeks in both groups (p<0.05). In the seaweed group, $TNF-{\alpha}$ was significantly decreased at two weeks post-therapy (p<0.05). In both groups, IGF-1 was significantly increased immediately post-therapy (p<0.05). There were no statistically significant differences after therapy between the groups in clinical scores and labs. Conclusions: Seaweed packs and mudpacks had similar positive effects for knee OA. Additionally, the seaweed pack group showed decreased levels of $TNF-{\alpha}$ at two weeks post-treatment, which may explain the reduced inflammatory reaction. For rehabilitation therapy, use of seaweed packs may serve as an alternative modality for the treatment of knee OA.

• Radiation Oncology Journal
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• 제21권3호
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• pp.207-215
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• 2003

목적: Extracellular signal-regulated kinase (ERK)는 mitogen-activated protein kinase cascade의 일원으로 다양한 세포독성 자극에 의해 유도되는 apoptosis에 반대되는 역할을 한다. 따라서 ERK의 억제는 항암제로서 유용하게 사용될 것으로 생각되어진다. 대상 및 방법: 마우스 간암인 HCa-I는 TCD50가 80 Gy 이상으로 강한 방사선 내성종양으로 알려져 있으며, 방사선 민감성의 증진을 위해 다양한 항암제가 실험되었으나 뚜렷한 효과를 나타내지 못했다. 이 실험을 통해 in vivo,, 특히 방사선 내성종양에서 ERK의 억제가 방사선에 의한 항암 작용을 증진시키는지 알아보고자 하였다. C3H/HeJ 마우스에 종양의 크기가 $7.5\~8\;mm$가 되었을 때 PD98059 ($0.16\;\mug/50\;\mul$로 종양에 직접 주사)를 처리하였다. 결과: 처리 1시간째에 p-ERK가 0.5배로 억제되었다. 종양 성장 지연 분석에서 증강 지수가 전 처리군과 후 처리군에서 각각 1.6과 1.87로 PD98059가 종양의 방사선 감수성을 증가시키는 것으로 관찰되었다. 25 Gy 방사선과 PD98059 복합처리 시 apoptosis가 크게 증가되었다. 각 실험군의 apoptosis 최대치는 방사선 조사군에서 $1.4\%$, PD98059 처리군에서 $0.9\%$ 복합처리군의 전 처리군과 후 처리군에서 각각 $4.9\%\;5.3\%$를 나타냈다. Apoptosis 조절 물질의 변화는, p53의 발현이 복합 처리군에서 PD98059 전 처리군과 후 처리군 모두에서 24시간까지 대조군에 비해 2.7배, 3.2배의 높은 발현 수준을 유지하여 처리 1시간째부터 발현 증가를 하여 24시간까지 지속되는 것이 관찰되었다. $p21^{WAF1/CIP1}$의 발현은 p53 발현 변화와 유사한 양상으로 특히 PD98059 후 처리군에서 방사선 조사군이나 PD98059 전 처리군과 비교하여 높은 발현수준을 보였으며, 24시간까지 3.2배의 높은 발현 수준을 유지하는 것으로 나타났다. Bcl-Xs는 25 Gy 방사선 조사군이나 PD98059 처리군에서는 뚜렷한 변화를 보이지 않았으나 복합 처리군중 전 처리군에서 4시간 째 대조군에 비해 1.93배 증가를 보였으며, 후 처리군에서는 1시간 후에 1.83배의 증가를 보였다. 모든 실험군에서 Bcl-2, $Bcl-X_L$, BaX는 뚜렷한 발현 변화를 보이지 않았다. 결론: 방사선 내성 종양인 간암에 MEK 억제제를 방사선 조사와 복합 처리하여 방사선 감수성을 향상시켜 치료 효율의 상승을 유도 할 수 있을 것으로 생각된다.

• 한국생물물리학회:학술대회논문집
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• 한국생물물리학회 2003년도 정기총회 및 학술발표회
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• pp.80-80
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• 2003

${\beta}$-lapachone(${\beta}$-Lap), a natural o-naphthoquinone, presents in the bark of the Lapacho tree. ${\beta}$-Lap is cytotoxic against a variety of human cancer cells and it potentiates the anti-tumor effect of Taxol. In addition, ${\beta}$-Lap has been reported to radiosensitize cancer cells by inhibiting the repair of radiation-induced DNA damage.In the present study, we investigated the cytotoxicity of ${\beta}$-Lap against RKO human colorectal cancer cells as well as the combined effect of ${\beta}$-LaP and ionizing radiation. An incubation of RKO cells with 5 ${\mu}$M of ${\beta}$-Lap for 4 h killed almost 90％ of the clonogenic cells. An incubation of RKO cells with 5 ${\mu}$M of ${\beta}$-Lap for 4 h or longer also caused massive apoptosis. Unlike other cytotoxic agents, ${\beta}$-Lap did not increase the expression of p53 and p21 and it suppressed the NFkB expression. The expression of Caspase 9 and 3 was minimally altered by ${\beta}$-Lap. Radiation and ${\beta}$-Lap acted synergistically in inducing clonogenic cell death and apoptosis in RKO cells when ${\beta}$-Lap treatment was applied after but not before the radiation exposure of the cells. Interestingly, a 4 h treatment with 5 ${\mu}$M of ${\beta}$-Lap starting 5 h after irradiation was as effective as that starting immediately after irradiation. The mechanisms of ${\beta}$-Lap-induced cell killing is controversial but a recent hypothesis is that ${\beta}$-Lap is activated by NAD(P)H: quinone-onidoreductase (NQO1) in the cells followed by an elevation of cytosolic Ca$\^$2+/ level and activation of proteases leading to apoptosis. It has been reported that NQO1 level in cells is markedly up-regulated for longer than 10 h after irradiation. Indeed, using immunological staining of NQO1, we observed a significant elevation of NQO1 expression in RKO cells 5h after 2-4 Gy irradiation. Such a prolonged elevation of NQO1 level after irradiation may be the reasons why the ${\beta}$-Lap treatment applied S h after irradiation was as effective as that applied immediately after irradiation in killing the cells. In view of the fact that the repair of radiation-induced damage is usually completed within 1-2 h after irradiation, it is highly likely that the ${\beta}$-Lap treahment applied 5 h after irradiation could not inhibit the repair of radiation-induced damage. For in vivo study, RKO cells were injected S.C. into the hind-leg of Nu/Nu mice, and allowed to grow to 130 mm3 tumor. The mice were i.p. injected with ${\beta}$-lapachone or saline 2 h after irradiation of tumors with 10 Gy of X-rays. The radiation induced growth delay was increased by 2.4 $\mu\textrm{g}$/g of ${\beta}$-lapachone. Taken together, we may conclude that the synergistic interaction of radiation and ${\beta}$-Lap in killing cancer cells is not due to radiosensitization by ${\beta}$-Lap but to an enhancement of ${\beta}$-Lap cytotoxicity by radiation through an upregulation of NQO1. The fact that NQO1 is elevated in tumors and that radiation causes prolonged increase of the NQO1 expression may be exploited to preferentially kill tumor cells using ${\beta}$-Lap in combination with radiotherapy.