• Asian Pacific Journal of Cancer Prevention
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• v.15 no.17
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• pp.7037-7041
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• 2014

Background: 5-Fluorouracil (5-FU) is the most commonly used drug in colon cancer therapy. However, despite impressive clinical responses initially, development of drug resistance to 5-Fu in human tumor cells is the primary cause of failure of chemotherapy. In this study, we established a 5-Fu-resistant human colon cancer cell line for comparative chemosensitivity studies. Materials and Methods: Real time PCR and Western blotting were used to determine gene expression levels. Cell viability was measured by MTT assay. Glucose uptake was assess using an Amplex Red Glucose/Glucose Oxidase assay kit. Results: We found that 5-Fu resistance was associated with the overexpression of Glut1 in colon cancer cells. 5-Fu treatment at low toxic concentration induced Glut1 expression. At the same time, upregulation of Glut1 was detected in 5-Fu resistant cells when compared with their parental cells. Importantly, inhibition of Glut1 by a specific inhibitor, WZB117, significantly increased the sensitivity of 5-Fu resistant cells to the drug. Conclusions: This study provides novel information for the future development of targeted therapies for the treatment of chemo-resistant colon cancer patients. In particular it demonstrated that Glut1 inhibitors such as WZB117 may be considered an additional treatment options for patients with 5-Fu resistant colon cancers.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.2
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• pp.629-635
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• 2014

Background: Acute myeloid leukemia (AML) is a fatal hematological malignancy which is resistant to a variety of chemotherapy drugs. Myeloid cell leukemia-1 (Mcl-1), a death-inhibiting protein that regulates apoptosis, has been shown to be overexpressed in numerous malignancies. In addition, it has been demonstrated that the expression level of the Mcl-1 gene increases at the time of leukemic relapse following chemotherapy. The aim of this study was to target Mcl-1 by small interference RNA (siRNA) and analyze its effects on survival and chemosensitivity of acute myeloid leukemia cell line HL-60. Materials and Methods: siRNA transfection was performed with a liposome approach. The expression levels of mRNA and protein were measured by real-time quantitative PCR and Western blot analysis, respectively. Trypan blue assays were performed to evaluate tumor cell growth after siRNA transfection. The cytotoxic effects of Mcl-1 siRNA (siMcl-1) and etoposide were determined using MTT assay on their own and in combination. Apoptosis was quantified using a DNA-histone ELISA assay. Results: Transfection with siMcl-1 significantly suppressed the expression of Mcl-1 mRNA and protein in a time-dependent manner, resulting in strong growth inhibition and spontaneous apoptosis. Surprisingly, pretreatment with siMcl-1 synergistically enhanced the cytotoxic effect of etoposide. Furthermore, Mcl-1 down-regulation significantly increased apoptosis sensitivity to etoposide. No significant biological effects were observed with negative control siRNA treatment. Conclusions: Our results suggest that specific suppression of Mcl-1 by siRNA can effectively induce apoptosis and overcome chemoresistance of leukemic cells. Therefore, siMcl-1 may be a potent adjuvant in leukemia chemotherapy.

• Tuberculosis and Respiratory Diseases
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• v.62 no.1
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• pp.33-42
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• 2007

Background: Heme oxygenase-1 (HO-1) is known to modulates the cellular functions, including cell proliferation and apoptosis. It is known that a high level of HO-1 expression is found in many tumors, and HO-1 plays an important role in rapid tumor growth on account of its antioxidant and antiapoptotic effects. Cisplatin is a widely used anti-cancer agent for the treatment of lung cancer. However, the development of resistance to cisplatin is a major obstacle to its use in clinical treatment. We previously demonstrated that inhibiting HO-1 expression through the transcriptional activation of Nrf2 induces apoptosis in A549 cells. The aim of this study was to determine of the inhibiting HO-1 enhance the chemosensitivity of A549 cells to cisplatin. Materials and Methods: The human lung cancer cell line, A549, was treated cisplatin, and the cell viability was measured by a MTT assay. The change in HO-1, Nrf2, and MAPK expression after the cisplatin treatment was examined by Western blotting. HO-1 inhibition was suppressed by ZnPP, which is a specific pharmacologic inhibitor of HO activity, and small interfering RNA (siRNA). Flow cytometry analysis and Western blot were performed in to determine the level of apoptosis. The level of hydrogen peroxide ($H_2O_2$) generation was monitored fluoimetrically using 2',7'-dichlorofluorescein diacetate. Results: The A549 cells showed more resistance to the cisplatin treatment than the other cell lines examined, whereas cisplatin increased the expression of HO-1 and Nrf2, as well as the phosphorylation of MAPK in a time-dependent fashion. Inhibitors of the MAPK pathway blocked the induction of HO-1 and Nrf2 by the cisplatin treatment in A549 cells. In addition, the cisplatin-treated A549 cells transfected with dither the HO-1 small interfering RNA (siRNA) or ZnPP, specific HO-1 inhibitor, showed in a more significantly decrease in viability than the cisplatin-only-treated group. The combination treatment of ZnPP and cisplatin caused in a marked increase in the ROS generation and a decrease in the HO-1 expression. Conclusion: Cisplatin increases the expression of HO-1, probably through the MAPK-Nrf2 pathway, and the inhibition of HO-1 enhances the chemosensitivity of A549 cells to cisplatin.

• Tuberculosis and Respiratory Diseases
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• v.44 no.3
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• pp.525-533
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• 1997

Background : No ideal combination chemotherapy for lung cancer has been established even though lots of combination anticancer chemotherapies have been tried. For the combination of anticancer drugs, the interaction of anticancer drugs is very important but unpredictable factor. In this experiment, we designed and tested new experiment to measure the interaction of two anticancer drugs using MIT assay in an attempt to predict clinical response of the combination regimen. Methods : With human lung adenocarcinoma cell line (PC-14), the cytotoxic effect of cisplatin, adriamycin, mitomycin C and etoposide were measured by in vitro chemosensitivity test (MIT assay). The combined cytotoxic effects of combination of two drugs were also measured in every combination of the drug concentrations and analyzed the interaction by Anava analysis of two way factorial design. Results : Four individual drugs showed cytotoxic effects on PC-14 by dose dependent fashion. Comparison of two drug combinations revealed that mitomycin C + cisplatin and adriamycin + cisplatin combinations showed stronger synergistic cytotoxic effects. Conclusion : From this experiment, we suggest two combinations of mitomycin C + cisplatin and adriamycin + cisplatin as chemotherapeutic regimens for unresectable non-small cell lung cancer. Furthermore, this experimental design could be applied to other types of cancer requiring combination anticancer chemotherapy.

• Journal of Korean Academy of Oral and Maxillofacial Radiology
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• v.27 no.2
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• pp.91-103
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• 1997

The purpose of this study was to aid in the prediction of tumor cell tolerance to radiotherapy and/or chemotherapy. For this study, cell surviving curves were obtained for human epidermoid carcinoma A-253 cell line using semiautomated MTT assay. 2,4,6,8,10 Gy were irradiated at a dose rate of 210 cGy/min using /sup 60/Co Irradiator ALDORADO 8. After irradiation, A-253 cell lines(2×10⁴cells/mil were exposed to bleomycin or cisplatin for 1 hour. The viable cells were determined for each radiation dose with/without 2 /lg/mi of drug at the 3rd day. And they were compared to control values. The results were obtained as follows : 1. The surviving curve with gentle slope was obtained after irradiation of 2, 4, 6, 8, 10 Gy on A-253 cell line. 2. The cytotoxicity of bleomycin or cisplatin at the concentration of 2㎍/ml was great on A-253 cell line. But, there was no significant difference between the cytotoxicity of bleomycin and that of cisplatin. 3. There were significant differences of surviving fractions after irradiation with 2㎍/mi of bleomycin compared with irradiation only on A-253 cell line. 4. There were significant differences of surviving fractions after irradiation with 2㎍/ml of cisplatin compared with irradiation only on A-253 cell line. 5. There were no significant differences of surviving fractions between the groups of irradiation with bleomycin and the groups of irradiation with cisplatin on A-253 cell line.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.9
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• pp.4807-4814
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• 2012

Purpose: The chemoresistance of human hepatocellular carcinoma (HCC) to cytotoxic drugs, especially intrinsic or acquired multidrug resistance (MDR), still remains a major challenge in the management of HCC. In the present study, possible mechanisms involved in MDR of HCC were identified using a 5-fluorouracil (5-FU)-resistant human HCC cell line. Methods: BEL-7402/5-FU cells were established through continuous culturing parental BEL-7402 cells, imitating the pattern of chemotherapy clinically. Growth curves and chemosensitivity to cytotoxic drugs were determined by MTT assay. Doubling times, colony formation and adherence rates were calculated after cell counting. Morphological alteration, karyotype morphology, and untrastructure were assessed under optical and electron microscopes. The distribution in the cell cycle and drug efflux pump activity were measured by flow cytometry. Furthermore, expression of potential genes involved in MDR of BEL-7402/5-FU cells were detected by immunocytochemistry. Results: Compared to its parental cells, BEL-7402/5-FU cells had a prolonged doubling time, a lower mitotic index, colony efficiency and adhesive ability, and a decreased drug efflux pump activity. The resistant cells tended to grow in clusters and apparent changes of ultrastructures occurred. BEL-7402/5-FU cells presented with an increased proportion in S and G2/M phases with a concomitant decrease in G0/G1 phase. The MDR phenotype of BEL-7402/5-FU might be partly attributed to increased drug efflux pump activity via multidrug resistance protein 1 (MRP1), overexpression of thymidylate synthase (TS), resistance to apoptosis by augmentation of the Bcl-xl/Bax ratio, and intracellular adhesion medicated by E-cadherin (E-cad). P-glycoprotein (P-gp) might play a limited role in the MDR of BEL-7402/5-FU. Conclusion: Increased activity or expression of MRP1, Bcl-xl, TS, and E-cad appear to be involved in the MDR mechanism of BEL-7402/5-FU.

• BMB Reports
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• v.51 no.9
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• pp.474-479
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• 2018

Cisplatin is one of the most effective chemotherapeutic drugs used in the treatment of HCC, but many patients will ultimately relapse with cisplatin-resistant disease. Used in combination with cisplatin, resveratrol has synergistic effect of increasing chemosensitivity of cisplatin in various cancer cells. However, the mechanisms of resveratrol enhancing cisplatin-induced toxicity have not been well characterized. Our study showed that resveratrol enhances cisplatin toxicity in human hepatoma cells via an apoptosis-dependent mechanism. Further studies reveal that resveratrol decreases the absorption of glutamine and glutathione content by reducing the expression of glutamine transporter ASCT2. Flow cytometric analyses demonstrate that resveratrol and cisplatin combined treatment leads to a significant increase in ROS production compared to resveratrol or cisplatin treated hepatoma cells alone. Phosphorylated H2AX (${\gamma}H2AX$) foci assay demonstrate that both resveratrol and cisplatin treatment result in a significant increase of ${\gamma}H2AX$ foci in hepatoma cells, and the resveratrol and cisplatin combined treatment results in much more ${\gamma}H2AX$ foci formation than either resveratrol or cisplatin treatment alone. Furthermore, our studies show that over-expression of ASCT2 can attenuate cisplatin-induced ROS production, ${\gamma}H2AX$ foci formation and apoptosis in human hepatoma cells. Collectively, our studies suggest resveratrol may sensitize human hepatoma cells to cisplatin chemotherapy via gluta${\gamma}H2AX$mine metabolism inhibition.

• Journal of Pharmacopuncture
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• v.19 no.2
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• pp.129-136
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• 2016

Objectives: The crude extracts of Scutellaria barbata D. Don (SB) have traditionally demonstrated inhibitory effects on numerous human cancers both in vitro and in vivo. Gastric cancer is one of the most common types of cancer on world. The authors investigated the effects of an ethanol extract of Scutellaria barbata D. Don (ESB) on the growth and survival of MKN-45 cells (a human gastric adenocarcinoma cell line). Methods: The MKN-45 cells were treated with different concentrations of ESB, and cell death was examined using an MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) assay. Analyses of sub-G1 peaks, caspase-3 and -9 activities, and mitochondrial membrane depolarizations were conducted to determine the anti-cancer effects of SB on MKN-45 cells. Also, intracellular reactive oxygen species (ROS) generation was investigated. Results: ESB inhibited the growth of MKN-45 cells, caused cell cycle arrest, and increased the sub-G1 population. In addition, ESB markedly increased mitochondrial membrane depolarization and the activities of caspase-3 and -9. ESB exerted anti-proliferative effects on MKN-45 cells by modulating the mitogen-activated protein kinase (MAPK) signaling pathway and by increasing the generation of ROS. Furthermore, combinations of anti-cancer drugs plus ESB suppressed cell growth more than treatments with an agent or ESB, and this was especially true for cisplatin, etoposide, and doxorubicin. Conclusion: ESB has a dose-dependent cytotoxic effect on MKN-45 cells and this is closely associated with the induction of apoptosis. ESB-induced apoptosis is mediated by mitochondria-, caspase- and MAPK dependent pathways. In addition, ESB enhances ROS generation and increases the chemosensitivity of MKN-45 cells. These results suggest that treatment with ESB can inhibit the proliferation and promote the apoptosis of human gastric adenocarcinoma cells by modulating the caspase-, MAPK- and ROS-dependent pathway.

• Journal of Korean Academy of Oral and Maxillofacial Radiology
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• v.27 no.1
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• pp.43-53
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• 1997

The purpose of this study was to aid in the prediction of tumor cell tolerance to radiotherapy and/or chemotherapy. For this study, cell surviving curves were obtained for mouse lymphoma YAC-1 cell line using semi automated MTT assay. 2, 4,6, 8, 10Gy were irradiated at a dose rate of 210cGy/rnin using /sup 60/Co Irradiator ALDORADO 8. After irradiation, YAC-1 cell lines(3×10⁴cells/ml) were exposed to bleomycin or cisplatin for 1 hour. The viable cells were determined for each radiation dose and/or each concentration of drug at the 4th day. And they were compared to control values. The obtained results were as follows : 1. The surviving curve with gentle slope was obtained after irradiation of 2, 4, 6, 8, 10 Gy on YAC-1 cell line. 2. The cytotoxicity of bleomycin or cisplatin was increased significantly at all concentration of 0.2㎍/ml, 2㎍/ml and 20㎍/ml on YAC-1 cell line (P<0.01). And the cytotoxicity of cisplatin was greater than that of bleomycin at all concentration on YAC-1 cell line (P<0.01). 3. There were no significant differences of surviving fractions among 4Gy, 6Gy and 8Gy after irradiation of each radiation dose with 2㎍/ml of bleomycin compared with irradiation only on YAC-1 cell line. 4. There was significant difference of surviving fraction between 2Gy and 10Gy after irradiation of each radiation dose with 2㎍/ml of cisplatin compared with irradiation only on YAC-1 cell line(P<0.05). 5. There were significant differences of surviving fractions between the groups of irradiation only and the groups of irradiation with 2㎍/ml of bleomycin or cisplatin at all doses of 2, 4, 6, 8 and 10Gy on YAC-1 cell line(P<0.05).

• Biomedical Science Letters
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• v.2 no.2
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• pp.153-158
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• 1996

Verapamil, a potent calcium channel blocker, has been proved to be one of the modulators to overcome drug resistance in cancer chemotherapy. In the present experiment, the possibility of verapamil as a MDR modulator was investigated by using MTT assay. Sole treatment of verapamil on the HeLa and Caski cervical cancer cell line revealed dose dependent cytotoxicity within a range of tested dose. Combined treatment of verapamil with 5-FU, DDP on two human cervical cancer cell line led to a significant synergistic cytotoxicity. Therefore , these studies showed that verapamil had a possibility to be applicable to cancer chemotherapy in gynecological oncology.