Browsing by Author "Ko, Jen-Chung"

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Astaxanthin down-regulates Rad51 expression via inactivation of AKT kinase to enhance mitomycin C-induced cytotoxicity in human non-small cell lung cancer cells
(2016-04-01)
Ko, Jen-Chung
;
Chen, Jyh-Cheng
;
Wang, Tai-Jing
;
Zheng, Hao-Yu
;
Chen, Wen-Ching
Astaxanthin has been demonstrated to exhibit a wide range of beneficial effects, including anti-inflammatory and anti-cancer properties. However, the molecular mechanism of astaxanthin-induced cytotoxicity in non-small cell lung cancer (NSCLC) cells has not been identified. Rad51 plays a central role in homologous recombination, and studies show that chemo-resistant carcinomas exhibit high levels of Rad51 expression. In this study, astaxanthin treatment inhibited cell viability and proliferation of two NSCLC cells, A549 and H1703. Astaxanthin treatment (2.5-20 mu M) decreased Rad51 expression and phospho-AKT(Ser473) protein level in a time and dose-dependent manner. Furthermore, expression of constitutively active AKT (AKT-CA) vector rescued the decreased Rad51 mRNA and protein levels in astaxanthin-treated NSCLC cells. Combined treatment with phosphatidylinositol 3-kinase (PI3K) inhibitors (LY294002 or wortmannin) further decreased the Rad51 expression in astaxanthin-exposed A549 and H1703 cells. Knockdown of Rad51 expression by transfection with si-Rad51 RNA or cotreatment with LY294002 further enhanced the cytotoxicity and cell growth inhibition of astaxanthin. Additionally, mitomycin C (MMC) as an anti-tumor antibiotic is widely used in clinical NSCLC chemotherapy. Combination of MMC and astaxanthin synergistically resulted in cytotoxicity and cell growth inhibition in NSCLC cells, accompanied with reduced phospho-AKT(Ser473) level and Rad51 expression. Overexpression of AKT-CA or Flag-tagged Rad51 reversed the astaxanthin and MMC-induced synergistic cytotoxicity. In contrast, pretreatment with LY294002 further decreased the cell viability in astaxanthin and MMC co-treated cells. In conclusion, astaxanthin enhances MMC-induced cytotoxicity by decreasing Rad51 expression and AKT activation. These findings may provide rationale to combine astaxanthin with MMC for the treatment of NSCLC. (C) 2016 Elsevier Inc. All rights reserved.
Curcumin enhances the mitomycin C-induced cytotoxicity via downregulation of MKK1/2-ERK1/2-mediated Rad51 expression in non-small cell lung cancer cells
(2011-09-15)
Ko, Jen-Chung
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Tsai, Min-Shao
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Weng, Shao-Hsing
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Kuo, Ya-Hsun
;
Chiu, Yu-Fan
Curcumin (diferuloylmethane), a major active component of turmeric (Curcuma longa), has been reported to suppress the proliferation of a wide variety of tumor cells. Rad51 is a key protein in the homologous recombination (HR) pathway of DNA double-strand break repair, and HR represents a novel target for cancer therapy. A high expression of Rad51 has been reported in chemo- or radio-resistant carcinomas. Therefore, in the current study, we will examine whether curcumin could enhance the effects of mitomycin C (MMC), a DNA interstrand cross-linking agent, to induce cytotoxicity by decreasing Rad51 expression. Exposure of two human non-small lung cancer (NSCLC) cell lines (A549 and H1975) to curcumin could suppress MMC-induced MKK1/2-ERK1/2 signal activation and Rad51 protein expression. Enhancement of ERK1/2 activation by constitutively active MKK1/2 (MKK1/2-CA) increased Rad51 protein levels in curcumin and MMC co-treated human lung cells. Moreover, the synergistic cytotoxic effect induced by curcumin combined with MMC was decreased by MKK1-CA-mediated enhancement of ERK1/2 activation by a significant degree. In contrast, MKK1/2 inhibitor, U0126 was shown to augment the cytotoxicity of curcumin and MMC through downregulation of ERK1/2 activation and Rad51 expression. Depletion of endogenous Rad51 expression by siRad51 RNA transfection significantly enhanced MMC and/or curcumin induced cell death and cell growth inhibition. In contrast, an overexpression of Rad51 protected lung cancer cells from synergistic cytotoxic effects induced by curcumin and MMC. We concluded that Rad51 inhibition may be an additional action mechanism for enhancing the chemosensitization of MMC by curcumin in NSCLC. (C) 2011 Elsevier Inc. All rights reserved.
Inhibition of p38 MAPK-dependent MutS homologue-2 (MSH2) expression by metformin enhances gefitinib-induced cytotoxicity in human squamous lung cancer cells
(2013-12-01)
Ko, Jen-Chung
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Chiu, Hsien-Chun
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Wo, Ting-Yu
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Huang, Yi-Jhen
;
Tseng, Sheng-Chieh
Objectives: Gefitinib, a quinazoline-derived tyrosine kinase inhibitor, has anti-tumor activity in vivo and in vitro. Human MutS homologue-2 (MSH2) plays a central role in promoting genetic stability by correcting DNA replication errors. The present study investigated the effects of p38 mitogen-activated protein kinase (MAPK) signal on gefitinib-induced MSH2 expression in two human non-small cell lung squamous cancer cell lines. Materials and methods: After the gefitinib treatment, the expressions of MSH2 mRNA were determined by real-time PCR and RT-PCR analysis. Protein levels of MSH2, phospho-MKK3/6, phospho-p38 MAPK were determined by Western blot analysis. We used specific MSH2, and p38 MAPK small interfering RNA to examine the role of p38 MAPK-MSH2 signal in regulating the chemosensitivity of gefitinib. Cell viability was assessed by MTS assay, trypan blue exclusion, and colony-forming ability assay. Results: Exposure of gefitinib increased MSH2 protein and mRNA levels, which was accompanied by MKK3/6-p38 MAPK activation in H520 and H1703 cells. Moreover, blocking p38 MAPK activation by SB202190 significantly decreased gefitinib-induced MSH2 expression by increasing mRNA and protein instability. In contrast, enhancing p38 activation using constitutively active MKK6 (MKK6E) increased MSH2 protein and mRNA levels. Specific inhibition of MSH2 expression by siRNA enhanced gefitinib-induced cytotoxicity. Metformin, an anti-diabetic drug, might reduce cancer risk. In human lung squamous cancer cells, metformin decreased gefitinib-induced MSH2 expression and augmented the cytotoxic effect and growth inhibition by gefitinib. Transient expression of MKK6E or HA-p38 MAPK vector could abrogate metformin and gefitinib-induced synergistic cytotoxic effect in H520 and H1703 cells. Conclusion: Together, down-regulation of MSH2 expression can be a possible strategy to enhance the sensitivity of gefitinib to human lung squamous cancer cells. (C) 2013 Elsevier Ireland Ltd. All rights reserved.
Modulation of Rad51, ERCC1, and thymidine phosphorylase by emodin result in synergistic cytotoxic effect in combination with capecitabine
(2011-03-01)
Ko, Jen-Chung
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Tsai, Min-Shao
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Kuo, Ya-Hsun
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Chiu, Yu-Fan
;
Weng, Shao-Hsing
Thymidine phosphorylase (TP) is the rate-limiting enzyme for the activation of capecitabine (pro-drug of fluorouracil), and as a useful predictor of tumor response to capecitabine-based chemotherapy. Overexpression of Rad51 and ERCC1 induce resistance to chemotherapeutic agents. Emodin, one of the main bioactive anthraquinone derivatives in the roots and rhizomes of numerous plants, possesses potent antitumor effects. Accordingly, we aimed to explore the molecular mechanism of emodin enhances the capecitabine-induced cytotoxicity through controlling Rad51, ERCC1, and TP expression in human non-small cell lung cancer (NSCLC). The results show that capecitabine increases the phosphorylation of MKK1/2-ERK1/2 and protein levels of Rad51 and ERCC1 through enhancing the protein stability. Depletion of endogenous Rad51 or ERCC1 expression by specific small interfering RNA transfection significantly increases capecitabine-induced cell death and growth inhibition. Emodin enhances the capecitabine-induced cytotoxic effects through ERK1/2 inactivation and decreasing the Rad51 and ERCC1 protein levels induced by capecitabine. Enhancement of ERK1/2 signaling by constitutively active MKK1/2 (MKK1/2-CA) results in increasing Rad51 and ERCC1 protein levels and cell viability in NSCLC cell lines treated with emodin and capecitabine. Interestingly, emodin enhances TP mRNA and protein expression in capecitabine treated NSCLC cell lines, and depletion of the TP expression decreases the cytotoxic effects induced by capecitabine and emodin. We conclude that enhancing the cytotoxicity to capecitabine by emodin is mediated by down-regulation the expression of Rad51 and ERCC1 and up-regulation TP expression. (C) 2010 Elsevier Inc. All rights reserved.
Pemetrexed downregulates ERCC1 expression and enhances cytotoxicity effected by resveratrol in human nonsmall cell lung cancer cells
(2013-12-01)
Chen, Ruey-Shyang
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Ko, Jen-Chung
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Chiu, Hsien-Chun
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Wo, Ting-Yu
;
Huang, Yi-Jhen
The multitargeted antifolate pemetrexed has demonstrated certain clinical activities against nonsmall cell lung cancer (NSCLC). Resveratrol (3,5,4-trihydroxy-trans-stilbene) is a polyphenol found in grapes and other plants and has great potential as a preventative and therapeutic agent due to its anticarcinogenic activity. The efficacy of adding resveratrol to pemetrexed to prolong the survival of patients with NSCLC still remains unclear. The excision repair cross-complementation 1 (ERCC1) is a DNA repair gene coding 5' endonuclease in nucleotide excision repair and is overexpressed in chemo- or radioresistant carcinomas. In this study, resveratrol (10-50 mu M) inhibited cell survival in two NSCLC cells, H520 and H1975. Treatment with resveratrol increased ERCC1 messenger RNA and protein levels in a MKK3/6-p38 MAPK signal activation-dependent manner. Furthermore, blocking p38 MAPK activation by SB202190 or knocking down ERCC1 expression by transfection with small interfering RNA of ERCC1 enhanced the cytotoxicity of resveratrol. Combining resveratrol with pemetrexed resulted in a synergistic cytotoxic effect, accompanied with the reduction of phospho-p38 MAPK and ERCC1 protein levels, and a DNA repair capacity. Expression of constitutively active MKK6 (MKK6E) or HA-p38 MAPK vectors significantly rescued the decreased p38 MAPK activity, and restored ERCC1 protein levels and cell survival in resveratrol and pemetrexed cotreated NSCLC cells. In this study, for the first time, we have demonstrated the synergistic effect of combined treatment with resveratrol and pemetrexed in human NSCLC cells through downregulation of the MKK3/6-p38 MAPK-ERCC1 signal, suggesting a potential benefit of combining resveratrol and pemetrexed to treat lung cancer in the future.
Salinomycin enhances cisplatin-induced cytotoxicity in human lung cancer cells via down-regulation of AKT-dependent thymidylate synthase expression
(2016-12-15)
Ko, Jen-Chung
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Zheng, Hao-Yu
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Chen, Wen-Ching
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Peng, Yi-Shuan
;
Wu, Chia-Hung
Salinomycin, a polyether antibiotic, acts as a highly selective potassium ionophore and has anticancer activity on various cancer cell lines. Cisplatin has been proved as chemotherapy drug for advanced human non-small cell lung cancer (NSCLC). Thymidylate synthase (TS) is a key enzyme in the pyrimidine salvage pathway, and increased expression of TS is thought to be associated with resistance to cisplatin. In this study, we showed that salinomycin (0.5-2 mu mL) treatment down-regulating of TS expression in an AKT inactivation manner in two NSCLC cell lines, human lung adenocarcinoma A549 and squamous cell carcinoma H1703 cells. Knockdown of TS using small interfering RNA (siRNA) or inhibiting AKT activity with PI3K inhibitor LY294002 enhanced the cytotoxicity and cell growth inhibition of salinomycin. A combination of cisplatin and salinomycin resulted in synergistic enhancement of cytotoxicity and cell growth inhibition in NSCLC cells, accompanied with reduced activation of phospho-AKT, and TS expression. Overexpression of a constitutive active AKT (AKT-CA) expression vector reversed the salinomycin and cisplatin-induced synergistic cytotoxicity. In contrast, pretreatment with LY294002 further decreased the cell viability in salinomycin and cisplatin cotreated cells. Our findings suggested that the down-regulation of AKT-mediated TS expression by salinomycin enhanced the cisplatin-induced cytotoxicity in NSCLC cells. These results may provide a rationale to combine salinomycin with cisplatin for lung cancer treatment. (C) 2016 Elsevier Inc. All rights reserved.
Tamoxifen enhances erlotinib-induced cytotoxicity through down-regulating AKT-mediated thymidine phosphorylase expression in human non-small-cell lung cancer cells
(2014-03-01)
Ko, Jen-Chung
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Chiu, Hsien-Chun
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Syu, Jhan-Jhang
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Jian, Yi-Jun
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Chen, Chien-Yu
Tamoxifen is a triphenylethylene nonsteroidal estrogen receptor (ER) antagonist used worldwide as an adjuvant hormone therapeutic agent in the treatment of breast cancer. However, the molecular mechanism of tamoxifen-induced cytotoxicity in non-small cell lung cancer (NSCLC) cells has not been identified. Thymidine phosphorylase (TP) is an enzyme of the pyrimidine salvage pathway which is upregulated in cancers. In this study, tamoxifen treatment inhibited cell survival in two NSCLC cells, H520 and H1975. Treatment with tamoxifen decreased TP mRNA and protein levels through AKT inactivation. Furthermore, expression of constitutively active AKT (AKT-CA) vectors significantly rescued the decreased TP protein and mRNA levels in tamoxifen-treated NSCLC cells. In contrast, combination treatment with PI3K inhibitors (LY294002 or wortmannin) and tamoxifen further decreased the TP expression and cell viability of NSCLC cells. Knocking down TP expression by transfection with small interfering RNA of TP enhanced the cytotoxicity and cell growth inhibition of tamoxifen. Erlotinib (Tarceva, OSI-774), an orally available small molecular inhibitor of epidermal growth factor receptor (EGFR) tyrosine kinase, is approved for clinical treatment of NSCLC. Compared to a single agent alone, tamoxifen combined with erlotinib resulted in cytotoxicity and cell growth inhibition synergistically in NSCLC cells, accompanied with reduced activation of phospho-AKT and phospho-ERK1/2, and reduced TP protein levels. These findings may have implications for the rational design of future drug regimens incorporating tamoxifen and erlotinib for the treatment of NSCLC. (C) 2014 Elsevier Inc. All rights reserved.
Vorinostat enhances the cisplatin-mediated anticancer effects in small cell lung cancer cells
(2016-11-07)
Pan, Chun-Hao
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Chang, Ying-Fang
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Lee, Ming-Shuo
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Wen, B-Chen
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Ko, Jen-Chung
Background: Vorinostat, a histone deacetylase (HDAC) inhibitor, is a promising agent for cancer therapy. Combining vorinostat with cisplatin may relax the chromatin structure and facilitate the accessibility of cisplatin, thus enhancing its cytotoxicity. Studies have not yet investigated the effects of the combination of vorinostat and cisplatin on small cell lung cancer (SCLC). Methods: We first assessed the efficacy of vorinostat with etoposide/cisplatin (EP; triple combination) and then investigated the effects of cotreatment with vorinostat and cisplatin on H209 and H146 SCLC cell lines. The anticancer effects of various combinations were determined in terms of cell viability, apoptosis, cell cycle distribution, and vorinostat-regulated proteins. We also evaluated the efficacy of vorinostat/cisplatin combination in H209 xenograft nude mice. Results: Our data revealed that the triple combination engendered a significant reduction of cell viability and high apoptotic cell death. In addition, vorinostat combined with cisplatin enhanced cell growth inhibition, induced apoptosis, and promoted cell cycle arrest. We observed that the acetylation levels of histone H3 and a-tubulin were higher in combination treatments than in vorinostat treatment alone. Moreover, vorinostat reduced the expression of thymidylate synthase (TS), and TS remained inhibited after cotreament with cisplatin. Furthermore, an in vivo study revealed that the combination of vorinostat and cisplatin significantly inhibited tumor growth in xenograft nude mice (tumor growth inhibition T/C% = 20.5 %). Conclusions: Combined treatments with vorinostat promote the cytotoxicity of cisplatin and induce the expression of vorinostat-regulated acetyl proteins, eventually enhancing antitumor effects in SCLC cell lines. Triple combinations with a low dosage of cisplatin demonstrate similar therapeutic effects. Such triple combinations, if applied clinically, may reduce the undesired adverse effects of cisplatin. The effects of the combination of vorinostat and cisplatin should be evaluated further before conducting clinical trials for SCLC treatment.