Project title
核酸修補蛋白Rad51及ERCC1在大黃素抑制人類非小細胞肺癌細胞存活之角色探討
Internal ID
DOH99-HO-2020
Principal Investigator
Start Date
January 1, 2010
End Date
December 31, 2010
Organizations
Partner Organizations
Ministry of Health and Welfare
Investigation of the molecular mechanism of the synergistic effects by emodin and cisplatin in inhibiting the cell survival of human non-small cell lung cancer cells = 核酸修補蛋白Rad51及ERCC1在大黃素抑制人類非小細胞肺癌細胞存活之角色探討
Keywords
非小細胞肺癌
大黃素
ERCC1修補蛋白質
Rad51修補蛋白質
核酸修補
細胞毒性
Description
Emodin Emodin (1,3,8-trihydroxy-6-methylanthraquinone) is a naturally occurring anthraquinone present in the roots and rhizomes of numerous plants and Chinese herbs (Demirezer et al., 2001; Kuo et al., 2001). A number of studies have demonstrated that emodin is capable of inducing cell apoptosis and growth arrest in various cancer cells, such as human lung cancer cells (Su et al., 2005), HER-2/neu–overexpressed breast cancer cells (Zhang et al., 1998; Zhang et al., 1999), cervical cancer cells, leukemia cells, hepatoma cells, and prostate cancer cell lines (Chen et al., 2002; Shieh et al., 2004; Wang et al., 2007). In addition, emodin induces sub-G1 accumulation and G2/M phase arrest in hepatoma cells (Shieh et al., 2004). Emodin, as a tyrosine kinase inhibitor, can inhibit the kinase activity of HER-2/neu and suppress the proliferation of HER-2/neu–overexpressed non-small cell lung cancer (NSCLC) and breast cancer cells (Zhang et al., 1995; Zhang et al., 1999). Emodin has also been shown to inhibit p56lck (Zhang et al., 1995) and casein kinase 2 (CK2) activity (Battistutta et al., 2000). In addition, emodin can repress epidermal growth factor receptor (EGFR) tyrosine kinase activity at high concentrations (Zhang et al., 1999). The molecular mechanism by which emodin induces cell apoptosis is through inactivation of ERK and AKT and the decrease of anti-apoptotic protein Bcl2 levels in human A549 cells (Su et al., 2005). However, the effect of emodin on the ability of DNA repair to regulate cell survival in NSCLC cells has not yet been fully defined. Excision repair cross-complementary 1 (ERCC1) Excision repair cross-complementary 1 (ERCC1) has a leading role in the nucleotide excision repair (NER) process because of its involvement in the excision of DNA adducts (de Laat et al., 1998). The damaged DNA strand is cleaved by ERCC1-XPF (xeroderma pigmentosum-F) on the 5' side during NER in human cells (Evans et al., 1997). ERCC1 RNA levels are highly correlated with NER activity in blood lymphocytes (Vogel et al., 2000). The increased expression of ERCC1 is associated with clinical resistance to platinum-based chemotherapy in human NSCLC (Lord et al., 2002). Moreover, patients with ERCC1-negative tumors had a longer survival time than those with ERCC1-positive tumors in NSCLC (Olaussen et al., 2006). Rad51 Homologous recombination repair (HRR), a major pathway for the repair of DNA double-strand breaks (DSBs) in eukaryotic cells, is mediated by Rad51. The HRR process consists of assembly of Rad51 onto DNA substrates at the site of a DNA break to form a helical nucleoprotein filament, which catalyzes homologous pairing and joins in a hetero-duplex formation with the sister chromatid (West, 2003). Rad51 is overexpressed in many tumors (Raderschall et al., 2002; Xia et al., 1997), and its overexpression is related to the resistance of the tumor to chemotherapeutic agents or radiation (Bello et al., 2002; Slupianek et al., 2002). In NSCLC, high expression of Rad51 in tumor tissue is associated with an unfavorable prognosis (Qiao et al., 2005; Takenaka et al., 2007). On the other hand, inhibition of Rad51 expression has been shown to sensitize cancer cells to radiotherapy and chemotherapeutic agents (Ko et al., 2008a; Ohnishi et al., 1998). However, whether Rad51 and ERCC1 are targeted by emodin to induce cell death in human lung cancer cells is still unclear and remains to be determined. MKK1/2-ERK1/2 signaling pathway ERK1/2 is serine-threonine protein kinase and one of mitogen-activated protein kinases (MAPKs) (Chang and Karin, 2001; Garrington and Johnson, 1999; Kolch, 2000; Kolch, 2005; Nebreda and Porras, 2000). The ERK1/2 signaling activation are connected to cell surface receptors, such as EGFR and platelet-derived growth factor receptor (PDGFR), and activated via interaction with a family of small GTPases, such as Ras (Kolch, 2000). In general, ERK1/2 is involved in the control of cell proliferation, survival, differentiation, growth arrest, apoptosis, cell transformation and tumor invasion through phosphorylation of many downstream substrates including transcription factors, intracellular protein kinases, phosphatases and cytoskeletal proteins (Kolch, 2000; Kolch, 2005; Marshall, 1995). In addition, ERK1/2 signaling pathway activation plays a vital role in tumorigenesis (Weinstein-Oppenheimer et al., 2000) and promotes tumor cell growth in several cancer cell lines (Albanell et al., 2001; El Sheikh et al., 2004). Rationale Lung cancer remains the leading cause of cancer-related deaths in the world, and more than 85% of lung cancers have NSCLC (Landis et al., 1999). In this study, we investigated the role of emodin in suppressing cell viability in four NSCLC cell lines and examined possible molecular mechanisms for this activity. We also determine the roles of repair proteins ERCC1 and Rad51 in emodin-induced cytotoxicity. Emodin may be a novel and improved therapeutic modality for advanced lung cancer in the future, especially for patients in whom lung cancer cells are resistant to chemotherapeutic agents. Emodin Emodin (1,3,8-trihydroxy-6-methylanthraquinone) is a naturally occurring anthraquinone present in the roots and rhizomes of numerous plants and Chinese herbs (Demirezer et al., 2001; Kuo et al., 2001). A number of studies have demonstrated that emodin is capable of inducing cell apoptosis and growth arrest in various cancer cells, such as human lung cancer cells (Su et al., 2005), HER-2/neu–overexpressed breast cancer cells (Zhang et al., 1998; Zhang et al., 1999), cervical cancer cells, leukemia cells, hepatoma cells, and prostate cancer cell lines (Chen et al., 2002; Shieh et al., 2004; Wang et al., 2007). In addition, emodin induces sub-G1 accumulation and G2/M phase arrest in hepatoma cells (Shieh et al., 2004). Emodin, as a tyrosine kinase inhibitor, can inhibit the kinase activity of HER-2/neu and suppress the proliferation of HER-2/neu–overexpressed non-small cell lung cancer (NSCLC) and breast cancer cells (Zhang et al., 1995; Zhang et al., 1999). Emodin has also been shown to inhibit p56lck (Zhang et al., 1995) and casein kinase 2 (CK2) activity (Battistutta et al., 2000). In addition, emodin can repress epidermal growth factor receptor (EGFR) tyrosine kinase activity at high concentrations (Zhang et al., 1999). The molecular mechanism by which emodin induces cell apoptosis is through inactivation of ERK and AKT and the decrease of anti-apoptotic protein Bcl2 levels in human A549 cells (Su et al., 2005). However, the effect of emodin on the ability of DNA repair to regulate cell survival in NSCLC cells has not yet been fully defined. Excision repair cross-complementary 1 (ERCC1) Excision repair cross-complementary 1 (ERCC1) has a leading role in the nucleotide excision repair (NER) process because of its involvement in the excision of DNA adducts (de Laat et al., 1998). The damaged DNA strand is cleaved by ERCC1-XPF (xeroderma pigmentosum-F) on the 5' side during NER in human cells (Evans et al., 1997). ERCC1 RNA levels are highly correlated with NER activity in blood lymphocytes (Vogel et al., 2000). The increased expression of ERCC1 is associated with clinical resistance to platinum-based chemotherapy in human NSCLC (Lord et al., 2002). Moreover, patients with ERCC1-negative tumors had a longer survival time than those with ERCC1-positive tumors in NSCLC (Olaussen et al., 2006). Rad51 Homologous recombination repair (HRR), a major pathway for the repair of DNA double-strand breaks (DSBs) in eukaryotic cells, is mediated by Rad51. The HRR process consists of assembly of Rad51 onto DNA substrates at the site of a DNA break to form a helical nucleoprotein filament, which catalyzes homologous pairing and joins in a hetero-duplex formation with the sister chromatid (West, 2003). Rad51 is overexpressed in many tumors (Raderschall et al., 2002; Xia et al., 1997), and its overexpression is related to the resistance of the tumor to chemotherapeutic agents or radiation (Bello et al., 2002; Slupianek et al., 2002). In NSCLC, high expression of Rad51 in tumor tissue is associated with an unfavorable prognosis (Qiao et al., 2005; Takenaka et al., 2007). On the other hand, inhibition of Rad51 expression has been shown to sensitize cancer cells to radiotherapy and chemotherapeutic agents (Ko et al., 2008a; Ohnishi et al., 1998). However, whether Rad51 and ERCC1 are targeted by emodin to induce cell death in human lung cancer cells is still unclear and remains to be determined. MKK1/2-ERK1/2 signaling pathway ERK1/2 is serine-threonine protein kinase and one of mitogen-activated protein kinases (MAPKs) (Chang and Karin, 2001; Garrington and Johnson, 1999; Kolch, 2000; Kolch, 2005; Nebreda and Porras, 2000). The ERK1/2 signaling activation are connected to cell surface receptors, such as EGFR and platelet-derived growth factor receptor (PDGFR), and activated via interaction with a family of small GTPases, such as Ras (Kolch, 2000). In general, ERK1/2 is involved in the control of cell proliferation, survival, differentiation, growth arrest, apoptosis, cell transformation and tumor invasion through phosphorylation of many downstream substrates including transcription factors, intracellular protein kinases, phosphatases and cytoskeletal proteins (Kolch, 2000; Kolch, 2005; Marshall, 1995). In addition, ERK1/2 signaling pathway activation plays a vital role in tumorigenesis (Weinstein-Oppenheimer et al., 2000) and promotes tumor cell growth in several cancer cell lines (Albanell et al., 2001; El Sheikh et al., 2004). Rationale Lung cancer remains the leading cause of cancer-related deaths in the world, and more than 85% of lung cancers have NSCLC (Landis et al., 1999). In this study, we investigated the role of emodin in suppressing cell viability in four NSCLC cell lines and examined possible molecular mechanisms for this activity. We also determine the roles of repair proteins ERCC1 and Rad51 in emodin-induced cytotoxicity. Emodin may be a novel and improved therapeutic modality for advanced lung cancer in the future, especially for patients in whom lung cancer cells are resistant to chemotherapeutic agents.