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Selective killing of K-ras-transformed pancreatic cancer cells by targeting NAD(P)H oxidase.

Wang P, Sun YC, Lu WH, Huang P, Hu Y - Chin J Cancer (2015)

Bottom Line: Importantly, capsaicin preferentially inhibited the enzyme activity of NOX and induced severe ROS accumulation in K-ras-transformed cells compared with parental E6E7 cells.Furthermore, capsaicin effectively inhibited cell proliferation, prevented invasiveness of K-ras-transformed pancreatic cancer cells, and caused minimum toxicity to parental E6E7 cells.Our study provides a basis for developing a novel therapeutic strategy to effectively kill K-ras-transformed cells through a redox-mediated mechanism.

View Article: PubMed Central - PubMed

Affiliation: Sun Yat-sen University Cancer Center; State Key Laboratory of Oncology in South China; Collaborative Innovation Center for Cancer Medicine, Guangzhou, Guangdong, 510060, P.R. China. pengwangsysu@foxmail.com.

ABSTRACT

Introduction: Oncogenic activation of the K-ras gene occurs in >90% of pancreatic ductal carcinoma and plays a critical role in the pathogenesis of this malignancy. Increase of reactive oxygen species (ROS) has also been observed in a wide spectrum of cancers. This study aimed to investigate the mechanistic association between K-ras-induced transformation and increased ROS stress and its therapeutic implications in pancreatic cancer.

Methods: ROS level, NADPH oxidase (NOX) activity and expression, and cell invasion were examined in human pancreatic duct epithelial E6E7 cells transfected with K-ras (G12V) compared with parental E6E7 cells. The cytotoxic effect and antitumor effect of capsaicin, a NOX inhibitor, were also tested in vitro and in vivo.

Results: K-ras transfection caused activation of the membrane-associated redox enzyme NOX and elevated ROS generation through the phosphatidylinositol 3'-kinase (PI3K) pathway. Importantly, capsaicin preferentially inhibited the enzyme activity of NOX and induced severe ROS accumulation in K-ras-transformed cells compared with parental E6E7 cells. Furthermore, capsaicin effectively inhibited cell proliferation, prevented invasiveness of K-ras-transformed pancreatic cancer cells, and caused minimum toxicity to parental E6E7 cells. In vivo, capsaicin exhibited antitumor activity against pancreatic cancer and showed oxidative damage to the xenograft tumor cells.

Conclusions: K-ras oncogenic signaling causes increased ROS stress through NOX, and abnormal ROS stress can selectively kill tumor cells by using NOX inhibitors. Our study provides a basis for developing a novel therapeutic strategy to effectively kill K-ras-transformed cells through a redox-mediated mechanism.

No MeSH data available.


Related in: MedlinePlus

Proposed model of the selective killing ofK-ras–transformed cells by NOX inhibition. K-ras transformation causes the activation of the downstream effector, phosphatidylinositol 3′-kinase (PI3K). Phosphatidylinositol (3,4,5)-trisphosphate (PIP3), the lipid product of a PI3K-catalyzed reaction, enhances the guanine-nucleotide exchange factor (GEF) activity that mediates the exchange of Rac-GDP for Rac-GTP, and provides the lipid necessary for NOX subunit (p40phox) binding upon the translocation to the membrane, therefore causing the activation of NOX. NOX-induced ROS production stimulates cell proliferation and contributes to tumor progression. NOX inhibitor targets the activated NOX and causes excessive ROS generation, leading to cancer cell death. +, activation by the upstream effectors.
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Fig6: Proposed model of the selective killing ofK-ras–transformed cells by NOX inhibition. K-ras transformation causes the activation of the downstream effector, phosphatidylinositol 3′-kinase (PI3K). Phosphatidylinositol (3,4,5)-trisphosphate (PIP3), the lipid product of a PI3K-catalyzed reaction, enhances the guanine-nucleotide exchange factor (GEF) activity that mediates the exchange of Rac-GDP for Rac-GTP, and provides the lipid necessary for NOX subunit (p40phox) binding upon the translocation to the membrane, therefore causing the activation of NOX. NOX-induced ROS production stimulates cell proliferation and contributes to tumor progression. NOX inhibitor targets the activated NOX and causes excessive ROS generation, leading to cancer cell death. +, activation by the upstream effectors.

Mentions: It is noteworthy that NOX, the membrane-binding ROS-generating enzyme, has also been suggested to be involved in cell transformation [25,26]. It is known that the activation of NOX requires the proper assembly of multiple regulatory components, including p22phox, ph47phox, p40phox, and p67phox, and the small GTPase, Rac [10,19]. In the current study, NOX expression and enzyme activity were consistently up-regulated in K-ras–transformed pancreatic cancer cells. Previous studies have indicated that the triggers of NOX regulatory subunits involved protein kinases, lipid-metabolizing enzymes, and guanine-nucleotide exchange proteins that activate Rac [10]. It has also been known that phosphatidylinositol (3,4,5)-trisphosphate (PIP3), the lipid product of PI3K, activates Rac by binding to the pleckstrin homology (PH) domain of GEF, which mediates the exchange of Rac-GDP for Rac-GTP [27,28]. Because PI3K is one major downstream effector of K-ras, we postulated that K-ras induces the constitutive activation of NOX through the PI3K/PIP3/GEF/Rac pathway (Figure 6). Accordingly, we found that the levels of both phosphorylated p40phox and active GTP-binding Rac were significantly up-regulated in K-ras–transformed E6E7 cells. Interestingly, the binding of PtdIns(3)P, a product of PI3K, to the phox homology (PX) domain of p40phox was previously reported to lead to the stimulation of ROS formation [29], indicating that PI3K could also mediate the Ras-induced activation of NOX through p40phox.Figure 6


Selective killing of K-ras-transformed pancreatic cancer cells by targeting NAD(P)H oxidase.

Wang P, Sun YC, Lu WH, Huang P, Hu Y - Chin J Cancer (2015)

Proposed model of the selective killing ofK-ras–transformed cells by NOX inhibition. K-ras transformation causes the activation of the downstream effector, phosphatidylinositol 3′-kinase (PI3K). Phosphatidylinositol (3,4,5)-trisphosphate (PIP3), the lipid product of a PI3K-catalyzed reaction, enhances the guanine-nucleotide exchange factor (GEF) activity that mediates the exchange of Rac-GDP for Rac-GTP, and provides the lipid necessary for NOX subunit (p40phox) binding upon the translocation to the membrane, therefore causing the activation of NOX. NOX-induced ROS production stimulates cell proliferation and contributes to tumor progression. NOX inhibitor targets the activated NOX and causes excessive ROS generation, leading to cancer cell death. +, activation by the upstream effectors.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4593348&req=5

Fig6: Proposed model of the selective killing ofK-ras–transformed cells by NOX inhibition. K-ras transformation causes the activation of the downstream effector, phosphatidylinositol 3′-kinase (PI3K). Phosphatidylinositol (3,4,5)-trisphosphate (PIP3), the lipid product of a PI3K-catalyzed reaction, enhances the guanine-nucleotide exchange factor (GEF) activity that mediates the exchange of Rac-GDP for Rac-GTP, and provides the lipid necessary for NOX subunit (p40phox) binding upon the translocation to the membrane, therefore causing the activation of NOX. NOX-induced ROS production stimulates cell proliferation and contributes to tumor progression. NOX inhibitor targets the activated NOX and causes excessive ROS generation, leading to cancer cell death. +, activation by the upstream effectors.
Mentions: It is noteworthy that NOX, the membrane-binding ROS-generating enzyme, has also been suggested to be involved in cell transformation [25,26]. It is known that the activation of NOX requires the proper assembly of multiple regulatory components, including p22phox, ph47phox, p40phox, and p67phox, and the small GTPase, Rac [10,19]. In the current study, NOX expression and enzyme activity were consistently up-regulated in K-ras–transformed pancreatic cancer cells. Previous studies have indicated that the triggers of NOX regulatory subunits involved protein kinases, lipid-metabolizing enzymes, and guanine-nucleotide exchange proteins that activate Rac [10]. It has also been known that phosphatidylinositol (3,4,5)-trisphosphate (PIP3), the lipid product of PI3K, activates Rac by binding to the pleckstrin homology (PH) domain of GEF, which mediates the exchange of Rac-GDP for Rac-GTP [27,28]. Because PI3K is one major downstream effector of K-ras, we postulated that K-ras induces the constitutive activation of NOX through the PI3K/PIP3/GEF/Rac pathway (Figure 6). Accordingly, we found that the levels of both phosphorylated p40phox and active GTP-binding Rac were significantly up-regulated in K-ras–transformed E6E7 cells. Interestingly, the binding of PtdIns(3)P, a product of PI3K, to the phox homology (PX) domain of p40phox was previously reported to lead to the stimulation of ROS formation [29], indicating that PI3K could also mediate the Ras-induced activation of NOX through p40phox.Figure 6

Bottom Line: Importantly, capsaicin preferentially inhibited the enzyme activity of NOX and induced severe ROS accumulation in K-ras-transformed cells compared with parental E6E7 cells.Furthermore, capsaicin effectively inhibited cell proliferation, prevented invasiveness of K-ras-transformed pancreatic cancer cells, and caused minimum toxicity to parental E6E7 cells.Our study provides a basis for developing a novel therapeutic strategy to effectively kill K-ras-transformed cells through a redox-mediated mechanism.

View Article: PubMed Central - PubMed

Affiliation: Sun Yat-sen University Cancer Center; State Key Laboratory of Oncology in South China; Collaborative Innovation Center for Cancer Medicine, Guangzhou, Guangdong, 510060, P.R. China. pengwangsysu@foxmail.com.

ABSTRACT

Introduction: Oncogenic activation of the K-ras gene occurs in >90% of pancreatic ductal carcinoma and plays a critical role in the pathogenesis of this malignancy. Increase of reactive oxygen species (ROS) has also been observed in a wide spectrum of cancers. This study aimed to investigate the mechanistic association between K-ras-induced transformation and increased ROS stress and its therapeutic implications in pancreatic cancer.

Methods: ROS level, NADPH oxidase (NOX) activity and expression, and cell invasion were examined in human pancreatic duct epithelial E6E7 cells transfected with K-ras (G12V) compared with parental E6E7 cells. The cytotoxic effect and antitumor effect of capsaicin, a NOX inhibitor, were also tested in vitro and in vivo.

Results: K-ras transfection caused activation of the membrane-associated redox enzyme NOX and elevated ROS generation through the phosphatidylinositol 3'-kinase (PI3K) pathway. Importantly, capsaicin preferentially inhibited the enzyme activity of NOX and induced severe ROS accumulation in K-ras-transformed cells compared with parental E6E7 cells. Furthermore, capsaicin effectively inhibited cell proliferation, prevented invasiveness of K-ras-transformed pancreatic cancer cells, and caused minimum toxicity to parental E6E7 cells. In vivo, capsaicin exhibited antitumor activity against pancreatic cancer and showed oxidative damage to the xenograft tumor cells.

Conclusions: K-ras oncogenic signaling causes increased ROS stress through NOX, and abnormal ROS stress can selectively kill tumor cells by using NOX inhibitors. Our study provides a basis for developing a novel therapeutic strategy to effectively kill K-ras-transformed cells through a redox-mediated mechanism.

No MeSH data available.


Related in: MedlinePlus