NADPH oxidase and pancreatic cancer cell survival

描述(申请人提供)：胰腺癌如此具有侵袭性且对治疗无反应的一个原因是它对细胞凋亡的抵抗力。生长因子既能刺激增殖，又能保护胰腺癌(PACA)细胞免于死亡。尤其是胰岛素样生长因子-I(IGF-I)在PACA细胞存活中起着重要作用。我们发现，NADPH氧化酶产生的活性氧(ROS)介导了IGF-I在这些细胞中的促存活作用。该项目的总体目标是确定NADPH氧化酶在PACA细胞中促进生存的机制。我们将确定PACA细胞中NADPH氧化酶激活的机制，这对于非吞噬细胞仍然是一个普遍重要的问题。特别是，我们计划确定不同的亚基，如NOX4和p22的参与，以及转录调控亚基的表达在IGF-I诱导的NADPH氧化酶激活中的作用。我们推测，NADPH氧化酶产生的ROS增强和延长了PI3K/Akt的激活，PI3K/Akt是主要的抗凋亡途径。这一促进生存的途径受到蛋白酪氨酸磷酸酶(PTPs)和蛋白酪氨酸磷酸酶(PTPs)的严格调控。虽然对蛋白激酶的作用已经有了广泛的研究，但对于PTPs在生长因子的抗细胞凋亡作用中的作用却知之甚少。PTPs对ROS高度敏感，并受到ROS的可逆抑制。我们进一步假设，NADPH氧化酶产生的ROS抑制特定的PTPs，如LMW-PTP和PTP1B，负向调节Pisk/Akt通路。ROS对这些PTP的抑制对于促进和维持PI3K/Akt的激活是生长因子，特别是IGF-I所必需的。了解NADPH氧化酶在PACA细胞死亡中的保护和抗凋亡作用，将使我们能够为胰腺癌提出新的治疗策略。特别是，抑制NADPH氧化酶或相关信号通路可能是一种刺激细胞凋亡的策略，从而克服胰腺癌对化疗和放射治疗的耐药性。我们研究的具体目标是：(1)确定NADPH氧化酶亚基在IGF-I激活PACA细胞中的作用；(2)确定转录上调亚基在IGF-I诱导的PACA细胞NADPH氧化酶激活中的作用；(3)确定IGF-I对PACA细胞中LMW-PTP和PTP1B的影响，以及NADPH氧化酶对这些PTP的调节；(4)确定LMW-PTP和PTP1B在Pisk/Akt激活和IGF-I抑制细胞凋亡中的作用；(5)在胰腺癌体内模型中检测NADPH氧化酶抑制对肿瘤形成的影响。

DESCRIPTION (provided by applicant): One reason why pancreatic cancer is so aggressive and unresponsive to treatments is its resistance to apoptosis. Growth factors both stimulate proliferation and protect pancreatic cancer (PaCa) cells from death. In particular, insulin-like growth factor I (IGF-I) plays a major role in PaCa cell survival. We have found that reactive oxygen species (ROS) produced by NADPH oxidase mediate the pro-survival effect of IGF-I in these cells. The overall goal of the proposed project is to determine the mechanisms of the pro-survival effect of NADPH oxidase in PaCa cells. We will determine the mechanism of NADPH oxidase activation in PaCa cells, which remains an unresolved issue of general importance for non- phagocytic cells. In particular, we plan to determine the involvement of different subunits, such as Nox4 and p22, and the role of transcriptional regulation of subunits' expression in the IGF-I induced activation of NADPH oxidase. We hypothesize that ROS generated by NADPH oxidase enhance and prolong the activation of PI3K/Akt, a major anti-apoptotic pathway. This pro-survival pathway is tightly regulated by kinases and protein tyrosine phosphatases (PTPs). Although the role of kinases has been extensively studied, much less is known on the involvement of PTPs in the anti-apoptotic effects of growth factors. PTPs are highly sensitive to and undergo reversible inhibition by ROS. We further hypothesize that ROS produced by NADPH oxidase inhibit specific PTPs, such as LMW-PTP and PTP1B, which negatively regulate the PISK/Akt pathway. The inhibition of these PTPs by ROS is necessary for growth factors, in particular IGF-I, to enhance and maintain the activation of PI3K/Akt. An understanding of the protective, anti-apoptotic role of NADPH oxidase in PaCa cell death will allow us to propose novel therapeutic strategies for pancreatic cancer. In particular, inhibition of NADPH oxidase or related signaling pathways could be one strategy to stimulate apoptosis and thus overcome pancreatic cancer resistance to chemo- and radiation therapies. The specific objectives of our proposal are (1) determine the role of NADPH oxidase subunits in its activation by IGF-I in PaCa cells; (2) determine the role of subunits' transcriptional up-regulation in NADPH oxidase activation induced by IGF-I in PaCa cells; (3) determine the effect of IGF-I on LMW-PTP and PTP1B, and the regulation of these PTPs by NADPH oxidase in PaCa cells; (4) determine the roles of LMW-PTP and PTP1B in PISK/Akt activation and inhibition of apoptosis by IGF-I in PaCa cells; (5) determine the effects of NADPH oxidase inhibition on tumorigenesis in an in vivo model of pancreatic cancer.