cPLA2alpha, COX-2 and TGF-beta in Liver Cancer

描述(申请人提供)：原发性肝癌是人类常见的恶性肿瘤，死亡率高，其发病率在全球范围内呈上升趋势，尤其是在美国。它主要发生在先前存在的慢性炎症性肝病中，包括慢性肝炎和肝硬变。本实验室最近的研究表明，前列腺素(PG)代谢在肝脏炎症和癌变中起重要作用。在这项研究中，我们假设前列腺素信号的水平和激活状态是决定细胞对转化生长因子的反应的关键因素。(转化生长因子-？)具体地说，我们假设胞浆磷脂酶A2？(cPLA2？)而环氧合酶-2(COX-2)控制的PG信号转导通路破坏了转化生长因子-β介导的有丝分裂抑制，这一机制通过选择和扩增转化生长因子-β参与了肝细胞癌的发生。耐药的异型和肿瘤性上皮细胞，进展更快，向恶变和肿瘤发展。因此，阻断PG信号通路可能恢复转化生长因子-β的生长抑制作用，从而预防肝癌的发生。本申请提出了一系列实验来评估上述假设。CPLA2表达改变的人肝癌细胞？并用COX-2检测其对转化生长因子-β的反应。Smad2/3的siRNA将被导入稳定表达反义cPLA2？或COX-2，这些细胞将在体外和SCID小鼠体内进行增殖和凋亡分析。靶向表达cPLA2的转基因小鼠？而肝脏中的COX-2将被开发并用于确定转化生长因子？调节的Smad激活、有丝分裂抑制、细胞凋亡和肝癌的发生。CPLA2？COX-2转基因和基因敲除小鼠将与转化生长因子受体II型基因敲除小鼠杂交，以确定肝再生和DEN诱导的肝癌发生。最后，将利用培养的肝星状细胞和实验动物模型来评估我们的假设。而纤维化肝星状细胞中的PG信号在肝纤维化和癌变的发病机制中起重要作用。这些拟议研究的结果有望为人类肝癌的化学预防和治疗提供重要的治疗意义。公共卫生相关性：原发性肝癌是一种高度恶性的人类肿瘤，目前还没有有效的化学预防或系统治疗。这一应用是为了检验我们的假设，即前列腺素信号的水平和激活状态是决定细胞对转化生长因子-β的反应的关键因素。阻断前列腺素信号通路可能恢复转化生长因子-β的生长抑制作用。并预防肝癌的发生。将进行一系列实验来评估这一中心假设。拟议中的实验结果有望揭示转化生长因子-？和前列腺素信号通路在肝癌发生中的作用，并提供重要的治疗意义。

DESCRIPTION (provided by applicant): Primary liver cancer is the common malignant neoplasm in human with high mortality and its incidence is rising worldwide, especially in the United States. It occurs largely in the preexisting chronic inflammatory liver disorders, including chronic hepatitis and cirrhosis. Recent studies from our lab show that prostaglandin (PG) metabolism plays an important role in liver inflammation and carcinogenesis. In this grant we hypothesize that the level and activation status of prostaglandin signaling represents a key factor that determines the cellular response to transforming growth factor-? (TGF-?). Specifically, we postulate that enhanced cytosolic phospholipase A2? (cPLA2?) and cyclooxygenase-2 (COX-2) controlled PG signaling subverts TGF-?-mediated mitoinhibition and this mechanism is critically involved in liver carcinogenesis through selection and expansion of TGF-? resistant dysplastic and neoplastic epithelial cells that progress more rapidly toward malignant transformation and tumor development. Therefore, blocking PG signaling may restore the growth- inhibitory action of TGF-??and prevent hepatocarcinogenesis. This application proposes a series of experiments to evaluate the above hypotheses. Human liver cancer cells with altered expression of cPLA2? and COX-2 will be utilized to determine their response to TGF-?. siRNA for Smad2/3 will be introduced into human liver cancer cells stably expressing antisense cPLA2? or COX-2 and these cells will be analyzed for proliferation and apoptosis, in vitro and in SCID mice. Transgenic mice with targeted expression of cPLA2? and COX- 2 in the liver will be developed and utilized to determine TGF-?-regulated Smad activation, mitoinhibition, apoptosis, and hepatocarcinogenesis. The cPLA2? and COX-2 transgenic and knockout mice will be crossed with the TGF-?receptor type II knockout mice to determine liver regeneration and DEN-induced hepatocarcinogenesis. Finally, cultured hepatic stellate cells and experimental animal models will be utilized to evaluate our hypothesis that TGF-? and PG signaling in the fibrogenic hepatic stellate cells is critically involved in the pathogenesis of liver fibrosis and carcinogenesis. Results from the proposed studies are expected to provide important therapeutic implications for the chemoprevention and treatment of human liver cancer. PUBLIC HEALTH RELEVANCE: Primary liver cancer is a highly malignant neoplasm in human and currently there is no effective chemoprevention or systematic therapy. This application is proposed to examine our hypothesis that the level and activation status of prostaglandin signaling represents a key factor that determines the cellular response to TGF-? and that blocking prostaglandin signaling may restore the growth-inhibitory action of TGF-? and prevent hepatocarcinogenesis. A series of experiments will be performed to evaluate this central hypothesis. Results of the proposed experiments are expected to reveal an important link between TGF-? and prostaglandin signaling pathways in liver carcinogenesis and provide important therapeutic implications.