Prevention of Lung Cancer Growth by Angiotensin-(1-7)

描述（由申请人提供）： 肺癌是男性和女性死亡的主要原因，在美国每年约有160，000人死亡，全世界有超过1，000，000例新发肺癌病例。尽管有许多临床试验测试了各种化合物作为化学预防剂，但没有一种能有效降低肺癌的风险。然而，用血管紧张素转换酶（ACE）抑制剂治疗高血压患者可降低患癌症的风险，尤其是肺癌。抑制ACE活性可降低血管紧张素II（Ang II），并增加缓激肽和血管紧张素-（1-7）[Ang-（1-7）]的水平。我们发现，Ang-（1-7）在体外可减少血管生长，在体内可防止新生内膜形成。更重要的是，我们的初步研究表明，Ang-（1-7）在体内显著抑制人肺癌细胞的生长并减少肿瘤的生长。因此，我们提出对Ang-（1-7）的抗增殖反应超出其对血管细胞的作用，并且Ang-（1-7）抑制癌细胞的生长。本申请中概述的研究旨在确定Ang-（1-7）是否抑制人肺癌细胞的增殖并鉴定参与该调节的分子机制。建立的人肺癌细胞系将在用不同剂量的Ang-（1-7）处理后检查生长抑制。通过3 H-胸苷掺入增殖细胞、使用改良的MTT试验减少细胞数量以及通过流式细胞术分析细胞周期来评估细胞生长的减弱。将使用其它血管紧张素肽和受体拮抗剂测定对Ang-（1-7）的抗增殖反应的特异性。将在具有化学诱导的肺肿瘤的小鼠中检查Ang-（1-7）对肺癌细胞生长的减弱，以证明Ang-（1-7）预防或减少体内肺肿瘤形成。基因阵列分析确定了在促分裂原刺激人肺癌细胞后由Ang-（1-7）调节的候选基因，包括编码肿瘤抑制因子和细胞死亡信号的基因的上调和编码刺激细胞生长的信号通路的基因的下调。通过逆转录-聚合酶链反应（RT-PCR）分析和蛋白质印迹杂交来评估Ang-（1-7）对特定基因和蛋白质的调节，以确定它们在Ang-（1-7）介导的肺癌细胞生长抑制中的作用。这些研究的结果将确定Ang-（1-7）是否有效地减弱人肺癌细胞的增殖，并确定参与抑制反应的细胞途径。这项工作的最终目标是确定施用Ang-（1-7）或增加循环Ang-（1-7）的药剂是否是肺癌的有效化学预防治疗。

DESCRIPTION (provided by applicant): Lung cancer is a leading cause of death in both men and women, accounting for approximately 160,000 deaths annually in the United States with over 1,000,000 new cases of lung cancer worldwide. Despite a number of clinical trials testing various compounds as chemopreventive agents, none were effective in reducing the risk of lung cancer. However, treatment of hypertensive patients with angiotensin converting enzyme (ACE) inhibitors reduced the risk of cancer, especially lung cancer. Inhibition of ACE activity decreases angiotensin II (Ang II) and increases the levels of both bradykinin and angiotensin-(1-7) [Ang-(1-7)]. We showed that Ang-(1-7) reduces vascular growth in vitro and prevents neointimal formation in vivo. More importantly, our preliminary studies showed that Ang-(1-7) markedly inhibits the growth of human lung cancer cells and reduces tumor growth in vivo. Thus, we propose that the antiproliferative response to Ang-(1-7) extends beyond its effects on vascular cells and that Ang-(1-7) inhibits the growth of cancer cells. The studies outlined in this application are designed to determine whether Ang-(1-7) inhibits the proliferation of human lung cancer cells and identify the molecular mechanisms involved in this regulation. Established cell lines of human lung carcinomas will be examined for growth inhibition following treatment with various doses of Ang-(1-7). Attenuation of cell growth will be assessed by 3H-thymidine incorporation into proliferating cells, reductions in cell number using the modified MTT assay, and cell cycle analysis by flow cytometry. The specificity of the antiproliferative response to Ang-(1-7) will be determined using other angiotensin peptides and receptor antagonists. The attenuation of lung cancer cell growth by Ang-(1-7) will be examined in mice with chemically-induced lung tumors, to demonstrate that Ang-(1-7) prevents or reduces lung tumor formation in vivo. Gene array analysis identified candidate genes regulated by Ang-(1-7) following mitogen stimulation of human lung cancer cells, including up-regulation of genes encoding tumor suppressors and cell death signals and downregulation of genes encoding signaling pathways stimulating cell growth. Regulation of specific genes and proteins by Ang-(1-7) will be assessed by reverse transcriptase-polymerase chain reaction (RT-PCR) assays and Western blot hybridization, to determine their role in Ang-(1-7)-mediated lung cancer cell growth inhibition. The results of these studies will determine whether Ang-(1-7) effectively attenuates the proliferation of human lung cancer cells and identify the cellular pathways involved in the inhibitory response. The ultimate goal of this work is to determine whether administration of Ang-(1-7) or agents, which increase circulating Ang-(1-7) is an effective chemopreventive treatment for lung cancers.