Regulation of Par-4 Secretion in Normal Cells for Paracrine Action in Tumor Cells

描述(申请人提供)：肺癌是美国男性和女性癌症死亡的主要原因。导致RAS癌基因激活和p53抑癌基因失活的突变是肺癌中最常见的两种改变。由于化疗和放射治疗的肿瘤生长抑制作用一般需要野生型P53的功能，携带P53突变或缺失的肺癌患者会对治疗产生抵抗，最终导致患者死亡。此外，致癌的RAS激活和P53失活导致核因子-kB活性升高，从而促进细胞存活和治疗耐药。肿瘤生长受到正常细胞和癌细胞之间相互作用的复杂网络的调控。除了局部的相互作用，肿瘤的生长还可能受到宿主系统产生的生长促进因子和生长抑制因子的影响。因此，癌症治疗可以适当地用来调节系统环境，以便直接针对异常生长。了解正常细胞系统性分泌促凋亡蛋白的机制及其对肿瘤上皮细胞的影响是此类研究的关键方面。耳癌患者体内的大部分细胞是正常的，具有野生型P53状态。这项建议将探索正常细胞中的P53激活是否会触发P53缺失的肺肿瘤中旁分泌细胞的死亡。由于P53的激活只能部分抑制核因子-kB的活性，我们的初步研究利用正常细胞中P53的共同激活和核因子-kB的抑制来鉴定可导致P53缺失的肺癌细胞旁分泌的凋亡效应的分泌蛋白(S)。根据我们的发现，我们假设p53调节正常细胞分泌促凋亡肿瘤抑制蛋白PAR-4，以实现癌细胞的旁分泌作用，并且P53激活和抑制正常细胞的核因子-kB最大限度地诱导PAR-4蛋白的表达，应该能有效地诱导P53缺陷的小鼠肺肿瘤的凋亡。拟议的研究将通过追求两个具体目标来探索这一新调查领域的可行性。(目的1)探讨正常细胞中P53激活诱导肿瘤细胞凋亡的机制。我们将研究P53在不同正常细胞类型中诱导PAR-4分泌的动力学和机制，从而导致P53缺陷的肺癌细胞的凋亡。(目的2)探讨正常细胞中P53活化诱导的PAR-4分泌对P53缺陷型肺癌的旁分泌作用。在同时激活p53和抑制NF-kB活性后，我们将检测K-rasLSL-G12D、p53F1/F1小鼠全身PAR-4蛋白升高的动力学及其对细胞凋亡和肺癌消退的影响。此外，将进行机制研究，以更深入地了解PAR-4在小鼠这些肿瘤的旁分泌作用中的功能作用。总而言之，这些研究将揭示正常细胞中P53激活的旁分泌效应在P53缺陷肺癌的凋亡和消退中的可行性和机制。由于p53在不同的癌症中发生突变，这项研究的发现将具有广泛的翻译意义。

DESCRIPTION (provided by applicant): Lung cancer is a leading cause of cancer death in men and women in the United States. Mutations leading to activation of the Ras oncogenes and inactivation of the p53 tumor suppressor gene are two of the most common alterations in lung cancer. As the tumor growth inhibitory action of chemotherapy and radiation therapy generally requires wild-type p53 function, lung tumors with p53 mutations or deletions develop resistance to therapy resulting in ultimate death of the patients. Moreover, oncogenic Ras activation and p53 inactivation results in elevated NF-kB activity that contributes to cell survival and therapeutic resistance. Tumor growth is regulated by a complex network of interactions between normal cells and cancer cells. In addition to local interactions, tumor growth may be influenced by growth-promoting and growth-inhibiting factors produced systemically by the host. Accordingly, cancer therapeutics can be aptly utilized to regulate the systemic environment in order to directly target aberrant growth. Understanding the mechanisms associated with systemic secretion of pro-apoptotic proteins by normal cells and their effect on tumor epithelial cells is a critical facet of such investigations. A majority of the cells throughout the body of lug cancer patients are normal and have wild type p53 status. This proposal will explore whether p53 activation in normal cells triggers paracrine cell death in p53-deficient lung tumors. As p53 activation only partially inhibits NF-kB activity, our Preliminary Studies used co-parallel activation of p53 and inhibition of NF-kB in normal cells to identify secreted protein(s) that can cause a paracrine apoptotic effect in p53-deficient lung cancer cells. Based on our findings, we hypothesize that p53 regulates the secretion of the pro-apoptotic tumor suppressor protein Par-4 from normal cells for paracrine effect in cancer cells, and that maximum induction of Par-4 protein by p53 activation and NF-kB inhibition from normal cells should effectively induce apoptosis in p53-deficient lung tumors in mice. The proposed study will explore the feasibility of this novel area of investigation by pursuing two Specific Aims. (Aim 1) Determine the mechanism by which p53 activation in normal cells induces tumor cell apoptosis. We will study the kinetics and mechanism of Par-4 secretion induced by p53 in diverse normal cell types leading to apoptosis of p53-deficient lung cancer cells. (Aim 2) Determine the paracrine effect of Par-4 secretion, which is induced by p53 activation in normal cells, on p53-deficient lung tumors. After concomitant activation of p53 and inhibition of NF-kB activity, we will test the kinetics of systemic elevation of Par-4 protein and its effect on apoptosis and regression of lung tumors in K-rasLSL-G12D,p53Fl/Fl mice. Moreover, mechanistic studies will be undertaken to gain deeper insights into the functional role of Par-4 in the paracrine action on these tumors in mice. Collectively, these studies will uncover the feasibility and mechanism of the paracrine effects of p53 activation in normal cells for apoptosis and regression of p53-deficient lung cancer. As p53 is mutated in diverse cancers, the findings of this study will have broad translational implications.