Role of Tissue Transglutaminase in Ovarian Cancer

描述(由申请人提供)： 我们发现组织谷氨酰胺转氨酶(TG2)是一种在卵巢癌(OC)中上调的蛋白质，它通过增强2-连环蛋白细胞信号，调节卵巢癌细胞上皮向间充质转化(EMT)和卵巢癌干细胞的存活。TG2是一种在多种上皮性恶性肿瘤中过度表达的酶，可诱导钙依赖蛋白的翻译后修饰和交联。我们首次报道了TG2在OC中表达上调，我们实验室在当前的资助期间的工作表明，该酶对OC的转移过程起着关键的调节作用。我们发现，当注射到裸鼠的腹膜内或卵巢囊下时，与对照细胞相比，表达降低TG2水平的工程OC细胞引起的腹膜播散显著减少。我们还发现TG2可以诱导OC细胞的EMT，并调节OC细胞球体的形成。这些发现导致我们提出假设，即TG2在EMT过程和卵巢癌干细胞的生存中起关键作用，我们将通过三个目标进行测试：目的1：确定TG2调节2-连环蛋白稳定性的机制，并研究其对2-连环蛋白依赖基因转录的影响。我们将研究TG2基因敲除或抑制是否改变了2-连环素的细胞定位、磷酸化和信号转导，以响应Wnt信号，从而促进OC细胞的EMT。目的：探讨TG2诱导的子宫内膜异位转移是否受2-连环蛋白信号的调控。我们将研究2-连环蛋白下调是否抑制TG2诱导的EMT和转移，以及它的稳定是否诱导低水平TG2表达的细胞的EMT。目的：探讨TG2基因表达对卵巢癌干细胞存活和功能的影响。我们将测量和比较TG2在卵巢CSCs和整个肿瘤中的表达。我们将敲除TG2或抑制其酶活性，以阻断卵巢CSCs的功能(如存活、球体形成和致瘤性)。意义：我们是第一个研究TG2在卵巢癌中的作用的小组，重点是TG2调节转移和EMT的机制。这一建议是及时和高度相关的，因为EMT最近被认为与癌症干细胞表型有关。我们小组已经提出了在卵巢肿瘤中存在这种独特的细胞群的证据。这些细胞在标准治疗后的持久性很可能是化疗未能根除癌症的原因。因此，证明TG2促进卵巢CSCs的存活是高度相关的，因为它将确定一条新的靶向途径，其目标是根除耐药干细胞和阻断OC转移。 公共卫生相关性： 与退伍军人健康和保健问题的相关性：卵巢癌是最致命的妇科恶性肿瘤，腹部器官转移是最常见的死亡原因。鉴于美国武装部队和退伍军人中的女性人数不断增加，OC的破坏性并发症和有限的治疗选择，开发更好的治疗方法的问题与我们退伍军人的健康高度相关。美国武装部队中的女性人数从2%增加到15%，退伍军人医院服务的退伍军人中7.4%是女性。这项建议将研究组织转谷氨酰胺酶在卵巢癌干细胞转移和生存中的作用。这些干细胞的持久性很可能是化疗未能根除癌症的原因。因此，对VA患者来说，更好地了解调节这些细胞的关键分子是很重要的，这些细胞会导致转移、治疗耐药和癌症复发。

DESCRIPTION (provided by applicant): Abstract The overarching hypothesis for this proposal is that tissue transglutaminase (TG2), a protein we found to be up-regulated in ovarian cancer (OC) regulates epithelial to mesenchymal transition (EMT) in OC cells and the survival of ovarian cancer stem cells by enhancing 2-catenin cellular signaling. TG2, an enzyme overexpressed in several epithelial malignancies, induces Cadependent protein post-translational modifications and cross-linking. We reported for the first time that TG2 is up-regulated in OC and work in our laboratory during the current funding period demonstrated that the enzyme critically regulates the process of OC metastasis. We showed that when injected intraperitoneally or under the ovarian bursa of nude mice, OC cells engineered to express decreased levels of TG2 induced significantly less peritoneal dissemination compared to control cells. We also showed that TG2 induces EMT of OC cells and regulates formation of spheroids from OC cells. These discoveries led us to formulate the hypothesis that TG2 is critical to the process of EMT and the survival of ovarian cancer stem cells, which we will test by addressing three objectives: Aim 1: Identify the mechanisms by which TG2 regulates 2-catenin stability and study its effects on 2-catenin dependent gene transcription. We will study whether TG2 knockdown or inhibition alters 2-catenin cellular localization, phosphorylation and signaling in response to Wnt signals, contributing to EMT of OC cells. Aim 2: Determine whether TG2 induced EMT and metastasis are regulated by 2-catenin signaling. We will investigate whether 2-catenin knock-down inhibits TG2 induced-EMT and metastasis and whether its stabilization induces EMT in cells expressing low levels of TG2. Aim 3: Establish the effects of TG2 expression on the survival and functions of ovarian cancer stem cells. We will measure and compare TG2 expression in ovarian CSCs vs. whole tumors. We will knock-down TG2 or inhibit its enzymatic activity to block the functions of ovarian CSCs (e.g. survival, formation of spheroids and tumorigenicity). Significance: We are the first group studying the role of TG2 in ovarian cancer focusing on mechanisms of metastasis and EMT modulated by TG2. This proposal is timely and highly relevant as EMT has been recently linked to the cancer-stem cell phenotype. Our group has produced evidence for the presence of such a distinct population of cells in ovarian tumors. It is likely that persistence of these cells after standard treatment is responsible for the failure of chemotherapy to eliminate cancer. Therefore, demonstrating that TG2 facilitates the survival of ovarian CSCs is highly relevant, as it will identify a new pathway that can be targeted with the goal of eradicating treatment resistant stem cells and of blocking OC metastasis. PUBLIC HEALTH RELEVANCE: Relevance to Veterans Health and Health Care Issues: Ovarian cancer is the most fatal gynecological malignancy, metastasis to abdominal organs being the most common cause of mortality. Given the increasing numbers of women in the US Armed Forces and the VA, the devastating complications of OC and limited treatment options, the issue of developing better therapies is highly relevant to the health of our veterans. The number of women in the US Armed Forces has increased from 2% to 15% and 7.4% of the veterans served by VA Hospitals are women. This proposal will investigate the role of tissue transglutaminase in metastasis and the survival of ovarian cancer stem cells. It is likely that the persistence of these stem cells is responsible for the failure of chemotherapy to eliminate cancer. It is therefore important to VA patients to better understand the key molecules that regulate these cells, which give rise to metastases, treatment resistance and cancer recurrence.