Role of CITED2 in Breast Cancer Bone Metastasis

描述(申请人提供)：转移是乳腺癌患者死亡的最终原因，比任何其他部位更容易发生在骨骼中。此外，大约80%的晚期乳腺癌患者会发生骨转移，并导致相当大的发病率，表现为骨痛、病理性骨折、神经压迫和危及生命的高钙血症。可悲的是，骨转移通常是无法治愈的，确诊后的中位生存期只有2年。尽管正在进行研究，但调控骨转移和肿瘤诱导的骨溶解的分子和细胞机制仍然知之甚少。确定控制这些事件的因素将是对这种毁灭性疾病进行预防和治疗干预的理想目标。利用乳腺癌转移的小鼠模型，我们确定转录共激活因子CITED2是骨转移的潜在介质。降低CITED2在小鼠乳腺肿瘤细胞中的水平显著减少体内骨转移和骨溶解的建立。支持CITED2在人类乳腺癌中的作用，CITED2在人类原发乳腺肿瘤中的水平与生存呈负相关，在转移中显著升高，在骨转移中水平最高。此外，在初步研究中，人类乳腺癌细胞中CITED2水平的增加显著缩短了生存时间，增加了后肢瘫痪的发生率，并增加了小鼠的骨破坏。此外，CITED2还诱导了Runx2以及OCL激活因子IL-8、COX-2和PTHrP等Runx2靶点的表达，并定位于Runx2启动子，提示CITED2可能正向调控骨转移介导物Runx2的转录。基于这些初步数据，我们推测CITED2在人乳腺癌骨转移和病理性骨丢失中起关键作用。首先，我们将通过确定增加或减少CITED2表达对人乳腺癌细胞建立骨转移和诱导小鼠骨破坏能力的影响来初步验证我们的假设，这些影响包括：i)乳腺内生长，ii)进入血管系统，并直接进入骨骼。接下来，我们将通过一系列实验来研究CITED2在调控Runx2表达中的作用，以确定CITED2促进Runx2转录的辅因子(S)。随后，我们将通过检测这些因子表达的增加或减少对CITED2增强体外迁移和侵袭能力以及体内定植和骨破坏的影响来确定Runx2及其可能的靶点IL-8、COX-2和PTHrP是否介导了CITED2对人乳腺癌细胞的促转移作用。如果成功，这些研究将确定一种新的骨转移介质，并为其预防和治疗开辟新的途径。

DESCRIPTION (provided by applicant): Metastasis is the ultimate cause of mortality in breast cancer patients, developing in the bone more frequently than any other site. Moreover, bone metastasis occurs in approximately 80% of advanced breast cancer patients and causes considerable morbidity in the form of bone pain, pathological fractures, nerve compression, and life-threatening hypercalcemia. Sadly, bone metastasis is frequently incurable and the median survival time following diagnosis is just 2 years. Despite ongoing research efforts, the molecular and cellular mechanisms that regulate the establishment of bone metastasis and tumor-induced osteolysis remain poorly understood. Identification of factors regulating these events would be ideal targets for preventative and therapeutic interventions against this devastating disease. Using a mouse model of breast cancer metastasis, we identified the transcriptional co-activator CITED2 as a potential mediator of bone metastasis. Reducing CITED2 levels in mouse mammary tumor cells significantly reduced the establishment of bone metastasis and osteolysis in vivo. Supporting a role for CITED2 in human breast cancer, CITED2 levels in human primary breast tumors were inversely correlated with survival and were significantly elevated in metastasis, with highest levels in bone metastasis. Further, in preliminary studies, increased levels of CITED2 in human breast cancer cells significantly reduced survival time, increased incidence of hind limb paralysis, and elevated bone destruction in mice. In addition, CITED2 induced the expression of Runx2 as well as the putative Runx2 targets, OCL-activating factors IL-8, COX-2, and PTHrP, and was localized to the Runx2 promoter in human breast cancer cells, indicating that CITED2 may positively regulate transcription of Runx2, a reported mediator of bone metastasis. Based on these preliminary data, we hypothesize that CITED2 plays a key role in mediating human breast cancer metastasis to bone and pathological bone loss. We will initially investigate our hypothesis by determining the effect of increasing or decreasing CITED2 expression on the ability of human breast cancer cells to establish bone metastasis and induce bone destruction in mice following i) growth in the mammary gland, ii) administration into the vascular system, and direct administration into the bone. Next, we will examine the role of CITED2 in regulating Runx2 expression in a series of experiments identifying the co-factor(s) through which CITED2 enhances Runx2 transcription. Subsequently, we will determine whether Runx2 and its putative targets IL-8, COX-2, and PTHrP mediate the pro-metastatic effects of CITED2 on human breast cancer cells by examining the effect of increasing or decreasing expression of these factors on the ability of CITED2 to enhance in vitro migration and invasion, and in vivo colonization of bone and bone destruction. If successful, these studies will identify a novel mediator of bone metastasis and a new avenue for its prevention and treatment.