Role of CITED2 in Breast Cancer Bone Metastasis

描述（由申请人提供）：转移是乳腺癌患者死亡的最终原因，在骨中发生的频率高于任何其他部位。此外，骨转移发生在大约80%的晚期乳腺癌患者中，并以骨痛、病理性骨折、神经压迫和危及生命的高钙血症的形式引起相当大的发病率。不幸的是，骨转移通常是无法治愈的，诊断后的中位生存时间仅为2年。尽管正在进行的研究工作，调节骨转移和肿瘤诱导的骨质溶解的建立的分子和细胞机制仍然知之甚少。确定调节这些事件的因素将是针对这种毁灭性疾病的预防和治疗干预的理想目标。使用乳腺癌转移的小鼠模型，我们鉴定了转录共激活因子CITED 2作为骨转移的潜在介质。降低小鼠乳腺肿瘤细胞中的CITED 2水平显著减少了体内骨转移和骨质溶解的建立。支持CITED 2在人类乳腺癌中的作用，人类原发性乳腺肿瘤中的CITED 2水平与存活率呈负相关，并且在转移中显著升高，在骨转移中水平最高。此外，在初步研究中，人乳腺癌细胞中CITED 2水平的增加显著缩短了小鼠的存活时间，增加了后肢瘫痪的发生率，并增加了骨破坏。此外，CITED 2诱导Runx 2以及推定的Runx 2靶点OCL激活因子IL-8、考克斯-2和PTHrP的表达，并且在人乳腺癌细胞中定位于Runx 2启动子，表明CITED 2可能正调控Runx 2的转录，Runx 2是已报道的骨转移的介导物。基于这些初步数据，我们假设CITED 2在介导人乳腺癌骨转移和病理性骨丢失中起关键作用。我们将首先通过确定增加或减少CITED 2表达对人乳腺癌细胞在小鼠中建立骨转移和诱导骨破坏的能力的影响来研究我们的假设，所述小鼠在i）乳腺中生长，ii）施用到血管系统中，和直接施用到骨中。接下来，我们将在一系列实验中研究CITED 2在调节Runx 2表达中的作用，这些实验鉴定了CITED 2通过其增强Runx 2转录的辅因子。随后，我们将通过检测增加或减少这些因子的表达对CITED 2增强体外迁移和侵袭以及体内骨定殖和骨破坏的能力的影响，来确定Runx 2及其假定靶点IL-8、考克斯-2和PTHrP是否介导CITED 2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，并为其预防和治疗提供新的途径。

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.