Targeting ASAP1-Controlled Signal Pathways to Inhibit Uveal Melanoma Metastasis

项目摘要：“靶向ASAP 1控制的信号通路抑制葡萄膜黑色素瘤 转移” 葡萄膜黑色素瘤（UM）约占所有黑色素瘤的5%，发病率为每百万人4-11例 在西方国家，每年。尽管我们已经对信号通路有了很多了解， 控制UM肿瘤发生，对控制转移性疾病的途径知之甚少。预防或 有效地治疗转移性疾病是关键的，因为它发生在约50%的患者中（最常见于 一旦出现，预后很差。染色体8 q的扩增是一个强有力的 转移性疾病的预测因子。位于染色体8 q上的ASAP 1已被证明控制 乳腺癌的转移。ASAP 1在约40%的UM患者中扩增，并且增加ASAP 1 表达水平与增加的转移和不良预后相关。ASAP 1的过表达 在体外伤口愈合试验中，在低级别肿瘤细胞中增加细胞迁移。此外，ASAP 1 与几种癌症的转移有关， 控制细胞骨架结构的重塑。我们最近开始探索 控制葡萄膜黑色素瘤转移的途径，并有初步数据显示，ASAP 1控制 葡萄膜黑色素瘤细胞在人葡萄膜原位异种移植模型中的侵袭、增殖和转移 黑素瘤此外，我们已经鉴定了几种肿瘤相关的转录因子，包括STAT 4， COUP-TF I和COUP-TF II在UM细胞中被ASAP 1过表达激活，提示了一种新的 ASAP 1也可以控制UM转移的机制。因此，我们假设ASAP 1 通过激活诱导肿瘤相关的信号通路促进葡萄膜黑色素瘤转移 通过药理学抑制来抑制这些途径将显著降低 转移性疾病我们将通过追求两个目标来检验这一假设。在目标1中，我们将确定ASAP 1 促进UM细胞侵袭和增殖的结构域。这些研究将涉及表达全长 ASAP 1及其在ASAP 1敲低后的序列缺失构建体，以确定ASAP 1的作用 UM单元格中的域。在目标2中，我们将评估ASAP 1控制的转录因子在细胞凋亡中的功能。 葡萄膜黑色素瘤细胞的侵袭、增殖和肿瘤转移。细胞培养实验将涉及 基因表达的敲低或药理学抑制以确定这些转录的作用 UM细胞中的因子。体内实验将确定是否药理学抑制ASAP 1激活的 在人UM的异种移植模型中，转录因子减少肿瘤进展和转移。为这些 在这些研究中，我们将使用JAK抑制剂和CIA 1，它们分别抑制STAT 4和COUP-TF II。这 研究应该为我们提供新的见解，转移性扩散的分子机制， 并可能导致新的方法来预防或治疗转移性UM。

Project Summary: "Targeting ASAP1-Controlled Signal Pathways to Inhibit Uveal Melanoma Metastasis" Uveal melanoma (UM) accounts for about 5% of all melanomas and has an incidence of 4-11 per million people per year in western countries. Although much has been learned about the signaling pathways that control UM oncogenesis, far less is known about the pathways that control metastatic disease. Preventing or effectively treating metastatic disease is critical because it occurs in about 50% of patients (most often in the liver) and once present, the prognosis is dismal. Amplification of chromosome 8q is one of the strong predictors of metastatic disease. ASAP1, which is located on chromosome 8q, has been shown to control metastasis in breast cancer. ASAP1 is amplified in about 40% of UM patients, and increased ASAP1 expression levels are associated with increased metastasis and poor prognosis. Overexpression of ASAP1 in low-grade tumor cells increases cell migration in an in vitro wound-healing assay. Furthermore, ASAP1 has been implicated in the metastasis of several cancers and is thought to promote metastasis by directly controlling the remodeling of the cytoskeletal architecture. We have recently begun to explore the molecular pathways that control uveal melanoma metastasis and have preliminary data showing that ASAP1 controls uveal melanoma cell invasion, proliferation, and metastasis in an orthotopic xenograft model of human uveal melanoma. Furthermore, we have identified several tumor-associated transcription factors, including STAT4, COUP-TF I, and COUP-TF II, that are activated by overexpression of ASAP1 in UM cells, suggesting a novel mechanism by which ASAP1 may also control UM metastasis. Therefore, we hypothesize that ASAP1 promotes uveal melanoma metastasis by activating signaling pathways that induce tumor-associated transcription factors and that inhibiting these pathways by pharmacologic inhibition will significantly reduce metastatic disease. We will test this hypothesis by pursuing two aims. In Aim 1, we will identify ASAP1 domain(s) that promote UM cell invasion and proliferation. These studies will involve expressing full-length ASAP1 and its serial deletion constructs following ASAP1 knockdown to determine the role of ASAP1 domains in UM cells. In Aim 2, we will assess the function of ASAP1-controlled transcription factor(s) in uveal melanoma cell invasion, proliferation, and tumor metastasis. Cell culture experiments will involve either knockdown of gene expression or pharmacological inhibition to determine the role of these transcription factors in UM cells. In vivo experiments will determine whether pharmacologic inhibition of ASAP1- activated transcription factors reduce tumor progression and metastasis in a xenograft model of human UM. For these studies, we will use JAK inhibitors and CIA1, which inhibit STAT4 and COUP-TF II, respectively. This research should provide us with new insights into the molecular mechanisms underlying metastatic spread of UM and may lead to novel approaches to prevent or treat metastatic UM.