Molecular Regulation of Human Melanoma Metastasis

描述（由申请人提供）：本申请的长期目标是确定白介素(IL)-8在恶性黑色素瘤发病机制中的分子机制。IL-8是CXC趋化因子家族的一员，在恶性黑色素瘤中组成性表达，是一种自分泌/旁分泌生长、侵袭性和血管生成因子。IL-8的作用是通过在黑色素瘤细胞和内皮细胞上表达的两个高亲和力受体CXCR1和CXCR2介导的。IL-8与CXCR1和CXCR2结合可引发多种细胞反应，然而，IL-8激活CXCR1和/或CXCR2在恶性黑色素瘤中的确切作用尚不清楚。因此，本应用的具体目的是确定CXCR1和CXCR2在介导il -8诱导的调节黑色素瘤生长、侵袭、血管生成和转移的细胞反应中的独特功能作用。我们假设IL-8与CXCR1和CXCR2的相互作用激活了不同的下游信号通路，并在黑色素瘤的生长、侵袭、血管生成和转移中发挥了多种作用。我们将验证这一假设，并通过以下方式实现该应用的目标：1)确定IL-8激活CXCR1和/或CXCR2是否在调节与黑色素瘤生长、侵袭和转移相关的细胞表型中具有不同或重叠的作用；2)研究内皮细胞中CXCR1和CXCR2在il -8诱导的黑色素瘤血管生成反应中的不同作用；3)确定il -8诱导的CXCR1-和cxcr2依赖性调节黑色素瘤细胞生长、存活和侵袭反应的信号转导机制。我们将在体外和体内研究CXCR1和cxcr2依赖性il -8介导的作用。我们将在异种移植模型和mCXCR2敲除裸鼠中使用小分子拮抗剂和腺病毒sirna载体在体内特异性靶向CXCR1和/或CXCR2。利用途径特异性抑制剂、磷酸化特异性抗体、免疫印迹和免疫组织化学，我们将在体外和体内研究CXCR1和/或CXCR2激活后的下游信号通路。这些研究将确定CXCR1和CXCR2激活及其下游信号通路在介导il -8诱导的恶性黑色素瘤生长、血管生成和转移中的独特和/或重叠作用。我们预计，从这些研究中获得的知识将确定新的治疗靶点，以抑制配体结合和/或信号转导事件，并开发恶性黑色素瘤的治疗方法。

DESCRIPTION (provided by applicant): The long-term objective of this application is to determine the molecular mechanism(s) of interleukin (IL)-8 in malignant melanoma pathogenesis. IL-8, a member of the CXC chemokine family is constitutively expressed in malignant melanoma and functions as an autocrine/paracrine growth, invasive and angiogenic factor. The effect of IL-8 is mediated through two high affinity receptors, CXCR1 and CXCR2, expressed on melanoma cells as well as endothelial cells. Binding of IL-8 to CXCR1 and CXCR2 can initiate diverse cellular responses, however, the precise role of activation of CXCR1 and/or CXCR2 by IL-8 in malignant melanoma is not known. Therefore, the specific objective of this application is to determine the distinct functional role of CXCR1 and CXCR2 in mediating IL-8-induced cellular responses that regulate melanoma growth, invasion, angiogenesis, and metastasis. We hypothesize that interaction of IL-8 with CXCR1 and CXCR2 activates different downstream signaling pathways and plays a diverse role in melanoma growth, invasion, angiogenesis, and metastasis. We will test this hypothesis and accomplish the objective of this application by: 1) Determining whether activation of CXCR1 and/or CXCR2 by IL-8 result in distinct or overlapping roles in regulating cellular phenotypes associated with melanoma growth, invasion and metastasis; 2) Examining the diverse roles of CXCR1 and CXCR2 in endothelial cells in an IL-8-induced angiogenic response in melanoma; and 3) Identifying the signal transduction mechanism(s) involved in IL-8-induced CXCR1- and CXCR2-dependent modulation of melanoma cell growth, survival and invasive response. We will examine the CXCR1 and CXCR2-dependent IL-8-mediated effects in vitro and in vivo. We will use specific targeting of CXCR1 and/or CXCR2 in vivo using small molecule antagonists and adenoviral-siRNA vectors in xenograft models and mCXCR2 knockout nude mice. Using pathways specific inhibitors, phospho-specific antibodies, immunoblotting and immunohistochemistry, we will examine the downstream signaling pathways following CXCR1 and/or CXCR2 activation in vitro and in vivo. These studies will identify distinct and/or overlapping roles for CXCR1 and CXCR2 activation and their downstream signaling pathways in mediating IL-8-induced responses in malignant melanoma growth, angiogenesis, and metastasis. We anticipate that the knowledge gained from these studies will identify new therapeutic targets for inhibiting the ligand binding and/or signal transduction events and the development of therapeutics for malignant melanoma.