Mechanisms of KSHV-Induced Cellular Transformation

描述(申请人提供)：卡波西氏肉瘤(KS)是艾滋病相关的最常见的恶性肿瘤，也是艾滋病流行的非洲部分地区最常见的癌症。感染Kaposi肉瘤疱疹病毒(KSHV；又称HHV8)存在于梭形细胞、其EC前体细胞和KS病变的单核细胞中，以及循环中的EC前体细胞和PBMC中。KS的几个特征表明趋化因子及其受体(CR)在疾病中发挥作用。KS肿瘤表现出深刻的血管生成和广泛的炎性浸润，这两个特征都表明趋化因子/CR的活性。此外，KS的多灶性表明，KSHV感染的梭形细胞前体或白细胞聚集在允许的部位，以启动或维持肿瘤的发展。SDF-1/CXCR4轴是与肿瘤生长、血管生成和转移相关的主要趋化因子/受体相互作用之一。然而，SDF-1/CXCR4在KS中的作用仍然不清楚；迄今为止进行的有限研究仅显示这些分子存在于KS肿瘤中。我们已经证明，最近鉴定的SDF-1的替代受体RDC1(也称为CXCR7)在KSHV感染的EC中被强烈诱导。虽然RDC1在非恶性细胞中的表达有限，但在几种癌症和肿瘤相关的EC中都有很好的表达。我们最近发现RDC1在KS肿瘤组织中高表达，我们和其他人发现SDF-1信号通过RDC1在几种细胞类型中促进迁移、血管生成、生长和促生存活性。这些发现表明，SDF-1通过其受体CXCR4和RDC1中的任何一个来传递信号，在KS肿瘤的种植和随后的发展中发挥重要作用。我们假设RDC1和CXCR4在病理上是由KSHV在EC/梭形细胞中诱导或激活的，而肿瘤微环境中的SDF-1和/或EC/梭形细胞共表达的SDF-1激活了下游信号通路，导致了促血管生成、促生长、促生存的表型，并促进了细胞向病变发生部位的运输。为了了解SDF-1/CXCR4和SDF-1/RDC1在KS肿瘤生长、发生和发展中的作用，我们提出了三个特定的靶点。目的1研究SDF-1、CXCR4和RDC1(I)在体内KS肿瘤和(II)体外不同类型KSHV感染的内皮细胞中的表达模式。在目标2中，我们将确定(A)SDF-1/CXCR4和(B)SDF-1/RDC1信号在KSHV感染的EC中的性质和功能。在目标3中，我们将使用一组KSHV血清阳性患者的样本来计数CEC数量，并确定SDF-1、CXCR4和RDC1在这些细胞上的表达和功能。此外，还将对正常和队列CEC进行功能研究，这些CEC在体外扩增为血液生长的EC。CXCR4拮抗剂目前正处于临床前和临床开发阶段，用于治疗白血病、肺癌和乳腺癌。最近开发了特定的RDC1拮抗剂，并在动物模型中显示了抗肿瘤活性。因此，明确SDF-1/CXCR4和SDF-1RDC1轴在KS中的作用可能为治疗干预提供新的途径。 与公共健康相关：趋化因子是一种分泌蛋白，通过与反应细胞上的特定趋化因子受体(CR)结合来调节细胞功能，通过刺激新的血管生长(肿瘤血管生成)和指导恶性或癌前细胞的迁移，在人类肿瘤的发展、生长和转移中发挥重要作用。KS肿瘤细胞(梭形细胞)同时表达参与肿瘤转移和血管生成的关键趋化因子SDF-1/CXCL12的CRs(CXCR4和RDC1/CXCR7)，我们的初步数据表明KSHV诱导了这些趋化因子/CR对的表达和活性。在这项应用中，我们将研究KSHV诱导SDF-1、CXCR4和RDC1的机制和功能结果，这些信息将提高我们对KS发病机制的理解，并表明阻断趋化因子/CR相互作用的药物是否值得探索用于KS治疗。

DESCRIPTION (provided by applicant): Kaposi's sarcoma (KS) is the most common AIDS-associated malignancy and the most frequent cancer in parts of Africa where HIV is endemic. The infectious agent, Kaposi's sarcoma herpesvirus (KSHV; also HHV8) is found in spindle cells, their EC precursors and monocytes in KS lesions, as well as in circulating EC progenitors and PBMC. Several features of KS suggest that chemokines and their receptors (CR) play a role in the disease. KS tumors display profound angiogenesis and an extensive inflammatory infiltrate, both features indicative of chemokine/CR activity. Also, the multi-focal nature of KS suggests that KSHV-infected spindle cell precursors or leucocytes home to permissive sites to initiate or sustain tumor development. The SDF-1/CXCR4 axis is one of the main chemokine/CR interactions associated with tumor growth, angiogenesis and metastasis. The role of SDF-1/CXCR4 in KS however remains obscure; the limited studies performed to date reveal only that these molecules are present in KS tumors. We have shown that the recently characterized alternate receptor for SDF-1, RDC1 (also CXCR7), is strongly induced in KSHV-infected EC. While RDC1 expression is limited in non-malignant cells, it is well expressed by several cancers and tumor-associated EC. We recently found that RDC1 is highly expressed on KS tumor tissue and we, and others, have shown that SDF-1 signaling through RDC1 promotes migratory, angiogenic, growth and pro-survival activity in several cell types. These findings suggest that SDF-1, signaling through either of both of its receptors CXCR4 and RDC1, plays an important role in the seeding and subsequent development of KS tumors. We hypothesize that RDC1 and CXCR4 are pathologically induced or activated by KSHV in EC/spindle cells and that SDF-1 in the tumor microenvironment and/or co-expressed by EC/spindle cells activates downstream signaling pathways that lead to a pro-angiogenic, pro-growth, pro-survival phenotype and promote trafficking of cells to sites of lesion development. To understand the role of SDF-1/CXCR4 and SDF-1/RDC1 in KS tumor seeding, initiation and development, we propose three Specific Aims. In Aim 1 we propose to characterize the expression patterns of SDF-1, CXCR4 and RDC1 (i) in vivo, in KS tumors and (ii) in vitro, in different types of KSHV-infected endothelial cells. In Aim 2, we will determine the nature and function of (a) SDF-1/CXCR4 and (b) SDF-1/RDC1 signaling in KSHV-infected EC. In Aim 3, we will use samples from a cohort of KSHV seropositive patients with and without KS and HIV to enumerate CEC numbers and determine the expression and function of SDF-1, CXCR4 and RDC1 on these cells. Functional studies will also be conducted on normal and cohort CEC following their expansion in vitro as blood outgrowth EC. CXCR4 antagonists are currently in pre-clinical and clinical development for the treatment of leukemia, lung and breast cancer. Specific RDC1 antagonists have recently been developed and have shown anti-tumor activity in animal models. Thus, defining the role of SDF-1/CXCR4 and SDF-1RDC1 axes in KS may provide an avenue for therapeutic intervention. PUBLIC HEALTH RELEVANCE: Chemokines, secreted proteins that regulate cell function by binding to specific chemokine receptors (CR) on responsive cells, play an essential role in the development, growth and metastasis of human tumors by stimulating new blood vessel growth (tumor angiogenesis) and directing the migration of malignant or pre-malignant cells. KS tumor cells (spindle cells) express both of the CRs (CXCR4 and RDC1/CXCR7) for one of the key chemokines (SDF-1/CXCL12) involved in tumor metastasis and angiogenesis, and our preliminary data shows that KSHV induces the expression and activity of these chemokine/CR pairs. In this application we will investigate the mechanisms and functional outcomes of KSHV-induction of SDF-1, CXCR4 and RDC1, information that will both improve our understanding of KS pathogenesis and indicate if drugs that block chemokine/CR interactions are worth exploring for KS therapy.