MECHANISMS OF KSHV-INDUCED CELLULAR TRANSFORMATION

这个子项目是许多利用 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 卡波西肉瘤（KS）是一种皮肤和内脏器官的血管增生性肿瘤，虽然在免疫功能正常的个体中很少见，但却是HIV阳性患者中最常见的癌症。人类疱疹病毒8型（humanherpesvirus 8，HHV 8/KSHV）是KS的病原体，感染的是表征病变的非典型内皮细胞和梭形细胞，在驱动肿瘤发展中起积极作用。该项目的目标是表征KSHV将内皮细胞转化为梭形细胞并促进肿瘤形成的分子机制。 我们已经建立了一个基于KSHV感染真皮微血管内皮细胞（DMVEC）的体外模型。DMVEC可以在体外被KSHV感染，并且感染导致梭形细胞形成和获得转化表型。KSHV感染的DMVEC反映了KS肿瘤中的基因表达，其中大多数感染的细胞以潜伏状态携带病毒基因组。我们已经使用DNA微阵列分析，然后使用靶向药物，反义寡核苷酸或RNA干扰的基因沉默方法，以确定和验证有助于KS肿瘤发生的细胞基因。 基因表达谱分析表明，许多原癌基因，和其他潜在的致瘤细胞基因，诱导在DMVEC转化KSHV感染。酪氨酸激酶抑制剂Gleevec的可用性使我们能够证明原癌基因c-Kit在KSHV诱导的DMVEC转化中的作用。使用反义和RNA干扰技术沉默c-Kit类似地抑制DMVEC转化，并证实这些基因沉默技术可用于验证其他细胞基因的作用。我们正在研究c-Kit诱导的机制以及癌基因激活引起的下游变化。此外，我们正在使用一种系统的方法来确定和验证其他KSHV诱导的致瘤基因。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Kaposi's sarcoma (KS) is an angioproliferative tumor of the skin and visceral organs that, while rare in immunocompetent individuals, is the most frequent cancer developing in HIV-positive patients. Human herpesvirus 8 (HHV8/KSHV), the etiologic agent of KS, infects the atypical endothelial cells and spindle cells that characterize the lesion, and plays an active role in driving tumor development. The goal of this project is to characterize the molecular mechanisms through which KSHV transforms endothelial cells into spindle cells and contributes to tumor formation. We have developed an in vitro model based on KSHV infection of dermal microvascular endothelial cells (DMVEC). DMVEC can be infected with KSHV in vitro, and infection leads to spindle cell formation and acquisition of a transformed phenotype. The KSHV-infected DMVEC mirror the gene expression seen in KS tumors whereby the majority of infected cells harbor the viral genome in a latent state. We have used DNA microarray analysis, followed by a gene silencing approach using targeted drugs, antisense oligonucleotides or RNA interference, to identify and validate cellular genes that contribute to KS tumorigenesis. Gene expression profiling has shown that a number of proto-oncogenes, and other potentially tumorigenic cellular genes, are induced in DMVEC transformed by KSHV infection. The availability of the tyrosine kinase inhibitor Gleevec allowed us to demonstrate a role for the proto-oncogene c-Kit in KSHV-induced DMVEC transformation. Silencing of c-Kit using antisense and RNA interference technology similarly inhibited DMVEC transformation and confirmed that these gene silencing techniques could be used to validate the role of additional cellular genes. We are investigating the mechanisms of c-Kit induction as well as downstream changes resulting from oncogene activation. In addition, we are using a systematic approach to identify and validate additional KSHV-induced tumorigenic genes.