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.
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