KSHV-induced ligand mediated activation of PDGF receptor-alpha drives Kaposi's sarcomagenesis

Kaposi’s sarcoma (KS) herpesvirus (KSHV) causes KS, an angiogenic AIDS-associated spindle-cell neoplasm, by activating host oncogenic signaling cascades through autocrine and paracrine mechanisms. Tyrosine kinase receptor (RTK) proteomic arrays, identified PDGF receptor-alpha (PDGFRA) as the predominantly-activated RTK in KSHV-induced mouse KS-tumors. We show that: 1) KSHV lytic replication and the vGPCR can activate PDGFRA through upregulation of its ligands PDGFA/B, which increase c-myc, VEGF and KSHV gene expression in infected cells 2) KSHV infected spindle cells of most AIDS-KS lesions display robust phospho-PDGFRA staining 3) blocking PDGFRA-signaling with N-acetyl-cysteine, RTK-inhibitors Imatinib and Sunitinib, or dominant-negative PDGFRA inhibits tumorigenesis 4) PDGFRA D842V activating-mutation confers resistance to Imatinib in mouse-KS tumorigenesis. Our data show that KSHV usurps sarcomagenic PDGFRA signaling to drive KS. This and the fact that PDGFRA drives non-viral sarcomas highlights the importance for KSHV-induced ligand-mediated activation of PDGFRA in KS sarcomagenesis and shows that this oncogenic axis could be successfully blocked to impede KS tumor growth.Fil: Cavallin, Lucas E.. University of Miami; Estados UnidosFil: Ma, Qi. University of Miami; Estados UnidosFil: Naipauer, Julian. University of Miami; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Medicina Traslacional, Trasplante y Bioingeniería. Fundación Favaloro. Instituto de Medicina Traslacional, Trasplante y Bioingeniería; ArgentinaFil: Gupta, Sachin. University of Miami; Estados UnidosFil: Kurian, Mani. University of Miami; Estados UnidosFil: Locatelli, Paola. University of Miami; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Medicina Traslacional, Trasplante y Bioingeniería. Fundación Favaloro. Instituto de Medicina Traslacional, Trasplante y Bioingeniería; ArgentinaFil: Romanelli, Paolo. University of Miami; Estados UnidosFil: Nadji, Mehrdad. University of Miami; Estados UnidosFil: Goldschmidt Clermont, Pascal J.. University of Miami; Estados UnidosFil: Mesri, Enrique Alfredo. University of Miami; Estados Unido

In Vivo-Restricted and Reversible Malignancy Induced by Human Herpesvirus-8 KSHV: A Cell and Animal Model of Virally Induced Kaposi's Sarcoma

Transfection of a Kaposi's sarcoma (KS) herpesvirus (KSHV) Bacterial Artificial Chromosome (KSHVBac36) into mouse bone marrow endothelial lineage cells generates a cell (mECK36) that forms KS-like tumors in mice. mECK36 expressed most KSHV genes and were angiogenic, but didn't form colonies in soft agar. In nude mice, mECK36 formed KSHV-harboring vascularized spindle-cell sarcomas that were LANA+/podoplanin+, overexpressed VEGF and Angiopoietin ligands and receptors, and displayed KSHV and host transcriptomes reminiscent of KS. mECK36 that lost the KSHV episome reverted to non-tumorigenicity. siRNA suppression of KSHV vGPCR, an angiogenic gene up-regulated in mECK36 tumors, inhibited angiogenicity and tumorigenicity. These results show that KSHV malignancy is in vivo growth-restricted and reversible, defining mECK36 as a biologically sensitive animal model of KSHV-dependent KS

Unbiased Screening of Activated Receptor Tyrosine Kinases (RTKs) in Tumor Extracts Using a Mouse Phospho-RTK Array Kit

Kaposi's sarcoma (KS) herpesvirus (KSHV) is a virus that causes KS, an angiogenic AIDS-associated spindle-cell neoplasm, by activating host oncogenic signaling cascades through autocrine and paracrine mechanisms. Many host signaling cascades co-opted by KSHV including PI3K/AKT/mTORC, NFkB and Notch are critical for cell-specific mechanisms of transformation and their identification is paving the way to therapeutic target discovery. Analysis of the molecular KS signature common to human KS tumors and our mouse KS-like tumors showed consistent expression of KS markers VEGF and PDGF receptors with upregulation of other angiogenesis ligands and their receptors . This points to the autocrine and paracrine activation of various receptor tyrosine kinase (RTK) signaling axes. Hereby we describe a protocol to screen for activated receptor tyrosine kinase of KSHV-induced KS-like mouse tumors using a Mouse Phospho-RTK Array Kit and its validation by RTK western blots. We showed that this method can be successfully used to rank the tyrosine kinase receptors most activated in tumors in an unbiased manner. This allowed us to identify PDGFRA as an oncogenic driver and therapeutic target in AIDS-KS

Abstract 2710: KSHV-induced PDGF signaling dyregulation drives a Rac1 and ROS-mediated paracrine tumorigenic mechanism necessary for Kaposi's sarcoma

Abstract Kaposi's sarcoma (KS), caused by the Kaposi's sarcoma-associated herpesvirus (KSHV), is a major AIDS-associated malignancy characterized by angiogenesis and proliferation of spindle cells. KSHV-infected KS lesions are composed of latently-infected cells and a minority of cells expressing lytic genes such as vGPCR, which are implicated the development of KS angioproliferative phenotype via a paracrine mechanism. A key unsolved question is how this minor population of lytically-infected cells expressing vGPCR transduce an angio-proliferative phenotype to latently-infected cells. Here, we describe a Rac1 and ROS driven mechanism of paracrine amplification of viral oncogenesis in the KSHV-induced tumor model mECK36. It is triggered by vGPCR-induced PDGF secretion, which in the presence of high expression of PDGF receptors and NADPH oxidase family members caused by latent KSHV infection, leads to STAT3-mediated transcriptional activation of c-Myc, VEGF and KSHV latent genes. This paracrine loop, driven by PDGF signaling and regulated by Rac1 and ROS, amplifies vGPCR oncogenesis by driving proliferation and angiogenesis in latently-infected cells. Inhibtion of this molecular pathway by either the antioxidant N-acetylcysteine (NAC) or PDGF receptor tyrosine kinase inhibitors impair KSHV-induced tumorigenesis, angiogenesis and lymphangiogenesis, further validating this paracrine oncogenesis mechanism and its components as anti-KS tumor targets. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2710. doi:10.1158/1538-7445.AM2011-2710</jats:p

Molecular mechanisms, therapeutic targets, and clinically tested drugs in AIDS-KS paracrine viral oncogenesis.

<p>In KS spindle cells lytically infected with KSHV cells or latently infected spindle cells expressing early lytic genes, KSHV genes such vGPCR, K1, and ORF45 constitutively trigger signaling cascades, leading to mTOR and ROS activation, which induce transcription and translation of PDGF and VEGF. These secreted growth factors can act in a paracrine manner to activate the same signaling cascades in latently infected cells expressing VEGF and PDGF receptors to drive KS cell proliferation and angiogenesis. Rapamycin (RAPA), which inhibits mTOR, and Imatinib (IMA), which inhibits PDGFR, can interrupt this paracrine loop to target KSHV tumorigenesis. Both drugs have shown efficacy in AIDS-KS clinical trials.</p