KSHV microRNAs in cellular transformation and tumorigenesis

描述（由申请人提供）：卡波西氏肉瘤相关疱疹病毒（KSHV）是卡波西氏肉瘤（KS）的病因，KS是一种常见于艾滋病患者的血管癌。尽管进行了广泛的研究，但kshv诱导的细胞转化和肿瘤发生的机制仍不明确。这一严重的知识差距阻碍了有效干预措施的制定。我们发现KSHV能有效感染和转化原代大鼠胚胎后肾间充质前体细胞。kshv感染的MM （KMM）在裸鼠体内诱导ks样肿瘤。通过反向遗传学，我们发现KSHV诱导的细胞转化和肿瘤发生需要10个KSHV前microrna （pre-miRs）集群。受这些发现的鼓舞，我们最近在用KSHV感染和转化人原代细胞方面取得了重大进展。具体来说，我们已经发现KSHV还可以感染、永生化和转化人类原代间充质干细胞（MSC），即人类版本的MM。南加州大学和南京医科大学之间的这项合作应用提议扩展这些令人兴奋的范式转变发现，目的是完善MSC模型，并使用MM和MSC模型来识别介导KSHV细胞转化和肿瘤发生的特异性miRs。并确定这些mir靶向的基本细胞基因和途径。基于我们的初步结果，我们提出了一个工作假设，即KSHV可以感染和转化MSC，特异性KSHV miRs操纵必要的细胞生长和存活途径，对KSHV诱导的细胞转化和肿瘤发生起关键作用。因此，拟议的项目将完善人类原代MSC的KSHV细胞转化模型（Aim 1）；通过基因互补确定单个病毒前miRs和miRs对kshv诱导的细胞转化和肿瘤发生的贡献（目标2）；确定单个病毒前miRs和miRs对细胞转化的影响，并确定那些调节细胞生长、存活、血管生成和侵袭的miRs（目标3）；并通过确定基本的直接靶点来描述病毒miRs介导的kshv诱导的细胞转化和肿瘤发生的机制（Aim 4）。这个合作项目利用了中美两国实验室的专业知识。这些工作具有重要的创新意义，因为它们将首次表明KSHV可以真正转化人类原代细胞，确定KSHV miRs在细胞转化和肿瘤发生中的功能和作用机制，并为开发创新的预后和治疗方法确定潜在的新靶点。该研究还将建立一个由病毒颠覆miR通路介导的肿瘤发生的新范式，从而为其他癌症的肿瘤发生提供见解。

DESCRIPTION (provided by applicant): Kaposi's sarcoma-associated herpesvirus (KSHV) is the etiologic agent of Kaposi's sarcoma (KS), a vascular cancer commonly found in AIDS patients. Despite extensive studies, the mechanism of KSHV-induced cellular transformation and tumorigenesis remains undefined. This critical gap of knowledge has impeded the development of effective intervention measures. We have shown that KSHV can efficiently infect and transform primary rat embryonic metanephric mesenchymal precursor cells (MM). KSHV-infected MM (KMM) induce KS-like tumors in nude mice. Using reverse genetics, we have found that a cluster of 10 KSHV pre-microRNAs (pre-miRs) is required for KSHV-induced cellular transformation and tumorigenesis. Encouraged by these findings, we have recently made significant progresses in infecting and transforming human primary cells with KSHV. Specifically, we have found that KSHV can also infect, immortalize and transform human primary mesenchymal stem cells (MSC), the human version of MM. This collaborative application between the University of Southern California and Nanjing Medical University proposes to extend these exciting paradigm-shifting discoveries with the objective to refine the MSC model, and use both the MM and MSC models to identify the specific miRs that mediate KSHV cellular transformation and tumorigenesis, and define the essential cellular genes and pathways that are targeted by these miRs. Based on our preliminary results, we have formulated a working hypothesis that KSHV can infect and transform MSC, and specific KSHV miRs manipulate essential cell growth and survival pathways, contributing critically to KSHV-induced cellular transformation and tumorigenesis. Therefore, the proposed project will refine the model of KSHV cellular transformation of human primary MSC (Aim 1); determine the contribution of individual viral pre-miRs and miRs to KSHV-induced cellular transformation and tumorigenesis by genetic complementation (Aim 2); determine the effect of individual viral pre-miRs and miRs on cellular transformation, and identify those that regulate cell growth, survival, angiogenesis and invasion (Aim 3); and delineate the mechanism of viral miRs- mediated KSHV-induced cellular transformation and tumorigenesis by identifying the essential direct targets (Aim 4). This collaborative project takes advantage of the expertise of the US and China laboratories. These works are significant and innovative because they will, for the first time, show that KSHV can truly transform human primary cells, define the functions and mechanisms of action of KSHV miRs in cellular transformation and tumorigenesis, and identify potential novel targets for developing innovative prognostic and therapeutic approaches. The study will also establish a novel paradigm of oncogenesis mediated by viral subversion of the miR pathway, thus providing insights into the oncogenesis of other cancers.