Molecular Pathways in T Cell Development and T-ALL

在过去的五年中，我们已经确定了T细胞分化的发展阶段，在这个阶段，由逆转录病毒插入含有ICN-1的载体引起的恶性转化首先表现为TCR前信号之后的CD4-8+TCRα/β负阶段。因此，肿瘤在TCRβ重排方面是单克隆性的，但表达不同的TCRAlpha链。肿瘤表现出正常的核型和基因组稳定性(经Sky和CGH验证)，但其特征是癌基因和调控生存和增殖的基因表达异常。因为测序还没有揭示基因 为了在逆转录病毒转导2-3周后早期发现不稳定和恶性细胞，我们将重点放在逆转录病毒载体的插入突变作为一种协同事件，因为ICN1过表达本身不会导致肿瘤，但与表型相同的正常细胞相比，多克隆的非致瘤细胞的存活和增殖略有增加。我们还证实了肿瘤表现出异常的miRNA表达模式。因此，我们将解决插入突变和异常表达的miRNA导致恶性肿瘤的假说，并将试图干预 通过使用反配子和miRNA的模拟物，以及通过击倒与恶性生长有关的异常过度表达的基因，来对抗恶性生长。 目的：确定逆转录病毒载体在肿瘤和非恶性ICN1高表达细胞中的整合位点，分析插入突变在肿瘤发生中的作用。 目的：表观遗传学分析T-ALL与表型相似但正常或ICN1过度表达的非致瘤细胞。 AIM3：miRNA在恶性转化中的作用。 目的4：SH RNA介导的T-ALL特异性过表达基因的敲除和肿瘤特异性抑制基因的过度表达。

In the past five years we have identified the developmental stage of Tcell differentiation at which malignant transformation caused by retroviral insertion of a vector containing ICN 1 becomes first apparent as the CD4-8+TCRalpha /beta minus stage that follows pre-TCR signaling. Consequently, the tumors are monoclonal with regard to TCR beta rearrangement but express diverse TCRalpha chains. The tumors exhibit a normal karyotype and genomic stability (verified by SKY and CGH),but are characterized by dysregulated expression of oncogenes and genes regulating survival and proliferation. Since sequencing has not revealed genetic instability and malignant cells can be detected early, 2-3 weeks after retroviral transduction , we will focus on insertional mutagenesis by the retroviral vector as a synergizing event since ICN1 overexpression alone does not result in tumors but in polyclonal, non-tumorigenic cells with slightly increased survival and proliferation when compared to phenotypically identical normal cells. We also established tht tumors exhibit an abnormal pattern of miRNA expression. We therefore will address the hypothesis that insertional mutagenesis and abnormally expressed miRNA contribute to the malignancy and will attempt to interfere with malignant growth by using antagomirs and mimics of miRNA and by knocking down abnormally overexpressed genes that are implicated in malignant growth. AIMI: Determine integration sites of retroviral vector in tumor and non-malignant ICN1 overexpressing cells to analyze contribution of insertional mutagenesis to tumor development. AIM2: Epigenetic analysis of T-ALL versus phenotypically similar but normal or ICN1 overexpressing nontumorigenic cells. AIM3: Contribution of miRNA to malignant transformation. AIM 4: Sh RNA mediated knockdown of genes specifically overexpressed in T-ALL and overexpression of genes specifically repressed in tumors.