Basis for Lymphomagenesis in Akt2 Transgenic Mice

摘要 AKT在肿瘤发生中起核心作用，并且在人类淋巴瘤中频繁激活。v-akt是一个 逆转录病毒携带的致癌基因，可诱导小鼠和转基因小鼠的胸腺T细胞淋巴瘤 在未成熟T细胞中特异性表达Akt的组成型活性形式（Myr-Akt）， 淋巴瘤发现Myr-Akt 2小鼠模型的多发性胸腺淋巴瘤具有新的倒位 6号染色体inv（6），在T细胞受体β链基因座（Tcrb）和Dss 1基因中具有断点。一 未检测到融合蛋白，但重排将Tcrb增强子置于几个基因附近， 这些非编码RNA一直被上调。上调这些候选基因之一，编码 转录因子Dlx 5最近在几种常见的人类癌症中被报道，这表明 当异常表达时，该同源异型盒基因可能是致癌的。这一广泛而长期的目标 该项目是为了提高我们对Akt 2在淋巴瘤发生中的作用的理解，并阐明其机制， Dlx 5的上调与Akt 2信号传导会聚以促进肿瘤形成。具体目标是： 1)评估上调的Dlx 5与淋巴瘤形成的相关性。我们将确定细胞增殖 和/或具有inv（6）的胸腺淋巴瘤细胞的存活力被Dlx 5的敲低抑制。此外，骨 将进行骨髓嵌合体实验以确定在非恶性骨髓中Dlx 5的敲低是否 来自inv（6）阳性淋巴瘤高发病率的创始细胞系的细胞抑制受体中的肿瘤形成 野生型同窝仔。我们还将检查是否转基因小鼠在胸腺中过表达Dlx 5， 发展成自发性T细胞淋巴瘤2)使用直接遗传方法，测试Dlx 5是否合作 与胸腺肿瘤发展的关系。Myr-Akt 2小鼠将与Dlx 5转基因小鼠或条件性小鼠杂交。 Dlx 5敲除小鼠，以分别确定肿瘤发展是否被加速或抑制。我们还将 通过评估细胞生长来鉴定Akt 2与Dlx 5在肿瘤发生中协同作用的机制， 来自单独表达Dlx 5或Myr-Akt 2的转基因小鼠的原代胸腺细胞的增殖和存活 而不是两者结合。3)确定Akt 2信号传导的过度激活是否是淋巴瘤和非淋巴瘤所必需的。 发展和维持已建立的肿瘤。荧光原位杂交、PCR和磁共振 将采用共振显微成像分析来确定在肿瘤发生过程中inv（6）何时出现。 发展;然后将使用早期干预策略来确定是否抑制Akt 2信号传导 预防或延缓淋巴瘤的发展。此外，将开发诱导型Myr-Akt 2模型 以确定在已建立的胸腺淋巴瘤中Akt 2的失活是否导致肿瘤消退。该项目将 增强我们对Akt 2介导的淋巴瘤发生以及Akt 2与一种新的 假定的癌基因Dlx 5，其激活也可能有助于各种人类癌症的发病机制。

ABSTRACT AKT plays a central role in tumorigenesis and is frequently activated in human lymphoma. v-akt is an oncogene harbored by a retrovirus that induced thymic T-cell lymphomas in mice, and transgenic mice expressing constitutively active forms of Akt (Myr-Akt) specifically in immature T cells develop thymic lymphomas. Multiple thymic lymphomas from a Myr-Akt2 mouse model were found to harbor a novel inversion of chromosome 6, inv(6), with breakpoints in the T-cell receptor beta chain locus (Tcrb) and Dss1 gene. A fusion protein was not detected, but the rearrangement places the Tcrb enhancer near several genes and noncoding RNAs that are consistently up regulated. Up regulation of one of these candidate genes, encoding the transcription factor Dlx5, has recently been reported in several common human cancers, suggesting that this homeobox gene may be oncogenic when expressed aberrantly. The broad, long-term objective of this project is to improve our understanding of the role of Akt2 in lymphomagenesis and elucidate mechanisms by which up regulation of Dlx5 converges with Akt2 signaling to promote tumor formation. The specific aims are: 1) Assess the relevance of up-regulated Dlx5 to lymphoma formation. We will determine if cell proliferation and/or viability of thymic lymphoma cells with the inv(6) are inhibited by knock down of Dlx5. In addition, bone marrow chimera experiments will be carried out to determine if knock down of Dlx5 in non-malignant marrow cells from a founder line with high incidence of inv(6)-positive lymphomas inhibits tumor formation in recipient wild-type littermates. We will also examine if transgenic mice engineered to overexpress Dlx5 in the thymus develop spontaneous T-cell lymphomas. 2) Using a direct genetic approach, test whether Dlx5 cooperates with Akt2 in thymic tumor development. Myr-Akt2 mice will be crossed with Dlx5 transgenic mice or conditional Dlx5 knockout mice to determine if tumor development is accelerated or inhibited, respectively. We will also identify mechanisms by which Akt2 cooperates with Dlx5 in tumorigenesis by assessing cell growth, proliferation and survival in primary thymocytes from transgenic mice expressing Dlx5 or Myr-Akt2 alone versus in combination. 3) Determine if hyperactivation of Akt2 signaling is required for both lymphoma development and maintenance of established tumors. Fluorescence in situ hybridization, PCR, and magnetic resonance microimaging analyses will be employed to establish when the inv(6) arises during tumor development; then an early intervention strategy will be used to determine if inhibition of Akt2 signaling prevents or delays the development of lymphoma. In addition, an inducible Myr-Akt2 model will be developed to ascertain if inactivation of Akt2 in established thymic lymphomas results in tumor regression. This project will enhance our understanding of Akt2-mediated lymphomagenesis and cooperation between Akt2 and a novel putative oncogene, Dlx5, whose activation may also contribute to the pathogenesis of various human cancers.