Basis for Lymphomagenesis in Akt2 Transgenic Mice

描述（由申请人提供）：AKT在肿瘤发生中起核心作用，在人类淋巴瘤中经常被激活。v-akt是逆转录病毒携带的致癌基因，其在小鼠中诱导胸腺T细胞淋巴瘤，并且在未成熟T细胞中特异性表达Akt的组成型活性形式（Myr-Akt）的转基因小鼠发展胸腺淋巴瘤。发现Myr-Akt 2小鼠模型的多发性胸腺淋巴瘤具有6号染色体的新倒位inv（6），在T细胞受体β链基因座（Tcrb）和Dss 1基因中具有断点。未检测到融合蛋白，但重排将Tcrb增强子放置在持续上调的几个基因和非编码RNA附近。这些候选基因之一，编码转录因子Dlx 5的上调，最近在几种常见的人类癌症中报道，这表明这种同源异型盒基因表达异常时可能是致癌的。该项目的广泛，长期的目标是提高我们对Akt 2在淋巴瘤发生中的作用的理解，并阐明Dlx 5的上调与Akt 2信号传导融合以促进肿瘤形成的机制。具体目的是：1）评估上调的Dlx 5与淋巴瘤形成的相关性。我们将确定具有inv（6）的胸腺淋巴瘤细胞的细胞增殖和/或活力是否被Dlx 5的敲低抑制。此外，还将进行骨髓嵌合体实验，以确定来自inv（6）阳性淋巴瘤发生率高的创始系的非恶性骨髓细胞中Dlx 5的敲低是否抑制受体野生型同窝仔中的肿瘤形成。我们还将检查是否在胸腺中过表达Dlx 5的转基因小鼠发生自发性T细胞淋巴瘤。2)使用直接遗传方法，测试Dlx 5是否与Akt 2在胸腺肿瘤发展中合作。Myr-Akt 2小鼠将与Dlx 5转基因小鼠或条件性Dlx 5敲除小鼠杂交，以分别确定肿瘤发展是否加速或抑制。我们还将通过评估来自单独表达Dlx 5或Myr-Akt 2的转基因小鼠与组合表达Dlx 5或Myr-Akt 2的转基因小鼠的原代胸腺细胞中的细胞生长、增殖和存活来鉴定Akt 2与Dlx 5在肿瘤发生中协作的机制。3)确定Akt 2信号传导的过度激活是否是淋巴瘤发展和已建立肿瘤的维持所必需的。将采用荧光原位杂交、PCR和磁共振显微成像分析来确定肿瘤发展过程中inv（6）何时出现;然后将使用早期干预策略来确定抑制Akt 2信号传导是否可以预防或延迟淋巴瘤的发展。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与一种新的推定癌基因Dlx 5之间的合作的理解，Dlx 5的激活也可能有助于各种人类癌症的发病机制。公共卫生相关性：细胞酶AKT 2在大多数人类癌症中过度活跃，并促进肿瘤细胞存活和对化疗的抵抗。我们在小鼠模型中的研究发现了一种新的基因（Dlx 5），当表达异常时，它与激活的Akt 2一起诱导胸腺的侵袭性淋巴瘤。人DLX 5最近已显示在一些淋巴瘤以及更常见的肿瘤如肺癌中以升高的水平表达，这表明DLX 5可作为人AKT 2相关恶性肿瘤的新治疗靶点。

DESCRIPTION (provided by applicant): 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. PUBLIC HEALTH RELEVANCE: The cellular enzyme AKT2 is hyperactive in most human cancers and promotes tumor cell survival and resistance to chemotherapy. Our studies in a mouse model have uncovered a novel gene (Dlx5) that, when expressed abnormally, works together with activated Akt2 to induce aggressive lymphomas of the thymus. Human DLX5 has recently been shown to be expressed at elevated levels in some lymphomas as well as more common tumors such as lung cancers, suggesting that DLX5 may serve as a new therapeutic target in human AKT2-related malignancies.