Studies of gene fusions in rhabdomyosarcoma

高级研究员于2011财年转移至国家癌症研究所，并在剩余时间内建立实验室。因此，2011财政年度期间未对该项目进行研究。然而，在宾夕法尼亚大学以前的研究中，高级研究员提出了以下观察结果。 我们利用人肌源性系统来研究PAX 3-FOXO 1融合在与肺泡横纹肌肉瘤（ARMS）更相关的细胞环境中。从用BMI 1和TERT永生化的人成肌细胞系开始，用PAX 3-FOXO 1和MYCN转导导致转化和致瘤活性。绝大多数转导细胞表达低PAX 3-FOXO 1水平，并且不具有转化或致瘤表型。相反，只有少数转导细胞表达高水平的PAX 3-FOXO 1，并被转化和致瘤。这些发现与PAX 3-FOXO 1高表达的毒性和细胞自主变化的发生一致，允许稀有细胞规避高水平表达的毒性作用。 为了研究在一个更动态的系统中表型和融合蛋白水平之间的关系，我们开发了一个4-羟基他莫昔芬诱导系统，其中PAX 3-FOXO 1连接到一个修饰的雌激素受体配体结合结构域。使用用MYCN和这种诱导型构建体转导的成肌细胞，我们发现PAX 3-FOXO 1诱导导致转化和致瘤活性，但也导致生长抑制增加。因此，该诱导系统将有助于我们研究PAX 3-FOXO 1诱导的毒性机制。

The Senior Investigator moved to the National Cancer Institute during FY2011 and has been setting up the laboratory during the remaining time. Therefore no research studies were performed on this project during FY2011. However, in previous studies at the University of Pennsylvania, the Senior Investigator made the following observations. We have utilized a human myogenic system to study the PAX3-FOXO1 fusion in a cellular environment more relevant to alveolar rhabodmyosarcoma (ARMS). Starting with a human myoblast line immortalized with BMI1 and TERT, transduction with PAX3-FOXO1 and MYCN resulted in transforming and tumorigenic activities. The far majority of transduced cells expressed low PAX3-FOXO1 levels and did not have a transformed or tumorigenic phenotype. In contrast, only a small number of transduced cells expressed high PAX3-FOXO1 levels, and were transformed and tumorigenic. These findings are consistent with the toxicity of high PAX3-FOXO1 expression and the occurrence of cell autonomous changes allowing rare cells to circumvent the toxic effects of high-level expression. To investigate the relationship between phenotype and fusion protein level in a more dynamic system, we developed a 4-hydroxytamoxifen inducible system in which PAX3-FOXO1 is joined to a modified estrogen receptor ligand-binding domain. Using myoblasts transduced with MYCN and this inducible construct, we found that PAX3-FOXO1 induction results in transforming and tumorigenic activities, but also causes increasing growth suppression. Therefore, the inducible system will be useful for our studies of the mechanism of PAX3-FOXO1-induced toxicity.