DISSECTING THE ROLES OF FAT1 AND NEAT1 IN SOFT TISSUE SARCOMA DEVELOPMENT AND METASTASIS USING NOVEL IN VIVO SARCOMA MODELS

软组织肉瘤（STS）是一种罕见的异质性间叶肿瘤，有超过75个亚型。STS是 研究不足的肿瘤，几乎没有建立研究模型，缺乏资金来源。几十年来， 对STS的治疗策略几乎没有改进，STS通常对当前的治疗有抵抗力 并且由于50%的患者在远处器官中发生转移而常常是致命的。为了解决这一未解决的临床问题， 准确概括这种癌症谱的体内模型为研究癌症提供了独特而有效的平台。 在新治疗策略转化为有限肉瘤人群之前的肉瘤生物学和临床前试验 患者然而，由于肿瘤抑制基因和致癌基因的缺失，可获得的体内肉瘤模型很少。 肉瘤发展和转移的驱动因素仍然未知。因此，我进行了全基因组体外遗传学 在野生型小鼠中筛选和指导体内CRISPR/Cas9敲除筛选，以鉴定其突变是 肉瘤发生所必需的。从这些筛选中，我产生了一种新的体内肉瘤模型，该模型由以下突变驱动： Fat 1在人STSS中经常发生突变。这是一个从头开始的体内模型，其概括了人类肿瘤的一个子集。 据我们所知，这是第一次确定Fat 1是人类STS中有效的肿瘤抑制因子。此外，委员会认为， 使用体内肉瘤模型和高通量RNA测序，我还鉴定了长链非编码RNA（lncRNA） Neat 1作为肉瘤转移的致癌驱动因子。这个K22奖将使我能够建立自己的研究平台， 进一步表征关键信号通路和靶基因在肉瘤的发展和转移，使用这些 独特的体内肉瘤模型。在具体目标1中，我们将剖析Hippo通路及其 效应子Yap 1/Taz通过Fat 1的突变驱动肉瘤。此外，我们将使用我的小说在体内肉瘤 模型来测试和优化抑制肉瘤肿瘤生长的最佳组合治疗策略。在特定 目的2：我们将确定lncRNA Neat 1驱动肉瘤转移的机制。我的初步结果 提示RNA剪接调控基因如Khsrp与Neat 1相互作用，促进肉瘤转移。我们将 使用我独特的体内肉瘤模型来剖析控制肉瘤转移的机制和 这些基因用于治疗转移性肉瘤患者的靶向疗法。最后，完成本提案 将决定编码基因Fat 1和非编码基因Neat 1在细胞中表达的功能后果。 并为设计有效的靶向治疗提供了新的候选途径和基因 治疗，以改善肉瘤患者的结果。

Soft tissue sarcomas (STSs) are rare heterogeneous mesenchymal tumors that have more than 75 subtypes. STSs are understudied tumors for which there are few established research models and a lack of funding sources. Over decades, there has been little improvement in the therapeutic strategies for STSs, which are often resistant to current therapies and can be frequently fatal as 50% of patients develop metastasis in distant organs. To solve this unmet clinical problem, in vivo models that accurately recapitulate this spectrum of cancers provide a unique and effective platform for studying sarcoma biology and preclinical trials before novel therapeutic strategies translate to limited population of sarcoma patients. However, there are very few in vivo sarcoma models available because the tumor suppressor and oncogenic drivers for sarcoma development and metastasis remain unknown. Therefore, I performed genome-wide in vitro genetic screens and direct in vivo CRISPR/Cas9 knockout screens in wild type mice to identify driver genes whose mutation is required for sarcoma initiation. From these screens, I generate a novel in vivo sarcoma model driven by the mutation of Fat1 which is frequently mutated in human STSs. This is a de novo in vivo model that recapitulates a subset of human STSs and, to our knowledge, the first determination that Fat1 is a potent tumor suppressor in human STSs. Furthermore, using in vivo sarcoma models and high throughput RNA sequencing, I also identified the long non-coding RNA (lncRNA) Neat1 as an oncogenic driver for sarcoma metastasis. This K22 award will allow me to build my own research platform to further characterize the critical signaling pathways and target genes in sarcoma development and metastasis using these unique in vivo sarcoma models. In Specific Aim 1, we will dissect the mechanism by which the Hippo pathways and their effectors Yap1/Taz drive sarcomas through the mutation of Fat1. In addition, we will use my novel in vivo sarcoma models to test and optimize the best combination treatment strategies that suppress sarcoma tumor growth. In Specific Aim 2, we will determine the mechanisms by which lncRNA Neat1 drives sarcoma metastasis. My preliminary results suggest that RNA splicing regulating genes, such as Khsrp, interact with Neat1 and promote sarcoma metastasis. We will use my unique in vivo sarcoma models to dissect the mechanisms governing sarcoma metastasis and the implications of these genes for targeted therapies in treating metastatic sarcoma patients. In conclusion, completion of this proposal will determine the functional consequences of expression of the coding gene Fat1 and the non-coding gene Neat1 in sarcoma development and metastasis and provide novel candidate pathways and genes for designing effective targeted therapies to improve outcomes for sarcoma patients.