Molecular and biochemical basis of Lymphangioleiomyomatosis

描述（由申请人提供）：肺淋巴管平滑肌瘤病（LAM）是一种进展缓慢的疾病，其特征是肺囊性破坏和最终的呼吸衰竭，每百万妇女中有3.4-7.8人发病。从生化角度来看，LAM是由结节性硬化症复合体（TSC）功能丧失引起的，似乎需要激活哺乳动物雷帕霉素复合体1 （mTORC1）信号通路的靶蛋白。尽管使用mTORC1抑制剂治疗LAM的临床试验结果令人鼓舞，但由于两个原因，这些药物受到限制：首先，基于雷帕霉素的治疗不能治愈，因为肿瘤在停止治疗后会重新生长；其次，长期使用雷帕霉素会产生巨大的副作用，包括免疫系统紊乱和糖尿病。为了解决这一目前尚未满足的临床需求，该项目将研究三个尚未解决的机制问题，以开发长效治疗LAM的长期目标。第一个目标将确定谷氨酰胺（Gln）代谢在AML和LAM细胞中的贡献及其在细胞存活中的重要性，因为mtorc1靶向治疗无法诱导毒性仍然是一个未解决的障碍。本研究将在体外和体内研究这些细胞中Gln代谢的机制，以及通过药理学和shRNA方法靶向AML和LAM细胞生长和存活的新途径。第二个目标是更好地了解mTORC1通路如何调节与LAM中“良性转移”表型相关的关键基因的选择性剪接。虽然mRNA剪接是高度调控和普遍存在的（发生在约95%的人类基因中，并且在癌症中经常解除管制），但导致解除管制的机制尚不清楚。利用我们最近完成的磷酸化蛋白质组学筛选的新发现，本目的将研究mTORC1途径如何调节与LAM“良性转移”表型相关的关键基因的剪接。最后，第三个目标将确定雌激素如何促进LAM发病机制的关键机制见解，这几乎只发生在育龄期的女性。目的是在mTORC1过度激活的情况下，研究雌激素受体信号通过ERK-Fra1-ZEB途径在调节LAM细胞存活、迁移和侵袭中的作用。除了解决LAM生物学的基本问题外，这些目标中的每一个都具有临床意义，并且在两个目标中，已经提出了动物研究将细胞生物学发现转化为相关的体内模型。综上所述，我们非常需要对LAM的生物学有更深入的了解，并期望通过鉴定新的生物标志物以及能够特异性消除TSC和LAM患者异常细胞生长、迁移和肿瘤形成的新药物，成功完成所提出的工作将影响LAM的治疗。

DESCRIPTION (provided by applicant): Pulmonary Lymphangioleiomyomatosis (LAM) is a slowly progressing disease characterized by cystic destruction of the lungs and eventual respiratory failure, and affects 3.4-7.8 per million women. Biochemically, LAM is caused by loss of function of the Tuberous Sclerosis Complex (TSC) and appears to require activation of the mammalian target of rapamycin complex 1 (mTORC1) signaling pathway. Despite encouraging clinical trial results from the use of mTORC1 inhibitors to treat LAM, these drugs are limited for two reasons: first, rapamycin-based treatments are not curative as tumors regrow following cessation of treatment; and second, long-term rapamycin usage has tremendous side effects including immune disorders and diabetes. To resolve this currently, unmet clinical need, this project will investigate three unresolved mechanistic questions, with the long-term goal of developing long-lasting therapies for LAM. The first aim will determine the contribution of glutamine (Gln) metabolism in AML and LAM cells and its importance in cellular survival as inability of mTORC1-targeted therapies to induce toxicity remains an unaddressed hurdle. This aim will investigate the mechanism of Gln metabolism in these cells as well as new approaches to target the growth and survival of AML and LAM cells via pharmacologic and shRNA methods, both in vitro and in vivo. The second aim will garner a greater understanding of how the mTORC1 pathway regulates the alternative splicing of key genes involved in the "benign metastasis" phenotype seen in LAM. While mRNA splicing is highly regulated and pervasive (occurring in ~95% of human genes and frequently deregulated in cancer), the mechanisms causing deregulation are unknown. Using novel hits from a phospho-proteomic screen we recently completed, this aim will investigate how the mTORC1 pathway regulates the splicing of key genes involved in the "benign metastasis" phenotype of LAM. Finally, the third aim will identify key mechanistic insights into how estrogen contributes to the pathogenesis of LAM, which occurs almost exclusively in females during childbearing years. The aim will investigate the role of estrogen receptor signaling via the ERK-Fra1-ZEB pathway, in the context of mTORC1 hyperactivation, in regulating LAM cell survival, migration and invasion. In addition to addressing basic questions of LAM biology, each of these aims has clinical implications, and in two of the aims animal studies have been proposed to translate the cell biology discoveries into relevant in vivo models. In conclusion, there's a great need for greater understanding of the biology of LAM, and the expectations are that successful completion of the proposed work will impact LAM treatment through identification of new biomarkers as well as novel drugs that can specifically eliminate abnormal cell growth, migration and tumor formation in TSC and LAM patients.