Deciphering the functional role of recurrent PPP2R1A mutations on endometrial metastasis

破译PPP2R1A复发突变对子宫内膜转移的功能作用

• 批准号: 10601651
• 负责人: Terrance James Haanen
• 金额: $ 4.08万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-01 至 2026-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10601651
• 关键词: 3-Dimensional; Biogenesis; CRISPR/Cas technology; Cancer Cell Growth; Cancer cell line; Carcinoma; Cell Line; Cell secretion; Cells; Cessation of life; Communication; Complement; Complex; Coupled; Data; Development; Diagnosis; Disease; Disease Progression; Distant; Dominant-Negative Mutation; Dose Limiting; Endometrial; Family; Genes; Genetic Transcription; Greater sac of peritoneum; Heterozygote; Holoenzymes; Immunocompromised Host; Impairment; In Vitro; Injections; Invaded; Investigation; Knock-in; Ligands; Link; Luciferases; Malignant Epithelial Cell; Malignant Neoplasms; Mediating; Modeling; Molecular; Mus; Mutation; Nature; Neoplasm Metastasis; Nodule; Pathway interactions; Patients; Phenotype; Phosphoric Monoester Hydrolases; Phosphorylation; Process; Protein Subunits; Protein phosphatase; Proteins; Recurrence; Regulation; Role; Serine; Serous; Signal Pathway; Signal Transduction; Site; Testing; Threonine; Transfection; Transforming Growth Factor beta; Tumor Promotion; Tumor Subtype; Tumor Suppressor Proteins; Tumorigenicity; United States; Uterine Cancer; Uterine Corpus Carcinosarcoma; Uterine Neoplasms; Uterus; WNT Signaling Pathway; Work; Xenograft Model; beta catenin; cancer cell; cancer type; disorder subtype; effective therapy; enhancing factor; gain of function; in vivo; intraperitoneal; leiomyosarcoma; mutant; neoplastic cell; neutralizing antibody; new therapeutic target; notch protein; overexpression; permissiveness; scaffold; small hairpin RNA; tumor; tumor initiation; tumor microenvironment; tumorigenesis; tumorigenic

Project Summary: Uterine serous carcinomas (USC), uterine carcinosarcomas (UCS), and leiomyosarcomas represent the most aggressive subtypes of uterine cancer and account for a disproportionate number of uterine cancer related deaths in the United States. The lethality of these high-grade uterine cancers is largely due to the propensity of these cancers to disseminate and metastasize, leaving patients with a lack of effective treatment options. Despite this, the molecular drivers of the metastatic potential of these disease subtypes remain incompletely understood. Previous work by our group and others have identified a recurrent mutational hotspot within the gene encoding for the A scaffolding subunit of the protein phosphatase 2A (PP2A), PPP2R1A. These mutations, P179R and S256F, occur exclusively within these high-grade subtypes of uterine cancer and are not found in any other cancer type. Protein phosphatase 2A (PP2A), is a family of serine/threonine phosphatases, where the active heterotrimeric form is comprised of the catalytic (C) subunit, the scaffolding (A) subunit, and a substrate directing regulatory (B) subunit. In aggregate, PP2A is considered a tumor suppressor protein, but certain heterotrimers have tumor promoting roles. We have shown that these mutations contribute to uterine cancer tumorigenesis by modulating PP2A function in a manner that impairs the formation of tumor suppressive heterotrimers, while leaving tumor promoting heterotrimeric complexes intact. My preliminary data show that the PP2A P179R and S256F mutations enhance tumor initiating capacity, and the ability to form 3D tumor spheres in vitro. Importantly, uterine cancer cells expressing these mutations also form intraperitoneal nodules at an increased rate in vivo, suggesting that these mutations may enhance the establishment of new tumor formation in the peritoneal cavity. Conditioned media from the mutant expressing uterine cancer cells promoted tumor sphere formation of WT expressing uterine cancer cell growth, indicating that the mutant cells may create a more permissive tumor microenvironment for tumor formation. Preliminary data suggests that the mutant cells have increased -catenin phosphorylation and WNT ligand expression. The working hypothesis of this proposal is that the recurrent mutants, P179R and S256F, enhance uterine cancer metastasis through increased WNT signaling and secretion of WNT ligands. To test this hypothesis, we will develop PP2A A knock-in xenograft models to determine how these recurrent mutants contribute to enhanced metastasis in vivo (Specific Aim 1). Second, mechanistic studies will explore the secreted factors from the mutant uterine cancer cells, exploring WNT ligands, coupled with secretome analysis. These studies will also explore the link between loss of PP2A and WNT mediated EMT (Specific Aim 2). This proposal aims to decipher the functional role of recurrent PP2A A𝛼 mutations on uterine cancer metastasis, which may identify a new therapeutic target(s) of high-grade uterine cancers.

项目总结：子宫浆液性癌（USC）、子宫癌肉瘤（UCS）和平滑肌瘤 代表了子宫癌的最具侵袭性的亚型，并占子宫癌的不成比例的数量。 美国癌症相关死亡人数这些高级别子宫癌的致命性主要是由于 这些癌症有扩散和转移的倾向，使患者缺乏有效的治疗 选项.尽管如此，这些疾病亚型转移潜力的分子驱动因素仍然存在， 不完全理解。 我们小组和其他人以前的工作已经确定了基因内的一个重复突变热点 编码蛋白磷酸酶2A（PP 2A）的A支架亚基PPP 2 R1 A。这些突变， P179 R和S256 F仅发生在这些高级别子宫癌亚型中，在 任何其他癌症类型。蛋白磷酸酶2A（PP 2A）是丝氨酸/苏氨酸磷酸酶的家族，其中蛋白磷酸酶2A（PP 2A）是丝氨酸/苏氨酸磷酸酶的一部分。 活性异源三聚体形式由催化（C）亚基、支架（A）亚基和底物组成 定向调节（B）亚基。总的来说，PP 2A被认为是一种肿瘤抑制蛋白，但某些 异源三聚体具有肿瘤促进作用。我们已经证明，这些突变有助于子宫癌 通过调节PP 2A功能以损害肿瘤抑制因子的形成来抑制肿瘤发生， 异源三聚体，同时保持肿瘤促进异源三聚体复合物完整。 我的初步数据显示PP 2A P179 R和S256 F突变增强了肿瘤起始能力， 以及在体外形成3D肿瘤球的能力。重要的是，表达这些突变的子宫癌细胞 也在体内以增加的速率形成腹膜内结节，这表明这些突变可能会增强 在腹膜腔中建立新的肿瘤形成。来自突变体表达的条件培养基 子宫癌细胞促进表达WT的子宫癌细胞生长的肿瘤球形成，表明 突变细胞可能为肿瘤形成创造更宽松的肿瘤微环境。初步 数据提示突变细胞具有增加的β-连环蛋白磷酸化和WNT配体表达。 该建议的工作假设是，复发突变体P179 R和S256 F增强子宫内膜异位症。 通过增加的WNT信号传导和WNT配体的分泌来抑制癌症转移。为了验证这个假设，我们 将开发PP 2A A β基因敲入异种移植模型，以确定这些复发突变体如何有助于 增强体内转移（特异性目的1）。第二，机制研究将探索分泌因子， 突变的子宫癌细胞，探索WNT配体，结合分泌组分析。这些研究还将 探索PP 2A缺失与WNT介导的EMT（特异性目的2）之间的联系。这项提案旨在破译 复发性PP 2A A突变对子宫癌转移的功能作用，这可能会发现一种新的 高级别子宫癌的治疗靶点。