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.

项目摘要：子宫浆液癌（USC），子宫癌（UCS）和平滑肌肉瘤 代表了子宫癌最具侵略性的亚型，并解释了不成比例的子宫 美国与癌症有关的死亡。这些高级子宫癌的杀伤力主要是由于 这些癌症的倾向散布和转移，使患者缺乏有效的治疗 选项。尽管如此，这些疾病亚型的转移性潜力的分子驱动因素仍然 不完全理解。 我们小组和其他人的先前工作已经确定了基因内的复发突变热点 编码蛋白质磷酸酶2a（PP2A）的A支架亚基PPP2R1A。这些突变， P179R和S256F仅发生在这些高级子宫癌中，在 任何其他癌症类型。蛋白质磷酸酶2a（PP2A）是丝氨酸/苏氨酸磷酸酶的家族，其中 活性异三聚合物形式由催化（C）亚基，脚手架（a）亚基和底物组成 指导监管（b）亚基。总体而言，PP2A被认为是肿瘤抑制蛋白，但确定 异三聚体具有肿瘤促进作用。我们已经表明，这些突变有助于子宫癌 通过以损害抑制肿瘤形成的方式调节PP2A功能来调节肿瘤发生 异三聚体，留下促进异构体复合物完好无损的肿瘤。 我的初步数据表明，PP2A P179R和S256F突变增强了肿瘤的启动能力， 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 还以增加的体内速率形成腹膜内结节，表明这些突变可能会增强 在腹膜腔中建立新的肿瘤形成。来自表达突变体的条件培养基 子宫癌细胞促进了表达子宫癌细胞生长的WT的肿瘤球体，表明 突变细胞可能会产生更允许的肿瘤微环境以形成肿瘤。初步的 数据表明，突变细胞增加了-catenin磷酸化和Wnt配体表达。 该提议的工作假设是复发突变体P179R和S256F增强子宫 Wnt配体的WNT信号传导和分泌增加，癌症转移。为了检验这一假设，我们 将开发pp2aa敲入宏观模型，以确定这些复发突变体如何贡献 增强体内转移（特定目标1）。其次，机械研究将探索来自 突变的子宫癌细胞探索了Wnt配体，再加上分泌的分析。这些研究也将 探索pp2a和Wnt介导的EMT丢失之间的联系（特定目标2）。该建议旨在破译 复发性pp2a a𝛼突变在子宫癌转移中的功能作用，这可能鉴定出一种新的 高级子宫癌的治疗靶标。