Tissue-Regulated Lineage-Specific Splicing in Glioblastoma Pathogenesis

胶质母细胞瘤发病机制中组织调节的谱系特异性剪接

• 批准号: 10409836
• 负责人: Markus Bredel
• 金额: $ 34.9万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10409836
• 关键词: Address; Adult; Affect; Alternative Splicing; Amino Acids; Annexins; Attenuated; Binding; Brain; Brain Neoplasms; Cell Lineage; Cells; Clinical; Complex; Data; Development; Disease; Endocytosis; Epidermal Growth Factor Receptor; Exclusion; Exocytosis; Exons; Foundations; Future; Genetic; Glioblastoma; Glioma; Knowledge; Length; Malignant - descriptor; Malignant Glioma; Mediating; Membrane; Mission; Mitotic; Modeling; N-terminal; Neoplasms; Neurons; Nuclear Pore Complex; Oncogenic; PDGFRA gene; Pathogenesis; Pathway interactions; Patients; Pattern; Polypyrimidine Tract-Binding Protein; Prevalence; Process; Protein Isoforms; Protein Tyrosine Kinase; Public Health; RNA Binding; RNA Splicing; Receptor Protein-Tyrosine Kinases; Recycling; Regulation; Research; Role; Signal Transduction; Sorting - Cell Movement; Testing; Tissues; Tumor Suppressor Genes; Tumor Suppressor Proteins; Tumorigenicity; United States National Institutes of Health; Variant; Work; Xenograft procedure; base; cell growth; epidermal growth factor receptor VIII; exon skipping; extracellular; extracellular vesicles; gain of function; in vivo; knock-down; mRNA Precursor; neoplastic cell; novel; novel therapeutics; precursor cell; prevent; progenitor; prototype; receptor; receptor recycling; relating to nervous system; response; stem cells; targeted treatment; therapeutic target; trafficking; trait; treatment response; tumor; tumorigenesis; tumorigenic; vesicle transport; virtual

PROJECT SUMMARY Lineage-specific alternative splicing, where splicing occurs in a tissue-regulated manner involving evolutionary conserved alternative exons, has a determinative role in brain development. The role of tissue-specific alternative exons in malignant transformation and tumor development is undefined. The impact of lineage- specific splicing on glioblastoma (GBM) pathogenesis remains unclear but is conceptually attractive given the prevalence of this process in determining the fate of ancestral cells of potential GBM origin. We found that brain-enriched splice factor PTBP1 mediates lineage-specific alternative splicing of the ANXA7 tumor suppressor gene. PTBP1 is expressed in neural and glial precursor cells (NPCs/GPCs) and binds ANXA7 pre-mRNA to skip exon 6; this produces spliced ANXA7 isoform 2 (I2). During lineage specification, PTBP1 is downregulated and this allows the expression of unspliced ANXA7 I1 (I1), which includes exon 6. We determined that the patterned expression of the I2 splice variant in the brain is restricted to lineages that represent potential GBM cells of origin but that I1 is virtually absent in these cells. Our preliminary data illustrate that lineage-specific splicing can augment genetic mechanisms to deregulate oncogenic pathways. Specifically, we showed ANXA7 I1, but not I2, targets oncogenic receptor tyrosine kinases (RTKs) such as EGFR, MET and PDGFRA for endosomal degradation. In the adult brain, PTBP1 is nearly absent; in GBM, PTBP1 aberrantly persists and RTK levels are elevated. However, we demonstrate that upon PTBP1 knockdown and/or, ANXA7 I1 re-expression, RTK signaling is reduced, and GBM tumorigenicity is diminished. Therefore, we hypothesize that GBMs benefit from persistent PTBP1 expression as this suppresses ANXA7 I1, prevents RTK endosomal degradation, and sustains RTK signaling in GBM. Our Specific Aims are: 1. Define the Mechanism by which ANXA7 I1 Regulates Endosomal and Cellular Trafficking of EGFR. Herein, we will determine the detailed mechanism by which ANXA7 I1 interacts with and regulates endosomal and extracellular vesicle transport of EGFR as a prototype RTK. 2. Demonstrate ANXA7 I1 is a Signaling Modulator for Multiple RTKs and RTK-Targeted Therapies by Perpetuating Endosomal Degradation. Herein, we will demonstrate that ANXA7 I1 is competent to attenuate signaling through multiple pro-tumorigenic RTKs, modulate TKI response, and thus has a broad impact on GBM in vivo. These mechanistic and focused aims are needed to better define the role of lineage-specific splicing processes in tumorigenesis, which remains understudied. Our results will determine that ANXA7 I1 is sufficient to simultaneously inhibit signaling by multiple RTKs and thus holds potential to reveal a new therapeutic direction. This work will define the role of aberrant lineage-specific alternative splicing in GBM pathogenesis, offer a broader understanding of this process in malignant transformation, and provide a potential foundation for future studies to therapeutically target developmentally regulated splicing factors.

项目摘要 谱系特异性替代剪接，其中剪接以组织调节的方式发生，涉及进化 保守的替代外显子在大脑发育中具有决定性的作用。组织特异性的作用 恶性转化和肿瘤发育中的替代外显子不确定。血统的影响 - 在胶质母细胞瘤（GBM）发病机理上的特定剪接尚不清楚，但在概念上具有吸引力 考虑到该过程在确定潜在GBM起源的祖先细胞的命运方面的流行。 我们发现，富含脑部的剪接因子PTBP1介导了Anxa7的谱系特异性替代剪接 肿瘤抑制基因。 PTBP1在神经和神经胶质前体细胞（NPC/GPC）中表达，并结合 Anxa7前MRNA跳过外显子6；这会产生剪接的Anxa7同工型2（I2）。在血统规范期间， ptbp1被下调，这允许表达未填充的anxa7 i1（i1），其中包括外显子6。 确定大脑中I2剪接变体的图案表达仅限于谱系 代表原始的潜在GBM细胞，但在这些细胞中实际上没有I1。我们的初步数据 说明谱系特异性的剪接可以增强遗传机制，从而消除过度调节 途径。具体而言，我们显示了Anxa7 I1，但不是I2，而是靶向致癌受体酪氨酸激酶（RTKS） 例如EGFR，MET和PDGFRA进行内体降解。在成年大脑中，PTBP1几乎不存在。在 GBM，PTBP1异常持续存在，RTK水平升高。但是，我们证明了PTBP1 敲低和/或ANXA7 I1重新表达，RTK信号降低，GBM肿瘤性降低。 因此，我们假设GBM受益于持续的PTBP1表达，因为这抑制了 AnxA7 I1可防止RTK内体降解，并在GBM中保持RTK信号传导。我们的具体目标 为：1。定义Anxa7 I1调节内体和细胞运输的机制 egfr。在此，我们将确定Anxa7 I1与之相互作用并调节的详细机制 EGFR作为原型RTK的内体和细胞外囊泡转运。 2。证明Anxa7 I1是一个 多个RTK和RTK靶向疗法的信号调节剂通过永存 降解。在此，我们将证明Anxa7 I1有能力通过多个来减弱信号 促肿瘤的RTK，调节TKI响应，因此对体内GBM产生了广泛的影响。这些 需要机械和集中的目标来更好地定义谱系特异性剪接过程的作用 肿瘤发生，静止不动。我们的结果将确定Anxa7 I1足以 同时抑制多个RTK的信号传导，因此具有揭示新的治疗方向的潜力。 这项工作将定义异常谱系特异性替代剪接在GBM发病机理中的作用， 在恶性转变中对这一过程有更广泛的了解，并提供潜力 未来研究的基础，以治疗靶向发育调节的剪接因素。