Role of Splicing Factors in Breast Cancer

剪接因子在乳腺癌中的作用

• 批准号: 8722508
• 负责人: OLGA ANCZUKOW-CAMARDA
• 金额: $ 10.6万
• 依托单位: COLD SPRING HARBOR LABORATORY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-01 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8722508
• 关键词: 3-Dimensional; Address; Alternative Splicing; Animal Model; Antisense Oligonucleotides; Apoptotic; Award; BCL2L11 gene; Biological; Breast; Breast Cancer Cell; Breast Cancer Model; Breast Epithelial Cells; Cancer cell line; Candidate Disease Gene; Cell Culture System; Cell Culture Techniques; Cell Line; Cell Proliferation; Cell model; Cells; Complement; Data; Databases; Development; Environment; Epithelial; Event; Exhibits; Faculty; Future; Gene Expression; Goals; Health; Human; In Vitro; Laboratories; Lead; Literature; MCF10A cells; Malignant Neoplasms; Mammary Neoplasms; Mammary gland; Mediating; Mentors; Mentorship; Molecular; Monitor; Neoplasm Metastasis; Oncogene Proteins; Oncogenic; Phase; Phenotype; Play; Positioning Attribute; Process; Production; Protein Family; Protein Isoforms; RNA Sequences; RNA Splicing; Regulation; Reporting; Research; Resistance; Role; Serine; Technology; Therapeutic; Training; Translating; Tumor Suppressor Genes; Tyrosine Kinase Inhibitor; anticancer research; base; cancer cell; cancer initiation; cancer therapy; career; cell transformation; human ARMET protein; in vivo; malignant breast neoplasm; member; migration; mouse model; next generation; next generation sequencing; novel; novel therapeutics; overexpression; prevent; programs; therapeutic target; transcriptome sequencing; tumor; vector

DESCRIPTION (provided by applicant): Cancer cells often display aberrant profiles of alternative splicing, leading to the production of protein isoforms that can increase cell proliferation, migration, and apoptotic resistance. My long-term goal is to establish an independent research lab, where I will elucidate molecular mechanisms by which alternative splicing misregulation plays a role in cancer by altering the expression of various oncogenes and tumor-suppressor genes. Importantly, these findings will translate into the development of novel therapeutic strategies. The K99/R00 career award will help in achieving this goal by advancing my training in: antisense oligonucleotide technologies under the guidance of my primary mentor Dr. Adrian Krainer; next-generation sequencing under the co-mentorship of Dr. Michael Schatz; and metastatic breast cancer models under the co-mentorship of Dr. Mikala Egeblad. This training will complement my previous expertise in breast cancer research and RNA splicing mechanisms. The very stimulating scientific environment at Cold Spring Harbor Laboratory will not only provide me with the expertise and facilities necessary for the completion of the mentored phase of this project, but will also prepare me to transition smoothly into an independent faculty position. During the K99 mentored phase, I will define the role of splicing factors and splicing misregulation in breast cancer. In the subsequent R00 independent phase, I will indentify oncogenic splicing events that could constitute therapeutic targets to pursue during my future independent research. We have previously demonstrated that overexpression of the splicing factor SRSF1 can transform human mammary epithelial cells in vitro and in vivo. We have also shown that SRFS1 levels are directly regulated by the MYC oncoprotein. However, additional splicing factors are also overexpressed in human breast tumors, suggesting that they may also play a role in breast cancer. In Aim 1, during the K99 phase, I will determine the role of specific splicing factors in breast cancer using relevant cell and animal models that mimic the biological context in which the tumors arise. This includes: (i) identifying changes in alternative splicing events underlying splicing-factor- mediated transformation by next-generation RNA-sequencing in 3-D cultures of human mammary epithelial cells; and (ii) defining the metastatic potential of these oncogenic splicing factors using in vivo mouse models. In Aim 2, during the K99 phase, I will determine the role of MYC in the regulation of alternative splicing in breast cancer, by identifying changes in both splicing-factor expression and in alternative splicing profiles by next-generation RNA-sequencing in a MYC-inducible cell culture system. In Aim 3, during the R00 phase, I will determine the role of splicing factors in acquisition of resistance to tyrosine kinase inhibitors in breast cancer, by identifying changes in splicing-factor levels and i alternative splicing events. Finally, starting in the K99 phase and leading into the R00 phase, I will determine the therapeutic potential of using antisense oligonucleotides to modulate specific oncogenic alternative splicing events identified in Aims 1-3. The proposed research will identify not only splicing factors involved in breast cancer but also their regulators and specific targets. This plan will establish the basis for my independent research program, in which I plan to contribute to the development of new cancer therapies based on modulating the expression and activity of splicing factors or their targets.

描述（由申请人提供）：癌细胞通常表现出异常的选择性剪接特征，导致产生可增加细胞增殖、迁移和凋亡抗性的蛋白质亚型。我的长期目标是建立一个独立的研究实验室，在那里我将阐明选择性剪接错误调节通过改变各种癌基因和肿瘤抑制基因的表达在癌症中发挥作用的分子机制。重要的是，这些发现将转化为新型治疗策略的开发。 K99/R00 职业奖将通过推进我在以下方面的培训来帮助实现这一目标： 在我的主要导师 Adrian Krainer 博士的指导下反义寡核苷酸技术；在 Michael Schatz 博士的共同指导下进行下一代测序；以及在 Mikala Egeblad 博士的共同指导下的转移性乳腺癌模型。这次培训将补充我之前在乳腺癌研究和 RNA 剪接机制方面的专业知识。冷泉港实验室非常刺激的科学环境不仅将为我提供完成该项目指导阶段所需的专业知识和设施，而且还将为我顺利过渡到独立教员职位做好准备。在 K99 指导阶段，我将定义剪接因子和剪接失调在乳腺癌中的作用。在随后的 R00 独立阶段，我将鉴定致癌剪接事件，这些事件可能构成在 我未来的独立研究。我们之前已经证明剪接因子SRSF1的过度表达可以在体外和体内转化人乳腺上皮细胞。我们还表明 SRFS1 水平直接受 MYC 癌蛋白调节。然而，其他剪接因子在人类乳腺肿瘤中也过度表达，表明它们也可能在乳腺癌中发挥作用。在目标 1 中，在 K99 阶段，我将确定以下角色： 使用模拟肿瘤发生的生物学背景的相关细胞和动物模型研究乳腺癌中的特定剪接因子。这包括： (i) 确定替代方案的变化 通过新一代 RNA 测序在人乳腺上皮细胞 3D 培养物中进行剪接因子介导的转化的剪接事件； (ii) 使用体内小鼠模型确定这些致癌剪接因子的转移潜力。在目标 2 中，在 K99 阶段，我将通过 MYC 中的下一代 RNA 测序来识别剪接因子表达和选择性剪接图谱的变化，从而确定 MYC 在乳腺癌选择性剪接调节中的作用-诱导细胞培养系统。在目标 3 中，在 R00 阶段，我将确定剪接因子在获得抗性中的作用 通过识别剪接因子水平的变化和选择性剪接事件，研究乳腺癌中的酪氨酸激酶抑制剂。最后，从 K99 阶段开始并进入 R00 阶段，我将确定使用反义寡核苷酸调节目标 1-3 中确定的特定致癌选择性剪接事件的治疗潜力。拟议的研究不仅将确定与乳腺癌有关的剪接因子，还将确定其调节因子和特定靶点。 该计划将为我的独立研究计划奠定基础，在该计划中，我计划为基于调节剪接因子或其靶标的表达和活性的新癌症疗法的开发做出贡献。