Conditional Mouse Model of Alveolar Rhabdomyosarcoma

条件性小鼠肺泡横纹肌肉瘤模型

• 批准号: 6847450
• 负责人: CHARLES KELLER
• 金额: $ 13.77万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCIENCE CENTER
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-09-07 至 2006-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6847450
• 关键词: angiogenesis; apoptosis; chromosome translocation; developmental genetics; disease /disorder model; enzyme activity; genetic transcription; genetic translation; laboratory mouse; lung alveolus; lung neoplasms; mammalian embryology; model design /development; myogenesis; neoplasm /cancer genetics; pediatric neoplasm /cancer; recombinase; rhabdomyosarcoma; transcription factor; transfection /expression vector

Alveolar rhabdomyosarcoma is an aggressive childhood muscle cancer associated with significant morbidity and mortality. Eighty-five percent of alveolar rhabdomyosarcomas demonstrate a common genetic alteration, the translocation-mediated fusion of a developmentally regulated Pax transcription factor (either Pax3 or Pax7) to the Forkhead transcription factor, Fkhr. An understanding of the molecular events underlying the development of alveolar rhabdomyosarcomas will improve our ability to stratify patients to risk-based therapies and to develop new therapies. The goals of this project are to investigate whether the postembryonic, translocation-mediated fusion of Pax3 to Fkhr initiates an aberrant embryonic muscle development program which is involved in the genesis of alveolar rhabdomyosarcoma, and to investigate whether the early and intermediate steps in the activation of oncogenic properties contributing to the genesis of alveolar rhabdomyosarcoma occur in a predictable sequence. The specific aims of this proposal are (I) to examine whether a genomic single copy-number of Pax3:Fkhr is sufficient and necessary to cause both aberrant Pax3 pathway activation and the initiation of alveolar rhabdomyosarcoma, (II) to study whether genetic, transcriptional, or translational contributions to common oncogenic mechanisms (eg. insensitivity to antigrowth signals, evasion of apoptosis) occur in a predictable sequence in alveolar rhabdomyosarcomas, and (III) to determine whether Cre recombinase-mediated translocations between heterologous chromosomes can be induced somatically at a frequency sufficient for the initiation of alveolar rhabdomyosarcoma. For the first and second aims, a conditional, knock-in mouse model of alveolar rhabdomyosarcoma will be created which initially harbors two functionally normal Pax3 alleles. Conditional, somatic expression of the site-specific DNA recombinase Cre will mediate a genomic rearrangement converting a single Pax3 allele to a single Pax3:Fkhr allele in skeletal muscle only. Animals will be serially sacrificed to analyze the ensuing sequence of intracellular changes in cells harboring Pax3:Fkhr. Analysis will be performed at the genomic, mRNA, and protein level with an emphasis on known markers of tumor progression. For the final aim, a conditional, somatic translocation between the Pax3 and Fkhr loci will be attempted using Cre-mediated rearrangement of heterologous chromosomes. Dr. Keller's research training with Dr. Capecchi and the support of the NCI will prepare Dr. Keller for an independent research career investigating developmental aspects of pediatric cancer. Ultimately, a detailed knowledge of dysregulated developmental pathways in pediatric malignancies may improve our ability to treat childhood cancer.

肺泡型横纹肌肉瘤是一种侵袭性的儿童期肌肉癌，发病率和死亡率都很高。85%的肺泡型横纹肌肉瘤表现出一种常见的遗传改变，即易位介导的发育调节的Pax转录因子(Pax3或Pax7)与Forkhead转录因子Fkhr的融合。对肺泡型横纹肌肉瘤发生的分子事件的了解将提高我们对患者进行基于风险的治疗和开发新疗法的能力。本项目的目的是研究胚胎后转位介导的Pax3到Fkhr的融合是否启动了参与肺泡型横纹肌肉瘤发生的异常胚胎肌发育程序，以及在参与肺泡型横纹肌肉瘤发生的致癌特性激活的早期和中期步骤是否以可预测的顺序发生。这一建议的具体目的是：(I)研究基因组中Pax3：Fkhr的单个拷贝数是否充分且必要，从而导致Pax3途径的异常激活和肺泡型横纹肌肉瘤的启动，(Ii)研究是否遗传、转录或翻译对常见的致癌机制(例如，Pax3：Fkhr)有贡献。目的：(I)在肺泡型横纹肌肉瘤中，(Iii)确定Cre重组酶介导的异源染色体易位能否以足以引发肺泡型横纹肌肉瘤的频率在躯体上被诱导。对于第一个和第二个目标，将创建一个条件的、敲入的肺泡型横纹肌肉瘤小鼠模型，该模型最初含有两个功能正常的Pax3等位基因。位点特异性DNA重组酶Cre的条件性体细胞表达将仅在骨骼肌中介导将单个Pax3等位基因转化为单个Pax3：Fkhr等位基因的基因组重排。动物将被连续处死，以分析随后携带Pax3的细胞内变化的序列：Fkhr。将在基因组、信使核糖核酸和蛋白质水平进行分析，重点放在肿瘤进展的已知标记物上。为了达到最终目的，将尝试使用Cre介导的异源染色体重排，在Pax3和Fkhr基因座之间进行有条件的体细胞易位。凯勒博士与卡佩奇博士一起接受的研究培训以及美国国家癌症研究所的支持，将为凯勒博士从事研究儿科癌症发展方面的独立研究生涯做好准备。最终，对儿童恶性肿瘤异常发育途径的详细了解可能会提高我们治疗儿童癌症的能力。