Conditional Mouse Model of Alveolar Rhabdomyosarcoma

条件性小鼠肺泡横纹肌肉瘤模型

• 批准号: 6522657
• 负责人: CHARLES KELLER
• 金额: $ 7.01万
• 依托单位: UNIVERSITY OF UTAH
• 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/6522657
• 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.

腺泡状横纹肌肉瘤是一种侵袭性的儿童肌肉癌，发病率和死亡率都很高。85%的腺泡型横纹肌肉瘤表现出一种常见的遗传改变，即发育调节的Pax转录因子（Pax3或Pax7）与Forkhead转录因子Fkhr的易位介导的融合。了解肺泡型横纹肌肉瘤发生的分子机制将提高我们对患者进行风险治疗和开发新疗法的能力。本项目的目的是研究Pax3与Fkhr的胚后易位介导的融合是否启动了一个参与腺泡状横纹肌肉瘤发生的异常胚胎肌肉发育程序，并研究导致腺泡状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。该提案的具体目的是（I）检查基因组单个拷贝数的Pax3：Fkhr是否足以和必要地引起Pax3通路的异常激活和腺泡状横纹肌肉瘤的起始，（II）研究基因、转录或翻译是否对常见的致癌机制（例如：对抗生长信号不敏感，逃避凋亡）发生在一个可预测的序列在腺泡状横纹肌肉瘤，和（III）以确定是否Cre重组酶介导的异源染色体之间的易位可以诱导体细胞在一个频率足以启动腺泡状横纹肌肉瘤。对于第一个和第二个目标，将创建一个有条件的、基因敲入的腺泡状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介导基因组重排，仅在骨骼肌中将单个Pax3等位基因转化为单个Pax3：Fkhr等位基因。将连续处死动物，以分析携带Pax3：Fkhr的细胞中随后的细胞内变化序列。将在基因组、mRNA和蛋白质水平进行分析，重点是肿瘤进展的已知标志物。为了最终的目的，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。Keller博士与Capecchi博士的研究培训和NCI的支持将为Keller博士的独立研究生涯做好准备，调查儿科癌症的发展方面。最终，详细了解儿童恶性肿瘤中失调的发育途径可能会提高我们治疗儿童癌症的能力。