Mouse models of Pik3ca brain overgrowth disorders

项目摘要 这一建议概述了一个全面的系列实验，以评估过度生长表型的基础 与PIK3CA中的功能获得突变相关，使用小鼠作为模型系统。虽然好 在癌症研究中，最近的数据揭示了PIK3CA突变在几种发育障碍中的作用 包括MCAP、DMEG、CLOVES综合征、表皮痣和脂溢性角化病。这里将评估 由3种不同的Pik3ca功能获得突变引起的发育和信号传导途径中断， 特别强调大脑的发育。迄今为止，所有报道的PIK3CA突变都是合子后或嵌合的。 而不是种系突变。因此，我们假设PIK3CA表型与PIK3CA和PIK3CA的表达相关。 突变的严重程度和镶嵌现象的水平（和分布）。为了验证这个假设和其他假设，我们将 使用标准的条件遗传学方法在胚胎中表达3种患者相关的Pik3ca gof突变， CNS神经元祖细胞及其后代。然后我们将产生胚胎干细胞嵌合体， 组成型表达相同的突变到野生型胚泡中以评估表型嵌合性 大脑内部和整个身体的关系。我们还建议在子宫内使用 电穿孔技术，以更特异性地将嵌合表达靶向发育中的大脑。我们的研究 将评估大脑病理学的发育发病机制，并描述相关的癫痫， 迫切需要临床相关表型。最后，我们将使用药理学方法来评估 在出生后小鼠中驱动急性Pik3ca依赖性癫痫发作的潜在机制。这些测定代表了 在PIK3CA节段性脑过度生长中开发分子理性癫痫治疗的第一步 综合征患者。

PROJECT SUMMARY This proposal outlines a comprehensive series of experiments to assess the basis of overgrowth phenotypes associated with gain of function mutations in PIK3CA, using the mouse as a model system. Although well studied in cancer, recent data has revealed a role for PIK3CA mutation in several developmental disorders including MCAP, DMEG, CLOVES syndrome, epidermal nevi and seborrheic keratosis. Here will assess the developmental and signaling pathway disruptions caused by 3 different Pik3ca gain of function mutations, with particular emphasis on the developing brain. To date, all reported PIK3CA mutations are postzygotic or mosaic rather than germline mutations. Accordingly, we hypothesize that PIK3CA phenotypes correlate with both the severity of the mutation and level (and distribution) of mosaicism. To test this and other hypotheses, we will use standard conditional genetic approaches to express 3 patient-related Pik3ca gof mutations in embryonic CNS neuronal progenitors and their descendants. We will then generate ES cell chimeras, injecting ES cells constitutively expressing the same mutations into wild-type blastocysts to assess phenotype-mosaicism relationships both within the brain and throughout the entire body. We also propose to use in utero electroporation technology to more specifically target mosaic expression to the developing brain. Our studies will assess the developmental pathogenesis of brain pathology and characterize the associated epilepsy, a pressing clinically relevant phenotype. Finally, we will use pharmacological approaches to assess the underlying mechanisms driving acutely Pik3ca-dependent seizures in post-natal mice. These assays represent the first step toward developing molecularly rational epilepsy therapy in PIK3CA segmental brain overgrowth syndrome patients.