Stimulating SMN2 exon 7 inclusion with short RNAs

描述(申请人提供)：脊髓性肌萎缩症(SMA)是一种常染色体隐性遗传性神经退行性疾病，是导致婴儿死亡的最常见的遗传原因。SMA决定基因位于染色体5q上，称为存活运动神经元-1(SMN1)。值得注意的是，存在一种几乎相同的复制基因，称为SMN2。与SMN1基因相比，该基因具有编码相同蛋白质的能力，然而，由于单个沉默的非多态核苷酸差异，大多数SMN2衍生的转录本被选择性剪接，并编码一种被截断的、具有生化缺陷的蛋白质SMN？7。 到目前为止，SMN2是唯一的SMA修饰基因。较温和的表型与SMN2拷贝数的增加相关，通常在2到4个拷贝之间。SMA的遗传背景使这种疾病特别适合于治疗干预，包括：SMN2在基本上所有SMA患者中保留；SMN2在所有组织中普遍表达；SMN2保留编码正常的全长SMN蛋白的能力。因此，SMN2已被确定为潜在的SMA治疗的主要靶点。最有吸引力的可能性包括刺激SMN 2总转录和/或调节SMN 2选择性剪接事件。 为了利用独特的SMA遗传背景，这项应用的目标是开发调节SMN2剪接的新型RNA。通过使用病毒传递系统(目标1)，将在各种实验环境中对这些RNA进行检查，以确定诱导全长SMN2表达水平最高的RNA(目标1和2)。然后，将在SMA的小鼠模型中检查排名第一的候选RNA，以确定RNAs是否可以调节生物体中的SMN2，以及预期的全长SMN2表达的增加是否会减少转基因小鼠中众所周知的温和SMA表型(目标3)。 虽然本申请中描述的实验对SMA疗法的开发具有直接影响，但这些结果可以用作广泛遗传疾病的模型，在这些疾病中，纠正剪接缺陷将恢复致病基因的功能。

DESCRIPTION (provided by applicant): Spinal muscular atrophy (SMA) is an autosomal recessive neurodegenerative disorder and is the most common genetic cause of infantile death. The SMA-determining gene is located on chromosome 5q, and is called survival motor neuron-1 (SMN1). Remarkably, a nearly identical copy gene is present called SMN2. This gene has the capacity to encode an identical protein compared to SMN1, however, due to a single silent non-polymorphic nucleotide difference, the majority of SMN2-derived transcripts are alternatively spliced and encode a truncated and biochemically defective protein called SMN?7. To date, SMN2 is the only SMA modifying gene. Milder phenotypes correlate with an increase in the number of SMN2 copies, typically ranging from two to four copies. The genetic context of SMA makes this disease especially amenable to therapeutic intervention including: SMN2 is retained in essentially all SMA patients; SMN2 is ubiquitously expressed in all tissues; and SMN2 retains the capacity to encode a normal, full-length SMN protein. Therefore, SMN2 has been identified as a major target for a potential SMA therapies. The most attractive possibilities include stimulating total SMN2 transcription and/or modulation of the SMN2 alternative splicing event. To take advantage of the unique SMA genetic context, the goal of this application is to develop novel RNAs that modulate SMN2 splicing. Through the use of a viral delivery system (Aim 1), these RNAs will be examined in a variety of experimental contexts designed to identify RNAs that induces the highest level of full-length SMN2 expression (Aim 1 and 2). The top candidate RNAs will then be examined in a murine model of SMA to determine whether the RNAs can modulate SMN2 in an organism and whether this expected increase in full-length SMN2 expression lessens the well described mild SMA phenotype in transgenic mice (Aim 3). While the experiments described in this application have immediate implications for the development of a SMA therapy, the results could be used as a model for a broad range of genetic disorders in which correcting a splicing defect would restore functionality to a disease-causing gene.