Orai1 as a Therapeutic Target for Muscular Dystrophy

钙池操纵性钙内流（SOCE）是2001年首次报道的一种较新的骨骼肌钙内流现象， 肌肉. SOCE通过两种蛋白质之间的偶联来协调：肌浆中的STIM 1钙传感器 网（SR）和钙渗透性Orai 1通道在横小管（TT）膜。SOCE增强 肌肉生长/发育，限制疲劳，并促进抗疲劳I型纤维规格。上 另一方面，SOCE功能障碍导致衰老中的肌肉无力/疲劳，加重肌肉无力/疲劳， STIM 1和Orai 1基因的突变导致衰弱性肌病。反映的情况 STIM 1/Orai 1依赖性SOCE活性的严格调节对于最佳肌肉性能至关重要， SOCE活性的增加或减少可导致肌肉疲劳、肌肉减少症和肌病。因此，在本发明中， Orai 1依赖性SOCE代表了肌营养不良症的潜在治疗靶点。我们最近证明，SOCE促进骨骼肌生长，限制肌肉疲劳，并加剧肌营养不良症的严重程度，在肌营养不良蛋白（mdx）和δ-肌聚糖缺陷（sgcd-/-）小鼠。对于这种R21应用，我们开发了他莫昔芬诱导的肌肉特异性Orai 1敲除小鼠，以确定在mdx和sgcd-/-小鼠中观察到的营养不良表型中骨骼肌中Orai 1依赖性Ca 2+进入的特定作用，mdx和sgcd-/-小鼠分别建立了Duchene肌营养不良和肢带肌营养不良的小鼠模型。我们还与CalciMedica Inc.建立了合作关系。评价全身递送4种有效的新的研究性SOCE通道抑制剂在减轻mdx和sgcd-/-小鼠的肌病表型中的功效。我们将使用这些新的研究工具和合作，以及全面的多学科实验方法，来评估抑制Orai 1依赖性SOCE作为肌营养不良症治疗干预的有效性。基于我们已发表的和初步的数据，我们假设，部分抑制Orai 1依赖性的钙离子进入骨骼肌提供保护，防止肌病在小鼠模型的肌营养不良症，而不增加易感性肌肉疲劳。这一中心假设的有效性将在两个具体目标中得到严格的检验。目的1将使用他莫昔芬诱导的，肌肉特异性Orai 1基因敲除小鼠，以确定部分发育后，肌肉特异性减少SOCE对肌营养不良症的影响。目的2将确定通过与CalciMedica Inc.合作获得的新一代Orai 1通道抑制剂的全身给药的治疗效果。以减少mdx和sgcd-/-小鼠的营养不良表型。该项目的结果将提供有关靶向Orai 1依赖性Ca 2+进入作为肌营养不良症治疗的治疗潜力所需的临床前证据。

Being first reported in 2001, store-operated Ca2+ entry (SOCE) is a relatively new phenomenon in skeletal muscle. SOCE is coordinated by coupling between two proteins: STIM1 calcium sensors in the sarcoplasmic reticulum (SR) and Ca2+-permeable Orai1 channels in the transverse tubule (TT) membrane. SOCE enhances muscle growth/development, limits fatigue, and promotes fatigue-resistant type I fiber specification. On the other hand, SOCE dysfunction contributes to muscle weakness/fatigue in aging, exacerbates muscular dystrophy, and mutations in STIM1 and Orai1 genes result in debilitating myopathies. The picture that emerges is that tight regulation of STIM1/Orai1-dependent SOCE activity is critical for optimal muscle performance such that increases or decreases in SOCE activity can lead to muscle fatigue, sarcopenia, and myopathy. Thus, Orai1-dependent SOCE represents a provocative potential therapeutic target for muscular dystrophy. We recently demonstrated that SOCE promotes skeletal muscle growth, limits muscle fatigue, and exacerbates the severity of muscular dystrophy in dystrophin- (mdx) and δ-sarcoglycan-deficient (sgcd-/-) mice. For this R21 application, we developed tamoxifen-inducible, muscle-specific Orai1 knockout mice in order to determine the specific role of Orai1-dependent Ca2+ entry in skeletal muscle in the dystrophic phenotypes observed in mdx and sgcd-/- mice, established mouse models of Duchene Muscular Dystrophy and Limb Girdle Muscular Dystrophy, respectively. We also established a collaboration with CalciMedica Inc. to evaluate the efficacy of systemic delivery of 4 potent new investigational SOCE channel inhibitors in mitigating the myopathic phenotypes of mdx and sgcd-/- mice. We will use these new research tools and collaborations, together with a comprehensive multi-disciplinary experimental approach, to evaluate the efficacy of inhibiting Orai1-dependent SOCE as a therapeutic intervention for muscular dystrophy. Based on our published and preliminary data, we hypothesize that partial inhibition of Orai1-dependent Ca2+ entry in skeletal muscle provides protection against myopathy in mouse models of muscular dystrophy without enhancing susceptibility to muscle fatigue. The validity of this central hypothesis will be rigorously tested in two Specific Aims. Aim 1 will use tamoxifen-inducible, muscle-specific Orai1 knockout mice to determine the impact of partial post-developmental, muscle-specific reduction of SOCE on muscular dystrophy. Aim 2 will determine the therapeutic efficacy of systemic administration of new generation Orai1 channel inhibitors obtained through a collaboration from CalciMedica Inc. to reduce the dystrophic phenotypes of mdx and sgcd-/- mice. The results of this project will provide needed preclinical evidence regarding the therapeutic potential of targeting Orai1-dependent Ca2+ entry as a treatment for muscular dystrophy.