Pathogenesis and Treatment of Bone Disease in the Mucopolysaccharidoses

摘要 粘多糖病(MPS)是一种遗传性、溶酶体储存障碍家族，其特征是 降解糖胺多聚糖(GAG)的酶缺乏。患有MPS的患者患有残疾 对目前的治疗没有反应的骨骼异常。MPS VII呈现出一种特别严重的 骨骼表型，患者表现为进行性后凸畸形和脊髓受压 导致慢性疼痛和瘫痪。MPS VII是由β-葡萄糖苷酸酶活性不足引起的，导致 多种口吃类型的堆积。将这种GAG堆积与细胞联系起来的分子机制 人们对功能障碍和骨骼疾病知之甚少，阻碍了有效治疗的发展。我们的 实验室使用了一种与临床相关的自然发生的MPS vii犬模型，它非常接近于 发生在人类患者身上的骨骼疾病的进展。在以前的工作中，我们证明了MPS VII 狗的椎骨中有软骨损伤，损害了椎间关节的稳定性。这些 损伤是由出生后生长过程中软骨转化为骨的失败引起的。在初步研究中，我们 已经确定了MPS VII犬首次出现异常骨化的确切发育窗口 这可以追溯到驻留的软骨细胞在肥大成熟过程中未能进展。 我们还发现，从早期开始，MPS VIII的骨痂软骨中就有异常的Gag积聚。 年龄，并且，通过全转录组测序，存在Wnt/β-连环蛋白信号的失调 在疾病发展的这个关键时刻。WNT生长因子是关键的调节因素 已知软骨细胞分化和GAG是Wnt分布和活性的重要调节因素， 这表明GAG的积累和这一途径的失调之间存在联系。这样做的目的是 建议研究MPS VII中失败的骨形成机制，并建立改进的 使用临床相关犬类模型的治疗范例。我们的中心假设是 MPS-VII骨痂软骨中GAG的积累扰乱了必要的信号通路 启动并维持软骨细胞肥大分化。此外，我们假设 要有效地治疗MPS VII中的骨病，既要使GAG转换正常化，又要 在骨痂软骨中激活必要的成骨信号通路。在目标1中，我们将定义时间 生长激素蓄积与骨痂软骨细胞分化潜能的空间关系 MPS VII犬从出生到骨骼成熟的骨病进展的关键阶段。在《目标2》中我们将 Wnt/β-catenin信号转导异常在MPS骨形成延迟中的关键作用 七条狗。在目标3中，我们将建立Wnt/β-连环蛋白信号的治疗靶点，单独和联合使用 配合酶替代疗法，可促进MPS VII犬的骨形成。

Abstract The mucopolysaccharidoses (MPS) are a family of genetic, lysosomal storage disorders characterized by deficiencies in enzymes that degrade glycosaminoglycans (GAGs). Patients with MPS suffer from crippling skeletal abnormalities that are unresponsive to current treatments. MPS VII presents with a particularly severe skeletal phenotype, where patients exhibit progressive kyphoscoliotic deformity and spinal cord compression resulting in chronic pain and paralysis. MPS VII is caused by deficient beta-glucuronidase activity, leading to accumulation of multiple GAG types. The molecular mechanisms linking this GAG accumulation to cellular dysfunction and skeletal disease are poorly understood, impeding development of effective therapies. Our laboratory uses a clinically-relevant, naturally-occurring canine model of MPS VII that closely mimics the progression of skeletal disease that occurs in human patients. In previous work we demonstrated that MPS VII dogs have cartilaginous lesions in the vertebrae that compromise the stability of the intervertebral joint. These lesions are caused by failed conversion of cartilage to bone during postnatal growth. In preliminary studies, we have identified the precise developmental window when abnormal ossification first manifests in MPS VII dogs and that this can be traced to a failure of resident chondrocytes to progress through hypertrophic maturation. We have also shown that there is abnormal GAG accumulation in MPS VII epiphyseal cartilage from an early age, and, using whole transcriptome sequencing, that there is dysregulation of the Wnt/β-catenin signaling pathway at this crucial juncture in the disease progression. Wnt growth factors are critical regulators of chondrocyte differentiation, and GAGs are known to be important regulators of Wnt distribution and activity, suggesting a link between GAG accumulation and dysregulation of this pathway. The objectives of this proposal are to investigate mechanisms of failed bone formation in MPS VII and establish improved treatment paradigms using a clinically-relevant canine model. Our central hypothesis is that abnormal accumulation of GAGs in MPS VII epiphyseal cartilage disrupts the signaling pathways necessary to initiate and sustain chondrocyte hypertrophic differentiation. Further, we hypothesize that to effectively treat bone disease in MPS VII, it will be necessary to both normalize GAG turnover and activate requisite osteogenic signaling pathways in epiphyseal cartilage. In Aim 1 we will define temporal and spatial relationships between GAG accumulation and epiphyseal chondrocyte differentiation potential at key stages of bone disease progression in MPS VII dogs, from birth to skeletal maturity. In Aim 2 we will establish the critical role of Wnt/β-catenin signaling dysregulation in delayed epiphyseal bone formation in MPS VII dogs. In Aim 3 we will establish if therapeutic targeting of Wnt/β-catenin signaling, alone and in combination with enzyme replacement therapy, can enhance bone formation in MPS VII dogs.