Pharmacological enhancement of mutated α-glucosidase activity in fibroblasts from patients with Pompe disease
Pharmacological enhancement of mutated α-glucosidase activity in fibroblasts from patients with Pompe disease
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DOI：
10.1038/sj.mt.6300074
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发表时间：
2007-03-01
影响因子：
12.4
通讯作者：
Andria, Generoso
中科院分区：
文献类型：
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作者：
Parenti, Giancarlo;Zuppaldi, Alfredo;Andria, Generoso
我们研究了使用药物伴侣治疗庞贝氏症，这是一种代谢性肌病，由于编码溶酶体水解酶α-葡萄糖苷酶（GAA）的基因突变，其特征是心脏和骨骼肌中的广义糖原蓄积。我们研究了两种亚氨基糖，脱氧野尻霉素（DNJ）和N-丁基脱氧野尻霉素（NB-DNJ），对残留的GAA活性的成纤维细胞从8例不同形式的庞贝氏症（两个经典的婴儿，两个非经典的婴儿发病，四个晚发型形式），并与不同的GAA基因突变。我们证明了在来自携带突变L552 P（三名患者）和G549 R（一名患者）的患者的成纤维细胞中，亚氨基糖处理后GAA活性显著增加（1.3 - 7.5倍）。在HEK 293 T细胞中证实了GAA增强，其中相同的突变过表达。对于其他突变，未观察到GAA活性增加。Western印迹分析表明，亚氨基糖增加了成熟GAA分子形式的量。在HEK 293 T细胞中过表达L552 P突变的免疫荧光研究表明，在亚氨基糖处理后，突变酶向溶酶体的运输得到改善。这些结果为庞贝氏症的替代治疗提供了理论依据，而不是酶替代。
We investigated the use of pharmacological chaperones for the therapy of Pompe disease, a metabolic myopathy due to mutations of the gene encoding the lysosomal hydrolase alpha-glucosidase (GAA) and characterized by generalized glycogen storage in cardiac and skeletal muscle. We studied the effects of two imino sugars, deoxynojirimycin (DNJ) and N-butyldeoxynojirimycin (NB-DNJ), on residual GAA activity in fibroblasts from eight patients with different forms of Pompe disease ( two classic infantile, two non-classic infantile onset, four late-onset forms), and with different mutations of the GAA gene. We demonstrated a significant increase of GAA activity (1.3 - 7.5-fold) after imino sugar treatment in fibroblasts from patients carrying the mutations L552P ( three patients) and G549R ( one patient). GAA enhancement was confirmed in HEK293T cells where the same mutations were overexpressed. No increase of GAA activity was observed for the other mutations. Western blot analysis showed that imino sugars increase the amount of mature GAA molecular forms. Immunofluorescence studies in HEK293T cells overexpressing the L552P mutation showed an improved trafficking of the mutant enzyme to lysosomes after imino sugar treatment. These results provide a rationale for an alternative treatment, other than enzyme replacement, to Pompe disease.