Therapeutic Strategies for Gaucher Disease: Miglustat (NB-DNJ) as a Pharmacological Chaperone for Glucocerebrosidase and the Different Thermostability of Velaglucerase Alfa and Imiglucerase
Therapeutic Strategies for Gaucher Disease: Miglustat (NB-DNJ) as a Pharmacological Chaperone for Glucocerebrosidase and the Different Thermostability of Velaglucerase Alfa and Imiglucerase
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DOI：
10.1021/mp200313e
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发表时间：
2011-11-01
影响因子：
4.9
通讯作者：
Sancho, Javier
中科院分区：
文献类型：
--
作者：
Abian, Olga;Alfonso, Pilar;Sancho, Javier
戈谢病（GD）是一种由溶酶体糖脑苷酶（GlcCerase）活性缺乏引起的鞘糖脂代谢紊乱，由于构象或功能缺陷变异，导致糖基神经酰胺在巨噬细胞中进行性沉积。葡萄糖类似物N-butyldeoxynojirimycin （NB-DNJ, miglustat）是神经酰胺特异性糖基转移酶的抑制剂，该酶催化鞘糖脂生物合成的第一步，目前被批准用于口服治疗1型GD。在之前的工作中，我们发现NB-DNJ存在时细胞培养中的GlcCerase活性增加，这可能意味着该化合物不仅是底物减减剂，而且是GlcCerase降解的药理学伴侣或抑制剂。在这项工作中，我们比较了imiglucerase（目前用于替代治疗的酶）和velaglucerase alfa（一种新的治疗酶形式）在构象稳定性和酶活性方面的差异，以及NB-DNJ对它们的影响。用等温滴定量热法研究了这些酶与NB-DNJ的相互作用。我们的研究结果表明，尽管velaglucerase alfa和imiglucerase表现出非常相似的活性谱，但velaglucerase alfa具有更高的体外热稳定性，并且不易聚集/沉淀，这可能有利于储存和临床给药。此外，我们发现在中性pH下，NB-DNJ结合并增强了这两种酶的稳定性，而在轻度酸性溶酶体条件下，它不与它们结合。这些结果支持NB-DNJ作为药物伴侣的潜在作用，在未来可能成为GD药物配方或联合治疗的一部分。
Gaucher disease (GD) is a disorder of glycosphingolipid metabolism caused by deficiency of lysosomal glucocerebrosidase (GlcCerase) activity, due to conformationally or functionally defective variants, resulting in progressive deposition of glycosylceramide in macrophages. The glucose analogue, N-butyldeoxynojirimycin (NB-DNJ, miglustat), is an inhibitor of the ceramide-specific glycosyltransferase, which catalyzes the first step of glycosphingolipid biosynthesis and is currently approved for the oral treatment of type 1 GD. In a previous work, we found a GlcCerase activity increase in cell cultures in the presence of NB-DNJ, which could imply that this compound is not only a substrate reducer but also a pharmacological chaperone or inhibitor for GlcCerase degradation. In this work we compare imiglucerase (the enzyme currently used for replacement therapy) and velaglucerase alfa (a novel therapeutic enzyme form) in terms of conformational stability and enzymatic activity, as well as the effect of NB-DNJ on them. The interaction between these enzymes and NB-DNJ was studied by isothermal titration calorimetry. Our results reveal that, although velaglucerase alfa and imiglucerase exhibit very similar activity profiles, velaglucerase alfa shows higher in vitro thermal stability and is less prone to aggregation/precipitation, which could be advantageous for storage and clinical administration. In addition, we show that at neutral pH NB-DNJ binds to and enhances the stability of both enzymes, while at mildly acidic lysosomal conditions it does not bind to them. These results support the potential role of NB-DNJ as a pharmacological chaperone, susceptible of being part of pharmaceutical formulation or combination therapy for GD in the future.