Lee L, Abe A, Shayman J A
Division of Nephrology, Department of Internal Medicine, University of Michigan Medical Center, Ann Arbor, Michigan 48109, USA.
J Biol Chem. 1999 May 21;274(21):14662-9. doi: 10.1074/jbc.274.21.14662.
Previous work has led to the identification of inhibitors of glucosylceramide synthase, the enzyme catalyzing the first glycosylation step in the synthesis of glucosylceramide-based glycosphingolipids. These inhibitors have two identified sites of action: the inhibition of glucosylceramide synthase, resulting in the depletion of cellular glycosphingolipids, and the inhibition of 1-O-acylceramide synthase, resulting in the elevation of cell ceramide levels. A new series of glucosylceramide synthase inhibitors based on substitutions in the phenyl ring of a parent compound, 1-phenyl-2-palmitoylamino-3-pyrrolidino-1-propanol (P4), was made. For substitutions of single functional groups, the potency of these inhibitors in blocking glucosylceramide synthase was primarily dependent upon the hydrophobic and electronic properties of the substituents. An exponential relationship was found between the IC50 of each inhibitor and the sum of derived hydrophobic (pi) and electronic (sigma) parameters. This relationship demonstrated that substitutions that increased the electron-donating characteristics and decreased the lipophilic characteristics of the homologues enhanced the potency of these compounds in blocking glucosylceramide formation. A novel compound was subsequently designed and observed to be even more active in blocking glucosylceramide formation. This compound, D-threo-4'-hydroxy-P4, inhibited glucosylceramide synthase at an IC50 of 90 nM. In addition, a series of dioxane substitutions was designed and tested. These included 3',4'-methylenedioxyphenyl-, 3',4'-ethylenedioxyphenyl-, and 3'4'-trimethylenedioxyphenyl-substituted homologues. D-threo-3', 4'-Ethylenedioxy-P4-inhibited glucosylceramide synthase was comparably active to the p-hydroxy homologue. 4'-Hydroxy-P4 and ethylenedioxy-P4 blocked glucosylceramide synthase activity at concentrations that had little effect on 1-O-acylceramide synthase activity. These novel inhibitors resulted in the inhibition of glycosphingolipid synthesis in cultured cells at concentrations that did not significantly raise intracellular ceramide levels or inhibit cell growth.
先前的研究已鉴定出葡萄糖神经酰胺合酶的抑制剂，该酶催化基于葡萄糖神经酰胺的糖鞘脂合成中的第一步糖基化反应。这些抑制剂有两个已确定的作用位点：抑制葡萄糖神经酰胺合酶，导致细胞糖鞘脂耗竭；抑制1-O-酰基神经酰胺合酶，导致细胞神经酰胺水平升高。基于母体化合物1-苯基-2-棕榈酰氨基-3-吡咯烷基-1-丙醇（P4）苯环上的取代基，制备了一系列新的葡萄糖神经酰胺合酶抑制剂。对于单官能团取代，这些抑制剂阻断葡萄糖神经酰胺合酶的效力主要取决于取代基的疏水和电子性质。发现每种抑制剂的IC50与衍生的疏水（π）和电子（σ）参数之和之间存在指数关系。这种关系表明，增加同系物供电子特性并降低其亲脂特性的取代基增强了这些化合物阻断葡萄糖神经酰胺形成的效力。随后设计并观察到一种新型化合物在阻断葡萄糖神经酰胺形成方面更具活性。这种化合物，D-苏式-4'-羟基-P4，以90 nM的IC50抑制葡萄糖神经酰胺合酶。此外，设计并测试了一系列二氧六环取代物。这些包括3',4'-亚甲基二氧苯基-、3',4'-亚乙基二氧苯基-和3'4'-三亚甲基二氧苯基取代的同系物。D-苏式-3',4'-亚乙基二氧-P4抑制葡萄糖神经酰胺合酶的活性与对羟基同系物相当。4'-羟基-P4和亚乙基二氧-P4在对1-O-酰基神经酰胺合酶活性影响很小的浓度下阻断葡萄糖神经酰胺合酶活性。这些新型抑制剂在不显著提高细胞内神经酰胺水平或抑制细胞生长的浓度下，导致培养细胞中糖鞘脂合成受到抑制。
