Doxorubicin attached to HPMA copolymer via amide bond modifies the glycosylation pattern of EL4 cells
Doxorubicin attached to HPMA copolymer via amide bond modifies the glycosylation pattern of EL4 cells
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DOI：
10.1007/s13277-010-0019-7
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发表时间：
2010-08-01
期刊：
影响因子：
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通讯作者：
Rihova, Blanka
中科院分区：
文献类型：
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作者：
Kovar, Lubomir;Etrych, Tomas;Rihova, Blanka
为了避免抗癌药物阿霉素（Dox）的副作用，我们将该药物与N-（2-羟丙基）甲基丙烯酰胺（HPMA）共聚物骨架结合。Dox通过酰胺键（Dox-HPMA（AM），PK 1）或腙pH敏感键（Dox-HPMA（HYD））缀合。与Dox和Dox-HPMA（HYD）相反，Dox-HPMA（AM）在细胞的细胞内膜内积累，包括高尔基复合体和内质网的细胞内膜，两者都参与蛋白质糖基化。流式细胞术用于测定凝集素结合和细胞死亡，免疫印迹用于表征CD 7、CD 43、CD 44和CD 45的存在，高效阴离子交换色谱法与脉冲安培检测器分析用于表征质膜糖组成。与Dox或Dox-HPMA（HYD）相比，用Dox-HPMA（AM）缀合物孵育EL 4细胞增加了膜表面相关糖蛋白的量，以及由花生凝集素、鸡冠刺桐或半乳糖凝集素-1凝集素识别的糖部分。只有Dox-HPMA（AM）增加了高度糖基化的膜糖蛋白CD 43的表达，而其他（CD 7，CD 44和CD 45）的表达不受影响。半乳糖凝集素-1的结合位点存在于CD 43分子上。此外，我们提出用Dox-HPMA（AM）处理的EL 4具有增加的对半乳糖凝集素-1诱导的凋亡的敏感性。在这项研究中，我们证明了Dox-HPMA（AM）治疗改变EL 4 T细胞淋巴瘤表面的糖基化，并使细胞对半乳糖凝集素-1诱导的细胞凋亡敏感。
To avoid the side effects of the anti-cancer drug doxorubicin (Dox), we conjugated this drug to a N-( 2-hydroxypropyl)methacrylamide (HPMA) copolymer backbone. Dox was conjugated via an amide bond (Dox-HPMA(AM), PK1) or a hydrazone pH-sensitive bond (Dox-HPMA(HYD)). In contrast to Dox and Dox-HPMA(HYD), Dox-HPMA(AM) accumulates within the cell's intracellular membranes, including those of the Golgi complex and endoplasmic reticulum, both involved in protein glycosylation. Flow cytometry was used to determine lectin binding and cell death, immunoblot to characterize the presence of CD7, CD43, CD44, and CD45, and high-performance anion exchange chromatography with pulsed amperometric detector analysis for characterization of plasma membrane saccharide composition. Incubation of EL4 cells with Dox-HPMA(AM) conjugate, in contrast to Dox or Dox-HPMA(HYD), increased the amounts of membrane surface-associated glycoproteins, as well as saccharide moieties recognized by peanut agglutinin, Erythrina cristagalli, or galectin-1 lectins. Only Dox-HPMA(AM) increased expression of the highly glycosylated membrane glycoprotein CD43, while expression of others (CD7, CD44, and CD45) was unaffected. The binding sites for galectin-1 are present on CD43 molecule. Furthermore, we present that EL4 treated with Dox-HPMA(AM) possesses increased sensitivity to galectin-1-induced apoptosis. In this study, we demonstrate that Dox-HPMA(AM) treatment changes glycosylation of the EL4 T cell lymphoma surface and sensitizes the cells to galectin-1-induced apoptosis.