Development of IL-1 with selective activities by introduction of carbohydrates

通过引入碳水化合物开发具有选择性活性的 IL-1

• 批准号: 07557159
• 负责人: ONOZAKI Kikuo
• 金额: $ 4.8万
• 依托单位: Faculty of Pharmaceutical Sciences, Nagoya City University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (A)
• 财政年份: 1995
• 资助国家: 日本
• 起止时间: 1995 至 1996
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-07557159/
• 关键词: neoglycoprotein; cytokine; interleukin 1; glycotechnology; arteficial glycoprotein; インターロイキン; 糖

IL-1 is potentially useful for therapy because of its antitumor effect and protective effect on animals against infection, radiation and chemotherapy. However, IL-1 exhibits serious deleterious effects. In this study we attempted to develop IL-1 with less deleterious effects by chemically introducing oligosaccharides to recombinant human IL-1alpha.1) Tissue distribution of mannose dimer-introduced IL-1We have previously succeeded in the synthesis of mannose dimer-introduced IL-1alpha with selective activities in vivo. In this study the effect of glycosylation on tissue distribution was examined. Mannose dimer-introduced IL-1 was labeled with ^<125>I,and then examined its tissue distribution after i.p.or i.v.injection into mice. Compared to the nonglycosylated IL-1, the glycosylated IL-1 distributed more in the liver, and less in the kidney.2) Analysis of glycosylated site in mannose dimer-introduced IL-1Mannose dimer-introduced IL-1 was dijested with lysylendopeptidase, and glycosylate … More d sites were analyzed by TOF-MASS.The conserved regions among IL-1alpha and IL-1beta were not glycosylated. However, the regions exposed to the surface of IL-1alpha molecule were glycosylated.3) Synthesis, purification and biological activities of galactose-introduced IL-1D-Galactose monosaccharide was coupled to IL-1alpha by the acyl azide method, and was purified by anion-exchange column chromatography. Galactose introduced per molecule of IL-1 was estimated to be 9. We examined a variety of IL-1 activities in vitro, including proliferative effect on T cells, antiproliferative effect on myeloid leukemia cells and melanoma cells, stimulatory effects on IL-6 syntheisi by melanoma cells and PGE2 synthesis by fibroblast cells. The glycosylated IL-1 exhibited reduced activities from ten to thousand times compared with nonglycosyslated IL-1 in all the activities performed in vitro. The glycosylated IL-1 exhibited reduced binding affinities to type I and II IL-1 receptors, suggesting that the reduced activities are due, at least partially, to the decrease in the binding affinities. Less

IL-1由于其抗肿瘤作用和对动物抵抗感染、放射和化学疗法的保护作用而潜在地用于治疗。然而，IL-1表现出严重的有害作用。在这项研究中，我们试图通过将寡糖化学引入重组人IL-1 α来开发具有较少有害作用的IL-1。1）甘露糖二聚体引入的IL-1的组织分布我们先前已经成功地合成了具有体内选择性活性的甘露糖二聚体引入的IL-1 α。在这项研究中，糖基化对组织分布的影响进行了检查。将甘露糖二聚体引入的IL-1用13 I标记<125>，然后在腹膜内或静脉内注射到小鼠中后检查其组织分布。与非糖基化IL-1相比，糖基化IL-1在肝脏中分布较多，在肾脏中分布较少。2）甘露糖二聚体导入IL-1糖基化位点分析 ...更多信息 用TOF-MASS分析IL-1 α和IL-1 β之间的保守区，发现IL-1 α和IL-1 β之间的保守区没有糖基化。IL-1 α分子表面暴露的区域被糖基化。3）半乳糖导入IL-1D的合成、纯化及生物学活性通过酰基叠氮法将半乳糖单糖偶联到IL-1 α上，并通过阴离子交换柱层析进行纯化。每分子IL-1引入的半乳糖估计为9。本实验检测了IL-1的多种体外活性，包括对T细胞的增殖作用、对髓系白血病细胞和黑色素瘤细胞的抗增殖作用、对黑色素瘤细胞合成IL-6的刺激作用和对成纤维细胞合成PGE_2的刺激作用。糖基化的IL-1在体外进行的所有活性中，与非糖基化的IL-1相比，表现出降低的活性从十倍到千倍。糖基化的IL-1表现出降低的I型和II型IL-1受体的结合亲和力，这表明活性降低至少部分是由于结合亲和力的降低。少

项目成果

T.Takii, N.Niki, D.Yang, H.Kimura, A.Ito, H.Hayashi and K.Onozaki: "Type I and type II interferons up-regulate functional type I interleukin 1 receptor in a human fibroblast cell line TIG-1." J.Interferon and Cytokine Res.15. 1065-1072 (1995)

T.Takii、N.Niki、D.Yang、H.Kimura、A.Ito、H.Hayashi 和 K.Onozaki：“I 型和 II 型干扰素上调人成纤维细胞系中功能性 I 型白细胞介素 1 受体

D.Yang,H.Hayashi,Y.Hiyama T.Takii,K.Onozaki: "Transfection of human melanomacells with typeI IL-2 receptor cDNA rendered them IL-1-responsive and revealed the importance of ODC activity dowk-regulation in IL-1 inducd growth inhibition." Journal of Biochem

D.Yang、H.Hayashi、Y.Hiyama T.Takii、K.Onozaki：“用 I 型 IL-2 受体 cDNA 转染人类黑色素瘤细胞，使其具有 IL-1 响应性，并揭示了 ODC 活性 dowk 调节在 IL 中的重要性

D.Yang,H.Hayashi,Y.Hiyama T.Takii,K.Onozaki: "Transfectionon of human melanoma cells with ytpe I interleukin-1(IL-1)receptor cDNA them IL-1 responsive and revealed the improtance of ODC activity down-regulation in IL-1-induced growth inhibition." Journal

D.Yang、H.Hayashi、Y.Hiyama T.Takii、K.Onozaki：“用 ytpe I 白细胞介素 1 (IL-1) 受体 cDNA 转染人类黑色素瘤细胞，它们对 IL-1 产生反应，并揭示了 ODC 活性的重要性

Y.Takei,D.Yang,T.Chiba,S.Nabeshima M.Naruoka,K.Wada,K.Onozaki: "D-mannose dimer introduced human recombinant interleukin 1α,neo IL-1α,exhibits altered tissue distribution in mice." Journal of Interferon and Cytokine Research. 16. 333-336 (1996)

Y. Takei、D. Yang、T. Chiba、S. Nabeshima M. Naruoka、K. Wada、K. Onozaki：“D-甘露糖二聚体引入人重组白细胞介素 1α、neo IL-1α，在小鼠中表现出组织分布的改变。 “干扰素和细胞因子研究杂志。16. 333-336 (1996)

S.Ito, H.Hayashi, N.Watanabe, Y.Kobayashi, T.Takii and K.Onozaki: "Interleukin 1 (IL-1) production is not essential for acquired resistance of human melanoma cells A375 to anti-proliferative effect of IL-1." Int.J.Cancer. 65. 805-811 (1996)

S.Ito、H.Hayashi、N.Watanabe、Y.Kobayashi、T.Takii 和 K.Onozaki：“白细胞介素 1 (IL-1) 的产生对于人黑色素瘤细胞 A375 的抗增殖作用的获得性抵抗并不重要。