Enhanced circulating half-life and hematopoietic properties of a human granulocyte colony-stimulating factor/immunoglobulin fusion protein
Enhanced circulating half-life and hematopoietic properties of a human granulocyte colony-stimulating factor/immunoglobulin fusion protein
复制标题
DOI：
10.1016/j.exphem.2004.01.012
复制
发表时间：
2004-05-01
影响因子：
2.6
通讯作者：
Doherty, DH
中科院分区：
文献类型：
--
作者：
Cox, GN;Smith, DJ;Doherty, DH
客观的。本研究的目的是确定包含人粒细胞集落刺激因子 (G-CSF) 与人免疫球蛋白 G1 和 G4（IgG1 和 IgG4）Fc 和 C-H 结构域连接的融合蛋白是否具有生物活性，并具有改善的体内药代动力学和造血特性。 材料和方法。构建了编码人 G-CSF 的嵌合基因，该基因融合到人 IgG1 和 IgG4 的 Fc 和 CH 结构域的 N 端，并用于转染猴 COS 细胞。通过蛋白A亲和层析从条件培养基中纯化融合蛋白。通过 G-CSF 依赖性体外生物测定来测量蛋白质的生物活性。在正常大鼠中测量 G-CSF/IgG1-Fc 融合蛋白的药代动力学和粒细胞生成特性。结果。 G-CSF/IgG-Fc 和 G-CSF/IgG-C-H 融合蛋白主要作为二硫键连接的同二聚体从转染的 COS 细胞中分泌。在摩尔基础上，纯化的G-CSF/IgG-Fc融合蛋白在体外生物测定中与G-CSF一样具有活性，而纯化的G-CSF/IgG-CH融合蛋白的生物活性降低了3至4倍。与G-CSF相比，G-CSF/IgG1-Fc融合蛋白在对大鼠进行静脉和皮下给药后表现出较慢的血浆清除率，并且刺激循环中性粒细胞和白细胞的增加更大且更持久。结论。 G-CSF 与人 IgG 结构域的融合导致同源二聚体融合蛋白具有高体外生物活性、长循环半衰期和增强的体内造血特性。 (C) 2004 年国际实验血液学学会。由爱思唯尔公司出版
Objective. The aim of this study was to determine whether fusion proteins comprising human granulocyte colony-stimulating factor (G-CSF) joined to human immunoglobulin G1 and G4 (IgG1 and IgG4) Fc and C-H domains are biologically active and have improved pharmacokinetic and hematopoietic properties in vivo.Material and Methods. Chimeric genes encoding human G-CSF fused to the N-termini of the Fc and CH domains of human IgG1 and IgG4 were constructed and used to transfect monkey COS cells. The fusion proteins were purified from the conditioned media by protein A affinity chromatography. Bioactivities of the proteins were measured in a G-CSF-dependent in vitro bioassay. Pharmacokinetic and granulopoietic properties of the G-CSF/IgG1-Fc fusion protein were measured in normal rats.Results. The G-CSF/IgG-Fc and G-CSF/IgG-C-H fusion proteins were secreted from transfected COS cells primarily as disulfide-linked homodimers. On a molar basis, the purified G-CSF/IgG-Fc fusion proteins were as active as G-CSF in in vitro bioassays, whereas bioactivities of the purified G-CSF/IgG-CH fusion proteins were decreased 3- to 4-fold. The G-CSF/IgG1-Fc fusion protein displayed a slower plasma clearance rate and stimulated greater and longer lasting increases in circulating neutrophils and white blood cells than G-CSF following intravenous and subcutaneous administration to rats.Conclusion. Fusion of G-CSF to human IgG domains results in homodimeric fusion proteins possessing high in vitro bioactivities, long circulating half-lives, and enhanced hematopoietic properties in vivo. (C) 2004 International Society for Experimental Hematology. Published by Elsevier Inc.