微型双功能抗体在裸鼠体内介导人T细胞杀伤白血病细胞

Specific targeted cytotoxicity to resistant leukemia cells mediated by anti-CD3/anti-CD19 diabody

目的:研究抗CD3/抗CD19微型双功能抗体介导人T细胞对白血病细胞的特异性靶向杀伤活性。方法:利用Ficoll-Hypaque法分离外周血淋巴细胞(PBL),流式细胞术从中分选T淋巴细胞,FACS检测T细胞表面标记CD25和CD69激活后的表达变化,实时定量PCR检测各组穿孔素(Perforin)和颗粒酶A(Granzyme A)的释放,建立BALB/c裸鼠Raji细胞移植瘤模型,测定该双功能抗体介导的体内靶向杀伤活性。结果:激活的T细胞,双功能抗体组CD25和CD69的表达以及释放Perforin和Granzyme A的量均明显高于对照组,在对人白血病裸鼠移植瘤模型的生物治疗中,抗CD3/抗CD19微型双功能抗体能有效抑制白血病移植瘤的生长。结论:抗CD3/抗CD19微型双功能抗体在体外及动物肿瘤模型实验中能介导人T细胞有效杀伤白血病细胞,具有潜在的临床应用前景。

Objective：To study the specific targeted cytotoxicity to resistant leukemia cells medidated by anti-CD3/anti-CD19 diabody.Methods：Human peripheral blood lymphocytes（PBL） were isolated using Ficoll-Hypaque density-gradient centrifugation from heparinized blood of healthy volunteers and monocytes were depleted by adherence to plastic flasks.Also releasing of perforin and granzyme A,expressing the membrane molecular CD25 and CD69 on activated T cells of the same group,and apotosis of these T cells were analyzed by qPCR or FACS respectively.The effect of the anti-CD3/anti-CD19 diabody mediated lysis of Raji cells was assayed by human leukemia nude mice xenograft model in vivo.Results：Diabody increased the cytotoxity of T cells in vitro apparently than the control groups（P〈0.05）.so the secretion of the perforin and Granzyme A,CD25 and CD69 expressed on activated T cells were increased much more than in the other groups（P〈0.05）.Furthermore,the Diabody showed improved tumor inhibiting activities（P〈0.05） and improved antitumor activity in nude mice bearing Raji xenografts.Conclusion：The diabody is proved to be a potent agent for mediating T lymphocyte cytotoxicity to leukemia cells in vitro and in vivo.