【作者】 杨华；

【导师】 李恒；

【作者基本信息】 华中科技大学 ， 外科学， 2010， 硕士

【摘要】 目的对中国医药集团武汉生物制品研究所自主研发的生物制剂2类新药“注射用鼠抗人T淋巴细胞CD25抗原单克隆抗体（WuTac）”进行I期临床肾移植病人多次给药耐受性和药代动力学试验（SFDA批件号:2003L03465）,评价WuTac在肾移植病人应用的安全性,对疗效进行初步探索,为制定本药物Ⅱ期临床试验给药方案提供安全的剂量范围。方法选择18例亲属活体供肾肾移植受者,分0.1 mg/kg（n=8）和0.2 mg/kg（n=10）两个剂量组,肾移植术前两小时给药1次,术后连续7天每天给药1次,静脉滴注WuTac 0.1 mg·kg^-1·d^-10或0.2 mg·kg^-1·d^-10;基础免疫抑制采用环孢素（CsA）+霉酚酸酯（MMF）+泼尼松三联免疫抑制方案。评价肾移植病人对WuTac的最大耐受剂量和不良事件的发生特点;双抗体夹心ELISA法检测血药浓度,药理学软件DAS 2.0处理血药浓度数据并计算药代动力学参数,评价WuTac在肾移植病人体内的药代动力学特征;间接ELISA法检测术后人抗鼠抗体（HAMA）的滴度,监测肾移植病人产生HAMA的规律;流式细胞仪连续检测外周血淋巴细胞亚群,初步探索WuTac免疫抑制作用的特点。数据录入、核查、分析采用EpiData 3.1、SPSS 15.0软件,显著性水平α选用0.05,Origin Pro8.0软件绘图。结果两组所有受试者均未出现首剂反应、细胞因子释放综合征和急性排斥反应。0.1 mg/kg剂量组纳入的8名受试者肾移植术后恢复顺利,无严重不良事件发生;按照试验对WuTac最大耐受剂量确证的需要,0.2 mg/kg剂量组共纳入10名受试者,其中一例受试者因用药观察期出现血清肌酐升高,于术后第3天退出试验,停用WuTac并辅以血液透析治疗后移植肾功能恢复;在完成试验的9名受试者中,发生移植肾功能延迟恢复1例,用药观察期出现血清肌酐升高1例,此两例受试者均辅以血液透析治疗后移植肾功能恢复;用药观察期发生肺部烟曲霉菌感染1例,对症治疗并调整免疫抑制剂后肺部感染治愈;此3例受试者和中途退出试验的1例受试者体重均大于50 kg,每日WuTac用药剂量超过10 mg,0.2 mg/kg组其余受试者肾移植术后恢复顺利。药代动力学参数t1/2、、tmax、Vd、CL、AUCss/D、AUC0-t/D、AUC0-∞/D两组无差异（P>0.05）,Cmax/D有统计学差异（P<0.05）。当WuTac血药浓度大于1000μg·L-1时,WuTac能明显抑制CD25+淋巴细胞,而对外周血T淋巴细胞总数（CD3+CD19-T淋巴细胞）、辅助T淋巴细胞（CD3+CD4+T淋巴细胞）、细胞毒T淋巴细胞（CD3+CD8+T淋巴细胞）、B淋巴细胞（CD3-CD19+淋巴细胞）、NK细胞（CD3-CD16+CD56+细胞）比例无影响。按0.1 mg·kg^-1·d^-10给药,WuTac与活化T淋巴细胞的结合发挥封闭作用约为10¹4天;按0.2 mg·kg^-1·d^-10给药,WuTac与活化T淋巴细胞的结合发挥封闭作用约为14²1天;0.1 mg/kg组有1例受试者术后第14天HAMA检测呈阳性,0.2 mg/kg组有1例受试者术后第35天HAMA检测呈阳性。结论WuTac术前两小时给药一次,术后连续7天每天给药一次,最大耐受剂量为0.2 mg·kg^-1·d^-10,且每日总剂量不超过10 mg;在推荐的安全剂量范围内,WuTac不增加不良事件的发生率。药代动力学试验证实:WuTac分布在血浆中,与组织或血浆蛋白结合较少。WuTac与活化T淋巴细胞的结合发挥封闭作用,而不是杀伤或清除作用。WuTac对肾移植病人的有效治疗浓度为1000μg·L-1,当WuTac血药浓度大于1000μg·L-1时,就能完全和稳定地阻断IL-2受体;当WuTac血药浓度小于1000μg·L-1时,CD25+淋巴细胞比例开始回升,1²周恢复至治疗前水平。肾移植病人使用WuTac后产生HAMA比例低,不影响WuTac的疗效。更多还原

【Abstract】 Objective WuTac is a murine monoclonal antibody against CD25 antigen, The CD25 also named interleukin 2 [alpha] chain receptor （Tac） which exists on the surface of activated human T lymphocytes. This anti-CD25 monoclonal antibody is manufactured by Wuhan Institute of Biologic Products in China. This study examined the safety and tolerance, pharmacokinetics, and pharmacodynamics multiple doses of WuTac injection in renal transplant patients. The range of safety dose is provided in this study for phaseⅡclinical trial of WuTac. Methods The 18 recipients of living related transplants were matched for this study, recipients were divided 0.1 mg/kg dose（n=8） and 0.2 mg/kg dose（n=10）. Triple combined immunesuppressive protocols consisted of cyclosporine A （CsA）, mycophenolate mofetil （MMF） and prednisone. The first infusion of WuTac (0.1 mg·kg^-1 or 0.2 mg·kg^-1) was administered on day 0 approximately 2 hours before transplantation and received infusion once-daily for 7 days after transplantation. All patients were monitored until day 40 by serial laboratory tests, clinical examinations, and recording of adverse events. Flow cytometry enumeration of lymphocyte subsets bearing the surface markers CD3, CD4, CD8, CD16CD56, CD19, CD25. Double-antibody-sandwich ELISA was used to quantitate WuTac in serum. Parameters of pharmacokinetics were calculated by software DAS 2.0（Drug and Statistics Software）, the law of HAMA in renal transplant recipients is explained by indirect ELISA. All data are inputted and analysised by EpiData 3.1, SPSS 15.0,α=0.05,graphics were done using Origin Pro 8.0 program. Results The clinical trial was successfully performed in 17 of the 18 renal transplant recipients, all participants in two doses didn’t have acute rejection in the observation period. 8 participants in 0.1 mg/kg dose recovered successfully. 10 renal transplant recipients participated in the trial of 0.2 mg/kg dose, participants GZ06 in 0.2 mg/kg dose quited from this study because serum creatinine increasing, so 9 participants accomplished trial, one case of 0.2 mg/kg dose suffered delayed renal graft function, one case occurred serum creatinine increasing, pneumonia developed in another case, the weight of those three cases and the patient who quited this program all exceeded 50 kg, the daily dose is greater than 10 mg, others participants in 0.2 mg/kg dose recovered smoothly. First-dose reaction and cytokine release syndrome were not observed in all participants, the incidence of adverse reactions was not statistically different in the two doses in 40 days after renal transplantation. Parameters of pharmacokinetics: t1/2, tmax, Vd, CL, AUCss/D, AUC0-t/D, AUC0-∞/D are no significant difference between two doses （P>0.05）, except Cmax/D （P<0.05）. Enzyme-linked immunosorbent assay （ELISA） of WuTac indicate that circulating through blood level exceed effective concentration (1000μg·L^-1) and peaked at day 7 after renal transplantation. T cell （CD3+CD19-）, Th cell （CD3+CD4+）, CTL（CD3+CD8+）, B cell（CD3-CD19+）, NK cell（CD3-CD16+CD56+） was not statistically different between preoperative and postoperative in the two doses（P>0.05）, except CD25+ lymphocyte cell decreased significantly after the first dose in the patients who received WuTac, the level of the cells that expressed CD25 was significantly decreased about 10 to 14 days and 14 to 21 days in the two doses after transplantation. Both doses with one patient developed HAMA after transplantation. Conclusion The maximum dose of WuTac is 0.2 mg·kg^-1·d^-1 and daily dose is not more than 10 mg. The biological tolerance of WuTac is well tolerated and safety by participants in recommended safe and tolerable dose. WuTac distribute in the blood plasma in the body. Complete and consistent binding to IL-2Rαin adults is maintained as long as serum WuTac levels exceed 1000μg·L^-1. As concentrations fall below this threshold, the IL-2Rαsites are no longer fully bound and the number of T-cells expressing unbound IL-2Rαreturns to pretherapy values within 1-2 weeks. Fewer of HAMA in renal transplant recipients did not influence curative effect of WuTac.更多还原