摘要
报道了血友病B基因治疗临床Ⅰ期试验的安全性研究,包括反转录病毒基因的DNA聚合酶链式反应(PCR)、反转录PCR(RT/PCR)、Southern杂交以及neo基因和lacZ基因的补救分析(resoueassay).从DNA水平。RNA水平和病毒活体水平对野生型病毒进行了检测,没有检测到反转录病毒,并对兔和人转基因细胞进行形态学观察、染色体核型分析、软琼脂实验、裸鼠接种实验、兔和裸鼠的病理检测及电镜分析以检测恶性细胞的存在与否,同时进行支原体检测、内毒素和过敏原测定,结果均为阴性.建立了有复制能力的野生型反转录病毒(RCR)的检测系统,为基因治疗临床试验的安全性提供了保证.
This paper deals with safety assessments required for animal test and clinical trial in haemophilia B gene therapy. Based on ourprevious works on gene transfer therapy for hemophilia B, a series of safety assessments were complered before clinical trial. Firstly, the existence of replication-completent retrovirus (RCR)were sensitively detected by Neo gene and lac Z gene rescue assay on the cellular level, and PCR, RT/PCR with Southern blot and dot blot on DNA and RNA level,results were found to be negative. Secondly, malignant transformation was followed on cells transduced with F ac cDNA by various techipues including cell morphology, histochemistry, chromsome karyotyping, soft-agar test, nude mice test and electromicroscopy, no abnormal changes were observed. Lastly, presence of bacteria, mold, endotoxin, toxin, allergen was rotinely done for culture medium and collagen and others used in clinical trial. All these safety assessments demonstrate that the safety of implantion of transduced cells is guaranteed.
出处
《复旦学报(自然科学版)》
CAS
CSCD
北大核心
1994年第5期559-568,共10页
Journal of Fudan University:Natural Science
基金
国家科委863高科技项目
关键词
基因疗法
血友病B
安全性
病毒
野生型
replication-competent retrovitus (RCR), gnen therapy, hemophilia B, rescue assay, RT/PCR