Hojman P, Gissel H, Gehl J
Laboratory of the Department of Oncology, University of Copenhagen at Herlev Hospital, Herlev, Denmark.
Gene Ther. 2007 Jun;14(12):950-9. doi: 10.1038/sj.gt.3302951. Epub 2007 Apr 5.
Electroporation-based gene transfer (electro gene transfer (EGT)) is gaining increasing momentum, in particular for muscle tissue, where long-term high-level expression is obtainable. Induction of expression using the Tet-On system was previously established; however, attempts to reach a predefined target dose - a prescription, have not been reported. We set three target haemoglobin levels (10, 12 and 14 mmol/l, base level was 8.2 mmol/l) and aimed at them by transferring the erythropoietin (EPO) gene to mouse tibialis cranialis (TC) muscle, and varying (1) DNA amount, (2) muscle mass transfected and (3) induction with the Tet-On system. Results showed that (a) using GFP, luciferase and EPO low DNA amounts were needed. In fact, 0.5 microg of DNA to one TC muscle led to significant Hgb elevation - this amount extrapolates to 1.4 mg of DNA in humans, (b) three prescribers hit the targets with average Hgb of 10.5, 12.0 and 13.7 mmol/l, (c) different approaches could be used, (d) undershooting could be corrected by retransferring, and (e) overshooting could be alleviated by reducing dose of inducer (doxycycline (dox)). In conclusion, this study shows that using EGT to muscle, a preset level of protein expression can be reached. This is of great interest for future clinical use.
基于电穿孔的基因转移（电基因转移（EGT））正越来越受到关注，特别是在肌肉组织中，能够实现长期高水平表达。此前已利用Tet-On系统诱导表达；然而，尚未有关于达到预定目标剂量（一种处方）的报道。我们设定了三个目标血红蛋白水平（10、12和14 mmol/L，基础水平为8.2 mmol/L），并通过将促红细胞生成素（EPO）基因转移至小鼠颅侧胫骨肌（TC），并改变（1）DNA量、（2）转染的肌肉质量和（3）用Tet-On系统诱导来实现这些目标。结果表明：（a）使用绿色荧光蛋白（GFP）、荧光素酶和EPO时，所需的DNA量较低。事实上，向一条TC肌肉注射0.5 μg DNA可使血红蛋白显著升高——此量推算至人体为1.4 mg DNA，（b）三位“处方制定者”达到了目标，平均血红蛋白水平分别为10.5、12.0和13.7 mmol/L，（c）可以采用不同方法，（d）未达到目标可通过再次转移来纠正，（e）超过目标可通过减少诱导剂（强力霉素（dox））剂量来缓解。总之，本研究表明，利用EGT作用于肌肉可达到预设的蛋白质表达水平。这对未来的临床应用具有重要意义。
