Yagi K, Ohishi N, Hamada A, Shamoto M, Ohbayashi M, Ishida N, Nagata A, Kanazawa S, Nishikimi M
Institute of Applied Biochemistry, Yagi Memorial Park, Mitake, Gifu, Japan.
Hum Gene Ther. 1999 Aug 10;10(12):1975-82. doi: 10.1089/10430349950017338.
For gene therapy of human malignant glioma, we adopted positively charged multilamellar liposomes entrapping the human interferon beta (hIFN-beta) gene. One week after the transplantation of human malignant glioma U251-SP cells to produce glioma in nude mouse brain, the liposomes entrapping the gene (500 ng of DNA per 25 nmol of lipids per 2 microl) were injected into the same site of the cell transplantation once every second day for a total of five injections; and by this means the tumor completely disappeared. To confirm the antiproliferative effect of hIFN-beta, we performed an in vitro study using a plasmid containing a secretion signal sequence-deleted hIFN-beta gene and one containing the hIFN-beta gene inserted in reverse. In both cases, there was no hIFN-beta release into the medium and no growth inhibition effect. On addition of anti-hIFN-beta antibody to the medium, the growth inhibition effect was abolished. As this cell line expresses IFN-alpha/beta receptor, the hIFN-beta produced in the transfected cells could be released and acted in a paracrine manner. For 120 days the body weight change of normal mice treated by the same procedure as used in the curing experiment was not significant among the groups injected with empty liposomes, plasmid only, and liposomes entrapping the gene. In all of these three groups, death, abnormal behavior, and significant histological changes were not observed.
对于人类恶性胶质瘤的基因治疗，我们采用了包裹人干扰素β（hIFN-β）基因的带正电荷的多层脂质体。将人类恶性胶质瘤U251-SP细胞移植到裸鼠脑内以产生胶质瘤一周后，将包裹基因的脂质体（每2微升中含500纳克DNA和25纳摩尔脂质）每隔一天注射到细胞移植的同一部位，共注射五次；通过这种方法肿瘤完全消失。为了证实hIFN-β的抗增殖作用，我们使用含有缺失分泌信号序列的hIFN-β基因的质粒和含有反向插入的hIFN-β基因的质粒进行了体外研究。在这两种情况下，均没有hIFN-β释放到培养基中，也没有生长抑制作用。向培养基中加入抗hIFN-β抗体后，生长抑制作用消失。由于该细胞系表达IFN-α/β受体，转染细胞中产生的hIFN-β可以释放并以旁分泌方式发挥作用。在按照治愈实验中使用的相同程序处理的正常小鼠中，注射空脂质体、仅注射质粒以及注射包裹基因的脂质体的组之间，120天内体重变化不显著。在所有这三组中，均未观察到死亡、异常行为和明显的组织学变化。
