Gene therapy for malignant brain tumor by infusion of antisense oligonucleotide

1.恶性胶质瘤的遗传学特征我们在31例人脑胶质瘤中发现了6例10号染色体微卫星不稳定性。恶性胶质瘤中细胞凋亡较少。这些结果提示该基因异常与胶质瘤的恶性转化密切相关.建立定量检测脑、肝、肾组织中反义寡核苷酸的方法。DNA片段在丙烯酰胺凝胶上电泳并转移到膜上。用放射性同位素标记的正义寡核苷酸对反义寡核苷酸进行定量检测.经颈动脉灌注缓激肽选择性递送反义寡核苷酸我们从荷瘤大鼠颈内动脉注射反义寡核苷酸。通过上述方法测量递送的寡核苷酸。与对照研究相比，反义寡核苷酸选择性地进入脑肿瘤中的数量增加了1.4倍.颈动脉内输注缓激肽对犬生命体征的影响：我们用20只犬研究了颈动脉内输注缓激肽对生命体征的影响。在1和2.5伽马剂量下，颈动脉内缓激肽输注没有显著改变生命体征。另一方面，在5和10伽马剂量下，其在输注开始时改变了包括脑血流量在内的生命体征。

1. Genetic characterization of malignant gliomasWe recognized the microsatellite instability in the chromosome 10 in 6 out of 31 humanﾟCgliomas. And rare apoptosis was recognized in malignant gliomas. These findingsﾟCsuggested that the genetic abnormality strongly associated with the malignantﾟCtransformation of the gliomas.2. Establishment of quantitative measurement of antisenseoligonucleotide in theﾟCorgans.After obtaining the organs, brain, liver, and kidney, DNA fractions was extracted using centrifugation and filters. The DNA fractions were electrophorated on the acrylamide gel and trasbiotted to the membrane. A quantitative measurement of antisenseoligonucleotide was performed using RI-labeled senseoligonucleotide.3. Selective delivery of antisenseoligonucleotide by intracarotid bradykinin infusion We injected antisenseoligonucleotide from internal carotid artery in tumor-bearing rat. The delivered oligonucleotide was measured by the methods described above. The antisenseoligonucleotide was selectively delivered into the brain tumor by 1.4 fold, compared with the control study.3. Effect of intracarotid bradykinin infusion on vital signs in dogs.Using 20 dogs, we examined the effect of intracarotid bradykinin infusion on vital signs. At the dose of 1 and 2.5 gamma, intracarotid bradykinin infusion did not change the vital signs significantly. On the other hand, at the dose of 5 and 10 gamma, it changed the vital signs including cerebral blood flow at the beginning of the infusion.