Okada H, Miyamura K, Itoh T, Hagiwara M, Wakabayashi T, Mizuno M, Colosi P, Kurtzman G, Yoshida J
Department of Neurosurgery, Nagoya University School of Medicine, Japan.
Gene Ther. 1996 Nov;3(11):957-64.
The efficacy of gene therapy for glioma was examined using adeno-associated virus (AAV)-based vectors to deliver genes to experimental tumors in mice. Stereotactic injection of 2 x 10(5) U-251SP human glioma cells into the brains of nude mice produced tumors of 19.06 +/- 1.79 mm2 17 days after injection. Employing a high titer preparation of AAV vector containing the gene for beta-galactosidase (AAV-lacZ), dose-dependent transduction of U-251SP cells was seen in vitro. When 1.6 x 10(10) AAV-lacZ particles were directly injected into tumors in vivo, 30-40% of the cells along the needle track expressed beta-galactosidase. Transduction of U-251SP cells in vitro with an AAV vector containing a bicistronic gene encoding both herpes simplex thymidine kinase and human interleukin-2 (AAV-tk-IRES-IL2) rendered them sensitive to the cytocidal effects of ganciclovir (GCV) and IL-2 was produced in a dose-dependent manner. Cocultures of AAV-tk-IRES-IL2 transduced cells and nontransduced cells proved highly sensitive to GCV indicating the contribution of the bystander effect. Stereotactic delivery of 6 x 10(10) AAV-tk-IRES-IL2 particles into day 7 tumors in nude mice followed by administration of GCV for 6 days, resulted in a 35-fold reduction in the mean volume of tumors compared with controls. Normal brains did not suffer from any toxic effect of the administration of AAV-tk-IRES-IL2 and GCV. These results indicate that high titer AAV vector treatment may be safe and effective for in vivo gene therapy of human brain tumors.
使用基于腺相关病毒（AAV）的载体将基因导入小鼠实验性肿瘤中，以检测基因治疗对胶质瘤的疗效。将2×10⁵个U - 251SP人胶质瘤细胞立体定向注射到裸鼠脑内，注射后17天产生面积为19.06±1.79 mm²的肿瘤。使用含有β - 半乳糖苷酶基因（AAV - lacZ）的高滴度AAV载体制剂，在体外观察到U - 251SP细胞的剂量依赖性转导。当将1.6×10¹⁰个AAV - lacZ颗粒直接体内注射到肿瘤中时，沿针道的30 - 40%的细胞表达β - 半乳糖苷酶。用含有编码单纯疱疹胸苷激酶和人白细胞介素 - 2的双顺反子基因的AAV载体（AAV - tk - IRES - IL2）在体外转导U - 251SP细胞，使其对更昔洛韦（GCV）的杀细胞作用敏感，并且以剂量依赖性方式产生IL - 2。AAV - tk - IRES - IL2转导细胞与未转导细胞的共培养物对GCV高度敏感，表明存在旁观者效应。将6×10¹⁰个AAV - tk - IRES - IL2颗粒立体定向递送至裸鼠第7天的肿瘤中，随后给予GCV 6天，与对照组相比，肿瘤平均体积减少了35倍。正常脑未受到AAV - tk - IRES - IL2和GCV给药的任何毒性作用。这些结果表明，高滴度AAV载体治疗对于人脑肿瘤的体内基因治疗可能是安全有效的。
