Herrlinger U, Jacobs A, Quinones A, Woiciechowsky C, Sena-Esteves M, Rainov N G, Fraefel C, Breakefield X O
Neurology Service, Massachusetts General Hospital and Harvard Medical School, Charlestown 02129, USA.
Hum Gene Ther. 2000 Jul 1;11(10):1429-38. doi: 10.1089/10430340050057503.
Subcutaneous vaccination therapy with glioma cells, which are retrovirally transduced to secrete granulocyte-macrophage colony-stimulating factor (GM-CSF), has previously proven effective in C57BL/6 mice harboring intracerebral GL261 gliomas. However, clinical ex vivo gene therapy for human gliomas would be difficult, as transgene delivery via retroviral vectors occurs only in dividing cells and ex vivo glioma cells have a low growth fraction. To circumvent this problem, a helper virus-free herpes simplex virus type 1 (HSV-1) amplicon vector was used. When primary cultures of human glioblastoma cells were infected with HSV-1 amplicon vectors at an MOI of 1, more than 90% of both dividing and nondividing cells were transduced. When cells were infected with an amplicon vector, HSVGM, bearing the GM-CSF cDNA in the presence of Polybrene, GM-CSF secretion into the medium during the first 24 hr after infection was 1026 ng/10(6) cells, whereas mock-infected cells did not secrete detectable GM-CSF. Subcutaneous vaccination of C57BL/6 mice with 5 x 10(5) irradiated HSVGM-transduced GL261 cells 7 days prior to intracerebral implantation of 10(6) wild-type GL261 cells yielded 60% long-term survivors (>80 days), similar to the 50% long-term survivors obtained by vaccination with retrovirally GM-CSF-transduced GL261 cells. In contrast, animals vaccinated with the same number of nontranduced GL261 cells or with GL261 cells infected with helper virus-free packaged HSV-1 amplicon vectors carrying no transgene showed only 10% long-term survivors. In conclusion, helper virus-free HSV-1 amplicon vectors appear to be effective for cytokine-enhanced vaccination therapy of glioma, with the advantages that both dividing and nondividing tumor cells can be infected, no viral proteins are expressed, and these vectors are safe and compatible with clinical use.
用逆转录病毒转导以分泌粒细胞巨噬细胞集落刺激因子（GM-CSF）的胶质瘤细胞进行皮下接种治疗，先前已证明对患有脑内GL261胶质瘤的C57BL/6小鼠有效。然而，人类胶质瘤的临床离体基因治疗将很困难，因为通过逆转录病毒载体的转基因递送仅发生在分裂细胞中，并且离体胶质瘤细胞的生长分数较低。为了解决这个问题，使用了无辅助病毒的单纯疱疹病毒1型（HSV-1）扩增载体。当人胶质母细胞瘤细胞的原代培养物以1的感染复数用HSV-1扩增载体感染时，超过90%的分裂和非分裂细胞都被转导。当细胞用携带GM-CSF cDNA的扩增载体HSVGM在聚凝胺存在下感染时，感染后最初24小时内GM-CSF分泌到培养基中的量为1026 ng/10(6)细胞，而模拟感染的细胞不分泌可检测到的GM-CSF。在脑内植入10(6)个野生型GL261细胞前7天，用5×10(5)个经辐射的HSVGM转导的GL261细胞对C57BL/6小鼠进行皮下接种，产生了60%的长期存活者（>80天），类似于用逆转录病毒GM-CSF转导的GL261细胞接种获得的50%的长期存活者。相比之下，用相同数量的未转导的GL261细胞或用感染了无辅助病毒包装的不携带转基因的HSV-1扩增载体的GL261细胞接种的动物仅显示10%的长期存活者。总之,无辅助病毒的HSV-1扩增载体似乎对胶质瘤的细胞因子增强接种治疗有效，其优点是分裂和非分裂肿瘤细胞均可被感染，不表达病毒蛋白，且这些载体安全并与临床应用兼容。
