Rainov N G, Kramm C M, Banning U, Riemann D, Holzhausen H J, Heidecke V, Burger K J, Burkert W, Körholz D
Department of Neurosurgery, Martin-Luther-University Halle-Wittenberg, Halle, Germany.
Gene Ther. 2000 Nov;7(21):1853-8. doi: 10.1038/sj.gt.3301311.
This study was conducted to investigate immunological components of somatic gene therapy for primary glioblastoma multiforme (GBM) in adults. It involved 13 patients treated by surgical resection of tumor with subsequent radiation therapy. Seven of them received additional herpes simplex virus thymidine kinase/ganciclovir (HSV-tk/GCV) gene therapy by direct intracerebral injection of retrovirus (RV) vector producing cells (VPC) during tumor surgery and subsequent systemic administration of GCV. Peripheral blood for FACS immunophenotyping, isolation of peripheral mononuclear cells (PMNC), and serum ELISA assays for IL-12 and soluble Fas ligand (sFasL) was collected daily during the first 4 post-operative weeks. Tumor specimens were obtained at primary or recurrent surgery and at autopsy. Tumors from gene therapy patients showed varying degrees of peritumoral necrosis around the former tumor resection cavity. Numbers of tumor-infiltrating lymphocytes found weeks after gene therapy were not significantly increased compared with primary tumors. Mitotic tumor cells were sparse close to the VPC injection sites, but abundant in brain areas somewhat distant from these sites. Serum ELISA revealed significantly increased sFasL and IL-12 levels in the gene therapy group compared with controls. Immunophenotyping of PMNC did not show a significant activation of T cells or NK cells during gene therapy. Interferon gamma secretion was evaluated by ELISPOT assays employing PMNC cocultivated with autologous tumor cells. It demonstrated an antitumor immune response in the gene therapy group, but not in the control group. These findings support the concept of in vivo induction of a systemic immune response by local intracerebral HSV-tk/GCV pharmacogene therapy for primary human GBM.
本研究旨在调查成人原发性多形性胶质母细胞瘤（GBM）体细胞基因治疗的免疫成分。研究涉及13例接受肿瘤手术切除并随后进行放射治疗的患者。其中7例患者在肿瘤手术期间通过直接脑内注射产生逆转录病毒（RV）载体的细胞（VPC）接受了额外的单纯疱疹病毒胸苷激酶/更昔洛韦（HSV-tk/GCV）基因治疗，并在术后进行了更昔洛韦的全身给药。在术后的前4周每天采集外周血用于流式细胞术免疫表型分析、分离外周单核细胞（PMNC）以及进行白细胞介素-12和可溶性Fas配体（sFasL）的血清ELISA检测。在初次或复发性手术以及尸检时获取肿瘤标本。接受基因治疗患者的肿瘤在前肿瘤切除腔周围显示出不同程度的瘤周坏死。与原发性肿瘤相比，基因治疗数周后发现的肿瘤浸润淋巴细胞数量没有显著增加。有丝分裂的肿瘤细胞在VPC注射部位附近稀疏，但在距离这些部位稍远的脑区丰富。血清ELISA显示，与对照组相比，基因治疗组的sFasL和IL-12水平显著升高。PMNC的免疫表型分析未显示基因治疗期间T细胞或NK细胞有显著激活。通过使用与自体肿瘤细胞共培养的PMNC的ELISPOT检测评估干扰素γ分泌。结果表明基因治疗组有抗肿瘤免疫反应，而对照组没有。这些发现支持了通过局部脑内HSV-tk/GCV药物基因治疗对原发性人类GBM进行体内系统性免疫反应诱导的概念。
