E.J. Hidvégi

Detailed characterization of the mouse glioma 261 tumor model for experimental glioblastoma therapy

Mouse glioma 261 (Gl261) cells are used frequently in experimental glioblastoma therapy; however, no detailed description of the Gl261 tumor model is available. Here we present that Gl261 cells carry point mutations in the K‐ras and p53 genes. Basal major histocompatibility complex (MHC)I, but not MHCII, expression was detected in Gl261 cells. The introduction of interferon‐γ‐encoding genes increased expression of both MHCI and MHCII. A low amount of B7‐1 and B7‐2 RNA was detected in wild‐type cells, but cytokine production did not change expression levels. Gl261 cells were transduced efficiently by adenoviral vectors; the infectivity of retroviral vectors was limited. Low numbers of transplanted Gl261 cells formed both subcutaneous and intracranial tumors in C57BL/6 mice. The cells were moderately immunogenic: prevaccination of mice with irradiated tumor cells 7 days before intracranial tumor challenge prevented tumor formation in approximately 90% of mice. When vaccination was carried out on the day or 3 days after tumor challenge, no surviving animals could be found. In vitro‐growing cells were radiosensitive: less than 2 Gy was required to achieve 50% cell mortality. Local tumor irradiation with 4 Gy X‐rays in brain tumor‐bearing mice slowed down tumor progression, but none of the mice were cured off the tumor. In conclusion, the Gl261 brain tumor model might be efficiently used to study the antitumor effects of various therapeutic modalities, but the moderate immunogenicity of the cells should be considered. (Cancer Sci 2006; 97: 546 – 553)

Local tumor irradiation augments the antitumor effect of cytokine-producing autologous cancer cell vaccines in a murine glioma model

The combined therapeutic effect of cytokine-producing cancer cell vaccines and local radiotherapy was studied in a mouse glioma 261 (Gl261) brain tumor model. Brain tumor–bearing mice were treated with cytokine (IL-4, IL-6, IL-7, GM-CSF, TNF-α, LIF, LT) producing vaccines made by in vitro transduction of Gl261 cells with the corresponding adenoviral vectors. Vaccines producing either IL-4 or GM-CSF cured 20–40% of mice. The antitumor effect strongly depended on the secreted cytokine level. Vaccination therapy induced specific activation of cytotoxic T lymphocytes measured by cell-mediated cytotoxicity assay. Brain tumors were heavily infiltrated by CD4+ lymphocytes after treatment with IL-4– or GM-CSF–secreting cells. GM-CSF vaccination induced moderate CD8+ infiltration, as well. Depleting either CD4+ or CD8+ lymphocyte subsets abolished the anticancer effect of GM-CSF–expressing cells. Strong synergism was observed by combining cytokine vaccination (GM-CSF, IL-4, IL-12) with local tumor irradiation: about 80–100% of the glioma-bearing mice was cured. The high efficiency of combined treatment was maintained even under suboptimal conditions when neither of the modalities cured any of the mice alone. This suggests that vaccination therapy might open a new potential in the clinical treatment of high-grade gliomas when applied as adjuvant to existing treatment modalities.

Decreased weight, DNA, RNA and protein content of the brain after neutron irradiation of the 18-day mouse embryo.

Pregnant mice were irradiated with 0.5 Gy fission neutrons on the eighteenth day of their gestation. The average litter size at birth was unchanged but mortality increased 5-6 fold in the first 3 days. The irradiated mice were the same weight as control mice at birth but showed a progressively increasing weight deficiency up to at least 36 days as compared to controls. Brain weight was 37, 45 and 25 per cent less in 2-, 3- and 52-week old irradiated animals, respectively, and the ratio of brain weight to body weight was 25, 27 and 13 per cent less. The concentrations of DNA, RNA and protein (mg/g wet tissue) were the same in irradiated and control mice in both brain and liver at all three ages. Total DNA, RNA and protein contents of whole brain after irradiation were 56-75 per cent of the control levels. No definite decrease was observed in liver. Histological study at 6 hours after irradiation showed nuclear pyknosis in the central nervous system from definite to very severe according to the part examined. It is concluded that damage to the central nervous system of the 18-day mouse foetus after neutron irradiation is mainly due to killing and/or inhibition of the differentiation of neuroblasts.

Carcinogenic alterations in murine liver, lung, and uterine tumors induced by in utero exposure to ionizing radiation

The atomic bombing of Hiroshima and Nagasaki and the nuclear accident at Chernobyl raised the question of prenatal sensitivity to ionizing radiation–induced cancer. In this study, mice were exposed to single doses of γ‐radiation (0.2–2.0 Gy) at different embryonic stages. The tumor incidence increased with dose from 15% in control mice to 35% in mice irradiated with 2.0 Gy on 18 d of prenatal life. Various oncogenic events were investigated in lymphoid, liver, lung, and uterine tumors. We observed threefold to fivefold increases in myc expression in 25% of the lymphomas, and the expression of Ha‐ras and p53 genes decreased in 40% and 60% of the lung tumors by twofold to fivefold. Point mutations were tissue specific: Ha‐ras codon 61 mutations were found in about 40% of the liver adenocarcinomas, Ki‐ras codon 12 mutations in about 17% of lung tumors, and p53 mutations in about 15% of the lymphomas. Amplification and rearrangement of the p53, myc, and Ha‐, Ki‐ and N‐ras genes were not detected. Loss of heterozygosity on chromosome 4 at the multiple tumor suppressor 1 and 2 genes was observed in all types of malignancies. Allelic losses on chromosome 11 at the p53 locus were found in lymphoid, liver, and lung tumors, but they were absent from uterine tumors. Multiple oncogenic changes were often detected. The frequency of carcinogenic alterations was similar in spontaneous and radiation‐induced lymphoid, liver, and uterine tumors. In radiation‐induced lung adenocarcinomas, however, the incidences of many oncogenic changes were different from those found in their spontaneous counterparts. This suggests that different oncogenic pathways are activated during spontaneous and in utero γ‐radiation–induced murine lung carcinogenesis. Mol. Carcinog. 21:100–110, 1998.

Brain Tumor Treatment with IL-2 and IL-12 Producing Autologous Cancer Cell Vaccines

In humans, gliomas constitute about 30-45% of all intracranial tumours. Conventional therapy for high-grade gliomas consists of surgery that is followed by radiation therapy and/or by chemotherapy. The life expectancy of patients with glioblastoma is less than one year from the time of diagnosis’, therefore development of new therapeutic approaches is necessary. Animal models have demonstrated the efficacy of cytokine gene therapy of melanomas using autologous cancer cells producing various cytokines2’3. We decided to study the therapeutic effect of cytokine expressing cancer cell vaccines for the treatment of high-grade gliomas.

Oncogenes and tumor suppressor genes in murine tumors induced by neutron- or gamma-irradiation in utero

Abstract Half a century after the A-bomb explosions, the cancer mortality is still increasing in prenatally exposed survivors. In this study, mice were exposed to neutron- or γ-irradiation of different embryonic stages. The tumor incidence increased to 35% after 2.0 Gy γ-irradiation on the 18th day of gestation, compared with 15% in the unirradiated controls. Most of the tumors developed in the lymphoid system, liver, lung and uterus. Some brain tumors were observed in mice irradiated in utero both with neutron and γ-rays, but none were found in the unirradiated controls. The expression of the p53 gene decreased by 2–5-fold in 60% of lung tumors. Allelic losses on chromosome 11 at the p53 locus were observed in 30–40% of the spontaneous and radiation-induced lymphoid tumors and liver adenocarcinomas. In lung tumors, about one-third of the spontaneous adenocarcinomas exhibited allelic losses at the p53 locus not associated with point mutation. This fact may suggest that the loss at one of p53 alleles is an earlier event during carcinogenesis than development of point mutations.