zinostatin and Glioma

Topics: Animals; Brain Neoplasms; Dogs; Glioma; Humans; Meningeal Neoplasms; Meningioma; Neoplasm Metastasis; Neoplasm Seeding; Rabbits; Rats; Subarachnoid Space; Zinostatin

Neocarzinostatin (NCS) is a member of enediyne antibiotics with high anticancer potential. Our study was performed to explore the synergistic anti-glioma effects of NCS and paclitaxel (PTX) in vitro and in vivo. By 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, the cytotoxicities of the drugs to human glioma cells U87MG and rat glioma cells C6 were evaluated. The results showed that the combinations of NCS and PTX can synergistically inhibit glioma cells survival. Cell apoptosis was detected by flow cytometry, and the results showed that the combinations of NCS and PTX synergistically enhanced apoptosis ratio of glioma cells. Western blot revealed that the cell signaling pathways of proliferation and apoptosis were synergistically regulated, in which Akt was synergistically inactivated, p53 was up-regulated with down-regulation of bcl-2. Meanwhile, with the subcutaneous model of U87MG cells and intracerebral implantation model of C6 cells, the combination strategy could synergistically delay the glioma growth and significantly prolong the survival of rats bearing orthotopic glioma. This study demonstrates that the combination of NCS and PTX can potentiate the effect on survival and apoptosis of glioma cells via suppression of Akt, bcl-2, and activations of p53; Meanwhile, the in vivo studies also confirmed that the combination of NCS and PTX synergistically inhibit the gliom growth. Our data about the combinational effects of NCS with PTX may provide an alternative strategy for glioma therapy.

Topics: Animals; Antibiotics, Antineoplastic; Antineoplastic Agents, Phytogenic; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Brain Neoplasms; Cell Line, Tumor; Cell Proliferation; Drug Synergism; Glioma; Humans; Male; Mice, Nude; Paclitaxel; Proto-Oncogene Proteins c-akt; Proto-Oncogene Proteins c-bcl-2; Rats, Wistar; Tumor Burden; Tumor Suppressor Protein p53; Zinostatin

Gliomas are resistant to radiation therapy, as well as to TNFα induced killing. Radiation-induced TNFα triggers Nuclear factor κB (NFκB)-mediated radioresistance. As inhibition of NFκB activation sensitizes glioma cells to TNFα-induced apoptosis, we investigated whether TNFα modulates the responsiveness of glioma cells to ionizing radiation-mimetic Neocarzinostatin (NCS). TNFα enhanced the ability of NCS to induce glioma cell apoptosis. NCS-mediated death involved caspase-9 activation, reduction of mitochondrial copy number and lactate production. Death was concurrent with NFκB, Akt and Erk activation. Abrogation of Akt and NFκB activation further potentiated the death inducing ability of NCS in TNFα cotreated cells. NCS-induced p53 expression was accompanied by increase in TP53-induced glycolysis and apoptosis regulator (TIGAR) levels and ATM phosphorylation. siRNA-mediated knockdown of TIGAR abrogated NCS-induced apoptosis. While DN-IκB abrogated NCS-induced TIGAR both in the presence and absence of TNFα, TIGAR had no effect on NFκB activation. Transfection with TIGAR mutant (i) decreased apoptosis and γH2AX foci formation (ii) decreased p53 (iii) elevated ROS and (iv) increased Akt/Erk activation in cells cotreated with NCS and TNFα. Heightened TIGAR expression was observed in GBM tumors. While NCS induced ATM phosphorylation in a NFκB independent manner, ATM inhibition abrogated TIGAR and NFκB activation. Metabolic gene profiling indicated that TNFα affects NCS-mediated regulation of several genes associated with glycolysis. The existence of ATM-NFκB axis that regulate metabolic modeler TIGAR to overcome prosurvival response in NCS and TNFα cotreated cells, suggests mechanisms through which inflammation could affect resistance and adaptation to radiomimetics despite concurrent induction of death.

Topics: Adenosine Triphosphate; Antibiotics, Antineoplastic; Apoptosis; Apoptosis Regulatory Proteins; Ataxia Telangiectasia Mutated Proteins; Brain Neoplasms; Caspase 9; Cell Cycle Proteins; Cell Line, Tumor; DNA-Binding Proteins; Glioma; Glycolysis; Humans; Intracellular Signaling Peptides and Proteins; Lactic Acid; Microtubule-Associated Proteins; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; NF-kappa B; Phosphoric Monoester Hydrolases; Phosphorylation; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins c-akt; Reactive Oxygen Species; RNA Interference; RNA, Small Interfering; Tumor Necrosis Factor-alpha; Tumor Suppressor Protein p53; Tumor Suppressor Proteins; Zinostatin

To identify and mathematically model molecular predictors of response to the enediyne chemotherapeutic agent, neocarzinostatin, in nervous system cancer cell lines.. Human neuroblastoma, breast cancer, glioma, and medulloblastoma cell lines were maintained in culture. Content of caspase-3 and Bcl-2, respectively, was determined relative to actin content for each cell line by Western blotting and optical densitometry. For each cell line, sensitivity to neocarzinostatin was determined. Brain tumor cell lines were stably transfected with human Bcl-2 cDNA cloned into the pcDNA3 plasmid vector.. In human tumor cell lines of different tissue origins, sensitivity to neocarzinostatin is proportional to the product of the relative contents of Bcl-2 and caspase-3 (r (2) = 0.9; P < 0.01). Neuroblastoma and brain tumor cell lines are particularly sensitive to neocarzinostatin; the sensitivity of brain tumor lines to neocarzinostatin is enhanced by transfection with an expression construct for Bcl-2 and is proportional in transfected cells to the product of the relative contents of Bcl-2 and caspase-3 (r (2) = 0.7).. These studies underscore the potential of molecular profiling in identifying effective chemotherapeutic paradigms for cancer in general and tumors of the nervous system in particular.

Topics: Antibiotics, Antineoplastic; Biomarkers, Tumor; Caspase 3; Cell Line, Tumor; Cell Survival; Drug Resistance; Drug Screening Assays, Antitumor; Gene Expression Profiling; Glioma; Humans; Medulloblastoma; Nervous System Neoplasms; Neuroblastoma; Predictive Value of Tests; Proto-Oncogene Proteins c-bcl-2; Zinostatin

Neocarzinostatin (NCS) linked to the thiol group on the hinge region of the Fab' fragment of GA-17, a murine monoclonal antibody reacting with tyrosine-specific phosphorylated antigens, which are exclusively expressed on the cell surface of human astrocytomas, was evaluated for in vivo activity. GA-17-NCS immunoconjugates significantly suppressed the growth of human malignant glioma cell line U87-MG subcutaneous xenografts in nude mice until day 50 when administered intravenously into the tail vein. Disulphide- and thioether-linked GA-17-NCS were nearly equipotent immunoconjugates, but thioether-linked GA-17-NCS was more effective than disulphide-linked conjugates with 250 U/kg NCS content on day 50 (P < 0.05). Thioether-linked GA-17-NCS was significantly more effective on day 50 than free NCS with 500 U/kg or 250 U/kg NCS content (P < 0.05, P < 0.01, respectively). These results suggest that GA-17-NCS may prove useful in the treatment of human malignant gliomas.

Topics: Animals; Antibodies, Monoclonal; Astrocytoma; Cell Survival; Drug Screening Assays, Antitumor; Glioma; Immunotoxins; Mice; Mice, Nude; Zinostatin

Topics: Animals; Brain Neoplasms; Glioma; Infusions, Intravenous; Maleic Anhydrides; Neoplasm Transplantation; Polystyrenes; Rats; Rats, Inbred Strains; Survival Rate; Zinostatin

Because of technical difficulties, the pharmacokinetics of neocarzinostatin (NCS) have not been thoroughly evaluated in patients with malignant glioma. The authors produced anti-NCS antibody by immunizing rabbits with NCS and established a means of quantifying tissue levels of NCS with enzyme-linked immunosorbent assay. In one patient given a bolus injection of 1 mg of NCS intra-arterially, the concentration of drug in neoplastic tissue at 25 minutes (0.1136 micrograms/g) was higher than that in blood at 20 minutes and was retained for a longer period. Rapid entry of NCS into the tumor cavity was observed at 5 minutes. In two postoperative cases, NCS applied topically to the tumor site (50 and 100 micrograms) was retained at high levels (0.2941 and 3.33 micrograms/ml) even after 48 hours, although no NCS was detected in blood after 60 minutes. NCS concentrations as low as 1 microgram/ml demonstrated cytocidal effects, and a delay in tumor growth was observed even at an NCS level of 0.1 microgram/ml, despite the fact that the half-life of NCS is extremely short (3 seconds in serum). Because its cytotoxic effect seems to be very rapid, it appears more important to obtain a high initial NCS concentration than to maintain a constant blood level.

Topics: Animals; Antibiotics, Antineoplastic; Brain Neoplasms; Drug Screening Assays, Antitumor; Enzyme-Linked Immunosorbent Assay; Glioma; Humans; Rats; Tissue Distribution; Tumor Cells, Cultured; Zinostatin

The authors studied the localization of neocarzinostatin (NCS) in cultured cells and in tumor-bearing rats by means of immunofluorescent staining. Anti-NCS antibody was obtained through immunization of rabbits with NCS. Cellular uptake of NCS was dose-dependent (1.0 to 1000 micrograms/ml) in 9L rat gliosarcoma cells in monolayer. In monolayer cells of 9L, KMG-4 (derived from human glioblastoma), and KMS II (human ependymoma) treated with 1 mg/ml of NCS, drug uptake occurred within a few seconds. Accumulation was much higher in the cytoplasm than in the nucleus and, although nuclear uptake increased slightly over time, there appeared to be no increase in total cellular uptake. Mitotic cells, which were spherical in culture, showed greater intracellular accumulation than other cells. There was no significant difference in uptake among non-mitotic cells. Cells surviving 20 hours of treatment retained accumulation as high as that in killed cells. In KMG-4 monolayers, cytoplasmic and nuclear NCS distribution still differed, whereas 9L monolayers exhibited more even intracellular distribution. In 9L spheroid models treated with 1 mg/ml of NCS, the drug permeated almost all layers within 10 minutes, and at 120 minutes had heavily accumulated in the central necrotic areas. In rats with transplanted brain tumors, NCS selectively accumulated in neoplastic tissues following intra-arterial administration. However, NCS uptake by arterial endothelium was also seen, which suggests the potential for vascular toxicity. The therapeutic value of NCS is discussed in terms of its pharmacokinetic characteristics.

Topics: Animals; Antibiotics, Antineoplastic; Cell Division; Glioma; Humans; Immunohistochemistry; Neoplasm Transplantation; Rats; Tumor Cells, Cultured; Zinostatin

Meningeal gliomatosis (MG), pathologically, is caused by the diffuse dissemination or infiltration of glioma cells in the subarachnoid space, for which an effective, systematic treatment has not been contrived. Although, in the case of malignant leptomeningeal tumor, in general, intrathecal chemotherapy with such anticancer drugs as methotrexate and cytosine arabinoside (Ara-C) has been applied, the effect of this kind of treatment is limited, especially on MG. Therefore, the development of a new type of treatment is urgently needed. According to recent reports, neocarzinostatin (NCS) has been disclosed to have a strong cytocidal effect on glioma cells instead of injuring normal glia cells, and the intrathecal injection of NCS is suggested to be effective on MG. In order to evaluate the efficacy of intrathecal treatment with NCS on MG, a rat MG model using C 6 glioma cells has been produced and intrathecal chemotherapy with NCS was performed on this MG model. In MG rats which were treated intrathecally with NCS (1 microgram/kg) 1 day after tumor inoculation, the survival time was significantly prolonged by this treatment, where % ILS was 52.1%. Furthermore, it was more significantly prolonged with 10 micrograms/kg NCS, where 108.5% of % ILS was obtained. Contrary to these effects, this prolongation of the survival time of MG rats by the treatment with NCS showed a tendency to decrease in MG rats treated with NCS 3 days after tumor inoculation. No chemotherapeutic effect was observed in MG rats treated with even 100 micrograms/kg NCS 5 days after tumor inoculation. In conclusion, intrathecal chemotherapy with a low dose of NCS was proved to be effective in the early stages of MG.

Topics: Animals; Antibiotics, Antineoplastic; Disease Models, Animal; Glioma; Male; Meningeal Neoplasms; Neoplasm Transplantation; Rats; Rats, Inbred Strains; Zinostatin

Topics: Adult; Aged; Antibiotics, Antineoplastic; Astrocytoma; Brain Neoplasms; Carotid Arteries; Combined Modality Therapy; Female; Glioma; Humans; Injections, Intra-Arterial; Male; Middle Aged; Zinostatin

Topics: Adolescent; Adult; Antibiotics, Antineoplastic; Astrocytoma; Brain Neoplasms; Carotid Artery, Internal; Chemotherapy, Cancer, Regional Perfusion; Eye Diseases; Female; Glioma; Humans; Male; Middle Aged; Zinostatin

Topics: Adult; Aged; Antibiotics, Antineoplastic; Astrocytoma; Brain Neoplasms; Child; Female; Glioma; Humans; Male; Zinostatin

A pharmacokinetic one-compartment model for the cerebrospinal infusion for the brain tumor chemotherapy is described together with various parameters used for computer simulation. An antitumor protein, neocarzinostatin (NCS) as a prototype drug, has been utilized since it was found effective against glioblastoma cells at extremely low concentration (less than 5 ng/ml) and it is readily inactivated by serum. A very slow infusion velocity was found necessary for an appropriate dose regimen; for example, 0.25mg of the drug should be infused into CSF for about 40 min to attain a drug level of 8 ng/ml.

Topics: Antibiotics, Antineoplastic; Brain Neoplasms; Cell Line; Computers; Glioma; Humans; Kinetics; Models, Biological; Zinostatin

Neocarzinostatin as previously reported, appeared to exhibit an intense cytotoxicity to the glioblastoma cells and some other malignant brain tumor cells, such as pineal germinoma or medulloblastoma, which are notoriously known to disseminate into the cerebrospinal fluid space. In vitro study, the minimum susceptibility of glioblastoma cells to neocarzinostatin was found to be below 0.005 microgram/ml, whereas normal glia cells were not affected at 0.3 microgram/ml. This study indicated that neocarzinostatin was extremely effective in the treatment of malignant brain tumor without affecting normal neural tissue. Pharmacokinetic study was performed in order to establish intermittent intrathecal perfusion therapy and to prevent subarachnoid dissemination of the brain tumor cells. Experimental results were applied to the treatment of 12 patients with brain tumor, who had shown positive cytology of the cerebrospinal fluid. Follow-up investigation showed quite a favorable result and it was considered that prophylactic irradiation to the entire spinal column could be replaced with intrathecal administration of neocarzinostatin. During clinical application no noticeable side effect was encountered and active stimulation of macrophages, which were mobilized into the CSF space, was another unexpected advantage of this treatment.

Topics: Adolescent; Adult; Aged; Antibiotics, Antineoplastic; Astrocytoma; Brain Neoplasms; Cerebrospinal Fluid Shunts; Child; Child, Preschool; Female; Glioma; Humans; Injections, Intraventricular; Injections, Spinal; Kinetics; Male; Medulloblastoma; Middle Aged; Pinealoma; Zinostatin