Human glioma-associated antigens detected by monoclonal antibodies (original) (raw)
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Production and Characterization of Two Human Glioma Xenograft-localizing Monoclonal Antibodies
Cancer Research
Multiple fusions following immunization of athymic mice with the extensively characterized human glioma cell line D-54 MG resulted in the selection of several antibodies (Mabs) highly reactive with tumors of neuroectodermal origin and unreactive with normal nervous system tissue. Two Mabs, C12 and D12, which localized specifically to tumors in athymic mouse-human glioma xenograft paired label localization assays, are IgG3 antibodies; both bind readily to staphylococcal protein A in column purification and radioimmunoprecipitation procedures. Both iodinate via the chloramine-T method yielding I2*l-immunoreactive product by direct cell surface radioimmunoassay and absorption assay. By indirect cell surface radioimmunoassay, a cultured cell line panel consisting of 17 gliomas, 3 medulloblastomas, 2 neuroblastomas, 2 melanomas, and 2 fetal and 2 adult brain-derived cell lines was examined; the two Mabs were highly similar but distinct in their reactivity profiles. Each was positive with >47% of the gliomas tested (C12, 9 of 17; D12, 8 of 17); and with 1 of 3 medulloblastomas, 1 of 2 melanomas, and cell lines derived from 12-and 16-week-gestation human fetal brain. No reactivity was observed with neuroblastoma or adult brain-derived cell lines or with neutral glycolipids and gangliosides extracted from D-54 MG xenografts or human glioma cell lines. Notable extraneuroectodermal reactivity included that of Mab D12 for splenic trabeculae and the spermatids and Sertoli cells in the testes. Following immunoprecipitation of ('Hjleucine labeled cell membrane preparations, Mabs C12 and D12 have consis tently yielded unique bands in the M, 180,000 and M, 88,000 regions respectively. When used in paired label localization experiments in s.c. D-54 MG xenograft-bearing athymic mice, Mabs C12 and D12 demon strate similar localization patterns, attaining peak localization indices at day 3 (D12) or 4 (C12); the maximum percentage of injected Mab bound to tumor ranged from 57°(D12) to 8% (C12). The peak tumor/normal brain localization ratios (167-181) attained by these Mabs at days 1-2 followed by their rapid clearance suggest that these Mabs are potentially useful imaging and therapeutic agents for further investigation. . 3The abbreviations used are: Mab, monoclonal antibody; CS-R1A, cell surface radioimmunoassay; FCS. fetal calf serum; and ZO, zinc option medium supple mented with glutamine and 10-20% PCS as detailed in the text; ABC, avidinbiotin complex; LI, localization index. Immunization, Fusion, Cloning, and Immunoglobulin Preparation.
Journal of Neuroimmunology, 1986
Summaff 69−c15isahighlyimmunogeniccelllinederivedfromanaviansarcomavirus(ASV)−inducedastrocytomainF−344rats.Monoclonalantibody(Mab)productionwasattemptedbyfusingF−344ratsplenocytesandmouseP3×63/Ag8.653myelomacellsafterasyngeneicimmunizationprotocol.336fusioncloneswerescreenedbycellsurfaceradioimmunoassay(CS−RIA)againsttheimmunizingline69-c15 is a highly immunogenic cell line derived from an avian sarcoma virus (ASV)-induced astrocytoma in F-344 rats. Monoclonal antibody (Mab) production was attempted by fusing F-344 rat splenocytes and mouse P3 × 63/Ag8.653 myeloma cells after a syngeneic immunization protocol. 336 fusion clones were screened by cell surface radioimmunoassay (CS-RIA) against the immunizing line 69−c15isahighlyimmunogeniccelllinederivedfromanaviansarcomavirus(ASV)−inducedastrocytomainF−344rats.Monoclonalantibody(Mab)productionwasattemptedbyfusingF−344ratsplenocytesandmouseP3×63/Ag8.653myelomacellsafterasyngeneicimmunizationprotocol.336fusioncloneswerescreenedbycellsurfaceradioimmunoassay(CS−RIA)againsttheimmunizingline69-c15, rat kidney fibroblast line 203−cllandWalkerratcarcinomaline.Mabs7G4,9F1,10E3and10E7whichreactedonlywith203-cll and Walker rat carcinoma line. Mabs 7G4, 9F1, 10E3 and 10E7 which reacted only with 203−cllandWalkerratcarcinomaline.Mabs7G4,9F1,10E3and10E7whichreactedonlywith69-c15 were chosen. Further analysis demonstrated that these Mabs reacted only with rat (13/23 astrocytomas, 2/4 gliomas, 1/11 neurinomas) or mouse (2/10 astrocytomas) neurogenic tumor cells induced by both viral and chemical agents. Reciprocal competition assays suggested that 7G4, 9F1 and 10E3 recognized the same epitope and that 10E7 reacted with a spatially close determinant. Antigen activity could not be found in adult rat tissues (brain, heart, lung, liver, kidney, spleen, thymus, intestine, muscle and peripheral nerve) and fetal brain (8, 12, 20 days gestation) by either absorption analysis or tissue staining. Preliminary characterization indicated that the epitope may be polypeptide-associated. Further antigen purification and tumor localization can be attempted with these Mabs.
Immunobiology of primary intracranial tumors
Journal of Neurosurgery, 1983
o-Serial serological studies were carried out on 19 of 20 patients with malignant gliomas who were actively immunized with one of two human glioma tissue culture cell lines (D-54MG or U-251MG). Most patients mounted a significant serum reaction to histocompatibility antigens (HLA's), as well as an antibody response to fetal bovine serum (FBS) which was added to the glioma-cell inoculum. These two sources of antibody accounted for greater than 90% of the antibody induced by these inoculations. Two patients continued to have significant amounts of binding antibody to the original immunizing cell line following exhaustive absorptions of FBS and HLA antibodies. One of these two had all remaining significant antibody removed by further absorption of the serum against the 2-T osteogenic sarcoma tissue culture cell line known to possess antigens cross-reactive with human gliomas. One single patient continued to show significant antibody binding to the original glioma cell line following absorption against FBS, human platelets, and the 2-T cell line, and therefore seems to have produced glioma-distinctive antibodies in response to immunization. The antibody preparation from this patient was also cytotoxic against the original glioma cell line, as well as another recently cultured human glioblastoma cell line. The significance of these serological studies is discussed as it relates to immunological responses patients with gliomas may make to active immunization.
Journal of Neuroimmunology, 1995
A monoclonal antibody 6DS, against a human glioblastoma multiforme cell line U-87MG recognizes a tumor-specific, cell surface antigen of human glioblastoma cell lines. Partial cross-reactivity is observed with two human neuroblastoma cell lines, SK-N-SH and SK-N-MC, with little or no reactivity towards a rat glioma cell line C6 or normal human adult and fetal brain tissues. The antibody recognizes an antigen of molecular mass 38 kDa as inferred from Western blot analysis and sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the immunoprecipitate. The monoclonal antibody 6DS, inhibits both the attachment to substratum and growth of U-87MG cells. It strongly cross-reacts with xenotransplants of U-87MG cells and inhibits tumorigenesis (subcutaneous implants of U-87MG cells) in nude mice.
Immunotherapy of Malignant Gliomas Using Autologous and Allogeneic Tissue Cells
Anti-Cancer Agents in Medicinal Chemistry, 2010
Immunotherapy of brain tumors is rapidly emerging as a potential clinical option [1-3]. The quality and magnitude of immune responses evoked by the new generation anti-tumor vaccines is in general highly dependent on the source or choice of peptide antigens, and as well, a suitable immunopotentiator. Poorly immunogenic antigens, such as those present in tumor cell lysates, may not reliably provide stimulation like recombinant or DNA-encoded protein antigens might be expected to. In addition, the efficacy of the vaccine may depend on inherent counteracting measures of the tumor which dampen immune surveillance and immune effector activity triggered by immunization [4]. Our body has many means of limiting an immune response to our own (self) proteins. In particular, patients with gliomas exhibit a broad suppression of cell-mediated immunity [5-8]. Unfortunately, for most tumor vaccines the induction of local or systemic immune effector cells does not necessarily translate into objecti...
Anticancer research
To study the neuropathology and selected tumour markers of malignant gliomas, an animal glioma model was developed using the implantation of human glioblastoma clone U87 into rat brains. The U87 cell suspension, or precultured U87 tumour spheroids, were inoculated into the brain of 4-week-old cyclosporin A immunosuppressed Wistar rats. The resulting first generation tumours were then transferred, through serial transplantations to rats, to obtain second and third generation tumours. Brain tumour sections were examined for various known tumour markers by routine HE staining and immunohistochemical analyses. The immunohistochemical analyses showed that p53, S100 protein, glial fibrillary acidic protein (GFAP) and synaptophysin expressions, initially present in the tissue culture, were gradually lost in later tumour generations, whereas nestin and musashi expressions increased, possibly indicating progressive tumour cell dedifferentation. Persistent kallikrein, CD68 and vimentin expres...
Induction of human glioma tumor in sprague-dawley rat with intact immune system
Turkish Neurosurgery, 2016
it provides the possibility to investigate the histological and genetic characteristics of neoplasms (11). Commonly applied models that generate brain neoplasms in animals include: a) Chemical mutagen-induced models (36) b) Genetic modification-induced models (2) c) Xenograft induced models (20, 30). These models have led to better understanding of the mechanisms related to tumor progression. However, these models are not comprehensive, and each of the models has its own issues (18, 31). The major drawbacks of chemically induced brain tumors include their histological characteristics █ INTRODUCTION G lioblastoma (GBM) is a common and aggressive type of brain tumor in humans. Despite improvement in therapy techniques, patients have a short life expectancy (12-15 months) and GBM is an incurable disease (7, 32). The inefficacy of current treatment methods stimulates the researchers to seek novel therapeutic agents and strategies. So, development of laboratory and animal model is a substantial step for the advances in the treatment of GBM (19, 23). An animal model is an important tool for the understanding of complex phenomena involved in glioma generation; and AIm: Glioblastoma (GBM) is an aggressive brain tumor in humans. The median survival rate of patients is one year after the diagnosis. So, development of an animal model is necessary for the advances in the research treatment of GBM. The aim of this study was to investigate the capability of human glioma cells in inducing glioma tumors in rats with intact immune system. mATERIAl and mEThODS: U87 cells were implanted in the frontal lobe of rats without suppressing the immune system. We used magnetic resonance imaging (MRI), Hematoxylin-Eosin (H&E) and Immunohistochemical (IHC) staining to assess characteristics of tumor. RESUlTS: At the 10 th and 14 th days of tumor inoculation, MRI images contained the tumor areas in the brain. All tumor-bearing rats developed tumors. The rats retained the morphology and histological characteristics of human glioma. Animals mimic GBM characteristics, such as mitotic activity, invasion, neovascularization, necrosis and pseudopalisading cells. IHC staining revealed tumor growth and progression in the tumor-bearing rats. CONClUSION: This model is a standard system for studying the tumor phenotype, genotype, and for evaluating the efficacy of anti-cancer agents. It is a reliable, simple, inexpensive, and easily reproducible model, which may be a way for pre-clinical studies.
Characterization of an established human malignant glioma cell line: LN-18
Acta Neuropathologica, 1981
A human malignant glioma cell line, has been established in monolayer culture and subcultured for more than 115 passages. LN-18 cells grow in vitro as bipolar or stellate cells with pleomorphic nuclei, have a doubling time of about 72 h and a plating efficiency of 3 %. The glial nature of these cells has been assessed by ultrastructural examination. The synthesis of glial fibrillary acidic and S-100 proteins could not be demonstrated, although the initial biopsy tissue and the early cultures were positive for the former. The presence of [a-like antigens on the surface of these cells was demonstrated using allo and xeno antisera. LN-18 cells were also shown to synthesize large quantities of fibronectin. The injection of LN-18 cells into nude mice induced the formation of solid tumor masses that could be retransplanted every 3 weeks and showed a morphology comparable to that of the initial biopsy. Karyotype analysis revealed the presence of three marker chromosomes, constantly present before and after hetero-transplantation.