Differential spectrum of expression of neural cell adhesion molecule isoforms and L1 adhesion molecules on human neuroectodermal tumors.

A series of four medulloblastomas, seven neuroblastomas (Nb), two ependymomas, and three gliomas, human neuroectodermal tumors, were screened for their expression of adhesion molecules L1, carcinoembryonic antigen, neural cell adhesion molecule isoforms (N-CAM) and HNK1 epitope by Western blotting and double immunofluorescence labeling. All seven neuroblastomas, whatever their differentiated state, expressed L1, a neural cell surface developmental antigen, whereas all other tumors tested were negative. All tumors expressed N-CAM; however, a large diversity was observed among the isoforms. Low sialylated N-CAM 140 was present, with different intensity, in ependymomas and astrocytomas. High sialylated isoforms were detected by a monoclonal antibody (anti-MenB) specifically recognizing high polymers of alpha 2-8 linked neuraminic acid. They were expressed in all medulloblastomas studied (4 of 4), and in 4 of 7 Nbs examined. Negative cases corresponded to tumors having undergone chemotherapeutic treatment or to ganglioneuroma. The interconversion from high to low sialylated forms might reflect changes which are critical for the conversion of Nbs into benign ganglioneuromas. HNK1 epitope was expressed on a large diversity of molecules by nearly all tumors except two Nbs which were also negative with anti-MenB antibody. This simultaneous loss of carbohydrate epitopes could correlate with higher maturation states of the tumors. None of the tumors expressed carcinoembryonic antigen. Therefore, anti-L1 and anti-MenB antibodies define differentiation-related antigens that could differentiate between Nbs and other tumors and may prove helpful in diagnosis and understanding of the biological nature of neuroectodermal tumors. An immunodot assay was set up and allowed to titrate the presence of polysialic acid units in cerebrospinal fluid from patients presenting meningeal spread of medulloblastomas. It could help to assess metastasis and to monitor the effects of chemotherapeutic treatment on polysialylated N-CAM positive tumors.

Localization and characterization of endothelin receptors in human gliomas: a growth factor?

Localization and characterization of endothelin receptors in surgical specimens of human gliomas (6 benign astrocytomas and 7 glioblastomas multiforme) and in normal human cortices were studied using quantitative receptor autoradiographic methods. Low numbers of [125I]endothelin-1 [( 125I]ET-1) binding sites were detected in the gray matter of the human frontal cortex, with little binding in the white matter. Conversely, relatively high numbers of [125I]ET-1 binding sites were homogeneously present in tissue sections derived from astrocytomas, whereas higher numbers of [125I]ET-1 binding sites were heterogeneously located on groups of cells with a pseudopalisading appearance and pleomorphic astrocytes in glioblastoma multiforme. Necrotic areas within the tissue sections derived from glioblastoma were devoid of binding. Binding of [125I]ET-1 to gliomas and normal gray matter was specific. Unlabeled ET-1 and its natural analogs (ET-2 and ET-3) inhibited the binding of [125I]ET-1 to these lesions in a concentration-dependent manner and with similar high potencies. Possibly related substances, such as ion channel regulators (omega-conotoxin, apamin, and tetrodotoxin), a Ca2+ channel blocker (nicardipine), and growth factors (epidermal growth factor and insulin-like growth factor I), did not affect the binding to tissue sections derived from gliomas or from normal frontal cortices. Scatchard analysis revealed the presence of a single class and high-affinity binding sites for endothelin in normal cortex and in gliomas. There was no significant difference in the binding affinities: dissociation constants (Kd) were 2.1 +/- 0.5 nM in 6 astrocytomas, 2.5 +/- 0.4 nM in 7 glioblastomas, and 1.4 and 1.5 nM in two normal cortices.(ABSTRACT TRUNCATED AT 250 WORDS)

Multiparameter flow cytometric analysis of neoplasms of the central nervous system: correlation of nuclear antigen p105 and DNA content with clinical behavior.

Analysis of the DNA content of various solid tumors and hematological malignancies may provide useful prognostic information. To date, however, there has been a striking lack of correlation between DNA content in neoplasms of the central nervous system and clinical behavior. Simultaneous quantitation of DNA content and proliferation-associated nuclear antigen (p105) by flow cytometry was performed on paraffin-embedded tissues representing three major groups of central nervous system neoplasms--1) 21 astrocytic tumors, 2) 13 pituitary tumors, and 3) 19 meningiomas--and the results were correlated with clinical behavior. All 4 well-differentiated gliomas were diploid, while 3 of 9 anaplastic astrocytomas and 1 of 8 glioblastomas had a demonstrable aneuploid peak. Three of 13 pituitary tumors had an identifiable aneuploid peak, while only 2 of 19 meningiomas had an aneuploid DNA content. Cell-cycle analysis of the malignant gliomas revealed a significantly higher proliferative index (PI, %S + G2M) compared with the well-differentiated astrocytomas (P less than 0.05). Within the subgroup of diploid anaplastic astrocytomas, however, extended patient survival appeared to be associated with a higher PI. For diploid pituitary adenomas, the PI was consistently lower in the 3 tumors that recurred than it was in the remaining 8 adenomas. Nuclear antigen quantitation of diploid tumors showed a wide range of p105 expression in G0G1 cells, suggesting that, within each tumor, the cells are heterogeneous with respect to proliferative activity. Aneuploid nuclei of glial tumors showed enhanced expression of p105 relative to diploid cells of the same specimen. In pituitary tumors, the median G2M/G0G1 fluorescence ratio for p105 was significantly higher (P less than 0.05) for the 3 diploid recurrent tumors than for those that did not recur. These data support the assumption that the aggressive clinical course of malignant glial neoplasms may be related to an abnormal DNA stemline and/or an alteration in cell proliferative activity. Cell cycle analysis and measurement of p105 by this technique may provide information useful from both a prognostic standpoint and in directing adjuvant therapy.

Silver colloid staining technique for analysis of glioma malignancy.

A silver colloid staining technique for identifying nucleolar organizer region-associated proteins (Ag-NOR's) was applied to 51 human gliomas. These comprised 20 glioblastomas multiforme, 15 anaplastic astrocytomas, and 16 astrocytomas, in which the mean numbers of Ag-NOR's per cell (+/- standard error of the mean) were 2.51 +/- 0.12, 2.01 +/- 0.10, and 1.76 +/- 0.06, respectively. Significant differences among these were recognized, and the mean number of Ag-NOR's paralleled the degree of histopathological malignancy. In 16 cases, studies were performed of the number of Ag-NOR's and the S-phase fraction by in vitro labeling using antibromodeoxyuridine monoclonal antibody. A linear relationship was demonstrated between these two factors (r = 0.857, p less than 0.001), although some scatter was seen. In 32 adult patients, the correlation between the number of Ag-NOR's and the prognosis was estimated. The results demonstrated that the group containing patients with less than 1.80 Ag-NOR's per cell had a better prognosis than the group with 1.80 Ag-NOR's or more. Thus, the number of Ag-NOR's reflected the degree of histopathological malignancy, S-phase fraction, and prognosis. Silver colloid staining for Ag-NOR's is a simple, rapid, and reproducible method for estimating the proliferative potential of human gliomas without requiring a complicated technique.

Products of cells from gliomas: VIII. Multiple-well immunoperoxidase assay of immunoreactivity of primary hybridoma supernatants with human glioma and brain tissue and cultured glioma cells.

To test the feasibility of primary screening of hybridoma supernatants against human glioma tissue, over 5000 combinations of hybridoma supernatants with glioma tissue, cultured glioma cells, and normal central neural tissue were screened with a new multiple-well (M-well) screening system. This is an immunoperoxidase assay system with visual endpoints for screening 20-30 hybridoma supernatants per single microscope slide. There were extensive differences between specificities to tissue and to cultured glioma cells when both were screened with M-wells and when cultured cells were screened with standard semi-automated fluorescence. Primary M-well screening with glioma tissue detected seven hybridoma supernatants that specifically identified parenchymal cells of glioma tissue and that were not detected with cultured cells. Immunoreactivities of individual supernatants for vascular components (nine supernatants), necrosis (five supernatants), and nuclei (three supernatants) were detected. Other supernatants bound multiple sites on glioma tissue and/or subpopulations of neurons and glia of normal tissue. The results show that primary screening with glioma tissue detects a number of different specificities of hybridoma supernatants to gliomas not detected by conventional screening with cultured cells. These are potentially applicable to diagnosis and therapy.

Investigation of the expression of epidermal growth factor receptor and blood group A antigen in 110 human gliomas.

The presence of epidermal growth factor receptor (EGF-R) and blood group A antigen was studied immunohistochemically in a series of 110 malignant gliomas using monoclonal antibodies. Fifty-seven percent of the tumours strongly expressed EGF-R on the malignant cells. Although blood group A antigen is present on EGF-R of A431 cells (a cell line derived from a human epidermoid carcinoma), in gliomas it was found only on vascular endothelial cells of tumours from blood group A patients. The results suggest that the EGF-R present in gliomas differs from that in A431 cells in the type or amount of the carbohydrate chains. This is in contrast to previous reports which have suggested that A antigen is present on EGF-R in gliomas. This has relevance in the choice of monoclonal antibodies used to study the EGF-R, as those directed against the A antigen component of the A431 cell EGF-R will not recognize EGF-R elsewhere and may cause normal blood group A antigen to be mistaken for EGF-R.

[DAPI-DNA cytofluorometric study of glioma cells--application of DAPI-DNA cytofluorometry to paraffin embedded archival glioma tissue for nuclear DNA content analysis].

Using DAPI-DNA cytofluorometry, the author analyzed nuclear DNA content of formalin fixed, paraffin embedded, glioma material obtained from 14 glioma cases at surgery. Sections of 10 microns were deparaffinized. Following simultaneous DAPI (4,6-diamidino-2-phenylindole dihydroporphyrin chloride)/HP (hematoporphyrin) staining, DAPI binds DNA and DNA-DAPI complexes emit blue fluorescence when exited by ultraviolet (UV) light. Through Zeiss fluorescence microscope, the author measured nuclear fluorescence intensity with histological verification of glioma cells. A DNA histogram was obtained with fluorescence intensity recorded on the abscissa and number of cells plotted on the ordinate. Samples of 20 normal non-neoplastic astrocytes taken from apparently normal brain tissue included in the histological slide were used as diploid (2 C) control. Based on DNA content, tumor cells were classified into 4 groups: N-group composed of cells with 2 C DNA content (normoploid), S-group with less than 2 C (hypoploid), L-group more than 4 C (hypertetraploid), I-group between 2 C and 4 C (intermediate ploidy). Intermediate ploidy was significantly higher and normoploid was significantly lower in glioblastoma compared with those of benign astrocytoma. Thus, DNA content and histological malignancy were well correlated. Due to limitation of measuring diaphragm of turret in the microscope, some extra large cell could not be included in it and was excluded from the measurement.(ABSTRACT TRUNCATED AT 250 WORDS)

Attenuation of opioid receptor activity by phorbol esters in neuroblastoma x glioma NG108-15 hybrid cells.

Chronic opioid treatment of neuroblastoma x glioma NG108-15 cells induces desensitization of the opioid receptor and this may involve a change in membrane protein phosphorylation. In an attempt to mimic this possible mechanism, we studied effects of phorbol ester activation of protein kinase C on opioid receptor activity. Incubation of NG108-15 hybrid cells with the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) abolished up to 45% of opioid inhibition of cyclic AMP accumulation in intact cells, while basal accumulation and prostaglandin E1-stimulated cyclic AMP accumulation were unaltered. This decrease of opioid inhibition was dose- and time-dependent and the potency order of phorbol esters and apparent K activation (90 nM) for TPA were consistent with phorbol esters acting through the stimulation of protein kinase C. TPA also decreased the inhibition of cyclic AMP accumulation mediated through muscarinic and alpha-2 adrenergic receptors. These effects of TPA were best explained by a TPA-induced alteration of the inhibitory nucleotide-binding protein (Gi), the common transducer protein of these receptors. Impairment of Gi by TPA treatment was evidenced by a reduction in agonist-stimulated GTP hydrolysis and activation by GTP. Quantification of Gi by pertussis toxin-catalyzed ADP-ribosylation revealed that TPA decreased maximal labeling. In summary, phorbol esters appeared to attenuate opioid receptor activity by altering the activity of the transducer protein Gi.

Suramin inhibits proliferation of rat glioma cells and alters N-CAM cell surface expression.

Suramin, a drug used in the treatment of trypanosomiasis and onchocerciasis inhibits growth-factor-induced mitogenesis. We have investigated the effect of suramin on the growth rate and the morphology of C6 glioma cells cultured in the presence of serum or in a serum-free defined medium. Exponentially growing cells were seeded in multi-dish plates (5 x 10(4) cells/2 cm2 well) in DMEM supplemented with 5% fetal calf serum and were continuously exposed to 1 microgram/ml to 1,000 micrograms/ml suramin. Growth rate (determined 9 days after seeding) was reduced by 5%, 33%, 56% and 97%, respectively for suramin concentrations of 1, 10, 100 and 1000 micrograms/ml. Similar results were obtained in serum-free defined medium (DMEM/F12, 1:1, v:v, EGF 5 ng/ml, transferrin 5 micrograms/ml, selenium 10 ng/ml). Moreover, the concentration of suramin in the culture medium remained constant, demonstrating that the drug was not actively metabolized by the cells. Suramin also induced marked changes in cell morphology: the usual bipolar shape of C6 cells evolved toward a more differentiated appearance, with numerous cellular processes allowing a wide number of cell-cell contacts. In parallel, we monitored expression of an adhesion molecule (N-CAM) at both the mRNA and protein levels. Indirect immunofluoresence technique showed an important increase in cell surface N-CAM expression, starting from a dose of 10 micrograms/ml suramin, whereas total cellular content of N-CAM protein as well as its mRNA levels were unaffected. We also observed that the levels of expression of actin and N-CAM mRNAs decreased by a factor of two in cells maintained in defined medium. However, the relative ratio of N-CAM mRNA over actin mRNA was virtually unchanged following suramin treatment. Taken together, our results suggest that suramin (i) exerts a blocking effect of autocrine growth factors, (ii) interferes with the turn-over mechanisms of N-CAM expressed at the cell surface, either by impairing its endocytosis and/or the process of release of the N-CAM 120 isoform.

Cautionary note on the use of the B subunit of cholera toxin as a ganglioside GM1 probe: detection of cholera toxin A subunit in B subunit preparations by a sensitive adenylate cyclase assay.

The use of the B subunit of cholera toxin, a protein that binds specifically to ganglioside GM1, has provided a new paradigm for studying physiological functions of ganglioside GM1. The B subunit inhibited the growth of rat glioma C6 cells that had been pretreated with ganglioside GM1. In some preparations of the B subunit, the inhibition was independent of adenylate cyclase activation and was due to the binding of the B subunit to ganglioside GM1 inserted onto the cell surface. However, in other preparations of the B subunit, there was an additional inhibitory effect due to small contaminations with the A subunit, which caused increases in intracellular cyclic adenosine monophosphate (cAMP) levels and concomitant growth inhibition. This vanishingly small contamination with the A subunit could not be detected by conventional protein sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis but could be measured utilizing a sensitive adenylate cyclase activation assay. Thus caution must be used to ensure that any biological effects of the B subunit are not due to contaminating A subunit and are due solely to the binding of the B subunit to ganglioside GM1 exposed on the cell surface. This is especially important in cyclic nucleotide-sensitive systems.

Effect of dexamethasone on neurotransmitter amines in a rat glioma model.

Vasogenic brain edema was produced by transplantation of rat glioma C6 cells into rat brain. Using this model, we measured neurotransmitter concentrations and water content of regional brain tissue. In the tumor-implanted controls, monoamine neurotransmitters in the hypothalamus, cortex, and striatum significantly decreased. When treated with DEX, these monoamines tended to return to the levels of the sham-operated controls. Additionally, DEX markedly lowered tumoral monoamines. In the nonsurgical animals, DEX significantly increased norepinephrine but not DA. Regardless of treating with DEX or not, water content showed no changes in the hypothalamus and striatum in the tumor-implanted animals. However, the increase in water content in the cortex was significant, and this increase could be reduced by DEX to the levels of controls. Water content of tumor tissue could also be markedly reduced by DEX. In the nonsurgical animals, there were no changes in water content between DEX-treated and nontreated animals. In conclusion, brain edema produced by the brain tumor may reduce noradrenergic and dopaminergic activities. This is more likely due to compression anoxia caused by the tumor mass, glial hydrops, and edema fluid. It is presumed that the effect of DEX is due to reduction of water content of the tumor and peritumoral white matter as well as by increasing noradrenergic activity.

Sigma and opioid receptors in human brain tumors.

Human brain tumors (obtained as surgical specimens) and nude mouse-borne human neuroblastomas and gliomas were analyzed for sigma and opioid receptor content. Sigma binding was assessed using [3H]1,3-di-o-tolylguanidine (DTG), whereas opoid receptor subtypes were measured with tritiated forms of the following: mu, [D-ala2,mePhe4,gly-ol5]enkephalin (DAMGE); kappa, ethylketocyclazocine (EKC) or U69,593; delta, [D-pen2,D-pen5]enkephalin (DPDPE) or [D-ala2,D-leu5]enkephalin (DADLE) with mu suppressor present. Binding parameters were estimated by homologous displacement assays followed by analysis using the LIGAND program. Sigma binding was detected in 15 of 16 tumors examined with very high levels (pmol/mg protein) found in a brain metastasis from an adenocarcinoma of lung and a human neuroblastoma (SK-N-MC) passaged in nude mice. kappa opioid receptor binding was detected in 4 of 4 glioblastoma multiforme specimens and 2 of 2 human astrocytoma cell lines tested but not in the other brain tumors analyzed.

Isolation and partial purification of growth factors with TGF-like activity from human malignant gliomas.

The effect of concentrated conditioned medium from each of eight human malignant glioma cell lines on the growth of indicator cells (normal rat kidney fibroblasts (NRK), clone 14) was determined in monolayer and in soft agar assay systems. The conditioned medium from all cell lines was mitogenic in the monolayer assay, but only SF-210, U-343 MG-A, and U-251 MG produced soluble factors that caused NRK cells to grow in soft agar. The soluble growth-promoting factors from these three cell lines were acid- and heat-stable (60 degrees C for 30 minutes) but were inactivated by trypsin (100 microns/ml) and dithiothreitol (50 microM). The growth factors from SF-210 and U-343 MG-A were further purified by molecular-sieve chromatography. The partially purified growth factor from U-343 MG-A retained transforming growth factor (TGF)-like activity, had a molecular weight of 9 kD, was potentiated by TGF-beta in the soft agar assay, competed effectively with 125I-epidermal growth factor (EGF) radiolabeled for the EGF receptor on A 431 epidermoid carcinoma cells, and was completely inhibited by monoclonal antibodies to TGF-alpha. The partially purified growth factor from SF-210 had a molecular weight of 17 kD, was not inhibited by monoclonal antibodies to platelet-derived growth factor (PDGF) or TGF-alpha, and did not bind to a heparin-Sepharose column. These results imply that U-343 MG-A secretes a growth factor with TGF-alpha-like activity, and SF-210 secretes a TGF with neither TGF-alpha nor TGF-beta activity.

Immunosuppression and transforming growth factor-beta in glioblastoma. Preferential production of transforming growth factor-beta 2.

Transforming growth factor (TGF)-beta 1 is a polypeptide that is assumed to play a fundamental role in the growth of both normal and neoplastic cells. TGF-beta 2 is a closely related polypeptide, originally described as glioblastoma cell-derived T cell suppressor factor (G-TsF) due to its immunosuppressive activity. Expression of the genes for TGF-beta 1 and G-TsF/TGF-beta 2 was examined in tumor cells and was found to be different in several cell lines and tissues that were tested. Whereas two glioblastoma cell lines expressed both TGF-beta 1 and G-TsF/TGF-beta 2 mRNA, one melanoma and neuroblastoma cell lines showed only TGF-beta 1 mRNA which in the case of the neuroblastoma required cycloheximide treatment for its detection. The coordinate expression of the genes for TGF-beta 1 and G-TsF/TGF-beta 2 in glioblastoma was not paralleled by secretion of both polypeptides as only G-TsF/TGF-beta 2 but not TGF-beta 1 was identified in supernatants of glioblastoma cells. These data provide evidence for a post-transcriptional level of regulation for production of the two forms of TGF-beta. As mRNA for G-TsF/TGF-beta 2 was also identified in fresh surgically removed human glioblastoma tissue, G-TsF/TGF-beta 2 may also be secreted within the tumor in vivo. Unlike glioblastoma, human fetal brain tissues or adult brain specimens studied did not express detectable levels of TGF-beta mRNA. Impaired cell-mediated immunity is an established finding in patients with glioblastoma. Secretion of G-TsF/TGF-beta 2 by tumor cells in vivo may contribute to decreased immune surveillance for tumor development, as well as neovascularization of the tumor tissue.

Identification of functional beta-adrenergic receptors on AC glioma cells.

AC glioma cells, a clonal cell line derived from a rat glioma, responded to 1 mM dibutyryl-cyclic AMP and isobutylmethylxanthine with a change to stellate morphology. A concentration-related morphological change was induced by beta 1- and beta 2-adrenergic agonists with the order of potency being isoproterenol greater than soterenol greater than norepinephrine. Propranolol (nonselective, beta-antagonist), butoxamine (beta 2-antagonist) and metoprolol (beta 1-antagonist) significantly decreased the cell response to isoproternol. Schild analysis of the response, using the competitive antagonist metoprolol, gave pA2 values of 7.5 and 8.5 for the agonists norepinephrine and soterenol, respectively, with slopes of the curves being less than unity. These observations indicate that both beta 1- and beta 2-adrenergic receptors mediate the change in cellular morphology.

Immunolocalization of extracellular matrix proteins during brain tumor invasion in BD IX rats.

The distribution of several native extracellular matrix proteins (type I, III, and IV collagens and fibronectin) using immunofluorescent localization is described for in two different malignant gliomas (BT4A and BT4An). In addition, antibodies against denatured forms of type I and III collagens were used to localize areas of active degradation within the tumors. We have shown that both tumors express the native connective tissue components studied, although the distribution of these components within and between the tumors was different. In addition, native type I and III collagens and fibronectin were overexpressed in the tumors compared to the normal brain. Morphometry on immunostained type IV collagen sections showed an increase in vascular elements in both tumors compared to normal brain tissue. The BT4A tumor, which by light microscopy showed a degradative mode of invasion, expressed denatured type I and III collagens at the tumor-brain border zone, suggesting that this tumor has collagenolytic activity. The present article suggests that the distribution and changes in extracellular matrix protein synthesis and degradation may play an important role in the progressive growth of brain tumors in vivo.

Immunocytochemical characterization of extracellular matrix proteins expressed by cultured glioma cells.

The immunolocalization of type I, III and IV collagens and fibronectin in two rat glioma cell lines in vitro (BT4C and BT4Cn) is described. In addition, antibodies against denatured type I and III collagens were used to study breakdown products of native type I and III collagens. For the BT4C cells, the extracellular matrix expression in monolayer cultures and in multicellular tumor spheroids was compared. Type IV collagen was strongly expressed in BT4C tumor spheroids but was negative in the corresponding monolayer cultures. Denatured type I collagen was found both in monolayers and in spheroids of BT4C, suggesting either a rapid turnover (i.e., synthesis and immediate breakdown) of type I collagen or an altered collagen gene transcription. Both cell lines were negative for native type I and III and denatured type III collagen. Fibronectin was strongly expressed in both cell lines. Supporting the immunofluorescence data, the hydroxyproline content in the tumor spheroids was twice the amount found in monolayer cultures. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis combined with immunoblotting also verified the immunostaining experiments, showing that glioma spheroids and injected tumor cells have the potential for fibronectin and collagen production, given the appropriate growth conditions.

Labeling monoclonal antibodies and F(ab')2 fragments with the alpha-particle-emitting nuclide astatine-211: preservation of immunoreactivity and in vivo localizing capacity.

alpha-Particles such as those emitted by 211At may be advantageous for radioimmunotherapy since they are radiation of high linear energy transfer, depositing high energy over a short distance. Here we describe a strategy for labeling monoclonal antibodies and F(ab')2 fragments with 211At by means of the bifunctional reagent N-succinimidyl 3-(trimethylstannyl)benzoate. An intact antibody, 81C6, and the F(ab')2 fragment of Me1-14 (both reactive with human gliomas) were labeled with 211At in high yield and with a specific activity of up to 4 mCi/mg in a time frame compatible with the 7.2-hr half-life of 211At. Quantitative in vivo binding assays demonstrated that radioastatination was accomplished with maintenance of high specific binding and affinity. Comparison of the biodistribution of 211At-labeled Me1-14 F(ab')2 to that of a nonspecific antibody fragment labeled with 211At and 131I in athymic mice bearing D-54 MG human glioma xenografts demonstrated selective and specific targeting of 211At-labeled antibody in this human tumor model.

Growth factors derived from a human malignant glioma cell line, U-251MG.

A human malignant glioma cell line, U-251 Mg, cultured under serum free conditions, was shown to produce a growth factor for BALB/c 3T3 cells (glioma-derived growth factor-1, GDGF-1). The biological activity of GDGF-1 resided in a heat- and acid-resistant protein with a molecular weight (MW) of 25 kDa estimated by gel permeation chromatography. GDGF-1 activity was neutralized by a goat anti-human platelet derived growth factor (PDGF) antibody, indicating that the two factors were immunologically related. Furthermore, U-251 Mg cells constitutively expressed c-sis mRNA. When U-251 Mg cells were stimulated with bacterial lipopolysaccharide, 2 novel growth factors (GDGF-2 and GDGF-3) were produced in addition to the PDGF-like substance. GDGF-2 was determined to be greater than 100 kDa MW and was not neutralized by the goat anti-PDGF antiserum. The biological activity of GDGF-3 was also heat- and acid-resistant with an apparent 14 kDa MW. This factor also did not show any common antigenicity with PDGF. GDGF-2 and GDGF-3 are currently under investigation and evidence as to their natures will be published elsewhere. Our findings with this glioma cell line provide further evidence that inappropriate expression of growth factor-related genes could play important autocrine role(s) in the processes leading to malignant transformation and/or uncontrolled proliferation and may provide a paracrine stimulus for such processes as glioma neovascularization.