Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0017638 (glioma)
 30,880 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The cellular uptake of antisense oligodeoxynucleotides (ODNs) may be enhanced by the use of carriers such as cationic liposomes or lipoplexes, but little is known about the intracellular fate and subcellular trafficking of these systems in target cells. In this study, we report on the cellular uptake and biodistribution of ODNs in the presence and absence of optimised self-assembled cationic lipoplexes using the C6 glioma cell line as an in vitro model. Biotin or radiolabelled 15-mer phosphorothioate (PS) ODNs were synthesised and their cellular uptake and subcellular biodistribution characterised in the presence and absence of an optimised cationic lipoplex delivery system using studies ranging from cellular association, cellular efflux and transmission electron microscopy (TEM). Ultrastructural studies clearly showed PS ODNs in the absence of liposomal delivery to be sequestered within endosomal and lysosomal vesicular bodies indicative of endocytic uptake. ODNs were also visible, to a lesser extent, in the nucleus and cytoplasm. By employing DOSPA (2'-(1",2"-dioleoyloxypropyldimethyl-ammonium bromide)-N-ethyl-6-amidospermine tetra trifluoroacetic acid) and DOPE (dioleoylphosphatidylethanolamine) complex in a 3 : 1 ratio, as a delivery system for ODNs at a optimal lipid/DNA charge ratio of 1 : 1, the level of ODN cellular association was significantly increased by approximately 10-12 fold with a concomitant change in subcellular distribution of PS ODN. TEM studies indicated enhanced penetration of ODN within the cytosol and the cell nucleus with reduced presence in vesicular compartments. Efflux studies confirmed that cationic lipoplexes promoted entry of ODNs into 'deeper' cellular compartments, consistent with endosomal release. Optimised cationic lipoplexes improved cellular delivery of ODNs by enhancing cell association, uptake and by favourably modulating the intracellular trafficking and distribution of ODNs into non-vesicular compartments including the cytosol and nucleus.
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PMID:Studies on uptake, sub-cellular trafficking and efflux of antisense oligodeoxynucleotides in glioma cells using self-assembling cationic lipoplexes as delivery systems. 1072 99

Growth factors play an important role in proliferation and differentiation of malignant brain gliomas in humans. Glial cell line-derived neurotrophic factor (GDNF) has been shown recently to be highly expressed in human glioblastomas and in rat glial cell lines B49 and C6. The aim of the present study was to knockdown GDNF, its receptor GFR-alpha1, and the related family member persephin by using antisense oligonucleotides and to observe the effects on cell proliferation. To enhance cellular uptake into C6 glioma cells, 15-mer phosphorothioate oligonucleotides were complexed with the cationic lipid Lipofectamine. The complex was applied for 3 x 12 hours to C6 glioma cells, and cells were allowed to recover for 24 hours after each transfection and then analyzed. This protocol markedly reduced GDNF and GFR-alpha1 protein levels in C6 glioma cells compared with control oligonucleotides. Knockdown of C6 cells with GDNF and GFR-alpha1 but not with persephin antisense oligonucleotides significantly decreased the number of C6 glioma cells and also inhibited the incorporation of bromodeoxyuridine as a sign of reduced DNA synthesis. In conclusion, it is shown that GDNF but not persephin is a potent proliferation factor for rat glioma cells. Knockdown of GDNF using antisense oligonucleotides complexed with lipids as carriers may be useful in gene therapeutic approaches in vitro and possibly also in vivo.
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PMID:Glial cell line-derived neurotrophic factor (GDNF) is a proliferation factor for rat C6 glioma cells: evidence from antisense experiments. 1107 71

Antisense radiopharmaceuticals could be used to image gene expression in the brain in vivo, should these polar molecules be made transportable through the blood-brain barrier. The present studies describe an antisense imaging agent comprised of an iodinated peptide nucleic acid (PNA) conjugated to a monoclonal antibody to the rat transferrin receptor by using avidin-biotin technology. The PNA was a 16-mer antisense to the sequence around the methionine initiation codon of the luciferase mRNA. C6 rat glioma cells were permanently transfected with a luciferase expression plasmid, and C6 experimental brain tumors were developed in adult rats. The expression of the luciferase transgene in the tumors in vivo was confirmed by measurement of luciferase enzyme activity in the tumor extract. The [(125)I]PNA conjugate was injected intravenously in anesthetized animals with brain tumors and killed 2 h later for frozen sectioning of brain and film autoradiography. No image of the luciferase gene expression was obtained after the administration of either the unconjugated antiluciferase PNA or a PNA conjugate that was antisense to the mRNA of a viral transcript. In contrast, tumors were imaged in all rats administered the [(125)I]PNA that was antisense to the luciferase sequence and was conjugated to the targeting antibody. In conclusion, these studies demonstrate gene expression in the brain in vivo can be imaged with antisense radiopharmaceuticals that are conjugated to a brain drug-targeting system.
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PMID:Antisense imaging of gene expression in the brain in vivo. 1110 72

Malignant gliomas of astrocytic origin have commonly expressed several features such as alterations in the tumor-suppressor gene p53 or p16 or the acquisition of telomerase activity, which are distinctive from astrocytes. Therefore, restoration of the tumor-suppressor gene or telomerase inhibition is expected to provide a cure for malignant gliomas. We have recently demonstrated that the treatment with a 19-mer antisense oligonucleotide against human telomerase RNA linked to a 2',5'-oligoadenylate (2-5A-anti-hTR) inhibited the growth of malignant glioma cells. From a therapeutic point of view, it is very important to investigate the antitumor efficacy of 2-5A-anti-hTR combined with the restoration of p53 or p16 gene. In this study, we evaluated the antitumor effect of 2-5A-anti-hTR in combination with recombinant adenoviruses bearing p53, its associated p21WAF1/CIP1, or p16CDKN2 gene (Ad5CMV-p53, Ad5CMV-p21, or Ad5CMV-p16) against malignant glioma cells in vitro and in vivo. Five malignant glioma cell lines expressing the mutant p53 gene (A172, GB-1, T98G, U251-MG and U373-MG) were more sensitive to the combination of 2-5A-anti-hTR and Ad5CMV-p53 than to other combinations. The additive effect of the combination therapy was due to induction of caspase-dependent apoptosis and cell growth arrest. Furthermore, the 2-5A-anti-hTR treatment when combined with Ad5CMV-p53 showed greater efficacy against subcutaneous U251-MG tumors in nude mice. In contrast, U87-MG cells expressing the wild-type p53 gene were insensitive to Ad5CMV-p53, although the treatment with 2-5A-anti-hTR was significantly effective. These results indicate that combining 2-5A-anti-hTR with Ad5CMV-p53 has the most therapeutic potential for malignant gliomas with mutant p53. For tumors exhibiting wild-type p53, it may be useful to treat with 2-5A-anti-hTR. Gene Therapy (2000) 7, 2071-2079.
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PMID:Combination therapy of malignant glioma cells with 2-5A-antisense telomerase RNA and recombinant adenovirus p53. 1122 87

We reported previously that the production of urokinase-type plasminogen activator receptor (uPAR) protein is greater in high-grade glioblastomas than in low-grade gliomas. Transcriptional activation of the uPAR gene or increased stability of the uPAR mRNA that encodes this protein could cause the increased production of this protein in cell lines of different grades of gliomas. We found similar half-life of uPAR mRNA of 10-12 h in glioblastoma multiforme (UWR3) and anaplastic astrocytoma (SW1783) cells. However, the human uPAR promoter was up-regulated 6-8-fold in SW1783 cells and 11-13-fold in UWR3 cells as compared with its activity in low-grade gliomas, a finding that correlates well with previous findings of increases in uPAR mRNA and protein levels in higher-grade gliomas. uPAR mRNA level was increased 11-fold over a 24-h period in low-grade glioma cell lines after treatment with phorbol myristate acetate. The region spanning -144 to -123 bp of the human uPAR promoter that contains the Sp-1 site and a PEA-3 element and an AP-1 site at -184 plays major roles in uPAR promoter activity in glioblastoma cells. Specific antibodies used in an electrophoretic mobility shift assay identified fra-1, fra-2, Jun D, and c-Jun proteins in the nuclear protein complex that bind a 51-mer containing the AP-1 consensus sequence at -184 and its flanking sequences in the uPAR promoter. We further studied the inhibition of uPAR promoter by coexpression of a transactivation domain lacking C-Jun; a dominant-negative ERK1 and ERK2 mutant and a dominant-negative C-raf in glioblastoma cell lines showed the repressed uPAR promoter activity compared with the effect of the empty expression vector. We conclude from our findings that increased transcription is the more likely mechanism underlying the increase in uPAR production in high-grade gliomas.
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PMID:Regulation of the urokinase-type plasminogen activator receptor gene in different grades of human glioma cell lines. 1123 78

The DNA repair protein O6-methylguanine-DNA methyltransferase (MGMT) confers resistance to methylating and chloroethylating agents in pediatric medulloblastoma- and glioma-derived cell lines and xenografts. Here, we assayed MGMT activity in 110 pediatric brain tumors to establish correlates with patient and tumor characteristics. We also assayed MGMT in histologically normal brain adjacent to 22 tumors to characterize changes in activity accompanying neurocarcinogenesis. MGMT activity was detected in 94% of tumors, ranging ca. 1,500-fold from 0.34 to 498 fmol/10(6) cells (approximately 205-300,000 molecules/cell). Mean activity was 25 +/- 66 fmol/10(6) cells, including six specimens with undetectable activity (Mer- phenotype; <0.25 fmol/10(6) cells or 151 molecules/cell). MGMT content varied 10-fold among diagnostic groups and was associated with degree of malignancy, as evidenced by a 4-fold difference in activity between high- and low-grade tumors (P = 0.03). Tumor MGMT content was age dependent, being 5-fold higher in children 3-12 years old than in infants (P = 0.015) and adolescents (P = 0.015). Mean activity in tumors was 9-fold higher than in adjacent histologically normal brain (21 +/- 44 versus 2.4 +/- 4.0 fmol/10(6) cells; P = 0.05). By comparing tumor and adjacent normal tissue from the same patient, we found that 68% of cases exhibited an elevation of tumor activity that ranged from 2- to >590-fold. Moreover, 67% of Mer- normal tissue was accompanied by Mer+ tumor. These observations indicate that MGMT activity is frequently elevated during pediatric neurocarcinogenesis. Significantly, enhanced MGMT activity may heighten resistance to alkylating agents, suggesting a potential role for MGMT inhibitors in therapy.
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PMID:O6-Methylguanine-DNA methyltransferase in pediatric primary brain tumors: relation to patient and tumor characteristics. 1129 57

Fibroblast growth factor-2 (FGF-2) is involved as an autocrine growth factor in the autonomous proliferation of glioma cells. To develop a new strategy for treating patients with glioma, we studied the effect on human glioma cells of a 16-mer oligopeptide with conformational similarity to the putative receptor-binding domain of FGF-2. A synthesized oligonucleotide was assessed its receptor-binding activity by BIAcore instrument. Its biological effect on glioma cell lines was examined in vitro by MTT assay. The peptide suppressed the in vitro growth of human glioma cells U87MG, T98G and U251MG cells, but not of A431 cells whose growth is not dependent on FGF-2. Apoptotic bodies were noted after 24-h incubation in the presence of the peptide; Ac-YVAD-CHO, a caspase-3 inhibitor, suppressed apoptosis. Furthermore, we examined the modulation of the cytotoxic effect of anticancer drugs by the oligopeptide. The addition of this oligopeptide to the chemotherapeutic agents CDDP, ACNU and VP16 had additive effects in vitro. These results suggest that the pathway of the FGF-2 autocrine loop through the FGF receptor plays an important role in the proliferation of glioma cells. New drugs targeting this loop may be highly effective in treating FGF-2-dependent tumors. Our results suggest that its addition to the therapeutic arsenal may lead to improved treatment regimens for patients with FGF-2-dependent tumors.
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PMID:In vitro growth suppression of human glioma cells by a 16-mer oligopeptide: a potential new treatment modality for malignant glioma. 1282 20

Scanning oligodeoxynucleotide (ODN) arrays appear promising in vitro tools for the prediction of effective antisense reagents but their usefulness has not yet been reported in mammalian systems. In this study, we have evaluated the use of scanning ODN arrays to predict efficacious antisense ODNs targeting the human epidermal growth factor receptor (EGFR) mRNA in a human epidermoid cancer cell line and in primary human glioma cells. Hybridisation accessibility profile of the first 120nt in the coding region of the human EGFR mRNA was determined by hybridising a radiolabelled EGFR transcript to a scanning array of 2684 antisense sequences ranging from monomers to 27-mers. Two ODNs, AS1 and AS2, complementary to accessible sequences within the EGFR mRNA, were designed and their ability to hybridise to EGFR mRNA was further confirmed by in vitro RNase H-mediated cleavage assays. Phosphorothioate-modified 21-mer AS1 and AS2 ODNs inhibited the growth of an established human A431 cancer cell line as well as primary glioma cells from human subjects when delivered as cationic lipoplexes. In contrast, scrambled controls and AS3-an antisense ODN complementary to an inaccessible site in EGFR mRNA-were inactive. Western blots showed that AS1 ODN exhibited a dose-dependent inhibition of EGFR protein expression in A431 cells in the nanomolar range. Microarray-based gene expression profiling studies of A431 cells treated with the 21-mer phosphorothioate AS1 ODN demonstrated successful inhibition of downstream signalling molecules further confirming the effective inhibition of EGFR expression in human cancer cells by antisense ODNs designed by scanning ODN array technology.
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PMID:Messenger RNA expression profiling of genes involved in epidermal growth factor receptor signalling in human cancer cells treated with scanning array-designed antisense oligonucleotides. 1294 63

Optical spectroscopic techniques such as CD, Raman scattering, and fluorescence imaging allowed us to analyze the complex formation and vectorization of a single-stranded 20-mer phosphorothioate oligodeoxynucleotide with a 15-mer amphipathic peptide at molecular and cellular levels. Different solvent mixtures (methanol and water) and molecular ratios of peptide/oligodeoxynucleotide complexes were tested in order to overcome the problems related to solubility. Optimal conditions for both spectroscopic and cellular experiments were obtained with the molecular ratio peptide/oligodeoxynucleotide equal to 21:4, corresponding to a 7:5 ratio for their respective +/- charge ratio. At the molecular level, CD and Raman spectra were consistent with a alpha-helix conformation of the peptide in water or in a methanol-water mixture. The presence of methanol increased considerably the solubility of the peptide without altering its alpha-helix conformation, as evidenced by CD and Raman spectroscopies. UV absorption melting profile of the oligodeoxynucleotide gave rise to a flat melting profile, corresponding to its random structure in solution. Raman spectra of oligodeoxynucleotide/peptide complexes could only be studied in methanol/water mixture solutions. Drastic changes observed in Raman spectra have undoubtedly shown: (a) the perturbation occurred in the peptide secondary structure, and (b) possible interaction between the lysine residues of the peptide and the oligodeoxynucleotide. At the cellular level, the complex was prepared in a mixture of 10% methanol and 90% cell medium. Cellular uptake in optimal conditions for the oligodeoxynucleotide delivery with low cytotoxicity was controlled by fluorescence imaging allowing to specifically locate the compacted oligonucleotide labeled with fluorescein at its 5'-terminus with the peptide into human glioma cells after 1 h of incubation at 37 degrees C.
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PMID:Complex formation and vectorization of a phosphorothioate oligonucleotide with an amphipathic leucine- and lysine-rich peptide: study at molecular and cellular levels. 1504 76

Seven new macrolides having a 12-membered ring, which we termed pladienolides, were isolated from the fermentation broth of Streptomyces platensis Mer-11107. Six of the seven pladienolides inhibited hypoxia-induced reporter gene expression controlled by human VEGF promoter with IC50 values of 0.0018-2.89 microM. They also demonstrated growth-inhibitory activity against U251 human glioma cells in vitro. Pladienolides are highly potent inhibitors of both hypoxia signals and cancer cell proliferation, and thus may be useful as antitumor agents.
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PMID:Pladienolides, new substances from culture of Streptomyces platensis Mer-11107. I. Taxonomy, fermentation, isolation and screening. 1515 2


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