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ABSTRACT: Hypoxic EMT-6 tumour cells displayed a high level of inducible nitric oxide synthase (iNOS) and an increased radiosensitivity after a 16 h exposure to lipopolysaccharide, a known activator of nuclear factor-kappaB (NF-kappaB). Both iNOS activation and radioresponse were impaired by the NF-kappaB inhibitors phenylarsine oxide and lactacystin. Contrasting to other studies, our data show that inhibition of NF-kappaB may impair the radioresponse of tumour cells through downregulation of iNOS.
British Journal of Cancer 02/2003; 88(1):120-4. · 5.04 Impact Factor
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Journal of Clinical Oncology 09/2001; 19(15):3572-4. · 18.37 Impact Factor
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ABSTRACT: Chronic hypoxia up-regulated the mRNA and protein expression of inducible nitric oxide synthase (iNOS) in EMT-6 tumour cells exposed to interferon (IFN)-gamma and interleukin (IL)-I beta. Low concentrations of cytokines (1 unit ml(-1)) in 1% but not in 21% oxygen induced a remarkable increase in NO production and a 1.8-fold hypoxic cell radiosensitization. Therefore, chronic hypoxia may potentially be exploited to increase tumour cell radioresponse through the cytokine-inducible iNOS pathway.
British Journal of Cancer 05/2001; 84(8):1122-5. · 5.04 Impact Factor
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ABSTRACT: Radiotherapy, more then any other treatment modality, relies heavily and often exclusively on medical imaging to determine the extent of disease and the spatial relation between target region and neighbouring healthy tissues. Radically new approaches to radiation delivery are inspired on CT scanning and treat patients in a slice-by-slice fashion using intensity modulated megavoltage fan beams. For quality assurance of complex 3-D dose distributions, MR based 3-D verificative dosimetry on irradiated phantoms has been described. As treatment delivery becomes increasingly refined, the need for accurate target definition increases as well and sophisticated imaging tools like image fusion and 3-D reconstruction are routinely used for treatment planning. While in the past patients were positioned on the treatment machines based exclusively on surface topography and the well-known skin marks, such approach is no longer sufficient for high-accuracy radiotherapy and special imaging tools like on-line portal imaging are used to verify and correct target positioning. Much of these applications rely on digital image processing, transmission and storage, and the development of standards, like DICOM and PACS have greatly contributed to these applications. Digital imaging plays an increasing role in many areas in radiotherapy and has been fundamental in new developments that have demonstrated impact on patient care.
European Journal of Radiology 11/2000; 36(1):41-8. · 2.61 Impact Factor
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ABSTRACT: The radiosensitizing activity of S-nitroso-N-acetylpenicillamine (SNAP), a nitric oxide (NO) donor, was assessed in a model of non-metabolic hypoxia achieved in an atmosphere of 95% nitrogen-5% carbon dioxide. A 10 min preincubation of hypoxic EMT-6 cells (10 x 10(6) ml(-1)) with 0.1 and 1 mM SNAP before radiation resulted in an enhancement ratio of 1.6 and 1.7 respectively. The level of spontaneous NO release, measured by a NO specific microsensor, correlated directly with the concentration of SNAP and was enhanced 50 times in the presence of cells. Dilution of the cell suspension from 10 to 0.1 x 10(6) ml(-1) resulted in a 16-fold decline in NO release, but only a twofold decrease in radiosensitization was observed. Preincubation of hypoxic cells with SNAP for 3 min up to 30 min caused an increasing radiosensitizing effect. Extended preincubation of 100 min led to the loss of radiosensitization although the half-life of SNAP is known to be 4-5 h. Taken together, these observations suggest that SNAP generates NO predominantly by a bioreductive mechanism and that its biological half-life is unlikely to exceed 30 min. The lack of correlation between free NO radical and radiosensitizing activity may reflect a role of intracellular NO adducts which could contribute to radiosensitization as well.
British Journal of Cancer 04/1999; 79(7-8):1085-9. · 5.04 Impact Factor
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ABSTRACT: EMT-6 cells treated for 16 h with 1-10 units/ml IFN-gamma showed a gradual activation of inducible nitric oxide synthase (iNOS) in Western and Northern blots, a simultaneous raise in NO output, and an increase in hypoxic cell radiosensitivity almost to the level of aerobic cells. Both the NO signal and radiosensitization were counteracted by the NO scavenger oxyhemoglobin, by the specific iNOS inhibitor aminoguanidine, and by the L-arginine analogue N(G)-monomethyl-L-arginine. Collectively, these data demonstrate that IFN-gamma can radiosensitize EMT-6 cells through iNOS induction and that NO is the effector molecule responsible for radiosensitization. Compared with the spontaneous NO releaser (2)-1-[N-(3-ammoniopropyl)-N-(n-propyl)amino)diazen-1-ium -1,2-diolate], the iNOS-generated NO signal appeared to be 10 times lower yet resulting in the same enhancement ratio of 2.4. Direct stimulation of NO synthesis in tumor cells through the L-arginine/iNOS pathway represents a novel approach to exploit the radiosensitizing properties of NO.
Cancer Research 01/1999; 58(24):5646-8. · 7.86 Impact Factor
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ABSTRACT: A panel of eight human pancreatic tumour cell lines displayed high intrinsic radioresistance, with mean inactivation doses between 2.4 and 6.5 Gy, similar to those reported for melanoma and glioblastoma. The radiosensitising potency of sodium nitroprusside, a bioreductive nitric oxide donor, was assessed in a model of metabolism-induced hypoxia in a cell micropellet. Sodium nitroprusside at 0.1 mM revealed a radiosensitising effect with an overall enhancement ratio of 1.9 compared with 2.5 for oxygen. Radiosensitising activity correlated with the enhancement of single-strand DNA breakage caused by radiation. In suspensions with cell densities of between 3% and 30% (v/v), the half-life of sodium nitroprusside decreased from 31 to 3.2 min, suggesting a value of around 1 min for micropellets. Despite this variation, the radiosensitising activity was similar in micropellets and in diluted cell suspensions. S-nitroso-L-glutathione was found to possess radiosensitising activity, consistent with a possible role of natural thiols in the storing of radiobiologically active nitric oxide adducts derived from sodium nitroprusside. As measured by a nitric oxide-specific microsensor, activation of sodium nitroprusside occurred by bioreduction, whereas S-nitroso-L-glutathione showed substantial spontaneous decomposition. Both agents appear to exert radiosensitising action through nitric oxide as its scavenging by carboxy phenyltetramethylimidazolineoxyl N-oxide (carboxy-PTI0) and oxyhaemoglobin resulted in attenuated radiosensitisation. Sodium nitroprusside was at least 10-fold more potent than etanidazole, a 2-nitroimidazole used as a reference. Our data suggest that sodium nitroprusside, a drug currently used for the treatment of hypertension, is a potential tumour radioresponse modifier.
British Journal of Cancer 01/1997; 74(11):1734-42. · 5.04 Impact Factor
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ABSTRACT: The human pancreatic tumour cell line PSN1/ADR, stepwise selected in 17-510 nM doxorubicin, displayed a multidrug resistance not conferred by P-glycoprotein (P-gp). Resistance to 17-51 nM doxorubicin was accompanied by overexpression of the vesicular marker lung resistance-related protein (LRP). Further selection in 170 nM doxorubicin led to the activation of multidrug resistance-associated protein (MRP) and to the development of drug accumulation/retention defects sensitive to verapamil. In addition, these defects were reversible by the vesicular traffic inhibitors brefeldin A, fluoroaluminate and nocodazole. In contrast, in human ovarian H134AD cells that are resistant to 1700 nM doxorubicin and used as P-gp-positive controls, the drug efflux was inhibited only by verapamil. The tyrosine kinase inhibitor genistein was a potent blocker of doxorubicin efflux in the PSN1/ADR cells but showed no activity in the H134 AD cells. The doxorubicin cytotoxicity in the PSN1/ADR cells was enhanced both by verapamil and brefeldin A, whereas in the parental PSN1 cells they demonstrated the opposite effects, being respectively sensitising and protecting. The P-gp-negative PSN1/ADR cells adapted to 510 nM doxorubicin retained brefeldin A-sensitive doxorubicin accumulation defects while MRP declined. The persistence of brefeldin A-responsive phenotype on the background of variable MRP expression suggests this agent as a useful functional probe for non-P-gp-mediated resistance to plasma-achievable doxorubicin concentrations.
British Journal of Cancer 04/1996; 73(5):596-602. · 5.04 Impact Factor
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ABSTRACT: Hexadecylphosphocholine (HePC), an experimental and clinical antitumour agent of the alkyllysophospholipid group, was tested for its radiosensitising effect on a panel of nine human mammary cancer cell lines in vitro. Growth inhibition by ionising radiation and recovery from it were not influenced by pretreatment with HePC in most cases, except for two cell lines expressing an activated ras oncogene. In the latter we found an enhanced radioresistance that was abolished by pretreatment with HePC. Our results suggest that HePC may act as a radiosensitiser for cells carrying an activated ras oncogene.
European Journal of Cancer 02/1993; 29A(14):1958-63. · 5.54 Impact Factor
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ABSTRACT: A high-performance liquid chromatographic method is described for the determination of vinblastine in various normal mouse tissues, such as lung, heart, liver, kidney and muscles, and in implanted MO4 tumours. Vincristine was used as the internal standard. Freshly obtained mouse tissue or tumour tissue was frozen at -20 degrees C and then lyophilized. After lyophilisation, the dry tissues were pulverized and homogeneously mixed, and an aliquot was suspended in 0.1 M hydrochloric acid. The drugs of interest were then isolated from this suspension using ion-pair extraction at pH 3 with octylsulphate as counter-ion. The obtained extracts were analysed on a reversed-phase system with a cyanopropyl stationary phase. The detection limit was 1 ng/l in plasma and 10 ng/g in tissue. The extraction recoveries of vincristine and vinblastine were between 45 and 67%, and there were no interferences from blank components. Preliminary pharmacokinetic data for different mouse tissues and tumour implanted in muscle tissue are presented.
Journal of Chromatography 08/1992; 578(2):223-9. · 4.53 Impact Factor
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ABSTRACT: A high-performance liquid chromatographic method is described for separating and determining navelbine and possible metabolites in plasma, cell culture medium and MO4 cells. Navelbine is extracted from these fluids by ion-pair extraction with sodium octylsulphate as the counter-ion at pH 3. The system uses a cyano column as the stationary phase and a mobile phase of acetonitrile-0.12 M phosphate buffer (pH 3) (60:40, v/v). Application of the method to a study of the pharmacokinetic behaviour of navelbine in MO4 mouse fibrosarcoma cells is reported.
Journal of Chromatography 06/1992; 576(2):351-7. · 4.53 Impact Factor
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ABSTRACT: Estramustine (EM) is a conjugate of estradiol and nor-nitrogen mustard (nor-HN2), which is effective in the treatment of prostate cancer. We have compared the effect of EM with that of the known microtubule inhibitor vinblastine (VLB) on the following functions of malignant MO4 mouse cells and of DU-145 human prostate cancer cells in vitro: directional migration, invasion; and the organization and the assembly/disassembly equilibrium of microtubule complexes. The circular area covered by cells migrating from an aggregate explanted on a solid substrate was taken as an index of directional migration. Invasion was studied through confrontation of MO4 or DU-145 cells with fragments of embryonic chick heart in organ culture. Microtubules were investigated immunocytochemically and through immunodetection on protein blots. VLB and EM inhibited directional migration and invasion of MO4 and DU-145 cells in a dose-dependent manner; equimolar combinations of estradiol plus nor-nitrogen mustard did not mimic these effects. At anti-invasive concentrations VLB led to partial disassembly of microtubule complexes, whereas EM resulted in an abnormal pattern of microtubule complexes without alteration of the overall assembly/disassembly equilibrium. Combined treatment with VLB and EM resulted in an enhanced VLB effect, namely complete disassembly. In all tests DU-145 cells were more sensitive to both VLB and EM than were MO4 cells, and the effects were less reversible. The present experiments showed that EM shares an anti-invasive activity with other microtubule inhibitors.
Cancer Research 05/1988; 48(7):1842-9. · 7.86 Impact Factor
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ABSTRACT: The effect of the alkyl lysophospholipid racemic-1-O-octadecyl-2-O-methyl glycero-3-phosphocholine on the expression of cell surface carbohydrates of four matched pairs of normal and malignant cells was studied using chromatographic techniques. After treatment with alkyl lysophospholipid, glycopeptides proteolytically derived from normal and malignant cells displayed a shift in the size distribution profiles obtained by gel filtration. These drug-induced changes in molecular weight distribution were expressed most strongly in untransformed cells and resembled the carbohydrate alterations found after their malignant transformation. Desialylation abolished the effect of alkyl lysophospholipid, thus suggesting an increased amount of sialic acid in the surface carbohydrates of drug-treated cells. Chromatography of glycopeptides on concanavalin A-Sepharose, Ricinus communis agglutinin I-agarose, and Bio-Gel P-4 columns excluded a higher degree of branching but suggested addition of extra terminal sialic acid residues as the major cause of the observed alterations. Alkyl lysophospholipid stimulated glycoprotein sialylation of normal cells to the level observed in malignant cells, thus inducing a "malignant-like" surface phenotype. The drug-induced carbohydrate changes in normal chick heart tissue prevented its being invaded by tumor cells when tested in an organotypic assay. The alkyl lysophospholipid thus appears to modulate in a nontoxic fashion the expression of surface molecules implicated in various cellular interactions including invasiveness.
Cancer Research 03/1988; 48(4):977-82. · 7.86 Impact Factor
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ABSTRACT: Tumor cells grown in the presence of 1-O-alkyl-2-O-methylglycero-3-phosphocholine (AMG-PC) accumulated this ether lipid in their membranes. Depending on the cell type and the dose of the compound, up to 17% of the total phospholipids of the purified plasma membranes consisted of authentic AMG-PC. Extensive incorporation of the agent resulted in a decrease in plasma membrane fluidity and inhibition of tumor cell invasiveness in embryonic chick heart fragments. The extent of AMG-PC incorporation and fluidity change was not strictly correlated with the degree to which tumor cell invasion was inhibited.
Lipids 12/1987; 22(11):820-3. · 2.13 Impact Factor
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ABSTRACT: The antiinvasive activity on MO4 mouse cells of the following lipid derivatives was tested in vitro: an alkyl-lysophospholipid derivative (ET-18-OCH3), a thioether-phospholipid derivative (BM 41.440), an alkyl-linked lipoidal amine (CP-46,665) and a naturally occurring ester-linked phospholipid (2-LPC). In this test, BM 41.440 had the same antiinvasive potency as ET-18-OCH3, whereas CP-46,665 and 2-LPC had no effect on invasion. Comparison of the antiinvasive effect of ET-18-OCH3 on three types of cells showed the following ranking: 12R1C-RK rat kidney adenovirus type 12 transfected cells greater than MO4 mouse cells greater than LLC-H61 Lewis lung carcinoma cells. This ranking was not reflected in ET-18-OCH3-induced changes of cell surface exposed glycopeptides derived from the three types of cells metabolically labeled with radioactive fucose. The present and previous experiments suggested that changes in invasion caused by lipid derivatives depended upon relative cell surface fucosyl-glycopeptide alterations in both the invasive cells and the normal tissue.
Lipids 12/1987; 22(11):847-50. · 2.13 Impact Factor
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Journal of Chromatography 06/1987; 416(2):375-81. · 4.53 Impact Factor
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ABSTRACT: A liquid chromatographic method is described for separating and determining vinblastine, vincristine and vindesine in plasma and urine. The drugs are extracted from the biological material using an ion-pair extraction, with sodium octylsulphate as counter-ion at pH 3. The extracts are injected on a reversed-phase system with a cyano column as stationary phase and a mobile phase composed of acetonitrile-phosphate buffer, pH 3 (65:35, vol. %). Stability studies are carried out for stock solutions of the drugs in water at different temperatures and pH values. The stability of these compounds in plasma is also investigated in the presence of an antioxidant. The method is applied to determine drug levels of vindesine and vinblastine in preliminary pharmacokinetic studies, using vincristine as the internal standard.
Journal of Chromatography 01/1986; 345(2):309-21. · 4.53 Impact Factor
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ABSTRACT: We have tested the value of a computer-assisted image analysis program for the quantitative study of invasionin vitro using experiments that were previously described semi-quantitatively. Mouse fibrosarcoma cell (MO4) aggregates were confronted with precultured fragments of embryonic chick heart in organ culture. Confronting pairs were fixed after 1, 2, 3 and 4 days, and processed for paraffin sectioning and immunostaining with an antiserum against chick heart. The image analysis system allowed separate quantification of two aspects of invasion, namely, occupation of the heart tissue by MO4 cells and degeneration of the invaded heart tissue. Complex combination of occupation and invasion added a qualitative aspect of invasion that has not been described previously and that revealed qualitative differences in invasion under various circumstances.
Clinical and Experimental Metastasis 03/1985; 3(2):87-101. · 3.52 Impact Factor
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ABSTRACT: Alkyl-lysophospholipids have been shown to possess antitumoral activity in animal and in human tumors. Among them, racemic 1-O-octadecyl-2-O-methylglycero-3-phosphocholine (ET-18-OCH3) had an antimetastatic effect in experimental tumors. We investigated the effect of ET-18-OCH3 on invasion of MO4 mouse fibrosarcoma cells and on cellular activities possibly related to invasion in vitro. Ten micrograms of ET-18-OCH3 per ml permitted growth of MO4 cells to about 75% of controls and slightly reduced trypan blue exclusion. Directional migration inferred from the area covered by MO4 cells that had migrated from an aggregate on glass was not affected. Reassembly of microtubules after treatment with 1 microgram of Nocodazole per ml occurred normally in presence of ET-18-OCH3. Invasion was completely inhibited when MO4 cell aggregates were confronted with precultured fragments of embryonic chick cardiac muscle or with fresh embryonic chick lung fragments in culture on gyratory shaker in fluid medium with 10 micrograms of ET-18-OCH3 per ml. These experiments showed that ET-18-OCH3, in contrast with microtubule inhibitors, interfered with invasion of MO4 cells in vitro at concentrations that permitted growth and directional migration of MO4 cells. Fluorescence polarization studies with the lipophylic probe diphenylhexatriene indicated that the antiinvasive effect of ET-18-OCH3 was accompanied by an overall increase of membrane fluidity. We tentatively concluded that alterations of the MO4 cellular membranes are responsible for the antiinvasive effect of ET-18-OCH3.
Cancer Research 02/1985; 45(1):351-7. · 7.86 Impact Factor
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ABSTRACT: We have studied recovery from growth inhibition in irradiated M04 mouse fibrosarcoma spheroids using various post-irradiation culture conditions. Recovery after irradiation with 18 Gy was significantly higher in M04 spheroids confronting fragments of embryonic chick heart in suspension culture as compared to M04 spheroids confronting pieces of Gel-foam in suspension culture (p less than 0.0007) or to M04 spheroids explanted on coverslips in static culture (p less than 0.006). These observations suggest that the normal tissue might contribute to recovery from growth inhibition in irradiated M04 cell populations.
Cell Biology International Reports 04/1984; 8(3):247-55.
