RESUMO
Combustion of coal for energy generation has been a significant contributor to increased concentrations of atmospheric carbon dioxide. It is of interest to evaluate the potential of former coalfields for mitigating these increases by carbon sequestration and to compare different options to achieving this end. Here, carbon sequestration in residual coal seams and through reclamation of spoil tips is compared, and their carbon dioxide storage potential in the South Wales Coalfield estimated. Coal seam sequestration estimates come from an established methodology and consider the total unmined coal resource below 500 m deep with potential for carbon sequestration. The most likely effective deep seam storage capacity is 104.9 Mt carbon dioxide, taking account of reservoir conditions and engineering factors. Whilst many spoil tips in South Wales have been reclaimed, the focus has not been on carbon sequestration potential. Estimates of minesoil restoration sequestration capacity were based on a survey of restored minesoil and vegetation carbon stocks, mainly on sites 20-30 years after restoration; data from this survey were then extrapolated to the coalfield as a whole. Minesoil storage is estimated at 1.5 or 2.5 Mt (+2.2 Mt in tree biomass) carbon dioxide based on average grassland or woodland measurements, respectively; modelled data predicted equilibrium values of 2.9 and 2.6 Mt carbon dioxide respectively in grassland or woodland minesoils. If all sites achieved close to the maximum capacity in their land use class, minesoil storage capacity would increase to 2.1 or 3.9 Mt carbon dioxide, respectively. Combining the best woodland minesoil and standing biomass values, sequestration capacity increases to 7.2 Mt carbon dioxide. The wider social, economic, environmental and regulatory constraints to achieving this sequestration for each approach are discussed. Coal seam sequestration has a much higher capacity but sequestration in mine sites is less costly and has fewer regulatory constraints. Findings indicate a significant combined potential for carbon sequestration in the South Wales Coalfield and highlight challenges in achieving this potential. On a global scale, ex-coalfield sequestration could contribute to broader efforts to mitigate emissions.
Assuntos
Sequestro de Carbono , Carvão Mineral , Dióxido de Carbono , Árvores , Reino UnidoRESUMO
The opioid system is implicated in the hedonic and motivational processing of food, and in binge eating, a behaviour strongly linked to obesity. The aim of this study was to evaluate the effects of 4 weeks of treatment with the mu-opioid receptor antagonist GSK1521498 on eating behaviour in binge-eating obese subjects. Adults with body mass index ≥ 30 kg m(-2) and binge eating scale scores ≥ 19 received 1-week single-blind placebo run-in, and were then randomized to 28 days with either 2 mg day(-1) GSK1521498, 5 mg day(-1) GSK1521498 or placebo (N=21 per arm) in a double-blind parallel group design. The outcome measures were body weight, fat mass, hedonic and consummatory eating behaviour during inpatient food challenges, safety and pharmacokinetics. The primary analysis was the comparison of change scores in the higher-dose treatment group versus placebo using analysis of covariance at each relevant time point. GSK1521498 (2 mg and 5 mg) was not different from placebo in its effects on weight, fat mass and binge eating scores. However, compared with placebo, GSK1521498 5 mg day(-1) caused a significant reduction in hedonic responses to sweetened dairy products and reduced calorific intake, particularly of high-fat foods during ad libitum buffet meals, with some of these effects correlating with systemic exposure of GSK1521498. There were no significant effects of GSK1521498 2 mg day(-1) on eating behaviour, indicating dose dependency of pharmacodynamics. GSK1521498 was generally well tolerated and no previously unidentified safety signals were detected. The potential for these findings to translate into clinically significant effects in the context of binge eating and weight regain prevention requires further investigation.
Assuntos
Bulimia/tratamento farmacológico , Comportamento Alimentar/efeitos dos fármacos , Indanos/farmacologia , Receptores Opioides mu/antagonistas & inibidores , Triazóis/farmacologia , Adolescente , Adulto , Relação Dose-Resposta a Droga , Método Duplo-Cego , Feminino , Humanos , Indanos/administração & dosagem , Indanos/uso terapêutico , Masculino , Pessoa de Meia-Idade , Triazóis/administração & dosagem , Triazóis/uso terapêutico , Adulto JovemRESUMO
We report a case of an adult female with disseminated tuberculosis, cytomegalovirus viraemia and haemophagocytic-lymphohistiocystosis syndrome associated with neutralizing anti- interferon gamma (IFNγ) autoantibodies demonstrated by absent IFNγ stimulated STAT1 phosphorylation in the presence of patient sera. A brief review of immunodeficiency caused by anti-IFNγ autoantibodies is also described.
RESUMO
Endothelin-1 (EDN1) has been reported to be implicated in the pathophysiology of asthma. Literature results on the genetic association of EDN1 in asthma are inconsistent. Eleven single nucleotide polymorphisms in EDN1 were genotyped in 342 and 100 families from UK and Norway, respectively. Asthma, bronchial hyperreactivity (BHR) and atopic asthma phenotypes were analyzed for the family-based association. Five single nucleotide polymorphisms (SNPs) were associated with asthma (0.0017Assuntos
Asma/genética
, Endotelina-1/genética
, Genética Populacional
, Polimorfismo de Nucleotídeo Único/genética
, Adolescente
, Adulto
, Alelos
, Estudos de Casos e Controles
, Criança
, Interpretação Estatística de Dados
, Família
, Feminino
, Frequência do Gene
, Marcadores Genéticos
, Haplótipos
, Humanos
, Desequilíbrio de Ligação
, Masculino
, Noruega
, Estatística como Assunto
, Reino Unido
RESUMO
BACKGROUND: Exposure of Chinese hamster ovary (CHO) cells to fractionated x irradiation in vitro has led to expression of a distinctive multiple-drug-resistant phenotype. This phenotype is characterized by overexpression of P-glycoprotein without an increase in P-glycoprotein messenger RNA or gene amplification; increased glutathione S-transferase (GST) activity; and resistance to vincristine, colchicine, and etoposide but not to doxorubicin. PURPOSE: To investigate whether this phenotype is dominant or recessive, we established intraspecific hybrids by fusion of x-ray-treated, drug-resistant CHO cells (DXR-10I or DXR-10II) with drug-sensitive CHO cells (E29). METHODS: Drug resistance levels were determined in the wild-type CHO cell line AuxB1, the drug-sensitive E29 line, the x-ray-pretreated lines, and the hybrid lines by colony-forming assay of cells grown in increasing concentrations of colchicine, vincristine, or doxorubicin. The hybrids were characterized by analysis of DNA content, P-glycoprotein expression by Western blotting, GST activity by use of 1-chloro-2,4-dinitrobenzene as substrate, and sensitivity to reversal of resistance to vincristine by exposure to verapamil. RESULTS: These hybrids proved resistant to colchicine (two-fold) and vincristine (five- to seven-fold) but not to doxorubicin. After the hybrids were exposed to verapamil, vincristine cytotoxicity was increased 10- to 12-fold. The hybrid lines exhibited levels of P-glycoprotein comparable to those of the unfused x-ray-treated parent cell line, suggesting that P-glycoprotein overexpression is a dominant trait in these hybrid lines. Interpretation of the role of increased GST activity in these hybrids was inconclusive because of the very high levels of GST in the drug-sensitive cell-fusion partner. CONCLUSIONS: The multiple-drug-resistant phenotype following x-ray treatment of CHO cells in vitro was dominantly expressed. Overall, these data are consistent with the hypothesis that this phenotype is a consequence of the dominant genetic alteration resulting from exposure to x irradiation. IMPLICATIONS: This work adds weight to our hypothesis that there is a biological basis for the expression of clinical drug resistance in certain patients whose tumors have been previously irradiated.
Assuntos
Antineoplásicos/farmacologia , Resistência a Medicamentos/genética , Genes Dominantes , Radiogenética , Membro 1 da Subfamília B de Cassetes de Ligação de ATP , Animais , Western Blotting , Células CHO/efeitos da radiação , Fusão Celular , Colchicina/farmacologia , Ensaio de Unidades Formadoras de Colônias , Cricetinae , Doxorrubicina/farmacologia , Glutationa Transferase/metabolismo , Células Híbridas/enzimologia , Glicoproteínas de Membrana/biossíntese , Fenótipo , Verapamil/farmacologia , Vincristina/farmacologiaRESUMO
We observed that in vitro exposure of mammalian tumor cells to fractionated x irradiation results in the expression of drug resistance. The cause of this resistance was investigated in a series of Chinese hamster ovary cell lines that had survived exposure to multiple lethal doses of radiation. These cell lines had increased levels of P-glycoprotein (Pgp), the multidrug-resistance-associated membrane glycoprotein. Consistent with the classic multidrug resistance phenotype, they exhibited cross-resistance to multiple drugs, as well as sensitivity to reversal of vincristine resistance by verapamil. However, the cell lines showed no change in their sensitivity to x rays. Pgp overexpression occurred in these cells, despite a lack of Pgp gene amplification or of significant alteration in Pgp messenger RNA levels. Although the cause of increased Pgp levels is not yet known, these data suggest a biological basis for the clinical problem of drug resistance that can occur in previously irradiated tumors.
Assuntos
Antineoplásicos/farmacologia , Resistência a Medicamentos/genética , Glicoproteínas de Membrana/efeitos da radiação , Células Tumorais Cultivadas/metabolismo , Membro 1 da Subfamília B de Cassetes de Ligação de ATP , Animais , Regulação Neoplásica da Expressão Gênica/efeitos da radiação , Glutationa/metabolismo , Glutationa Transferase/metabolismo , Immunoblotting , Glicoproteínas de Membrana/biossíntese , Glicoproteínas de Membrana/genética , RNA Mensageiro/efeitos da radiação , RNA Neoplásico/efeitos da radiação , Células Tumorais Cultivadas/efeitos dos fármacos , Ensaio Tumoral de Célula-TroncoRESUMO
Two human ovarian tumor cell lines (SK-OV-3 and TR175), established from patients previously treated with alkylating agents, but not with cisplatin, expressed greater than 23-fold differences in cisplatin sensitivities in vitro. Cisplatin resistance in SK-OV-3 cells appeared to be associated with increased levels of glutathione and activities of glutathione reductase and glutathione peroxidase, with reduced catalase activity. No significant modification of drug uptake was noted and there was only marginally lower (16%) total platination of DNA, measured immunochemically, in these cells compared with the more sensitive TR175 cell line. SK-OV-3 cells, however, showed a significantly lower overall ability to remove drug-induced DNA damage, with an apparent inability to remove either the major DNA-DNA intrastrand cross-links in the sequence pGpG or the adducts cis-Pt(NH3)2d(GMP)2, although by alkaline elution repair of DNA-DNA interstrand cross-links was demonstrated. Significantly more of these interstrand cross-links were induced in these resistant cells. These data provide evidence for the involvement of altered glutathione metabolism and increased tolerance of certain types of drug-induced DNA damage as factors associated with the resistance phenotype of SK-OV-3 cells. Paradoxically, however, although the highly cisplatin-sensitive TR175 cells had lower glutathione levels this was not reflected in significantly higher total platination of DNA, and these cells appeared to be proficient in removing all the major platinum-DNA adducts quantitated in this study. Mechanisms responsible for this relative sensitivity to cisplatin remain to be identified.
Assuntos
Cisplatino/farmacologia , Reparo do DNA/efeitos dos fármacos , DNA de Neoplasias/metabolismo , Neoplasias Ovarianas/metabolismo , Platina/metabolismo , Butionina Sulfoximina , Cisplatino/metabolismo , DNA de Neoplasias/efeitos dos fármacos , Relação Dose-Resposta a Droga , Resistência a Medicamentos , Feminino , Glutationa/análise , Glutationa Transferase/análise , Humanos , Metionina Sulfoximina/análogos & derivados , Metionina Sulfoximina/farmacologia , Neoplasias Ovarianas/tratamento farmacológico , Neoplasias Ovarianas/enzimologia , Células Tumorais Cultivadas/metabolismoRESUMO
In vitro exposure of the TR170 ovarian carcinoma cell line to six intermittent 24-h treatments with a 90% inhibitory concentration of cisplatin (CDDP) (0.15 micrograms/ml; 0.5 microM) resulted in a 2-fold stably resistant subline designated TR170/CP+ (B.T. Hill et al., Int. J. Cancer, 39: 219-225, 1987). Resistance to CDDP in these CP+ cells has now been associated with reduced uptake of 195mCDDP (2-fold; P less than 0.01) and decreased removal of specific Pt-DNA adducts, quantitated immunochemically, indicative of an apparent increased tolerance of CDDP-induced DNA damage. Specifically these resistant cells appeared deficient in removal of the major cis-Pt-(NH3)2d(pGpG) adduct and the difunctional cis-Pt(NH3)2d(GMP)2 lesion, showed less efficiency in removing cis-Pt(NH3)2d(pApG) adducts, but proved as proficient as the parental cell line in removing DNA-DNA interstrand cross-links. Activities of DNA polymerase-alpha and -beta were comparable in both lines, and no significant alterations in glutathione metabolism were identified. Response to acute X-irradiation was not modified in these TR170/CP+ cells, but they showed marked (10-fold) cross-resistance to 5-fluorouracil and, unusually, proved collaterally sensitive (12-fold) to methotrexate. Resistance to 5-fluorouracil was associated with significantly increased thymidylate synthase activity (P less than 0.01), but this was not reflected in altered gene expression, while increased sensitivity to methotrexate was accompanied by increased drug uptake but by unaltered activity and expression of dihydrofolate reductase. These results indicate that exposure to CDDP can result in numerous alterations, both intracellularly and at the cellular membrane, reflected in significant changes in the tumor cells' responses to the cytotoxic effects of a range of antitumor drugs. The clinical relevance of these observations remains to be established.
Assuntos
Antineoplásicos/farmacologia , Cisplatino/farmacologia , Resistência a Medicamentos/fisiologia , Fluoruracila/farmacologia , Metotrexato/farmacologia , Catalase/metabolismo , Divisão Celular/efeitos dos fármacos , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Cisplatino/metabolismo , DNA Polimerase Dirigida por DNA/metabolismo , Feminino , Fluoruracila/metabolismo , Glutationa/metabolismo , Glutationa Peroxidase/metabolismo , Glutationa Redutase/metabolismo , Glutationa Transferase/metabolismo , Humanos , Metotrexato/metabolismo , NAD(P)H Desidrogenase (Quinona)/metabolismo , Neoplasias Ovarianas , Superóxido Dismutase/metabolismo , Raios XRESUMO
Exposure of the Chinese hamster ovarian AuxB1 cell line in vitro to fractionated X-irradiation generated sublines designated DXR-10, which proved resistant to multiple drugs and overexpressed P-glycoprotein (Pgp), as judged by Western blotting using the C219 monoclonal antibody. Further characterization of these irradiated DXR-10 sublines has provided evidence for: (i) the expression of cross-resistance to gramacidin D, taxol, puromycin and Navelbine, but not to daunomycin or mitoxantrone; (ii) overexpression of the class I Pgp, as judged by Western blotting using the C494 monoclonal antibody; (iii) decreased accumulation of 3H-vincristine, which could be enhanced by verapamil addition; (iv) unaltered accumulation and subcellular distribution of adriamycin; (v) significantly increased rhodamine 123 accumulation in the presence of verapamil; (vi) plasma-membrane ultrastructural modifications resulting in a significantly increased surface area; (vii) numerous clonal karyotypic alterations, with abnormalities involving the long arm of chromosome 1 being consistently identified; (viii) a lack of overexpression of sorcin; (ix) increased total glutathione levels and overexpression of glutathione S-transferase pi. The fact that only certain of these features are considered characteristic of the 'classic' multidrug-resistant CHRC5 cell line supports our earlier proposal that exposure to fractionated X-irradiation results in the expression of a unique drug-resistance phenotype.
Assuntos
Células CHO/metabolismo , Proteínas de Transporte/metabolismo , Glicoproteínas de Membrana/metabolismo , Membro 1 da Subfamília B de Cassetes de Ligação de ATP , Animais , Células CHO/efeitos da radiação , Células CHO/ultraestrutura , Proteínas de Ligação ao Cálcio/metabolismo , Bandeamento Cromossômico , Cricetinae , Interações Medicamentosas , Resistência a Medicamentos/genética , Glutationa/metabolismo , Cariotipagem , FenótipoRESUMO
About 40% (350 kb) of the human MHC class II region has been sequenced and a coordinated effort to sequence the entire MHC is underway. In addition to the coding information (22 genes/pseudogenes), the non-coding sequences reveal novel information on the organisation and evolution of the MHC as demonstrated here by the example of a 200 kb contig that has been analysed for local and global features. In conjunction with cross-species comparisons, our results present new evidence on the structure of isochores, the evolutionary dynamics of repeat-mediated recombination and its effect on certain MHC encoded genes, and a higher than average degree of natural polymorphism that has implications for sequencing the human genome. We also report the finding of a class I-related pseudogene (HLA-ZI) in the middle of the class II region, which provides the first direct evidence for DNA exchange between these two related regions in man.
Assuntos
Evolução Molecular , Genes MHC da Classe II/genética , Sequência de Aminoácidos , Animais , Composição de Bases , Sequência de Bases/genética , Humanos , Dados de Sequência Molecular , Polimorfismo Genético , Pseudogenes/genética , Sequências Repetitivas de Ácido Nucleico/genética , Análise de Sequência de DNA , Especificidade da EspécieRESUMO
We have established that drug resistance can be expressed following in vitro exposure of tumour cells not only to antitumor drugs but also to fractionated X-irradiation. These data therefore suggest a biological basis for the clinical problem of drug resistance that can occur in patients with previously irradiated tumors. These observations, if confirmed, have clinical implications for the combined modality approach and need to be considered when attempting to identify resistant tumour cells in clinical specimens with the aim of monitoring or identifying effective drug regimens.
Assuntos
Antineoplásicos/farmacologia , Células Tumorais Cultivadas/efeitos da radiação , Membro 1 da Subfamília B de Cassetes de Ligação de ATP , Antineoplásicos/farmacocinética , Cisplatino/farmacologia , DNA de Neoplasias/metabolismo , Resistência a Medicamentos/efeitos da radiação , Humanos , Glicoproteínas de Membrana/metabolismo , Doses de Radiação , Células Tumorais Cultivadas/efeitos dos fármacos , Células Tumorais Cultivadas/metabolismoRESUMO
In breast cancers and sarcomas, a variant polymorphism in the cell cycle inhibitor P21CIP1/WAF1 is under-represented in those individuals whose tumours contain mutated TP53. The aim of this study was to determine whether this variant polymorphism was also under-represented in those with ovarian carcinoma and TP53 mutations. We studied lymphocyte DNA from 104 women with ovarian carcinoma, 15 with borderline tumours and 16 with benign tumours, using a previously-reported PCR-RFLP technique. 96 of the ovarian carcinoma cases had been previously examined for mutations in TP53 and/or for overexpression of the TP53 protein. The variant allele was seen in 11 out of 104 women (10.6%) with ovarian carcinoma. There was no significant difference in the distribution of the variant allele in the women whose tumours had (7/47) or did not have (4/49) TP53 mutations (P = 0.523). It does not appear that the presence of this variant allele of P21CIP1/WAF1 has any aetiological role in ovarian carcinomas. Studies in other tumours support this finding.
Assuntos
Ciclinas/genética , Inibidores Enzimáticos , Genes p53 , Neoplasias Ovarianas/genética , Polimorfismo Genético , Alelos , Inibidor de Quinase Dependente de Ciclina p21 , Eletroforese em Gel de Poliacrilamida , Feminino , Humanos , Proteínas de Neoplasias/genéticaRESUMO
Examples of collateral sensitivity, even in experimental tumor systems, remain few. Preliminary data from this laboratory indicated that certain tumor cells expressed increased sensitivity to cisplatin after exposure in vitro to x-irradiation. To further clarify whether the type of fractionated radiation procedure used clinically can induce hypersensitivities to certain antitumor drugs we have pre-exposed the human ovarian carcinoma cell line JA-T/P derived from a tumor from an untreated patient to fractionated x-irradiation (total dose 50 Gy) in vitro. The resultant subline JA-T/DXR-10 expressed collateral sensitivity to cisplatin (CDDP), methotrexate (MTX) and fluorouracil (5-FU), but not to acute x-irradiation. Hypersensitivity to CDDP was associated with decreased activity of DNA polymerase beta (3.5-fold, P less than .01), but unaltered glutathione metabolism. Pre-incubation with cyclosporin A or with 3-aminobenzamide significantly enhanced (twofold, P less than .01) CDDP-induced cytotoxicity in JA-T/P cells, but not in the DXR-10 subline. Consistent with MTX hypersensitivity dihydrofolate reductase activity was significantly decreased (2.9-fold, P less than .01). Despite collateral sensitivity to 5-FU, however, thymidylate synthase activity was increased (twofold, P less than .05) suggesting alternative mechanisms for 5-FU-induced cytotoxicity in these JA-T/DXR-10 cells. These data demonstrate that DNA repair and associated reduced folate metabolism can be modified not only by drugs but also by fractionated x-irradiation.
Assuntos
Cisplatino/farmacologia , Cistadenocarcinoma/tratamento farmacológico , Fluoruracila/farmacologia , Metotrexato/farmacologia , Neoplasias Ovarianas/tratamento farmacológico , Terapia Combinada , Cistadenocarcinoma/enzimologia , Cistadenocarcinoma/radioterapia , Resistência a Medicamentos/efeitos da radiação , Feminino , Glutationa/análise , Humanos , Neoplasias Ovarianas/enzimologia , Neoplasias Ovarianas/radioterapia , Tetra-Hidrofolato Desidrogenase/análise , Timidilato Sintase/análise , Células Tumorais CultivadasRESUMO
In vitro exposure of a human testicular teratoma continuous cell line to fractionated X-irradiation resulted in the expression of resistance to cisplatin. In two independently-derived sublines, designated SUSA-DXR13 and SUSA-DXR10 resulting from treatment with either 13 fractions of 1.5 Gy (dose required to reduce survival by 1 log) or 10 fractions of 3 Gy (dose required to reduce survival by 2 logs) respectively, the IC50 values for cisplatin were 2- and 3.1-fold higher than that of the parental cell line. These sublines were cross-resistant to carboplatin (approximately 2-fold) but not to adriamycin and they showed unaltered radiosensitivities. The SUSA-DXR10 subline expressed some cross-resistance to mitomycin C and melphalan but none to Carmustine (BCNU). Total glutathione content was significantly reduced in both SUSA-DXR10 and SUSA-DXR13 cells, but the activities of associated enzymes, including the glutathione S-transferases, peroxidase and reductase were not modified significantly in the resistant sublines. Resistance in the SUSA-DXR10 subline was associated with significantly decreased 195mcisplatin uptake (p less than 0.01), but this was not reflected in a reduced level of drug bound to the DNA. The formation and removal of four platinum-DNA adducts were immunochemically quantitated. Immediately following drug treatment there was a higher level of total platination of the DNA in the resistant subline indicative of increased tolerance to DNA damage. After an 18 hr post treatment incubation, there was an indication of some repair capacity in this SUSA-DXR10 cell line, which was not apparent in the parental cells. Neither the parental nor the SUSA-DXR10 cell line was proficient in the repair of the major adduct Pt-GG, whereas both lines repaired the monofunctional adduct and the adduct Pt(GMP)2. SUSA-DXR10 cells were also able to repair the intrastrand adduct Pt-AG and interstrand crosslinks, unlike the repair deficient parental cells. Higher levels of interstrand crosslinks were characteristic of the SUSA-DXR10 subline. These observations therefore implicate both enhanced repair and increased tolerance of DNA damage as mechanisms of resistance to cisplatin resulting from in vitro exposure of a human teratoma cell line to fractionated X-irradiation.
Assuntos
Cisplatino/farmacologia , Adutos de DNA , Dano ao DNA , Reparo do DNA/efeitos da radiação , DNA/efeitos dos fármacos , Resistência a Medicamentos/efeitos da radiação , Teratoma/metabolismo , Neoplasias Testiculares/metabolismo , Células Tumorais Cultivadas/efeitos da radiação , Cisplatino/metabolismo , Cisplatino/farmacocinética , DNA/metabolismo , Relação Dose-Resposta a Droga , Glutationa/metabolismo , Glutationa Peroxidase/metabolismo , Glutationa Redutase/metabolismo , Glutationa Transferase/metabolismo , Humanos , Masculino , Dosagem Radioterapêutica , Teratoma/enzimologia , Teratoma/patologia , Neoplasias Testiculares/enzimologia , Neoplasias Testiculares/patologia , Fatores de Tempo , Células Tumorais Cultivadas/enzimologiaRESUMO
Three etoposide-selected resistant sublines of the SuSa testicular teratoma cell line expressing 9-, 21- and 33-fold levels of resistance, proved increasingly cross resistant to cisplatin with levels approximating to 3-, 4- and 6-fold in sublines VPC2, VPC3 and VPC4, respectively. Cisplatin resistance was not associated with any significant modifications in levels of total glutathione or associated enzyme activities. Decreased platinum (Pt) accumulation was detected, although this did not correlate either with total platination levels judged immunochemically or with peak induction of interstrand crosslinks (ISC) determined by alkaline elution. Following exposure to cisplatin in the least resistant subline, VPC2, total platination levels were markedly decreased (3-fold) relative to those of the parental cells, whilst peak ISC levels were markedly increased (4-fold). In the most highly resistant subline, VPC4, peak levels of ISCs were even higher (9-fold), although total platination levels remained comparable with those in parental cells. Both VPC2 and VPC4 cells appeared highly proficient in removing ISCs, unlike the parental cells. However, whilst VPC2 cells appeared to share deficient removal of the intrastrand platinated lesions with parental cells, VPC4 cells proved proficient in removing specific adducts in the sequence pApG. This unusual expression of cross resistance to cisplatin in a series of etoposide-selected resistant sublines derived from an inherently repair deficient parental cell line, SuSa, therefore appears to be associated with enhanced removal of the specific intrastrand crosslinks in the sequence pApG and/or of DNA-DNA ISCs. Similar mechanisms have been implicated in two other cisplatin resistant SuSa sublines selected following in vitro exposure to the drug itself or to fractionated X-irradiation.
Assuntos
Cisplatino/farmacologia , DNA de Neoplasias/efeitos dos fármacos , Etoposídeo/farmacologia , Teratoma/metabolismo , Neoplasias Testiculares/metabolismo , Dano ao DNA , Reparo do DNA , Resistência a Medicamentos , Ensaios de Seleção de Medicamentos Antitumorais , Glutationa/metabolismo , Humanos , Masculino , Platina/metabolismo , Teratoma/tratamento farmacológico , Teratoma/genética , Neoplasias Testiculares/tratamento farmacológico , Neoplasias Testiculares/genética , Células Tumorais CultivadasRESUMO
Glutathione and its associated enzyme activities have been quantitated in a series of human tumour continuous cell lines expressing a range of in vitro sensitivities to certain antitumour agents. Fourteen different parental lines and 15 various drug- and X-ray-selected resistant sublines have been studied. Quantitative relationships between total glutathione levels and related enzyme activities and sensitivities to six clinically-useful antitumour drugs or X-rays, as judged by colony forming assays, have been determined by linear regression analysis. A positive correlation has been identified between glutathione levels and sensitivities to cisplatin. Adriamycin, or to X-rays. In addition, positive correlations were noted between cisplatin sensitivities and glutathione peroxidase and reductase activities and for Adriamycin responses with respect to glutathione peroxidase activity, using cumene hydroperoxide as substrate. However, no positive correlations were noted for glutathione levels or these enzyme activities with differential methotrexate, etoposide, vincristine or 5-fluorouracil cytotoxicities. Furthermore, no direct relationship was apparent between total glutathione S-transferase activities and any of these drug or X-ray sensitivities in this series of cell lines. These data appear to provide further evidence linking altered glutathione metabolism with differential cytotoxicities of certain clinically-useful antitumour agents.
Assuntos
Antineoplásicos/farmacologia , Glutationa Peroxidase/metabolismo , Glutationa Redutase/metabolismo , Glutationa Transferase/metabolismo , Glutationa/metabolismo , Células Tumorais Cultivadas/efeitos dos fármacos , Doxorrubicina/farmacologia , Resistência a Medicamentos , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Compostos Organoplatínicos/farmacologia , Análise de RegressãoRESUMO
A series of five potential modulators of resistance were tested for their relative ability, as compared with verapamil, to sensitize CEM lymphoblastoid leukemia drug-resistant tumor sublines expressing either the classic or the atypical multidrug-resistance (MDR) phenotype to vinblastine or teniposide. Maximal non-cytotoxic concentrations of each modulator were tested and sensitization induces (SIs) were derived by comparing the drug concentration required to inhibit growth by 50% in their presence or absence. Like verapamil (10 microM) itself, three of the other modulators tested, namely, S9788 (4 microM), flunarizine (20 microM) and quinidine (30 microM), resulted in 2- to 3-fold sensitization of vinblastine against the parental CEM cells, and comparable effects were noted in the CEM/VM-1 cells, which were not cross-resistant to vinblastine. In contrast, cyclosporin A (0.5 microM) and B859-35 (2 microM) did not enhance vinblastine growth inhibition in these lines. However, the greatest sensitization with all the modulators was noted in the classic MDR VBL1000 cells, with SIs ranging from 40- to 350-fold, except for cyclosporin A, which proved ineffective at the concentration tested (SI, 2.6). The greatest extent of differential sensitization of these VBL1000 tumor cells occurred with quinidine or B859-35, which proved significantly more effective than verapamil alone. Combinations of modulators resulted in additive effects, with B859-35 plus cyclosporin A proving superior to B859-35 plus verapamil. In contrast, none of these compounds proved effective as a sensitizer to teniposide. The growth-inhibitory effects of this drug were not modified significantly in either the 92-fold teniposide-resistant VM-1 cells or in the parental cells. Addition of verapamil itself also failed to modulate teniposide growth inhibition in the VBL1000 cells, which express significant cross-resistance to this drug (36-fold). However, SI values of 3- to 5-fold were obtained using quinidine or B859-35. These results serve (a) to emphasise the need to monitor the effects of modulators not only on drug-resistant cells but also on their drug-sensitive counterparts so as to ensure differential sensitization such that normal sensitive tissues are not likely to be adversely influenced and (b) to highlight the observation that the extent of modulation differs depending not only on the antitumor drug used but also on the mechanism of drug resistance expressed. This in vitro model system appears to provide a useful screening system for resistance modulators and certainly could be used in attempts to identify alternative agents that may influence teniposide sensitivity in these drug-resistant sublines.
Assuntos
Leucemia de Células T/tratamento farmacológico , Teniposídeo/farmacologia , Verapamil/farmacologia , Vimblastina/farmacologia , Ciclosporina/farmacologia , Di-Hidropiridinas/farmacologia , Relação Dose-Resposta a Droga , Resistência a Medicamentos , Ensaios de Seleção de Medicamentos Antitumorais , Flunarizina/farmacologia , Humanos , Piperidinas/farmacologia , Quinidina/farmacologia , Triazinas/farmacologia , Células Tumorais CultivadasRESUMO
Using a range of cell lines of murine and human tumour origin in which relatively modest levels (2- to 17-fold) of drug resistance have been selected in vitro by exposure to a range of standard antitumour drugs, we compared the cytotoxic effects of doxorubicin (DOX) and mitoxantrone (MITO). In general, significantly lower concentrations of MITO than of DOX were required to achieve comparable cytotoxicity, confirming previously published data. MITO appears more generally effective against the murine L5178Y drug-resistant sublines than DOX, although there was no expression of collateral sensitivity to this newer agent. In the various human tumour lines there was a lack of cross-resistance to both DOX and MITO in two 5-fluorouracil (FU)-resistant lines and one of two cisplatin (CDDP)-resistant cells, but cross-resistance was expressed in one subline resistant to vincristine (VCR) and two etoposide (VP-16)-resistant sublines. One murine and two human DOX-resistant sublines were effectively killed by MITO, whilst DOX proved effective against the human MITO-resistant subline. This apparent lack of cross-resistance between DOX and MITO in these resistant sublines expressing low levels of resistance in vitro therefore appears to contrast with previous reports involving highly multidrug-resistant DOX-selected sublines. However, since the latter lines generally exhibited profound cross-resistance to VCR and definite cross-resistance to VP-16, this may at least in part dictate their responses to MITO. Therefore, attempts to use experimentally derived drug-resistant sublines for preclinical drug screening should be approached with caution, since patterns of drug response appear to be influenced by the level of drug resistance expressed. The need remains to determine which type of model system provides the most relevant clinical information.
Assuntos
Doxorrubicina/farmacologia , Mitoxantrona/farmacologia , Células Tumorais Cultivadas/efeitos dos fármacos , Animais , Resistência a Medicamentos , Fluoruracila/farmacologia , Humanos , Camundongos , Vincristina/farmacologiaRESUMO
We reported previously that Chinese hamster ovary (CHO) cells surviving exposure to repeated doses of 9 Gy of X-irradiation in vitro expressed a multiple drug resistance phenotype characterized by cross-resistance to epipodophyllotoxins and to Vinca alkaloids, and by P-glycoprotein (Pgp) overexpression (Hill et al. 1990). We have now shown that exposure of these CHO cells to a single 30-Gy X-ray dose similarly resulted in the survivors expressing resistance to vincristine and to etoposide and overexpressing Pgp. In agreement with data obtained on cells which received repeated X-ray exposures, this Pgp overexpression occurred in the absence of any significant elevation of Pgp mRNA. However, the reduced ability to accumulate rhodamine 123 identified in these sublines, and the ability of verapamil to reverse this accumulation defect, implies that the Pgp which was overexpressed was functional. These findings indicate that a series of X-ray exposures is not necessary for expression of this distinctive multiple drug resistance phenotype, suggesting that this results not from a general 'stress-type' response but rather more specifically from the radiation exposure itself, with both single-dose and repeated X-irradiation selecting for similar genetic mutants.