Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 26
Filtrar
1.
Cancer Res ; 54(16): 4355-61, 1994 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-7913875

RESUMO

Very low concentrations of paclitaxel, a clinically active anticancer agent isolated from the bark of the Pacific yew tree, were found to produce micronuclei in human colon carcinoma cells, suggesting inhibition of mitotic spindle assembly or function. The possibility that paclitaxel acts at the level of the mitotic spindle was investigated by evaluating its ability to inhibit the progression of mitotic cells to G1 phase. Paclitaxel inhibited mitotic progression with a median inhibitory concentration of 4 nM, a concentration equivalent to the median cytotoxic concentration, without arresting cells in mitosis. A direct correlation was shown to exist between the cytotoxic potency and ability to inhibit mitotic progression for analogues of paclitaxel and antimicrotubule agents but not for the topoisomerase II-active agents etoposide and teniposide. After release from the nocodazole block, cells synchronized in mitosis remained sensitive to very low concentrations of paclitaxel for < 30 min, the time required for spindle formation, yet remained sensitive to vinblastine for > 90 min. This result indicates that very low concentrations of paclitaxel inhibit formation of mitotic spindles in cells without affecting function of preformed spindles and without arresting cells in mitosis. Continuous exposure to low nanomolar concentrations of paclitaxel for more than one cell cycle resulted in cells with DNA contents > 4C and as much as 8C. These results support a hypothesis, that, by not being capable of segregating sister chromatids, paclitaxel-treated cells eventually reform nuclear membranes around individual or clusters of chromosomes, revert to G1 phase cells containing 4C DNA, and enter S phase, resulting in cells with as much as 8C DNA content. It is proposed that this is the primary cytotoxic mechanism of paclitaxel.


Assuntos
Anáfase/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Paclitaxel/farmacologia , Fuso Acromático/efeitos dos fármacos , Telófase/efeitos dos fármacos , Neoplasias do Colo/patologia , Ensaios de Seleção de Medicamentos Antitumorais , Fase G1 , Humanos , Nocodazol/farmacologia , Células Tumorais Cultivadas
2.
Cancer Res ; 47(19): 5141-8, 1987 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-2441861

RESUMO

An MCF-7 human breast cancer cell line was selected which was 200-fold more resistant to Adriamycin than the wild type cell line. This Adriamycin-resistant (AdrR) cell line exhibited a multidrug-resistant phenotype and was cross-resistant to a wide range of antineoplastic agents including Vinca alkaloids, anthracyclines, and epipodophyllotoxins. Cytogenetic analysis of the AdrR cell line showed the presence of homogeneously staining regions on several chromosomes which were not present in the parental cell line. Using the technique of in-gel renaturation, DNA sequences which were amplified 50- to 100-fold in the AdrR cell line and which covered a total of over 140 kilobases were isolated. In addition, AdrR cells were found to contain amplified and overexpressed sequences which were homologous to hamster P-glycoprotein gene sequences. A hamster cDNA P-glycoprotein gene probe was used to screen a lambda gt10 cDNA library made from human AdrR cell line mRNA and human cDNA sequences homologous to the P-glycoprotein gene were isolated. Hybridization studies with the cloned human cDNA (pADR1) showed that the AdrR MCF-7 cell line contained a 60-fold amplification of this DNA sequence and that polyadenylated mRNA from the AdrR cell line contained a 4.8-kilobase transcript which was overexpressed 45-fold. There was a direct correlation between DNA and RNA copy number of this sequence and level of resistance among several MCF-7 Adriamycin-resistant cell lines. In situ hybridization studies demonstrated that the human P-glycoprotein gene sequence was found on chromosome 7q21.1 in normal human lymphocytes and that amplified DNA sequences isolated from the AdrR MCF-7 cells by the in-gel hybridization technique were linked to the human P-glycoprotein sequences in the homogeneously staining regions in the AdrR cells.


Assuntos
Neoplasias da Mama/genética , DNA de Neoplasias/análise , DNA/análise , Doxorrubicina/farmacologia , Amplificação de Genes , Sequência de Bases , Neoplasias da Mama/tratamento farmacológico , Linhagem Celular , Resistência a Medicamentos , Feminino , Glicoproteínas/genética , Humanos , RNA/análise , Vincristina/farmacologia
3.
Cancer Res ; 51(6): 1638-44, 1991 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-1998955

RESUMO

In order to identify changes in 31P nuclear magnetic resonance (NMR) spectra associated with multiple drug resistance (MDR), a number of wild type and drug-resistant cancer cell lines were studied. The resistant cells included cells selected with various drugs, mainly Adriamycin, as well as cells transfected with the human multidrug resistance gene (MDR1 gene), which encodes P-glycoprotein. In most cases, 31P NMR spectra were significantly different from those of parental, drug-sensitive lines. The spectra of resistant cells generally indicated increased levels of ATP and phosphocreatine in the cytoplasm. These changes are compatible with the increased glucose utilization rate previously described for resistant cells. Major changes were also observed in the levels of glycerophosphocholine and glycerophosphoethanolamine. Changes in cellular metabolism reflected by 31P NMR spectra depend on the drug used to select the cells for MDR. The direction of these changes was not consistent for all cell lines studied and could not be directly attributed to expression of P-glycoprotein, suggesting that the changes may be related to alterations in metabolism and membrane function associated with other mechanisms of MDR. The results demonstrate the suitability of 31P NMR for studies of biochemical changes associated with MDR. The toxicity of 2-deoxyglucose, a glucose antimetabolite, was investigated in addition to the NMR studies and was found to be consistently higher in multidrug-resistant cells than in the parental drug-sensitive lines. For MCF-7 breast cancer cells, where several sublines with different levels of resistance were available, the toxicity was highest for the most resistant lines.


Assuntos
Desoxiglucose/farmacologia , Resistência a Medicamentos , Trifosfato de Adenosina/análise , Sobrevivência Celular/efeitos dos fármacos , Doxorrubicina/farmacologia , Resistência a Medicamentos/genética , Humanos , Espectroscopia de Ressonância Magnética , Fenótipo , Transfecção , Células Tumorais Cultivadas
4.
Cancer Res ; 58(6): 1111-5, 1998 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-9515790

RESUMO

Eleutherobin is a novel natural product isolated from a marine soft coral that is extremely potent for inducing tubulin polymerization in vitro and is cytotoxic for cancer cells with an IC50 similar to that of paclitaxel. This compound is cross-resistant along with other multidrug-resistant agents against P-glycoprotein-expressing cells and is cross-resistant with paclitaxel against a cell line that has altered tubulin. In mechanistic studies, eleutherobin shares with paclitaxel the ability to induce tubulin polymerization in vitro and is most likely cytotoxic by virtue of this mechanism. Human colon carcinoma cells exposed to eleutherobin contain multiple micronuclei and microtubule bundles, and they arrest in mitosis, depending on concentration, cell line, and length of exposure. These morphological abnormalities appearing in cultured cells are indistinguishable from those induced by paclitaxel. Electron microscopy reveals that eleutherobin induces homogeneous populations of long, rigid microtubules similar to those formed by paclitaxel. Thus, eleutherobin is a new chemotype with a mechanism of action similar to that of paclitaxel and, as such, has promising potential as a new anticancer agent.


Assuntos
Alcaloides/farmacologia , Diterpenos , Microtúbulos/efeitos dos fármacos , Animais , Antineoplásicos/farmacologia , Sítios de Ligação , Ligação Competitiva , Bovinos , Neoplasias do Colo/patologia , Feminino , Inibidores do Crescimento/farmacologia , Humanos , Neoplasias Ovarianas/patologia , Paclitaxel/farmacologia , Polímeros , Tubulina (Proteína)/metabolismo , Células Tumorais Cultivadas
5.
Cancer Res ; 61(20): 7507-17, 2001 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-11606387

RESUMO

BMS-214662 is a potent and selective inhibitor of farnesyltransferase (FTI). In rodent fibroblasts transformed by oncogenes, BMS-214662 reversed the H-Ras-transformed phenotype but not that of K-Ras or other oncogenes. In soft agar growth assays, BMS-214662 showed good potency in inhibiting H-ras-transformed rodent cells, A2780 human ovarian carcinoma tumor cells, and HCT-116 human colon carcinoma tumor cells. Inhibition of H-Ras processing in HCT-116 human colon tumor cells was more rapid than in H-Ras-transformed rodent fibroblast tumors. BMS-214662 is the most potent apoptotic FTI known and demonstrated broad spectrum yet robust cell-selective cytotoxic activity against a panel of cell lines with diverse histology. The presence of a mutant ras oncogene was not a prerequisite for sensitivity. Athymic and conventional mice were implanted s.c. with different histological types of human and murine tumors, respectively. BMS-214662 was administered both parenterally and p.o. and was active by all these routes. Curative responses were observed in mice bearing staged human tumor xenografts including HCT-116 and HT-29 colon, MiaPaCa pancreatic, Calu-1 lung, and EJ-1 bladder carcinomas. A subline of HCT-116, HCT-116/VM46, resistant to many standard cytotoxic agents by means of a multiple drug resistance mechanism, remained quite susceptible to BMS-214662, and borderline activity was achieved against N-87 human gastric carcinoma. Two murine tumors, Lewis lung carcinoma and M5076 sarcoma, were insensitive to the FTI. In a study performed using Calu-1 tumor-bearing mice, no obvious schedule dependency of BMS-214662 was observed. The FTI, BMS-214662, demonstrated broad spectrum activity against human tumors, but murine tumors were not as sensitive.


Assuntos
Alquil e Aril Transferases/antagonistas & inibidores , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Benzodiazepinas/farmacologia , Inibidores Enzimáticos/farmacologia , Imidazóis/farmacologia , Animais , Antineoplásicos/toxicidade , Benzodiazepinas/toxicidade , Bovinos , Esquema de Medicação , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores Enzimáticos/toxicidade , Farnesiltranstransferase , Humanos , Imidazóis/toxicidade , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBA , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/patologia , Ensaios Antitumorais Modelo de Xenoenxerto , Proteínas ras/antagonistas & inibidores , Proteínas ras/metabolismo
6.
Cancer Res ; 49(6): 1422-8, 1989 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-2466554

RESUMO

The development of multidrug resistance in MCF-7 human breast cancer cells and the acquisition of broad resistance to xenobiotics in rat hyperplastic nodules are both associated with increased P-glycoprotein (mdr) gene expression as well as changes in activities of intracellular detoxication enzymes; among these changes is a significant increase in the activity of the anionic isozyme of glutathione-S-transferase (GST). We have isolated a cDNA encoding the human anionic glutathione-S-transferase, GST pi-1, from a cDNA library constructed from multidrug-resistant MCF-7 cells. The deduced amino acid sequence of GST pi-1 shows that while the human anionic GST displays 85% nucleotide and amino acid sequence homology to the rat anionic isozyme, it is markedly less related to human basic GST isozymes. We have examined the expression of GST pi and P-glycoprotein in 170 specimens of human tissues and tumors. P-Glycoprotein RNA expression was positive in eight of 23 lymphomas and two of 12 colon tumors; however, many other normal and malignant tissues, including lung, bladder, and breast tumors, had low or undetectable levels of P-glycoprotein RNA expression. In contrast, GST pi was readily detected in a wide variety of normal and malignant tissues. The level of GST pi mRNA expression in normal tissues was heterogeneous, with lowest levels found in liver and the highest levels found in lung, esophagus, and placenta. GST pi was also variably expressed in human tumors, with the lowest relative levels occurring in lymphoma and breast cancer and the highest levels found in lung cancer and head and neck tumors. In addition, comparison of paired specimens from the same patient indicated that GST pi expression was increased in many tumors relative to matched normal tissue.


Assuntos
Glutationa Transferase/genética , Isoenzimas/genética , Glicoproteínas de Membrana/genética , Neoplasias/análise , RNA/análise , Membro 1 da Subfamília B de Cassetes de Ligação de ATP , Sequência de Bases , DNA/análise , Resistência a Medicamentos , Regulação da Expressão Gênica , Humanos , Dados de Sequência Molecular
7.
Clin Cancer Res ; 7(7): 2016-21, 2001 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-11448919

RESUMO

BMS-275183 is a taxane, the mechanism of action of which is like other known taxanes, and is the polymerization of tubulin. BMS-275183 given p.o. was as effective as i.v. paclitaxel in five tumor models [murine M109 lung and C3H mammary 16/C, and human A2780 ovarian (grown in mice and rats) and HCT/pk colon]. It was active in one other tumor model (human HCT-116 colon) but inferior to parenteral paclitaxel. BMS-275183 given p.o. was active in a human, hormone-dependent, prostate tumor model, CWR-22, and just as effective as anti-androgen chemotherapy. In a schedule dependency study, increasing the interval of time between oral administrations resulted in greater cumulative dose tolerance and improved therapeutic outcome. Oral BMS-275183 was evaluated as a combination therapy in conjunction with i.v. paclitaxel. Therapeutic advantages were evident for tumor-bearing mice that received the oral taxane either after induction chemotherapy or between courses of such treatment. BMS-275183 is currently in Phase I clinical trials at multiple sites.


Assuntos
Antineoplásicos/farmacologia , Hidrocarbonetos Aromáticos com Pontes/farmacologia , Administração Oral , Animais , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Feminino , Humanos , Concentração Inibidora 50 , Injeções Intravenosas , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C3H , Camundongos Endogâmicos DBA , Camundongos Nus , Transplante de Neoplasias , Neoplasias Experimentais/patologia , Neoplasias Experimentais/prevenção & controle , Paclitaxel/farmacologia , Ratos , Ratos Nus , Fatores de Tempo , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
8.
Clin Cancer Res ; 7(5): 1429-37, 2001 May.
Artigo em Inglês | MEDLINE | ID: mdl-11350914

RESUMO

BMS-247550, a novel epothilone derivative, is being developed by Bristol-Myers Squibb Company (BMS) as an anticancer agent for the treatment of patients with malignant tumors. BMS-247550 is a semisynthetic analogue of the natural product epothilone B and has a mode of action analogous to that of paclitaxel (i.e., microtubule stabilization). In vitro, it is twice as potent as paclitaxel in inducing tubulin polymerization. Like paclitaxel, BMS-247550 is a highly potent cytotoxic agent capable of killing cancer cells at low nanomolar concentrations. Importantly, BMS-247550 retains its antineoplastic activity against human cancers that are naturally insensitive to paclitaxel or that have developed resistance to paclitaxel, both in vitro and in vivo. Tumors for which BMS-247550 demonstrated significant antitumor activity encompass both paclitaxel-sensitive and -refractory categories, i.e., (a) paclitaxel-resistant: HCT116/VM46 colorectal (multidrug resistant), Pat-21 breast and Pat-7 ovarian carcinoma (clinical isolates; mechanisms of resistance not fully known), and A2780Tax ovarian carcinoma (tubulin mutation); (b) paclitaxel-insensitive: Pat-26 human pancreatic carcinoma (clinical isolate) and M5076 murine fibrosarcoma; and (c) paclitaxel sensitive: A2780 ovarian, LS174T, and HCT116 human colon carcinoma. In addition, BMS-247550 is p.o. efficacious against preclinical human tumor xenografts grown in immunocompromised mice or rats. Schedule optimization studies indicate that BMS-247550 is efficacious when administered frequently (every 2 days x 5) or intermittently (every 4 days x 3 or every 8 days x 2). These efficacy data demonstrate that BMS-247550 has the potential to surpass Taxol in both clinical efficacy and ease of use (i.e., less frequent treatment schedule and/or oral administration).


Assuntos
Antineoplásicos/farmacologia , Epotilonas , Compostos de Epóxi/farmacologia , Paclitaxel/farmacologia , Tiazóis/farmacologia , Administração Oral , Animais , Antineoplásicos/uso terapêutico , Neoplasias da Mama/tratamento farmacológico , Ciclo Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Neoplasias do Colo/tratamento farmacológico , Modelos Animais de Doenças , Resistencia a Medicamentos Antineoplásicos , Compostos de Epóxi/química , Compostos de Epóxi/uso terapêutico , Feminino , Humanos , Infusões Parenterais , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBA , Transplante de Neoplasias , Neoplasias Ovarianas/tratamento farmacológico , Paclitaxel/uso terapêutico , Neoplasias Pancreáticas/tratamento farmacológico , Sarcoma/tratamento farmacológico , Tiazóis/química , Tiazóis/uso terapêutico , Tubulina (Proteína)/genética , Tubulina (Proteína)/metabolismo , Células Tumorais Cultivadas , Ensaio Tumoral de Célula-Tronco , Ensaios Antitumorais Modelo de Xenoenxerto
9.
Int J Radiat Oncol Biol Phys ; 20(2): 361-7, 1991 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-1671383

RESUMO

Tumor cells exposed in tissue culture to one of several different classes of antineoplastic agents, including anthracyclines, vinca alkaloids, epipodophyllotoxins, and certain antitumor antibiotics, can develop resistance to the selecting agent and cross resistance to the other classes of agents. This phenomena of multidrug resistance is generally associated with decreased drug accumulation and overexpression of a membrane glycoprotein. This membrane protein, referred to as P-glycoprotein, apparently acts as an energy-dependent drug efflux pump. Multidrug resistance in human MCF-7 breast cancer cells selected for resistance to adriamycin (AdrR MCF-7) is associated with amplification and overexpression of the mdr1 gene which encodes P-glycoprotein. A number of other changes are also seen in this resistant cell line including alterations in Phase I and Phase II drug metabolizing enzymes. Similar biochemical changes occur in a rat model for hepatocellular carcinogenesis and are associated in that system with broad spectrum resistance to hepatotoxins. The similar changes in these two models of resistance suggests that these changes might be part of a battery of genes whose expression can be altered in response to cytotoxic stress, thus rendering the cell resistant to a wide variety of cytotoxic agents.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Neoplasias/tratamento farmacológico , Membro 1 da Subfamília B de Cassetes de Ligação de ATP , Animais , Resistência a Medicamentos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Glutationa Transferase/metabolismo , Humanos , Glicoproteínas de Membrana/genética , Neoplasias/enzimologia , Neoplasias/genética , Células Tumorais Cultivadas/efeitos dos fármacos , Células Tumorais Cultivadas/fisiologia
10.
J Med Chem ; 41(20): 3909-11, 1998 Sep 24.
Artigo em Inglês | MEDLINE | ID: mdl-9748366

RESUMO

The known 2-aminoimidazole alkaloid naamidine A (1) was isolated from a Fijian Leucetta sp. sponge as an inhibitor of the epidermal growth factor (EGF) receptor. The compound exhibited potent ability to inhibit the EGF signaling pathway and is more specific for the EGF-mediated mitogenic response than for the insulin-mediated mitogenic response. Evaluation in an A431 xenograft tumor model in athymic mice indicated that naamidine A exhibited at least 85% growth inhibition at the maximal tolerated dose of 25 mg/kg. Preliminary mechanism of action studies indicate that the alkaloid fails to inhibit the binding of EGF to the receptor and has no effect on the catalytic activity of purified c-src tyrosine kinase.


Assuntos
Alcaloides/farmacologia , Antineoplásicos/farmacologia , Receptores ErbB/antagonistas & inibidores , Imidazóis/farmacologia , Células 3T3 , Alcaloides/isolamento & purificação , Animais , Antineoplásicos/isolamento & purificação , Proteína Tirosina Quinase CSK , Carcinoma de Células Escamosas/patologia , Divisão Celular/efeitos dos fármacos , Inibidores Enzimáticos/isolamento & purificação , Inibidores Enzimáticos/farmacologia , Fator de Crescimento Epidérmico/metabolismo , Receptores ErbB/metabolismo , Imidazóis/isolamento & purificação , Camundongos , Camundongos Nus , Transplante de Neoplasias , Poríferos/química , Proteínas Tirosina Quinases/antagonistas & inibidores , Transplante Heterólogo , Quinases da Família src
11.
J Med Chem ; 44(26): 4577-83, 2001 Dec 20.
Artigo em Inglês | MEDLINE | ID: mdl-11741476

RESUMO

The preparation of C-7 paclitaxel ethers is described. Various substituted ethers were prepared via activation of the corresponding methylthiomethyl ether followed by alcohol addition. Variation of the C-7 ether group as well the 3' side chain position led to the discovery of a novel taxane, BMS-184476 (4), with preclinical antitumor activity superior to paclitaxel.


Assuntos
Antineoplásicos/síntese química , Paclitaxel/análogos & derivados , Paclitaxel/síntese química , Taxoides , Antineoplásicos/química , Antineoplásicos/farmacologia , Resistência a Múltiplos Medicamentos , Resistencia a Medicamentos Antineoplásicos , Ensaios de Seleção de Medicamentos Antitumorais , Éteres , Humanos , Espectroscopia de Ressonância Magnética , Paclitaxel/química , Paclitaxel/farmacologia , Relação Estrutura-Atividade , Transplante Heterólogo , Células Tumorais Cultivadas
12.
J Med Chem ; 41(19): 3715-26, 1998 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-9733497

RESUMO

The anticancer drug paclitaxel (Taxol) has been converted to a large number of 2-debenzoyl-2-aroyl derivatives by three different methods. The bioactivities of the resulting analogues were determined in both tubulin polymerization and cytotoxicity assays, and several analogues with enhanced activity as compared with paclitaxel were discovered. Correlation of cytotoxicity in three cell lines with tubulin polymerization activity showed reasonable agreement. Among the cell lines examined, the closest correlation with antitubulin activity was observed with a human ovarian carcinoma cell line.


Assuntos
Antineoplásicos Fitogênicos , Paclitaxel , Antineoplásicos Fitogênicos/síntese química , Antineoplásicos Fitogênicos/química , Antineoplásicos Fitogênicos/farmacologia , Biopolímeros , Catálise , Divisão Celular/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Paclitaxel/análogos & derivados , Paclitaxel/síntese química , Paclitaxel/química , Paclitaxel/farmacologia , Polietilenoglicóis/química , Relação Estrutura-Atividade , Tubulina (Proteína)/metabolismo , Células Tumorais Cultivadas
13.
Biochem Pharmacol ; 35(20): 3533-41, 1986 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-3533081

RESUMO

The delayed cytotoxicity of 6-thioguanine (TG) may relate to the arrest of cells in G2 upon completion of one cell cycle after drug exposure. In Chinese hamster ovary (CHO) cells, both the unilateral chromatid damage in G2 chromosomes, determined by induction of premature condensed chromosome condensation [Maybaum and Mandel, Cancer Res. 43, 3852 (1983)], and incorporation of TG into DNA resulting in DNA strand breakage [Christie et al., Cancer Res. 44, 3665 (1984)] were correlated with cytotoxicity. We have studied the correlation between strand breakage and unilateral chromatid damage in L1210 cells. DNA breaks were detected only when cells were treated with TG (0.25 microM) for one cell cycle time (12 hr) followed by 12 hr in drug-free medium containing [3H]thymidine (TdR) to label the DNA. After simultaneous incubation of cells with drug and label during the first or second 12-hr period, strand breaks were not found. Strand breaks increased with dose, which correlated with greater cytotoxicity (0.01 to 0.25 microM). Treatment of cells with 0.25 microM TG for 12 hr, and transfer to drug-free medium for 12 hr prior to making prematurely condensed chromosomes (PCC), resulted in unilateral chromatid damage. Prominent curving of G2 chromosomes with gapping and diffuse staining of one of the sister chromatids occurred. The 4-fold increase in the percentage of cells in G2 compared with control cells suggested G2 arrest. When cells were treated with TG for 12 hr and PCC made immediately, neither the arrest of cells in G2 nor unilateral chromatid damage was observed. These data suggest that strand breaks and unilateral chromatid damage occur in the second cell cycle after TG exposure and that this damage may be important in TG-delayed cytotoxicity.


Assuntos
Cromátides/efeitos dos fármacos , DNA/efeitos dos fármacos , Conformação de Ácido Nucleico/efeitos dos fármacos , Tioguanina/farmacologia , Animais , Linhagem Celular , Cricetinae , Cricetulus , Concentração de Íons de Hidrogênio , Interfase , Cariotipagem , Leucemia L1210/genética , Fatores de Tempo
14.
Biochem Pharmacol ; 42(2): 391-402, 1991 Jul 05.
Artigo em Inglês | MEDLINE | ID: mdl-1677569

RESUMO

The phenotypic expression of multidrug resistance by the doxorubicin-selected AdrR human breast tumor cell line is associated with overexpression of plasma membrane P-170 glycoprotein and increased cytosolic selenium-dependent GSH-peroxidase activity relative to the parental MCF-7 wild-type line (WT). To determine whether doxorubicin resistance by AdrR cells persists in vivo, and to further investigate the possibility of biochemical differences between WT and AdrR solid tumors, both tumor cell lines were grown as subcutaneous xenografts in athymic nude mice. Tumorigenicity depended upon cell inoculation burden, and tumor incidence was similar for both cell lines (greater than 80% tumor takes at 10(7) cells/mouse) at 14 days, provided 17 beta-estradiol was supplied to the animals bearing the WT tumors. However, the growth rate for the AdrR xenografts was only about half that of WT xenografts. Doxorubicin (2-8 mg/kg, i.p., injected weekly) significantly diminished the growth of the WT tumors, but AdrR solid tumors failed to respond to doxorubicin. The accumulation of 14C-labeled doxorubicin was 2-fold greater in WT xenografts that in AdrR, although there were no differences in host organ drug levels in mice bearing either type of tumors. Membrane P-170 glycoprotein mRNA was detected by slot-blot analysis in the AdrR tumors, but not in WT. Electron spin resonance 5,5-dimethylpyrroline-N-oxide-spin-trapping experiments with microsomes and mitochondria from WT and AdrR xenographs demonstrated a 2-fold greater oxygen radical (superoxide and hydroxyl) formation from activated doxorubicin with WT xenographs compared to AdrR. Selenium-dependent glutathione (GSH)-peroxidase, superoxide dismutase and GSH-S-aryltransferase activities in AdrR xenografts were elevated relative to WT. Although the activities of the latter two enzymes were similar to those measured in both tumor cell lines, GSH-peroxidase activities were elevated 70-fold (WT) and 10-fold (AdrR) in xenografts compared to tumor cells. In contrast, in both WT and AdrR solid tumors in vivo, catalase, NAD(P)H-oxidoreductases, and glutathione disulfide (GSSG)-reductase activities, and GSH and GSSG levels were not markedly different, and were essentially the same as in cells in vitro. Like the MDR cells in culture, AdrR tumor xenografts were extremely resistant to doxorubicin and retained most of the characteristics of the altered phenotype. These results suggest that WT and AdrR breast tumor xenografts provide a useful model for the study of biochemical and pharmacological mechanisms of drug resistance by solid tumors in vivo.


Assuntos
Neoplasias da Mama/tratamento farmacológico , Doxorrubicina/farmacologia , Membro 1 da Subfamília B de Cassetes de Ligação de ATP , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Linhagem Celular/enzimologia , Sistema Enzimático do Citocromo P-450/metabolismo , Doxorrubicina/administração & dosagem , Doxorrubicina/farmacocinética , Resistência a Medicamentos/genética , Tolerância a Medicamentos/genética , Espectroscopia de Ressonância de Spin Eletrônica , Glutationa/metabolismo , Humanos , Hidróxidos/metabolismo , Radical Hidroxila , Glicoproteínas de Membrana/genética , Camundongos , Camundongos Nus , Transplante de Neoplasias , Fenótipo , RNA Mensageiro/análise , Frações Subcelulares/metabolismo , Superóxidos/metabolismo
15.
Org Lett ; 3(17): 2693-6, 2001 Aug 23.
Artigo em Inglês | MEDLINE | ID: mdl-11506611

RESUMO

[reaction: see text]. A series of 12alpha,13alpha-aziridinyl epothilone derivatives were synthesized in an efficient manner from epothilone A. The final semisynthetic route involves a formal double-inversion of stereochemistry at both the C12 and C13 positions. All aziridine analogues were tested for effects on tubulin binding polymerization and cytotoxicity. The results indicate that the aziridine moiety is a viable isosteric replacement for the epoxide in the case of epothilones.


Assuntos
Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Aziridinas/síntese química , Aziridinas/farmacologia , Compostos de Epóxi/química , Antineoplásicos/química , Aziridinas/química , Relação Estrutura-Atividade
16.
Cancer Chemother Pharmacol ; 22(1): 26-32, 1988.
Artigo em Inglês | MEDLINE | ID: mdl-3396144

RESUMO

By inhibiting dihydrofolate reductase, methotrexate (MTX) depletes cellular stores of reduced folates, resulting in the inhibition of DNA and RNA synthesis. Inhibition of RNA synthesis arrests cells in the G1 phase of the cell cycle, preventing these cells from entering S phase and rendering them insensitive to MTX. Because MTX cytotoxicity can be enhanced by concurrent administration of hypoxanthine (HX), we examined the hypothesis that this modulation can allow normal rates of RNA synthesis and cell cycle progression from G1 to S phase. For L1210 cells exposed to MTX for 12 h or 24 h, the addition of HX enhanced the cytotoxicity of MTX; however, no enhancement was observed with a 6-h exposure. Inhibition of RNA synthesis by MTX was prevented by concurrent administration of HX. The effect of HX on cell cycle progression was first examined using flow cytometry, which indicated that MTX treatment alone or with concurrent HX caused a buildup of cells with a G1 content of DNA. Because this technique may fail to distinguish between cells in late G1 phase, the G1/S border, or early S, the method of premature chromosome condensation was used to determine cell cycle position based on chromatin morphology. A shift to a higher degree of chromatin decondensation was observed when HX was coadministered with MTX during a 12-h exposure, suggesting progression from G1 towards S. This correlated with the enhancement of MTX cytotoxicity by HX after 12 h exposure. The results of these studies suggest that HX potentiates MTX cytotoxicity by maintaining RNA synthesis, allowing cells that might ordinarily be arrested in G1 to progress into the cytotoxic S phase.


Assuntos
Hipoxantinas/farmacologia , Leucemia L1210/metabolismo , Metotrexato/farmacologia , Animais , Ciclo Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Cromossomos/efeitos dos fármacos , DNA de Neoplasias/biossíntese , Sinergismo Farmacológico , Citometria de Fluxo , Hipoxantina , RNA Neoplásico/biossíntese
18.
Mol Pharmacol ; 37(6): 801-9, 1990 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-1972772

RESUMO

The nucleotide sequence of the mdr1 gene encoding a putative drug efflux pump (P-glycoprotein) is homologous to a class of bacterial membrane-associated transport proteins. These bacterial proteins are part of a multicomponent system that includes soluble periplasmic proteins that bind substrates, channeling them through the membrane in an energy-dependent manner. We have investigated the possibility that a similar multicomponent transport system exists in a multidrug-resistant human MCF-7 breast cancer cell line that was initially selected for resistance to doxorubicin (AdrR MCF-7). AdrR MCF-7 cells overexpress both the mdr1 gene and the pi class isozyme of glutathione S-transferase (GST-pi) (EC 2.5.1.18). The latter is one of several isozymes known to have a ligand-binding function in addition to drug-metabolizing capabilities. Although we have recently shown that transfection of a functional GST-pi expression vector is insufficient to confer resistance to doxorubicin in cells that lack P-glycoprotein expression [Mol. Pharmacol. 36:22-28 (1989)], we examined the possibility that GST-pi interacts with P-glycoprotein to alter multidrug resistance. To do this, we have cloned cDNAs encoding these proteins from AdrR MCF-7 cells, constructed expression vectors containing these two genes, and transfected these vectors sequentially into drug-sensitive MCF-7 cells. The human mdr1 cDNA isolated from AdrR MCF-7 is a variant gene whose sequence differs from that isolated previously from vinblastine-resistant KB cells [Cell 53:519-529 (1989)], resulting in an amino acid substitution of alanine to serine at position 893 (mdr1/893ala). Transfection of eukaryotic expression vectors containing the mdr1 gene isolated from AdrR MCF-7 cells produced a multidrug-resistant phenotype in recipient cells, with a cross-resistance pattern similar to that in the AdrR MCF-7 cells. To determine whether GST-pi expression could augment resistance provided by mdr1, two clones transfected with mdr1, one with high levels (153% of mdr1 RNA in AdR MCF-7 cells) and one with low levels (10% of mdr1 RNA in AdrR MCF-7 cells), were subsequently cotransfected with a GST-pi expression vector and pSVNeo and selected for resistance to G418. Six of these clones contained levels of GST-pi that were 8- to 18-fold greater than GST levels found in mdr1-expressing clones transfected with nonspecific DNA. We found no difference in the degree of resistance to doxorubicin, actinomycin D, and vinblastine between the clones expressing mdr1 only and the clones expressing both mdr1 and GST-pi.(ABSTRACT TRUNCATED AT 400 WORDS)


Assuntos
Glutationa Transferase/genética , Glicoproteínas de Membrana/genética , Membro 1 da Subfamília B de Cassetes de Ligação de ATP , Animais , Linhagem Celular , DNA/genética , Resistência a Medicamentos , Expressão Gênica , Variação Genética , Glutationa Transferase/biossíntese , Humanos , Glicoproteínas de Membrana/biossíntese , RNA Mensageiro/metabolismo , Ratos , Transfecção
19.
J Biol Chem ; 268(11): 8290-7, 1993 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-8096520

RESUMO

The human mdr1 gene encodes a putative drug efflux pump (P-glycoprotein) whose overexpression is associated with the development of multidrug resistance (MDR). The promoter and 5'-flanking DNA of this gene were isolated from a human genomic DNA library and used to prepare a series of chloramphenicol acetyltransferase (CAT) fusion vectors under the transcriptional control of the mdr1 promoter (mdrCAT vectors). Transient transfection of these mdrCAT vectors produced CAT activities similar to those produced by transfection of CAT vectors containing viral promoters. The regulation of mdr1 expression was examined in two MDR tumor cell lines selected for resistance to doxorubicin and their corresponding parental cell lines. Although nuclear run-on analysis indicates that the expression of the mdr1 gene in these two MDR cell lines is regulated by transcriptional mechanisms, mdrCAT expression was not significantly increased in either of these lines relative to parental cells. Thus, the sequences involved in the transcriptional regulation in these cells are apparently not included in the constructs studied (-4741 to +286). Analyses of a series of deletion constructs show that the basal mdr1 promoter activity is encoded by sequences that span a region adjacent to the transcription start site (-134 to +286) and that sequences 3' to the start of mdr1 transcription are necessary for proper initiation of transcription in vivo. Structural and functional studies indicate that an initiator (Inr) sequence surrounding the major transcription start site governs accurate initiation of mdr1 transcription.


Assuntos
Resistência a Medicamentos/genética , Regulação da Expressão Gênica , Glicoproteínas de Membrana/genética , Regiões Promotoras Genéticas , Sequências Reguladoras de Ácido Nucleico , Transcrição Gênica , Membro 1 da Subfamília B de Cassetes de Ligação de ATP , Animais , Sequência de Bases , Linhagem Celular , Cloranfenicol O-Acetiltransferase/genética , Cloranfenicol O-Acetiltransferase/metabolismo , Células HeLa , Humanos , Dados de Sequência Molecular , Oligodesoxirribonucleotídeos , Reação em Cadeia da Polimerase/métodos , Proteínas Recombinantes/metabolismo , Deleção de Sequência , Homologia de Sequência do Ácido Nucleico , Transfecção
20.
Cancer Commun ; 3(6): 181-9, 1991 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-1675575

RESUMO

Cross-resistance to anticancer drugs, termed multidrug resistance (mdr), has been functionally associated with the expression of a plasma membrane energy-dependent efflux pump, termed P-glycoprotein, the product of the mdr1 gene. When MCF-7 breast carcinoma cells were transfected with the human mdr1 gene (BC-19 cells), they expressed levels of P-glycoprotein equivalent to those of cells selected for resistance to doxorubicin (MCF-7/ADR) but exhibited 10- to 50-fold less resistance to doxorubicin and vinblastine. We have now demonstrated that when BC-19 cells were stably transfected with protein kinase C alpha (PKC alpha), resistance to doxorubicin and vinblastine was increased; wild-type MCF-7 cells transfected with PKC alpha did not exhibit any change in drug resistance. Increased resistance in PKC alpha-transfected BC-19 cells was associated with enhanced PKC activity and phosphorylation of P-glycoprotein and decreased drug accumulation. The PKC activator, phorbol dibutyrate, further increased resistance to doxorubicin and stimulated P-glycoprotein phosphorylation. These results demonstrate that transfection of P-glycoprotein-expressing cells with PKC resulted in increased mdr and that PKC may have served as an important modulator of this process.


Assuntos
Glicoproteínas de Membrana/metabolismo , Proteína Quinase C/metabolismo , Transfecção , Membro 1 da Subfamília B de Cassetes de Ligação de ATP , Linhagem Celular , Doxorrubicina/metabolismo , Resistência a Medicamentos/genética , Fosforilação , Ensaio Tumoral de Célula-Tronco
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA