RESUMEN
Microvesicular steatosis is an important component of the overall pathogenesis of drug-mediated liver injury. Although mitochondrial damage has a role in the development of microvesicular steatosis, the consequences of fatty change for hepatic gene function are unclear. The present study was undertaken to evaluate hepatic cytochrome P-450 (CYP) function in a rat model of microvesicular steatosis produced by the intake of diets containing 1% orotic acid (OA) that were administered for 5, 10, or 21 days. Hepatic triglyceride levels were increased to 3-fold of control after 5 days and were elevated further at 10 and 21 days. Cholesterol and phospholipid contents were increased after 10 and 21 days but not by 5 days of feeding. Microsomal androst-4-ene-3,17-dione hydroxylation activities mediated by CYP2C11 (16alpha-hydroxylation) and CYP3A2 (6beta-hydroxylation) were decreased in liver from OA-fed rats for only 5 days, whereas CYP2A1/2-mediated steroid 7alpha-hydroxylation was decreased after 10 days; these observations were complemented by immunoblot analysis that demonstrated the impaired expression of the corresponding CYP proteins. CYP2C11 mRNA, the major CYP in male rat liver, was down-regulated in steatotic liver to 52 +/- 4% of control. Thus, microvesicular steatosis induced by short-term intake of OA-containing diets is histologically similar to that produced by hepatotoxic drugs and produces the rapid down-regulation of constitutive CYPs in rat liver. Analogous processes of lipid deposition in human liver after drug- or disease-related injury could precipitate adverse effects during subsequent drug therapy.
Asunto(s)
Hidrocarburo de Aril Hidroxilasas , Sistema Enzimático del Citocromo P-450/biosíntesis , Regulación hacia Abajo/efectos de los fármacos , Hígado Graso/inducido químicamente , Hígado Graso/enzimología , Microsomas Hepáticos/enzimología , Ácido Orótico , Esteroide 16-alfa-Hidroxilasa , Androstenodiona/metabolismo , Animales , Peso Corporal , Citocromo P-450 CYP2C8 , Citocromo P-450 CYP2C9 , Citocromo P-450 CYP3A , Familia 2 del Citocromo P450 , Dieta , Hidroxilación , Metabolismo de los Lípidos , Hígado/efectos de los fármacos , Hígado/metabolismo , Masculino , Proteínas de la Membrana , Microcirculación/enzimología , Microcirculación/patología , Microsomas Hepáticos/efectos de los fármacos , Microsomas Hepáticos/ultraestructura , NADPH-Ferrihemoproteína Reductasa/metabolismo , Sondas de Oligonucleótidos/farmacología , ARN Mensajero/biosíntesis , Ratas , Ratas Wistar , Esteroide Hidroxilasas/biosíntesis , Esteroides/metabolismoRESUMEN
The H82 "variant" and the H69 "classic" small cell lung cancer (SCLC) cell lines were treated with low levels of epirubicin (69 and 14 nM) which caused little cell death but produced the H82/E8 and H69/E8 extended-multidrug resistant sublines. Both were resistant to drugs associated with multidrug resistance (MDR), and to chlorambucil (9.5- and 5.6-fold, respectively) and cisplatin (2.3- and 8.5-fold, respectively). There was increased expression of the multidrug resistance-associated protein (MRP1) in the H82/E8 subline while P-glycoprotein expression was not detected in any cells or sublines. Treatment of the H82 cells for 1 hr with 69 nM epirubicin increased MRP1-mRNA expression within 4 hr and this was associated with an increase in the resistance to epirubicin, chlorambucil, cisplatin and paclitaxel. Further, a 1 hr treatment with non-cytotoxic doses of chlorambucil (2.5 microM), cisplatin (1.3 microM) or paclitaxel (13 nM), drugs not normally associated with MRP1-mediated MDR, also increased MRP1-mRNA expression in the H82 cells with paclitaxel causing the highest increase (4.5-fold). For chlorambucil treatment, this increased MRPI-mRNA expression was accompanied by increased drug resistance while paclitaxel treatment had no effect on drug resistance in the H82 cells. For the drug resistant H82/E8 subline, these drug treatments had no effect on the MRP1-mRNA expression and little effect on increasing the subline drug resistance. However, pretreatment with paclitaxel sensitised the H82/E8 subline to chlorambucil and cisplatin returning the subline to the sensitivity of the H82 cell line. We conclude that treatment with low levels of MDR and non-MDR drugs can induce extended-multidrug resistance in SCLC cells, a process that probably involves the co-ordinate upregulation of MRP1 and other resistance mechanisms. The results also suggest paclitaxel may have a role as a response modifier in the treatment of refractory SCLC.
Asunto(s)
Antineoplásicos Fitogénicos/farmacología , Proteínas Bacterianas , Carcinoma de Células Pequeñas/tratamiento farmacológico , Resistencia a Múltiples Medicamentos , Neoplasias Pulmonares/tratamiento farmacológico , Paclitaxel/farmacología , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/biosíntesis , Carcinoma de Células Pequeñas/metabolismo , Clorambucilo/farmacología , Cisplatino/farmacología , Interacciones Farmacológicas , Resistencia a Antineoplásicos , Ensayos de Selección de Medicamentos Antitumorales , Etopósido/farmacología , Humanos , Neoplasias Pulmonares/metabolismo , ARN Mensajero/metabolismo , Serina Endopeptidasas/biosíntesis , Células Tumorales Cultivadas/efectos de los fármacos , Vinblastina/farmacologíaRESUMEN
The effectiveness of N-[2-(dimethylamino)ethyl]acridine-4-carboxamide (DACA) relative to that of amsacrine, idarubicin, daunorubicin and paclitaxel against three different forms of multidrug resistance (MDR) was determined using two sublines of the CCRF-CEM human leukaemia cell line, the P-glyco-protein-expressing CEM/VLB100 subline and the MRP-expressing CEM/E1000 subline, and two extended-MDR sublines of the HL60 human leukaemia cell line, HL60/E8 and HL60/V8. DACA was effective against P-glycoprotein-mediated MDR and MRP-mediated MDR, whereas the extended-MDR phenotype showed only low levels of resistance (< 2-fold) to DACA. In comparison, idarubicin was ineffective against the MRP and extended-MDR phenotypes. Repeated exposure of the K562 human leukaemia cell line to DACA (55, 546 or 1092 nM for 3 days over 10 weeks) did not result in the development of any significant drug resistance. We conclude that DACA has the potential to treat refractory leukaemia.
Asunto(s)
Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/biosíntesis , Acridinas/toxicidad , Antineoplásicos/toxicidad , Supervivencia Celular/efectos de los fármacos , Resistencia a Múltiples Medicamentos , Amsacrina/toxicidad , Butionina Sulfoximina/farmacología , Línea Celular , Daunorrubicina/toxicidad , Células HL-60 , Humanos , Idarrubicina/toxicidad , Leucemia , Paclitaxel/toxicidad , Fenotipo , ARN Mensajero/biosíntesis , Transcripción Genética , Células Tumorales Cultivadas , Verapamilo/farmacologíaRESUMEN
The development of drug resistance was studied in a series of haemopoietic cells to determine its relationship to cell lineage. Treatment of the U937 monocytic cell line with epirubicin (15 ng/ml) or vinblastine (8 ng/ml) induced drug-resistant sublines with cross-resistance to epirubicin (8- and 16-fold respectively), vinblastine (5- and 20-fold), paclitaxel (15- and 42-fold) and etoposide (19- and 13-fold). However, sublines were also 3-5-fold resistant to the alkylating agent chlorambucil, cis-platinum and methotrexate, demonstrating an extended multidrug resistance (MDR) phenotype. These cells over-expressed P-glycoprotein, but decreased drug accumulation was not restored in the presence of verapamil, suggesting that the P-glycoprotein was not functional. Similar drug treatment of the HL60 promyelocytic cell line also produced sublines exhibiting an extended MDR phenotype. The KG1a and the HEL cell lines expressed functional P-glycoprotein and were resistant to the drug concentrations used for treatment. Multidrug resistance as mediated by P-glycoprotein cannot explain the resistance of CML patients to chemotherapy, especially in blast crisis. The induction of an extended MDR phenotype specifically in myeloid cells in response to drug treatment may explain the resistance observed in the treatment of CMI.
Asunto(s)
Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/metabolismo , Epirrubicina/uso terapéutico , Etopósido/uso terapéutico , Células Madre Hematopoyéticas/patología , Leucemia Mieloide/tratamiento farmacológico , Paclitaxel/uso terapéutico , Vinblastina/uso terapéutico , Western Blotting , Resistencia a Múltiples Medicamentos , Resistencia a Antineoplásicos , Células HL-60/metabolismo , Células HL-60/patología , Células Madre Hematopoyéticas/efectos de los fármacos , Humanos , Leucemia Mieloide/metabolismo , Leucemia Mieloide/patología , Células Tumorales CultivadasRESUMEN
In an attempt to mimic clinical conditions for the treatment of leukaemia, the HL60 promyelocytic cell line was treated for 18 h with low, clinically relevant, levels of the anthracycline epirubicin and the Vinca alkaloid vinblastine. The resulting drug-resistant sublines not only expressed P-glycoprotein and the MDR phenotype but were also cross-resistant to chlorambucil, methotrexate and cisplatinum, and had increased resistance to radiation. Development of resistance was associated with an aberrant differentiation phenotype with decreased expression of myeloid antigens and expression of glycophorin A, an antigen normally associated with erythroid differentiation. The ability of HL60 cells to terminally differentiate in response to all-trans-retinoic acid (vitamin A acid) was lost in the sublines. These results suggest that either a single novel mechanism is responsible for multiple drug resistance or the initial response to drug treatment is the co-induction of multiple mechanisms. These cells and the method by which they were generated therefore provide a clinically relevant model for the study of the initial events in the development of not only multidrug resistance but also the extended multiple drug resistance usually encountered in the treatment of leukaemia.