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1.
Mol Pharmacol ; 89(5): 492-504, 2016 May.
Artigo em Inglês | MEDLINE | ID: mdl-26893303

RESUMO

Breast cancer resistance protein (BCRP) is expressed in various tissues, such as the gut, liver, kidney and blood brain barrier (BBB), where it mediates the unidirectional transport of substrates to the apical/luminal side of polarized cells. Thereby BCRP acts as an efflux pump, mediating the elimination or restricting the entry of endogenous compounds or xenobiotics into tissues and it plays important roles in drug disposition, efficacy and safety. Bcrp knockout mice (Bcrp(-/-)) have been used widely to study the role of this transporter in limiting intestinal absorption and brain penetration of substrate compounds. Here we describe the first generation and characterization of a mouse line humanized for BCRP (hBCRP), in which the mouse coding sequence from the start to stop codon was replaced with the corresponding human genomic region, such that the human transporter is expressed under control of the murineBcrppromoter. We demonstrate robust human and loss of mouse BCRP/Bcrp mRNA and protein expression in the hBCRP mice and the absence of major compensatory changes in the expression of other genes involved in drug metabolism and disposition. Pharmacokinetic and brain distribution studies with several BCRP probe substrates confirmed the functional activity of the human transporter in these mice. Furthermore, we provide practical examples for the use of hBCRP mice to study drug-drug interactions (DDIs). The hBCRP mouse is a promising model to study the in vivo role of human BCRP in limiting absorption and BBB penetration of substrate compounds and to investigate clinically relevant DDIs involving BCRP.


Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Proteínas de Neoplasias/metabolismo , Xenobióticos/farmacocinética , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/antagonistas & inibidores , Transportadores de Cassetes de Ligação de ATP/química , Transportadores de Cassetes de Ligação de ATP/genética , Animais , Disponibilidade Biológica , Biotransformação/efeitos dos fármacos , Barreira Hematoencefálica/efeitos dos fármacos , Barreira Hematoencefálica/metabolismo , Interações Medicamentosas , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Técnicas de Introdução de Genes , Humanos , Absorção Intestinal/efeitos dos fármacos , Masculino , Moduladores de Transporte de Membrana/sangue , Moduladores de Transporte de Membrana/metabolismo , Moduladores de Transporte de Membrana/farmacocinética , Moduladores de Transporte de Membrana/farmacologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/química , Proteínas de Neoplasias/genética , Regiões Promotoras Genéticas/efeitos dos fármacos , RNA Mensageiro/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Distribuição Tecidual/efeitos dos fármacos , Xenobióticos/sangue , Xenobióticos/metabolismo , Xenobióticos/farmacologia
2.
Drug Metab Dispos ; 43(11): 1679-90, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-26265742

RESUMO

Variability in drug pharmacokinetics is a major factor in defining drug efficacy and side effects. There remains an urgent need, particularly with the growing use of polypharmacy, to obtain more informative experimental data predicting clinical outcomes. Major species differences in multiplicity, substrate specificity, and regulation of enzymes from the cytochrome P450-dependent mono-oxygenase system play a critical role in drug metabolism. To develop an in vivo model for predicting human responses to drugs, we generated a mouse, where 31 P450 genes from the Cyp2c, Cyp2d, and Cyp3a gene families were exchanged for their relevant human counterparts. The model has been improved through additional humanization for the nuclear receptors constitutive androgen receptor and pregnane X receptor that control the expression of key drug metabolizing enzymes and transporters. In this most complex humanized mouse model reported to date, the cytochromes P450 function as predicted and we illustrate how these mice can be applied to predict drug-drug interactions in humans.


Assuntos
Citocromo P-450 CYP3A/metabolismo , Modelos Animais , Preparações Farmacêuticas/metabolismo , Transdução de Sinais/fisiologia , Animais , Citocromo P-450 CYP3A/genética , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Microssomos Hepáticos/efeitos dos fármacos , Microssomos Hepáticos/enzimologia , Microssomos Hepáticos/metabolismo , Preparações Farmacêuticas/administração & dosagem , Transdução de Sinais/efeitos dos fármacos
3.
Drug Metab Dispos ; 42(6): 1022-30, 2014 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-24671958

RESUMO

In humans, 75% of all drugs are metabolized by the cytochrome P450-dependent monooxygenase system. Enzymes encoded by the CYP2C, CYP2D, and CYP3A gene clusters account for ∼80% of this activity. There are profound species differences in the multiplicity of cytochrome P450 enzymes, and the use of mouse models to predict pathways of drug metabolism is further complicated by overlapping substrate specificity between enzymes from different gene families. To establish the role of the hepatic and extrahepatic P450 system in drug and foreign chemical disposition, drug efficacy, and toxicity, we created a unique mouse model in which 30 cytochrome P450 genes from the Cyp2c, Cyp2d, and Cyp3a gene clusters have been deleted. Remarkably, despite a wide range of putative important endogenous functions, Cyp2c/2d/3a KO mice were viable and fertile, demonstrating that these genes have evolved primarily as detoxification enzymes. Although there was no overt phenotype, detailed examination showed Cyp2c/2d/3a KO mice had a smaller body size (15%) and larger livers (20%). Changes in hepatic morphology and a decreased blood glucose (30%) were also noted. A five-drug cocktail of cytochrome P450 isozyme probe substrates were used to evaluate changes in drug pharmacokinetics; marked changes were observed in either the pharmacokinetics or metabolites formed from Cyp2c, Cyp2d, and Cyp3a substrates, whereas the metabolism of the Cyp1a substrate caffeine was unchanged. Thus, Cyp2c/2d/3a KO mice provide a powerful model to study the in vivo role of the P450 system in drug metabolism and efficacy, as well as in chemical toxicity.


Assuntos
Sistema Enzimático do Citocromo P-450/genética , Sistema Enzimático do Citocromo P-450/metabolismo , Deleção de Genes , Intestino Delgado/enzimologia , Microssomos Hepáticos/enzimologia , Preparações Farmacêuticas/metabolismo , Animais , Intestino Delgado/efeitos dos fármacos , Masculino , Camundongos , Camundongos Knockout , Microssomos Hepáticos/efeitos dos fármacos , Preparações Farmacêuticas/administração & dosagem
4.
Drug Metab Dispos ; 42(8): 1301-13, 2014 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24855184

RESUMO

Organic anion transporting polypeptide (Oatp) 1a/1b knockout and OATP1B1 and -1B3 humanized mouse models are promising tools for studying the roles of these transporters in drug disposition. Detailed characterization of these models will help to better understand their utility for predicting clinical outcomes. To advance this approach, we carried out a comprehensive analysis of these mouse lines by evaluating the compensatory changes in mRNA expression, quantifying the amounts of OATP1B1 and -1B3 protein by liquid chromatography-tandem mass spectrometry, and studying the active uptake in isolated hepatocytes and the pharmacokinetics of some prototypical substrates including statins. Major outcomes from these studies were 1) mostly moderate compensatory changes in only a few genes involved in drug metabolism and disposition, 2) a robust hepatic expression of OATP1B1 and -1B3 proteins in the respective humanized mouse models, and 3) functional activities of the human transporters in hepatocytes isolated from the humanized models with several substrates tested in vitro and with pravastatin in vivo. However, the expression of OATP1B1 and -1B3 in the humanized models did not significantly alter liver or plasma concentrations of rosuvastatin and pitavastatin compared with Oatp1a/1b knockout controls under the conditions used in our studies. Hence, although the humanized OATP1B1 and -1B3 mice showed in vitro and/or in vivo functional activity with some statins, further characterization of these models is required to define their potential use and limitations in the prediction of drug disposition and drug-drug interactions in humans.


Assuntos
Inibidores de Hidroximetilglutaril-CoA Redutases/metabolismo , Transportadores de Ânions Orgânicos Sódio-Independentes/metabolismo , Transportadores de Ânions Orgânicos/metabolismo , Animais , Interações Medicamentosas/fisiologia , Fluorbenzenos/metabolismo , Hepatócitos/metabolismo , Humanos , Fígado/enzimologia , Fígado/metabolismo , Transportador 1 de Ânion Orgânico Específico do Fígado , Masculino , Camundongos , Pravastatina/metabolismo , Pirimidinas/metabolismo , RNA Mensageiro/genética , Rosuvastatina Cálcica , Membro 1B3 da Família de Transportadores de Ânion Orgânico Carreador de Soluto , Sulfonamidas/metabolismo
5.
Nat Med ; 29(4): 859-868, 2023 04.
Artigo em Inglês | MEDLINE | ID: mdl-36928816

RESUMO

One of the great challenges in therapeutic oncology is determining who might achieve survival benefits from a particular therapy. Studies on longitudinal circulating tumor DNA (ctDNA) dynamics for the prediction of survival have generally been small or nonrandomized. We assessed ctDNA across 5 time points in 466 non-small-cell lung cancer (NSCLC) patients from the randomized phase 3 IMpower150 study comparing chemotherapy-immune checkpoint inhibitor (chemo-ICI) combinations and used machine learning to jointly model multiple ctDNA metrics to predict overall survival (OS). ctDNA assessments through cycle 3 day 1 of treatment enabled risk stratification of patients with stable disease (hazard ratio (HR) = 3.2 (2.0-5.3), P < 0.001; median 7.1 versus 22.3 months for high- versus low-intermediate risk) and with partial response (HR = 3.3 (1.7-6.4), P < 0.001; median 8.8 versus 28.6 months). The model also identified high-risk patients in an external validation cohort from the randomized phase 3 OAK study of ICI versus chemo in NSCLC (OS HR = 3.73 (1.83-7.60), P = 0.00012). Simulations of clinical trial scenarios employing our ctDNA model suggested that early ctDNA testing outperforms early radiographic imaging for predicting trial outcomes. Overall, measuring ctDNA dynamics during treatment can improve patient risk stratification and may allow early differentiation between competing therapies during clinical trials.


Assuntos
Carcinoma Pulmonar de Células não Pequenas , DNA Tumoral Circulante , Neoplasias Pulmonares , Humanos , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , DNA Tumoral Circulante/genética , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Biomarcadores Tumorais/genética
6.
Mol Pharmacol ; 82(6): 1022-9, 2012 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-22918969

RESUMO

Compared with rodents and many other animal species, the human cytochrome P450 (P450) Cyp2c gene cluster varies significantly in the multiplicity of functional genes and in the substrate specificity of its enzymes. As a consequence, the use of wild-type animal models to predict the role of human CYP2C enzymes in drug metabolism and drug-drug interactions is limited. Within the human CYP2C cluster CYP2C9 is of particular importance, because it is one of the most abundant P450 enzymes in human liver, and it is involved in the metabolism of a wide variety of important drugs and environmental chemicals. To investigate the in vivo functions of cytochrome P450 Cyp2c genes and to establish a model for studying the functions of CYP2C9 in vivo, we have generated a mouse model with a deletion of the murine Cyp2c gene cluster and a corresponding humanized model expressing CYP2C9 specifically in the liver. Despite the high number of functional genes in the mouse Cyp2c cluster and the reported roles of some of these proteins in different biological processes, mice deleted for Cyp2c genes were viable and fertile but showed certain phenotypic alterations in the liver. The expression of CYP2C9 in the liver also resulted in viable animals active in the metabolism and disposition of a number of CYP2C9 substrates. These mouse lines provide a powerful tool for studying the role of Cyp2c genes and of CYP2C9 in particular in drug disposition and as a factor in drug-drug interaction.


Assuntos
Hidrocarboneto de Aril Hidroxilases/genética , Sistema Enzimático do Citocromo P-450/genética , Família Multigênica/genética , Animais , Hidrocarboneto de Aril Hidroxilases/metabolismo , Citocromo P-450 CYP2C9 , Inibidores das Enzimas do Citocromo P-450 , Sistema Enzimático do Citocromo P-450/metabolismo , Humanos , Hidroxilação , Inativação Metabólica , Fígado/efeitos dos fármacos , Fígado/enzimologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Família Multigênica/efeitos dos fármacos , Tolbutamida/metabolismo , Tolbutamida/farmacocinética
7.
Mol Pharmacol ; 81(1): 63-72, 2012 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-21989258

RESUMO

The highly polymorphic human cytochrome P450 2D6 enzyme is involved in the metabolism of up to 25% of all marketed drugs and accounts for significant individual differences in response to CYP2D6 substrates. Because of the differences in the multiplicity and substrate specificity of CYP2D family members among species, it is difficult to predict pathways of human CYP2D6-dependent drug metabolism on the basis of animal studies. To create animal models that reflect the human situation more closely and that allow an in vivo assessment of the consequences of differential CYP2D6 drug metabolism, we have developed a novel straightforward approach to delete the entire murine Cyp2d gene cluster and replace it with allelic variants of human CYP2D6. By using this approach, we have generated mouse lines expressing the two frequent human protein isoforms CYP2D6.1 and CYP2D6.2 and an as yet undescribed variant of this enzyme, as well as a Cyp2d cluster knockout mouse. We demonstrate that the various transgenic mouse lines cover a wide spectrum of different human CYP2D6 metabolizer phenotypes. The novel humanization strategy described here provides a robust approach for the expression of different CYP2D6 allelic variants in transgenic mice and thus can help to evaluate potential CYP2D6-dependent interindividual differences in drug response in the context of personalized medicine.


Assuntos
Citocromo P-450 CYP2D6/metabolismo , Células-Tronco Embrionárias/metabolismo , Modelos Animais , Alelos , Animais , Linhagem Celular , Interações Medicamentosas/fisiologia , Células-Tronco Embrionárias/enzimologia , Variação Genética , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos
8.
Mol Pharmacol ; 80(3): 518-28, 2011 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-21628639

RESUMO

Cytochrome P450 (P450) 3A4 is the predominant P450 enzyme expressed in human liver and intestine, and it is involved in the metabolism of approximately 50% of clinically used drugs. Because of the differences in the multiplicity of CYP3A genes and the poor correlation of substrate specificity of CYP3A proteins between species, the extrapolation of CYP3A-mediated metabolism of a drug from animals to man is difficult. This situation is further complicated by the fact that the predictability of the clinically common drug-drug interaction of pregnane X receptor (PXR)-mediated CYP3A4 induction by animal studies is limited as a result of marked species differences in the interaction of many drugs with this receptor. Here we describe a novel multiple humanized mouse line that combines a humanization for PXR, the closely related constitutive androstane receptor, and a replacement of the mouse Cyp3a cluster with a large human genomic region carrying CYP3A4 and CYP3A7. We provide evidence that this model shows a human-like CYP3A4 induction response to different PXR activators, that it allows the ranking of these activators according to their potency to induce CYP3A4 expression in the human liver, and that it provides an experimental approach to quantitatively predict PXR/CYP3A4-mediated drug-drug interactions in humans.


Assuntos
Citocromo P-450 CYP3A/metabolismo , Receptores de Esteroides/metabolismo , Animais , Citocromo P-450 CYP3A/efeitos dos fármacos , Interações Medicamentosas , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Receptor de Pregnano X , Receptores de Esteroides/efeitos dos fármacos
9.
J Clin Invest ; 118(9): 3228-39, 2008 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-18677425

RESUMO

The pregnane X receptor (PXR) and the constitutive androstane receptor (CAR) are closely related orphan nuclear hormone receptors that play a critical role as xenobiotic sensors in mammals. Both receptors regulate the expression of genes involved in the biotransformation of chemicals in a ligand-dependent manner. As the ligand specificity of PXR and CAR have diverged between species, the prediction of in vivo PXR and CAR interactions with a drug are difficult to extrapolate from animals to humans. We report the development of what we believe are novel PXR- and CAR-humanized mice, generated using a knockin strategy, and Pxr- and Car-KO mice as well as a panel of mice including all possible combinations of these genetic alterations. The expression of human CAR and PXR was in the predicted tissues at physiological levels, and splice variants of both human receptors were expressed. The panel of mice will allow the dissection of the crosstalk between PXR and CAR in the response to different drugs. To demonstrate the utility of this panel of mice, we used the mice to show that the in vivo induction of Cyp3a11 and Cyp2b10 by phenobarbital was only mediated by CAR, although this compound is described as a PXR and CAR activator in vitro. This panel of mouse models is a useful tool to evaluate the roles of CAR and PXR in drug bioavailability, toxicity, and efficacy in humans.


Assuntos
Avaliação Pré-Clínica de Medicamentos/métodos , Modelos Animais , Receptores Citoplasmáticos e Nucleares/metabolismo , Receptores de Esteroides/metabolismo , Fatores de Transcrição/metabolismo , Alelos , Animais , Química Farmacêutica/métodos , Receptor Constitutivo de Androstano , Avaliação Pré-Clínica de Medicamentos/instrumentação , Técnicas Genéticas , Humanos , Camundongos , Camundongos Knockout , Modelos Biológicos , Modelos Genéticos , Receptor de Pregnano X , RNA Mensageiro/metabolismo , Receptores Citoplasmáticos e Nucleares/genética
10.
Drug Metab Dispos ; 38(7): 1046-53, 2010 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-20354104

RESUMO

Dexamethasone (DEX) is a potent and widely used anti-inflammatory and immunosuppressant glucocorticoid. It can bind and activate the pregnane X receptor (PXR), which plays a critical role as xenobiotic sensor in mammals to induce the expression of many enzymes, including cytochromes P450 in the CYP3A family. This induction results in its own metabolism. We have used a series of transgenic mouse lines, including a novel, improved humanized PXR line, to compare the induction profile of PXR-regulated drug-metabolizing enzymes after DEX administration, as well as looking at hepatic responses to rifampicin (RIF). The new humanized PXR model has uncovered further intriguing differences between the human and mouse receptors in that RIF only induced Cyp2b10 in the new humanized model. DEX was found to be a much more potent inducer of Cyp3a proteins in wild-type mice than in mice humanized for PXR. To assess whether PXR is involved in the detoxification of DEX in the liver, we analyzed the consequences of high doses of the glucocorticoid on hepatotoxicity on different PXR genetic backgrounds. We also studied these effects in an additional mouse model in which functional mouse Cyp3a genes have been deleted. These strains exhibited different sensitivities to DEX, indicating a protective role of the PXR and CYP3A proteins against the hepatotoxicity of this compound.


Assuntos
Dexametasona/farmacologia , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Receptores de Esteroides/biossíntese , Receptores de Esteroides/efeitos dos fármacos , Animais , Hidrocarboneto de Aril Hidroxilases/biossíntese , Hidrocarboneto de Aril Hidroxilases/genética , Linhagem Celular Transformada , Citocromo P-450 CYP3A , Sistema Enzimático do Citocromo P-450/biossíntese , Sistema Enzimático do Citocromo P-450/genética , Família 2 do Citocromo P450 , Dexametasona/farmacocinética , Humanos , Fígado/efeitos dos fármacos , Fígado/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Receptor de Pregnano X , Rifampina/farmacologia , Esteroide Hidroxilases/biossíntese , Esteroide Hidroxilases/genética
11.
Mol Cell Biol ; 23(11): 3982-9, 2003 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-12748299

RESUMO

ES cell-tetraploid (ES) mice are completely derived from embryonic stem cells and can be obtained at high efficiency upon injection of hybrid ES cells into tetraploid blastocysts. This method allows the immediate generation of targeted mouse mutants from genetically modified ES cell clones, in contrast to the standard protocol, which involves the production of chimeras and several breeding steps. To provide a baseline for the analysis of ES mouse mutants, we performed a phenotypic characterization of wild-type B6129S6F(1) ES mice in relation to controls of the same age, sex, and genotype raised from normal matings. The comparison of 90 morphological, physiological, and behavioral parameters revealed elevated body weight and hematocrit as the only major difference of ES mice, which exhibited an otherwise normal phenotype. We further demonstrate that ES mouse mutants can be produced from mutant hybrid ES cells and analyzed within a period of only 4 months. Thus, ES mouse technology is a valid research tool for rapidly elucidating gene function in vivo.


Assuntos
Comportamento Animal/fisiologia , Embrião de Mamíferos/citologia , Engenharia Genética/métodos , Camundongos/genética , Células-Tronco/fisiologia , Animais , Blastocisto/citologia , Blastocisto/fisiologia , Análise Química do Sangue , Peso Corporal , Células Cultivadas , Metabolismo Energético/fisiologia , Feminino , Células Híbridas/fisiologia , Isoenzimas/metabolismo , Masculino , Camundongos Endogâmicos , Camundongos Mutantes , Morfogênese/fisiologia , Fenótipo , Poliploidia , Células-Tronco/citologia
12.
Nat Biotechnol ; 20(5): 455-9, 2002 May.
Artigo em Inglês | MEDLINE | ID: mdl-11981557

RESUMO

We have devised a general strategy for producing female mice from 39,X0 embryonic stem (ES) cells derived from male cell lines carrying a targeted mutation of interest. We show that the Y chromosome is lost in 2% of subclones from 40,XY ES cell lines, making the identification of targeted 39,X0 subclones a routine procedure. After gene targeting, male and female mice carrying the mutation can be generated by tetraploid embryo complementation from the 40,XY ES cell line and its 39,X0 derivatives. A single intercross then produces homozygous mutant offspring. Because this strategy avoids outcrossing and therefore segregation of mutant alleles introduced into the ES cells, the time and expense required for production of experimental mutant animals from a targeted ES cell clone are substantially reduced. Our data also indicate that ES cells have inherently unstable karyotypes, but this instability does not interfere with production of adult ES cell tetraploid mice.


Assuntos
Embrião de Mamíferos/citologia , Técnicas Genéticas , Células-Tronco/citologia , Alelos , Animais , Southern Blotting , Linhagem Celular , Clonagem de Organismos , Feminino , Genótipo , Homozigoto , Hibridização in Situ Fluorescente , Cariotipagem , Masculino , Metáfase , Camundongos , Mutação , Ploidias , Fatores Sexuais , Fatores de Tempo , Cromossomo X , Cromossomo Y
13.
Nucleic Acids Res ; 31(4): e12, 2003 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-12582257

RESUMO

We have generated an optimized inducible recombination system for conditional gene targeting based on a Cre recombinase-steroid receptor fusion. This configuration allows efficient Cre-mediated recombination in most organs of the mouse upon induction, without detectable background activity. An ES cell line, was established that carries the inducible recombinase and a loxP-flanked lacZ reporter gene. Out of this line, completely ES cell-derived mice were efficiently produced through tetraploid blastocyst complementation, without the requirement of mouse breeding. Our findings provide a new concept allowing the generation of inducible mouse mutants within 6 months, as compared to 14 months using the current protocol.


Assuntos
Engenharia Genética/métodos , Camundongos Knockout/genética , Animais , Linhagem Celular , Embrião de Mamíferos/citologia , Embrião de Mamíferos/metabolismo , Feminino , Expressão Gênica , Humanos , Integrases/genética , Integrases/metabolismo , Masculino , Camundongos , Camundongos Transgênicos , Receptores de Esteroides/genética , Receptores de Esteroides/metabolismo , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Recombinação Genética , Proteínas Virais/genética , Proteínas Virais/metabolismo
14.
Toxicol Sci ; 116(2): 452-66, 2010 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-20403969

RESUMO

Mouse nongenotoxic hepatocarcinogens phenobarbital (PB) and chlordane induce hepatomegaly characterized by hypertrophy and hyperplasia. Increased cell proliferation is implicated in the mechanism of tumor induction. The relevance of these tumors to human health is unclear. The xenoreceptors, constitutive androstane receptors (CARs), and pregnane X receptor (PXR) play key roles in these processes. Novel "humanized" and knockout models for both receptors were developed to investigate potential species differences in hepatomegaly. The effects of PB (80 mg/kg/4 days) and chlordane (10 mg/kg/4 days) were investigated in double humanized PXR and CAR (huPXR/huCAR), double knockout PXR and CAR (PXRKO/CARKO), and wild-type (WT) C57BL/6J mice. In WT mice, both compounds caused increased liver weight, hepatocellular hypertrophy, and cell proliferation. Both compounds caused alterations to a number of cell cycle genes consistent with induction of cell proliferation in WT mice. However, these gene expression changes did not occur in PXRKO/CARKO or huPXR/huCAR mice. Liver hypertrophy without hyperplasia was demonstrated in the huPXR/huCAR animals in response to both compounds. Induction of the CAR and PXR target genes, Cyp2b10 and Cyp3a11, was observed in both WT and huPXR/huCAR mouse lines following treatment with PB or chlordane. In the PXRKO/CARKO mice, neither liver growth nor induction of Cyp2b10 and Cyp3a11 was seen following PB or chlordane treatment, indicating that these effects are CAR/PXR dependent. These data suggest that the human receptors are able to support the chemically induced hypertrophic responses but not the hyperplastic (cell proliferation) responses. At this time, we cannot be certain that hCAR and hPXR when expressed in the mouse can function exactly as the genes do when they are expressed in human cells. However, all parameters investigated to date suggest that much of their functionality is maintained.


Assuntos
Clordano/toxicidade , Neoplasias Hepáticas Experimentais/induzido quimicamente , Fenobarbital/toxicidade , Receptores Citoplasmáticos e Nucleares/fisiologia , Receptores de Esteroides/fisiologia , Animais , Hidrocarboneto de Aril Hidroxilases/biossíntese , Proliferação de Células/efeitos dos fármacos , Receptor Constitutivo de Androstano , Citocromo P-450 CYP3A/biossíntese , Família 2 do Citocromo P450 , Humanos , Hiperplasia , Hipertrofia , Fígado/efeitos dos fármacos , Fígado/metabolismo , Fígado/patologia , Neoplasias Hepáticas Experimentais/patologia , Proteínas de Membrana/biossíntese , Camundongos , Camundongos Endogâmicos C57BL , Receptor de Pregnano X , Especificidade da Espécie , Esteroide Hidroxilases/biossíntese
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