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1.
Mol Pharmacol ; 76(5): 946-56, 2009 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-19633067

RESUMO

ABCG2 is an ATP-binding cassette transporter that counts multiple anticancer compounds among its substrates and is believed to regulate oral bioavailability as well as serve a protective role in the blood-brain barrier, the maternal-fetal barrier, and hematopoietic stem cells. We sought to determine whether novel compounds that interact with the transporter could be identified through analysis of cytotoxicity profiles recorded in the NCI Anticancer Drug Screen database. A flow cytometric assay was used to measure ABCG2 function in the 60 cell lines and generate a molecular profile for COMPARE analysis. This strategy identified >70 compounds with Pearson correlation coefficients (PCCs) >0.4, where reduced drug sensitivity correlated with ABCG2 expression, as well as >120 compounds with PCCs < -0.4, indicating compounds to which ABCG2 expression conferred greater sensitivity. Despite identification of known single nucleotide polymorphisms in the ABCG2 gene in a number of the cell lines, omission of these lines from the COMPARE analysis did not affect PCCs. Available compounds were subjected to validation studies to confirm interaction with the transporter, including flow cytometry, [(125)I]IAAP binding, and cytotoxicity assays, and interaction was documented in 20 of the 27 compounds studied. Although known substrates of ABCG2 such as mitoxantrone or topotecan were not identified, we characterized three novel substrates-5-hydroxypicolinaldehyde thiosemicarbazone (NSC107392), (E)-N-(1-decylsulfanyl-3-hydroxypropan-2-yl)-3-(6-methyl-2,4-dioxo-1H-pyrimidin-5-yl)prop-2-enamide (NSC265473), and 1,2,3,4,7-pentahydroxy-1,3,4,4a,5,11b-hexahydro[1,3]dioxolo[4,5-j]phenanthridin-6(2H)-one [NSC349156 (pancratistatin)]-and four compounds that inhibited transporter function-2-[methyl(2-pyridin-2-ylethyl)-amino]fluoren-9-one hydroiodide (NSC24048), 5-amino-6-(7-amino-5,8-dihydro-6-methoxy-5,8-dioxo-2-quinolinyl)-4-(2-hydroxy-3,4-dimethoxyphenyl)-3-methyl-2-pyridinecarboxylic acid, methyl ester (NSC45384), (17beta)-2,4-dibromo-estra-1,3,5(10)-triene-3,17-diol (NSC103054), and methyl N-(pyridine-4-carbonylamino)carbamodithioate (NSC636795). In summary, COMPARE analysis of the NCI drug screen database using the ABCG2 functional profile was able to identify novel substrates and transporter-interacting compounds.


Assuntos
Transportadores de Cassetes de Ligação de ATP/fisiologia , Ensaios de Seleção de Medicamentos Antitumorais/métodos , National Cancer Institute (U.S.) , Proteínas de Neoplasias/fisiologia , 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/genética , Antineoplásicos/química , Antineoplásicos/farmacologia , Linhagem Celular , Linhagem Celular Tumoral , Perfilação da Expressão Gênica , Variação Genética , Humanos , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/genética , Polimorfismo de Nucleotídeo Único , Especificidade por Substrato , Estados Unidos
2.
Drug Metab Dispos ; 36(4): 780-95, 2008 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-18180275

RESUMO

The hypothesis was tested that sequence diversity in breast cancer resistance protein (BCRP)'s cis-regulatory region is a significant determinant of BCRP expression. The BCRP promoter and intron 1 were resequenced in lymphoblast DNA from the polymorphism discovery resource (PDR) 44 subset. BCRP single nucleotide polymorphisms (SNPs) were genotyped in donor human livers, intestines, and lymphoblasts quantitatively phenotyped for BCRP mRNA expression. Carriers of the -15622C>T SNP had lower BCRP expression in multiple tissues. The intron 1 SNP 16702C>T was associated with high expression in livers; 1143G>A was associated with low expression in intestine; 12283T>C was associated with higher expression in the PDR44 and White livers. The -15994C>T promoter SNP was significantly associated with higher BCRP expression in multiple tissues. Patients with the -15994C>T genotype had substantially higher clearance of p.o. imatinib. We next determined whether BCRP expression was related to polymorphic alternative splicing or alternative promoter use. Liver polymorphically expressed an alternatively spliced mRNA [splice variant (SV) 1] skipping exon 2. Although SV1+ livers did not uniformly carry the exon 2 G34A allele, 90% of G34A livers expressed SV1 (versus 4% of 34GG livers). BCRP mRNA was significantly lower among Hispanic livers with the G34A variant genotype and may be due, in part, to polymorphic exon 2 splicing. Analysis of allele expression imbalance (AEI) showed that PDR44 samples with AEI had lower BCRP mRNA expression; however, no linked cis-polymorphisms were identified. BCRP used multiple promoters, and livers differentially using alternative exon 1b had lower BCRP. In conclusion, BCRP expression in lymphoblasts, liver, and intestine is associated with novel promoter and intron 1 SNPs.


Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Íntrons/genética , Proteínas de Neoplasias/genética , Fenótipo , Polimorfismo de Nucleotídeo Único/genética , Regiões Promotoras Genéticas/genética , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/biossíntese , Sequência de Aminoácidos/genética , Feminino , Frequência do Gene/genética , Humanos , Masculino , Dados de Sequência Molecular , Proteínas de Neoplasias/biossíntese
3.
Mol Endocrinol ; 21(8): 1769-80, 2007 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-17519356

RESUMO

The farnesoid X receptor (FXR or NR1H4) is an important bile-acid-activated, transcriptional regulator of genes involved in bile acid, lipid, and glucose homeostasis. Accordingly, interindividual variations in FXR expression and function could manifest as variable susceptibility to conditions such as cholesterol gallstone disease, atherosclerosis, and diabetes. We performed an FXR polymorphism discovery analysis of European-, African-, Chinese-, and Hispanic-Americans and identified two rare gain-of-function variants and a common single nucleotide polymorphism resulting in a G-1T substitution in the nucleotide adjacent to the translation initiation site (FXR*1B) with population allelic frequencies ranging from 2.5 to 12%. In cell-based transactivation assays, FXR*1B (-1T) activity was reduced compared with FXR*1A (-1G). This reduced activity for FXR*1B resulted from neither decreased translational efficiency nor the potential formation of a truncated translational variant. To further define the relevance of this polymorphism, gene expression was examined in a human liver bank to reveal that levels of the FXR target genes small heterodimer partner and organic anion transporting polypeptide 1B3 were significantly reduced in livers harboring an FXR*1B allele. These findings are the first to identify the presence of a common genetic variant in FXR with functional consequences that could contribute to disease risk or therapeutic outcomes.


Assuntos
Ácidos e Sais Biliares/fisiologia , Proteínas de Ligação a DNA/genética , Regulação da Expressão Gênica/genética , Fígado/metabolismo , Polimorfismo Genético , Receptores Citoplasmáticos e Nucleares/genética , Fatores de Transcrição/genética , Alelos , Sequência de Aminoácidos , Proteínas de Ligação a DNA/fisiologia , Humanos , Dados de Sequência Molecular , Isoformas de Proteínas/genética , Receptores Citoplasmáticos e Nucleares/fisiologia , Fatores de Transcrição/fisiologia
4.
Int Clin Psychopharmacol ; 28(1): 50-6, 2013 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-23104241

RESUMO

Fixed oral doses of clozapine produce up to 45-fold interindividual variability among its serum levels in patients with treatment-resistant schizophrenia. Although the relationship between serum clozapine level and its therapeutic response is uncertain, the presence of a therapeutic window and level-dependent adverse effects require the estimation of serum clozapine levels. As routine therapeutic drug monitoring of clozapine is not feasible in many clinical settings, identification of clinical predictors of serum clozapine levels is desirable. Hence, we aimed to evaluate the clinical variables associated with serum clozapine levels. We assessed the sociodemographic and clinical profiles, cognition, disability and psychopathology of 101 consecutive patients with treatment-resistant schizophrenia on a stable dose of clozapine, using standard assessment schedules. We determined their serum clozapine levels using high-performance liquid chromatography with ultraviolet detection. While employing multivariate robust regression models, oral clozapine dose (P<0.001), caffeine intake (P=0.04) and Valproate comedication (P=0.005) were associated with serum clozapine levels. Serum clozapine levels above 750 ng/ml increased the risk of seizures (odds ratio 5.15; P=0.03). Clinical variables are useful to model a dosing nomogram for serum clozapine levels. The importance of caffeine consumption and Valproate comedication should be considered during clozapine dose adjustments to enhance its therapeutic response and safety profile.


Assuntos
Antipsicóticos/sangue , Antipsicóticos/uso terapêutico , Clozapina/sangue , Clozapina/uso terapêutico , Resistência a Medicamentos , Esquizofrenia/tratamento farmacológico , Administração Oral , Adulto , Anticonvulsivantes/efeitos adversos , Antipsicóticos/administração & dosagem , Antipsicóticos/efeitos adversos , Antipsicóticos/farmacocinética , Cafeína/efeitos adversos , Estudos de Casos e Controles , Estimulantes do Sistema Nervoso Central/efeitos adversos , Distribuição de Qui-Quadrado , Cromatografia Líquida de Alta Pressão , Clozapina/administração & dosagem , Clozapina/efeitos adversos , Clozapina/farmacocinética , Estudos Transversais , Interações Medicamentosas , Monitoramento de Medicamentos/métodos , Feminino , Interações Alimento-Droga , Humanos , Análise dos Mínimos Quadrados , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Razão de Chances , Escalas de Graduação Psiquiátrica , Fatores de Risco , Esquizofrenia/sangue , Esquizofrenia/diagnóstico , Psicologia do Esquizofrênico , Espectrofotometria Ultravioleta , Falha de Tratamento , Ácido Valproico/efeitos adversos
5.
J Clin Oncol ; 26(7): 1119-27, 2008 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-18309947

RESUMO

PURPOSE: To assess the pharmacogenomic and pharmacokinetic determinants of skin rash and diarrhea, the two primary dose-limiting toxicities of the epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor erlotinib. PATIENTS AND METHODS: A prospective clinical study of 80 patients with non-small-cell lung cancer, head and neck cancer, and ovarian cancer was performed. Detailed pharmacokinetics and toxicity of erlotinib were assessed. Polymorphic loci in EGFR, ABCG2, CYP3A4, and CYP3A5 were genotyped, and their effects on pharmacokinetics and toxicities were evaluated. RESULTS: A novel diplotype of two polymorphic loci in the ABCG2 promoter involving -15622C/T and 1143C/T was identified, with alleles conferring lower ABCG2 levels associated with higher erlotinib pharmacokinetic parameters, including area under the curve (P = .019) and maximum concentration (P = .006). Variability in skin rash was best explained by a multivariate logistic regression model incorporating the trough erlotinib plasma concentration (P = .034) and the EGFR intron 1 polymorphism (P = .044). Variability in diarrhea was associated with the two linked polymorphisms in the EGFR promoter (P < .01), but not with erlotinib concentration. CONCLUSION: Although exploratory in nature, this combined pharmacogenomic and pharmacokinetic model helps to define and differentiate the primary determinants of skin and gastrointestinal toxicity of erlotinib. The findings may be of use both in designing trials targeting a particular severity of rash and in considering dose and schedule modifications in patients experiencing dose-limiting toxicities of erlotinib or similarly targeted agents. Further studies of the relationship between germline polymorphisms in EGFR and the toxicity and efficacy of EGFR inhibitors are warranted.


Assuntos
Receptores ErbB/genética , Neoplasias de Cabeça e Pescoço/genética , Neoplasias Pulmonares/genética , Neoplasias Ovarianas/genética , Farmacogenética , Polimorfismo Genético , Inibidores de Proteínas Quinases/farmacocinética , Quinazolinas/farmacocinética , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Citocromo P-450 CYP3A , Sistema Enzimático do Citocromo P-450/genética , Diarreia/induzido quimicamente , Receptores ErbB/antagonistas & inibidores , Cloridrato de Erlotinib , Exantema/induzido quimicamente , Feminino , Neoplasias de Cabeça e Pescoço/metabolismo , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Masculino , Pessoa de Meia-Idade , Proteínas de Neoplasias/genética , Neoplasias de Células Escamosas/genética , Neoplasias de Células Escamosas/metabolismo , Neoplasias Ovarianas/metabolismo , Estudos Prospectivos , Inibidores de Proteínas Quinases/efeitos adversos , Quinazolinas/efeitos adversos
6.
Cancer Res ; 68(13): 4983-9, 2008 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-18593894

RESUMO

Thiopurines are effective immunosuppressants and anticancer agents, but intracellular accumulation of their active metabolites (6-thioguanine nucleotides, 6-TGN) causes dose-limiting hematopoietic toxicity. Thiopurine S-methyltransferase deficiency is known to exacerbate thiopurine toxicity. However, many patients are highly sensitive to thiopurines for unknown reasons. We show that multidrug-resistance protein 4 (Mrp4) is abundant in myeloid progenitors and tested the role of the Mrp4, an ATP transporter of monophosphorylated nucleosides, in this unexplained thiopurine sensitivity. Mrp4-deficient mice experienced Mrp4 gene dosage-dependent toxicity caused by accumulation of 6-TGNs in their myelopoietic cells. Therefore, Mrp4 protects against thiopurine-induced hematopoietic toxicity by actively exporting thiopurine nucleotides. We then identified a single-nucleotide polymorphism (SNP) in human MRP4 (rs3765534) that dramatically reduces MRP4 function by impairing its cell membrane localization. This SNP is common (>18%) in the Japanese population and indicates that the increased sensitivity of some Japanese patients to thiopurines may reflect the greater frequency of this MRP4 SNP.


Assuntos
Citoproteção/genética , Resistencia a Medicamentos Antineoplásicos/genética , Doenças Hematológicas/induzido quimicamente , Proteínas Associadas à Resistência a Múltiplos Medicamentos/genética , Compostos de Sulfidrila/efeitos adversos , Alelos , Animais , Membrana Celular/metabolismo , Células Cultivadas , Citoproteção/efeitos dos fármacos , Doenças Hematológicas/mortalidade , Hematopoese/efeitos dos fármacos , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/genética , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Proteínas de Membrana Transportadoras/fisiologia , Mercaptopurina/efeitos adversos , Mercaptopurina/farmacologia , Mercaptopurina/uso terapêutico , Camundongos , Camundongos Knockout , Modelos Biológicos , Proteínas Associadas à Resistência a Múltiplos Medicamentos/metabolismo , Proteínas Associadas à Resistência a Múltiplos Medicamentos/fisiologia , Polimorfismo de Nucleotídeo Único/fisiologia , Purinas/efeitos adversos , Purinas/química , Purinas/uso terapêutico , Compostos de Sulfidrila/uso terapêutico , Análise de Sobrevida , Distribuição Tecidual
7.
Clin Chem ; 49(5): 777-81, 2003 May.
Artigo em Inglês | MEDLINE | ID: mdl-12709369

RESUMO

BACKGROUND: Inherited hemoglobin disorders represent the most common Mendelian disease worldwide. Prevention programs based on molecular diagnosis of heterozygous carriers and/or patients require the use of reliable mutation scanning methods in at-risk populations. METHODS: We developed a rapid and highly specific mutation-screening test based on temporal temperature gradient gel electrophoresis (TTGE). We analyzed 889 beta-thalassemia genes from homozygous beta-thalassemia patients and unrelated individuals with heterozygous beta-thalassemia. Previously reported common mutations were screened by reverse dot blots using allele-specific probes. The rare mutations were analyzed by TTGE. RESULTS: We found common mutations in 753 beta-thalassemia genes. TTGE analysis in the rest of the genes showed the presence of mutations in different regions of the beta-globin gene in 134 of them, and these mutations were characterized by DNA sequencing. In the two genes in which mutations were not identified, large deletions spanning beta-globin gene were suspected. CONCLUSIONS: Compared with other approaches for comprehensive mutation screening, the reported method is rapid, highly sensitive, cost-effective, and suitable for high-throughput screening of a large number of samples.


Assuntos
Globinas/genética , Eletroforese em Gel de Poliacrilamida , Humanos , Mutação , Desnaturação de Ácido Nucleico , Reação em Cadeia da Polimerase , Polimorfismo Genético , Temperatura
8.
Blood ; 104(5): 1574-7, 2004 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-15142875

RESUMO

Hepatic venoocclusive disease (HVOD) in bone marrow transplantation (BMT) is attributed to toxicity of cytoreductive agents, especially busulfan and cyclophosphamide, in the conditioning therapy. Busulfan, as well as the metabolites of cyclophosphamide, are conjugated with glutathione (GSH), catalyzed by enzymes of the glutathione S-transferase (GST) family. To assess the impact of polymorphisms of the GST genes, GSTM1 and GSTT1, on the risk of HVOD, we evaluated 114 consecutive patients with beta-thalassemia major undergoing BMT. There was a significantly increased incidence of HVOD in patients with the GSTM1-null genotype compared with those with the GSTM1-positive genotype (46.5% vs 18.3%; P =.001). Pharmacokinetic analysis in these patients showed that the clearance of busulfan was higher and first-dose steady-state concentration was lower among those with HVOD (0.403 +/- 0.06 vs 0.33 +/- 0.071 L/h/kg, Student t test P value =.000 01; and 508 +/- 125 vs 656 +/- 255 ng/mL, t test P value =.001, respectively). We conclude that the GSTM1-null genotype predisposes to HVOD, and the sinusoidal endothelial cells and hepatocyte damage may be mediated by metabolites of busulfan through depletion of the cellular GSH pool.


Assuntos
Transplante de Medula Óssea , Bussulfano/administração & dosagem , Glutationa Transferase/genética , Hepatopatia Veno-Oclusiva/genética , Imunossupressores/administração & dosagem , Talassemia beta/terapia , Adolescente , Bussulfano/efeitos adversos , Criança , Pré-Escolar , Ciclofosfamida/administração & dosagem , Ciclofosfamida/efeitos adversos , Predisposição Genética para Doença/epidemiologia , Genótipo , Hepatopatia Veno-Oclusiva/epidemiologia , Humanos , Imunossupressores/efeitos adversos , Lactente , Polimorfismo Genético , Fatores de Risco , Talassemia beta/epidemiologia , Talassemia beta/genética
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