RESUMO
Lung cancer is one of the most common cancers in men. Although many diagnostic and treatment regimens have been followed in the treatment for lung cancer, increasing mortality rate due to lung cancer is depressing and hence requires alternative plant based therapeutics with with less side-effects. Myrtenol exhibits anti-inflammatory and antioxidant properties. Hence we intended to study the effect of Myrtenol on B(a)P-induced lung cancer. Our study showed that B(a)P lowered hematological count, decreased phagocyte and avidity indices, nitroblue tetrazolium (NBT) reduction, levels of immunoglubulins, antioxidant levels, whereas Myrtenol treatment restored them back to normal levels. On the other hand, xenobiotic and liver dysfunction marker enzymes and pro-inflammatory cytokines were elevated on B(a)P exposure, which retuned back to normal by Myrtenol. This study thus describes the immunomodulatory and antioxidant effects of Myrtenol on B[a]P-induced immune destruction.
Assuntos
Monoterpenos Bicíclicos , Neoplasias Pulmonares , Humanos , Masculino , Camundongos , Animais , Neoplasias Pulmonares/induzido quimicamente , Neoplasias Pulmonares/tratamento farmacológico , Citocinas/metabolismo , Benzo(a)pireno/toxicidade , Antioxidantes/metabolismo , Antígeno Nuclear de Célula em Proliferação/metabolismo , Biomarcadores Tumorais/metabolismo , Pulmão/metabolismoRESUMO
Failure to demonstrate efficacy and safety issues are important reasons that drugs do not reach the market. An incomplete understanding of how drugs exert their effects hinders regulatory and pharmaceutical industry projections of a drug's benefits and risks. Signaling pathways mediate drug response and while many signaling molecules have been characterized for their contribution to disease or their role in drug side effects, our knowledge of these pathways is incomplete. To better understand all signaling molecules involved in drug response and the phenotype associations of these molecules, we created a novel method, PathFX, a non-commercial entity, to identify these pathways and drug-related phenotypes. We benchmarked PathFX by identifying drugs' marketed disease indications and reported a sensitivity of 41%, a 2.7-fold improvement over similar approaches. We then used PathFX to strengthen signals for drug-adverse event pairs occurring in the FDA Adverse Event Reporting System (FAERS) and also identified opportunities for drug repurposing for new diseases based on interaction paths that associated a marketed drug to that disease. By discovering molecular interaction pathways, PathFX improved our understanding of drug associations to safety and efficacy phenotypes. The algorithm may provide a new means to improve regulatory and therapeutic development decisions.
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Algoritmos , Desenvolvimento de Medicamentos/estatística & dados numéricos , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos , Biologia Computacional , Bases de Dados de Produtos Farmacêuticos , Tomada de Decisões , Aprovação de Drogas , Desenvolvimento de Medicamentos/legislação & jurisprudência , Desenvolvimento de Medicamentos/normas , Descoberta de Drogas/legislação & jurisprudência , Descoberta de Drogas/normas , Descoberta de Drogas/estatística & dados numéricos , Interações Medicamentosas , Reposicionamento de Medicamentos , Controle de Medicamentos e Entorpecentes , Humanos , Segurança , Resultado do Tratamento , Estados Unidos , United States Food and Drug AdministrationRESUMO
Inter-individual variation in gene regulatory elements is hypothesized to play a causative role in adverse drug reactions and reduced drug activity. However, relatively little is known about the location and function of drug-dependent elements. To uncover drug-associated elements in a genome-wide manner, we performed RNA-seq and ChIP-seq using antibodies against the pregnane X receptor (PXR) and three active regulatory marks (p300, H3K4me1, H3K27ac) on primary human hepatocytes treated with rifampin or vehicle control. Rifampin and PXR were chosen since they are part of the CYP3A4 pathway, which is known to account for the metabolism of more than 50% of all prescribed drugs. We selected 227 proximal promoters for genes with rifampin-dependent expression or nearby PXR/p300 occupancy sites and assayed their ability to induce luciferase in rifampin-treated HepG2 cells, finding only 10 (4.4%) that exhibited drug-dependent activity. As this result suggested a role for distal enhancer modules, we searched more broadly to identify 1,297 genomic regions bearing a conditional PXR occupancy as well as all three active regulatory marks. These regions are enriched near genes that function in the metabolism of xenobiotics, specifically members of the cytochrome P450 family. We performed enhancer assays in rifampin-treated HepG2 cells for 42 of these sequences as well as 7 sequences that overlap linkage-disequilibrium blocks defined by lead SNPs from pharmacogenomic GWAS studies, revealing 15/42 and 4/7 to be functional enhancers, respectively. A common African haplotype in one of these enhancers in the GSTA locus was found to exhibit potential rifampin hypersensitivity. Combined, our results further suggest that enhancers are the predominant targets of rifampin-induced PXR activation, provide a genome-wide catalog of PXR targets and serve as a model for the identification of drug-responsive regulatory elements.
Assuntos
Regulação da Expressão Gênica/efeitos dos fármacos , Fígado/efeitos dos fármacos , Fígado/fisiologia , Receptores de Esteroides/genética , Sequências Reguladoras de Ácido Nucleico , Células Cultivadas , Citocromo P-450 CYP3A/genética , Genoma Humano , Células Hep G2/efeitos dos fármacos , Hepatócitos/efeitos dos fármacos , Hepatócitos/fisiologia , Histonas/metabolismo , Humanos , Polimorfismo de Nucleotídeo Único , Receptor de Pregnano X , Regiões Promotoras Genéticas , Receptores de Esteroides/metabolismo , Reprodutibilidade dos Testes , Rifampina/farmacologia , Fatores de Transcrição de p300-CBP/metabolismoRESUMO
OBJECTIVE: Pharmacogenomic studies have established the important contribution of drug-metabolizing enzyme genotype toward drug toxicity and treatment failure; however, clinical implementation of pharmacogenomics has been slow. The aim of this study was to systematically review the information on drug-metabolizing enzyme pharmacogenomics available in the US drug labeling, practice guidelines, and recommendations. METHODS: Drug-metabolizing enzyme genotype and phenotype information was assessed in US FDA drug labeling, clinical practice guidelines, and independent technology assessors to evaluate the consistency in information sources for healthcare providers. RESULTS: Eighty four gene-drug pairs were identified as having drug-metabolizing enzyme genotype or phenotype information within the label. The manner in which pharmacogenomic information was presented was heterogeneous both within the label and between clinical practice recommendations. CONCLUSION: For proper implementation of pharmacogenomics in clinical practice, information sources for healthcare providers should relay consistent and clear information for the appropriate use of biomarkers.
Assuntos
Sistema Enzimático do Citocromo P-450/genética , Preparações Farmacêuticas/metabolismo , Farmacogenética , Guias de Prática Clínica como Assunto , Biomarcadores , Relação Dose-Resposta a Droga , Monitoramento de Medicamentos , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos , Genótipo , Humanos , FenótipoRESUMO
Statins are widely prescribed and highly susceptible to pharmacokinetic (PK)-based drug-drug interactions (DDIs). To date, there has not been a comprehensive analysis of the basis upon which statin DDI recommendations in US Food and Drug Administration (FDA) prescribing information (PI) are derived. We have conducted such an analysis. We also assessed the degree of concordance of statin DDI recommendations in FDA PI and those provided in common tertiary clinical resources. We catalogued statin DDI information, including PK data and management recommendations, for statin precipitant drugs approved from 2010 to 2021, available from FDA PI and tertiary clinical resource databases. Recommendations were categorized and mapped with associated PK data to assess consistency in the PK basis for labeling recommendations. From the 80 precipitant drugs evaluated, 180 statin DDIs were identified in FDA PI. Dedicated clinical DDI studies were conducted for 54% (n = 97) of these DDIs and 34% (n = 61) of DDI recommendations were extrapolated from clinical data with other statins. Overall, we found that PK-based statin recommendations were consistent across PI. These findings highlight regulatory precedence for translating information across statins without conducting dedicated clinical DDI studies, which may support future efforts toward streamlining the approach to investigation and labeling of statin DDIs. In addition, with the exception of some notable discrepancies, general concordance was observed between FDA and tertiary resources regarding "Dose Adjustment" and "Avoid Coadministration" recommendations. However, further analyses are warranted across other DDI pairs to determine whether discordance can routinely lead to different clinical recommendations depending on the drug information resource.
Assuntos
Interações Medicamentosas , Rotulagem de Medicamentos , Inibidores de Hidroximetilglutaril-CoA Redutases , United States Food and Drug Administration , Humanos , Estados Unidos , Rotulagem de Medicamentos/normas , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacocinética , Bases de Dados FactuaisRESUMO
Rifampin causes drug interactions by altering hepatic drug metabolism. Because microRNAs (miRNAs) have been shown to regulate genes involved in drug metabolism, we determined the effect of rifampin on the expression of hepatic miRNAs. Primary human hepatocytes from seven subjects were treated with rifampin, and the expression of miRNA and cytochrome P450 (P450) mRNAs was measured by TaqMan assays and RNA-seq, respectively. Rifampin induced the expression of 10 clinically important and 13 additional P450 genes and repressed the expression of 9 other P450 genes (P < 0.05). Rifampin induced the expression of 33 miRNAs and repressed the expression of 35 miRNAs (P < 0.05). Several of these changes were highly negatively correlated with the rifampin-induced changes in the expression of their predicted target P450 mRNAs, supporting the possibility of miRNA-induced regulation of P450 mRNA expression. In addition, several other miRNA changes were positively correlated with the changes in P450 mRNA expression, suggesting similar regulatory mechanisms. Despite the interindividual variability in the rifampin effects on miRNA expression, principal components analysis clearly separated the rifampin-treated samples from the controls. In conclusion, rifampin treatment alters miRNA expression patterns in human hepatocytes, and some of the changes were correlated with the rifampin-induced changes in expression of the P450 mRNAs they are predicted to target.
Assuntos
Hepatócitos/efeitos dos fármacos , MicroRNAs/genética , Rifampina/farmacologia , Células Cultivadas , Sistema Enzimático do Citocromo P-450/genética , Humanos , RNA Mensageiro/genéticaRESUMO
Quantitative and qualitative analysis of amino acids in biofluids offers relevant information in diagnosis of diseases, evaluation of nutritional state, and in elucidating metabolic influences on physiology. A simple, rapid, and robust procedure in terms of sample treatment, separation, and quantitation based on CE-LIF has been optimized for use in human plasma samples. Time required for derivatization was 15 min and analysis time was 35 min. 4-Fluoro-7-nitro-2,1,3-benzoxadiazole (NBD-F) was the labeling agent used for obtaining fluorescent derivatives. Electrophoretic conditions were: 175 mM borate buffer at pH 10.25 prepared with 12.5 mM ß-cyclodextrin. The voltage applied was +21 kV. Fourteen amino acids could be quantified: L-proline, L-phenylalanine, L-leucine, L-isoleucine, L-ornithine, D-ornithine, L-glutamine, L-alanine, L-threonine, glycine, L-serine, D-serine, taurine and L-glutamate. With this chiral CE-LIF method, L- and D-amino acids are adequately separated. The method was validated for a representative group of amino acids in human plasma: L-proline, L-isoleucine, L-ornithine, L-glutamine, L-alanine L-threonine, glycine, L-serine, D-serine, and glutamate. The method has been successfully applied to human plasma from patients with bipolar disorder, all of whom were taking lithium as a mood stabilizer. Eleven amino acids were quantified in plasma from nine patients, aged 24-55 years. The results were in accordance to published values for the bipolar patients. The method is useful particularly in studies where plasma amino acid levels can be used as biomarkers for diagnosis of diseases, evaluating the disease progression, and monitoring response to drug therapy.
Assuntos
Aminoácidos/sangue , Transtorno Bipolar/sangue , Eletroforese Capilar/métodos , Adulto , Feminino , Fluorescência , Humanos , Lasers , Masculino , Pessoa de Meia-Idade , Sensibilidade e Especificidade , Adulto JovemRESUMO
The prevalence of obesity has grown tremendously in recent years and this population has an increased risk of disease comorbidities. The presence of disease comorbidities requires treatment interventions and proper dosing guidelines. However, drug development programs often do not have adequate representation of individuals who are obese in clinical trials, leaving gaps in the understanding of treatment response leading to a lack of adequate individualization options. Based on a recent survey of approved drug product package inserts, very few approved products included specific dosing based on obesity, in both adults and pediatrics. Reasons for the limited information on patients who are obese may include the under-reporting of information regarding such patients and a lack of clinical trial diversity in enrolling patient groups in whom obesity or obesity-related comorbidities are more prevalent. An inadvertent impact of the practice of exclusion of subsets of patients with some comorbidities in clinical trials may play a role in the reduced enrollment of individuals who are obese. Recently, regulatory authorities have taken specific initiatives to promote clinical trial diversity, including engaging with stakeholders and publishing regulatory guidance. These guidance documents highlight the need to enroll diverse clinical trial populations and provide recommendations on concepts related to drug development for obese populations. Such efforts will help to address the gap in information regarding drug response and dosing in patients who are obese.
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Desenvolvimento de Medicamentos , Obesidade , Adulto , Humanos , Criança , Obesidade/complicações , Obesidade/tratamento farmacológico , Obesidade/epidemiologiaRESUMO
Although the population in the United States is diverse, there are disparities in healthcare outcomes in some populations, for example, based on characteristics such as race, ethnicity, sex, gender, age, socioeconomic status, and geographic location. Despite disproportionate healthcare outcomes, certain populations are frequently under-represented in clinical trials intended to support applications requesting US Food and Drug Administration (FDA) approval to market a drug or biologic. Additionally, safety and efficacy of therapeutic products may vary based on intrinsic (e.g., sex, age, race, and ethnicity) and/or extrinsic (e.g., drug interactions and medical practice) factors. Enrolling diverse populations in clinical trials can aid in addressing disparities and better inform the use of medical products in all patients who will use them upon approval. Herein, we outline a few initiatives and activities, such as policy development, regulatory review, regulatory research, and stakeholder engagement, that the FDA has undertaken to promote diversity in clinical trials, to support submission of such information in marketing applications for subgroup analyses, and to communicate information to the public.
Assuntos
Etnicidade , Marketing , Humanos , Estados Unidos , United States Food and Drug Administration , Aprovação de DrogasRESUMO
Many of the conditions for the safe and effective use of new molecular entities (NMEs) are understood at the time of initial drug approval. However, some remaining knowledge gaps can be addressed after drug approval through postmarketing requirements (PMRs) or commitments (PMCs) established by the US Food and Drug Administration (FDA). Our objective was to conduct an assessment of clinical pharmacology-related PMRs and PMCs established at the time of approval and evaluate the impact of fulfilled PMRs and PMCs on prescription information (PI). This analysis included clinical pharmacology-related PMRs and PMCs established for NMEs approved between 2009 and 2020. Of the 1171 PMRs and PMCs, over one-third were clinical pharmacology-related. Of these, 46% were to evaluate drug interactions, 16% were to evaluate drug dosing in patients with hepatic impairment, and 10% were related to dose. The majority (57%) of PMRs and PMCs were fulfilled at the time of analysis, with a median time to fulfillment of approximately 2.3 years. The majority (94%) of the fulfilled PMRs and PMCs, either with or without a PI revision, resulted in new or modified instructions for use or supported existing instructions for use. This is the first time that clinical pharmacology-related PMRs and PMCs have been catalogued and analyzed to understand their impact on PI. An understanding of the knowledge gaps that exist at the time of drug approval could inform the most effective and efficient methods for evidence generation prior to and after new drug approval.
Assuntos
Farmacologia Clínica , Estados Unidos , Humanos , United States Food and Drug Administration , Vigilância de Produtos Comercializados/métodos , Aprovação de Drogas/métodosRESUMO
Clinical trials are an integral aspect of drug development. Tremendous progress has been made in ensuring drug products are effective and safe for use in the intended pediatric population, but there remains a paucity of information to guide drug dosages in pediatric patients with obesity. This is concerning because obesity may influence the disposition of drug products. When pediatric patients with obesity are not enrolled in clinical trials, dosing options for use in this subpopulation may be suboptimal. Reliance on physiological-based dosing strategies that are not informed by evaluation of the pharmacokinetics of the drug product could lead to under- or over-dosing with ensuing therapeutic failure or toxicity consequences. Thus, representation of pediatric patients with obesity in clinical trials is crucial to understand the benefit-risk profile of drug products in this subpopulation. It is important to acknowledge that this is a challenging endeavor, but not one that is insurmountable. Collective efforts from multiple stakeholders including drug developers and regulators to enhance diversity in clinical trials can help fill critical gaps in knowledge related to the influence of obesity on drug disposition.
RESUMO
Deriving pediatric doses for locally acting drugs (LADs) presents a unique challenge because limited systemic exposure hinders commonly used approaches such as pharmacokinetic matching to adults. This study systematically evaluated drug development practices used for pediatric dose selection of LADs approved by the U.S. Food and Drug Administration from 2002 to 2020. The three study objectives were: (1) to determine the dose selection approach for the labeled pediatric dose, (2) to examine the studied pediatric dose(s), and (3) to evaluate the characteristics of the pediatric clinical programs used to support the labeled pediatric dose. A total of 187 pediatric submissions were characterized for the labeled and studied pediatric doses of LADs. The pediatric dose was predominantly labeled as a flat dose (91%) and at a single-dose level (67%) similar to adults. The majority (68.4%) of the submissions had the same labeled dose for pediatrics and adults. Independent pharmacodynamic/efficacy studies in pediatric patients commonly (64.2%) provided supportive evidence for the labeled pediatric dose. Inhalation, nasal, and injectable submissions had the highest number of clinical trials, lowest usage of an extrapolation of efficacy approach, and utilized diverse approaches in selecting the studied pediatric doses. This article highlights approaches for LAD dosing in pediatric patients and can be used to inform drug development of these products in the pediatric population.
Assuntos
Aprovação de Drogas , Desenvolvimento de Medicamentos , Adulto , Estados Unidos , Criança , Humanos , Preparações Farmacêuticas , United States Food and Drug Administration , Relação Dose-Resposta a DrogaRESUMO
Inhibition of cell proliferation by fenoterol and fenoterol derivatives in 1321N1 astrocytoma cells is consistent with ß(2)-adrenergic receptor (ß(2)-AR) stimulation. However, the events that result in fenoterol-mediated control of cell proliferation in other cell types are not clear. Here, we compare the effect of the ß(2)-AR agonists (R,R')-fenoterol (Fen) and (R,R')-4-methoxy-1-naphthylfenoterol (MNF) on signaling and cell proliferation in HepG2 hepatocarcinoma cells by using Western blotting and [(3)H]thymidine incorporation assays. Despite the expression of ß(2)-AR, no cAMP accumulation was observed when cells were stimulated with isoproterenol or Fen, although the treatment elicited both mitogen-activated protein kinase and phosphatidylinositol 3-kinase/Akt activation. Unexpectedly, isoproterenol and Fen promoted HepG2 cell growth, but MNF reduced proliferation together with increased apoptosis. The mitogenic responses of Fen were attenuated by 3-(isopropylamino)-1-[(7-methyl-4-indanyl)oxy]butan-2-ol (ICI 118,551), a ß(2)-AR antagonist, whereas those of MNF were unaffected. Because of the coexpression of ß(2)-AR and cannabinoid receptors (CBRs) and their impact on HepG2 cell proliferation, these Gα(i)/Gα(o)-linked receptors may be implicated in MNF signaling. Cell treatment with (R)-(+)-[2,3-dihydro-5-methyl-3-(4-morpholinylmethyl)pyrrolo[1,2,3-de]-1,4-benzoxazin-6-yl]-1-napthalenylmethanone (WIN 55,212-2), a synthetic agonist of CB(1)R and CB(2)R, led to growth inhibition, whereas inverse agonists of these receptors blocked MNF mitogenic responses without affecting Fen signaling. MNF responses were sensitive to pertussis toxin. The ß(2)-AR-deficient U87MG cells were refractory to Fen, but responsive to the antiproliferative actions of MNF and WIN 55,212-2. The data indicate that the presence of the naphthyl moiety in MNF results in functional coupling to the CBR pathway, providing one of the first examples of a dually acting ß(2)-AR-CBR ligand.
Assuntos
Agonistas de Receptores Adrenérgicos beta 2/farmacologia , Apoptose/fisiologia , Carcinoma Hepatocelular/metabolismo , Fenoterol/farmacologia , Neoplasias Hepáticas/metabolismo , Receptores de Canabinoides/metabolismo , Apoptose/efeitos dos fármacos , Carcinoma Hepatocelular/patologia , Fenoterol/química , Células Hep G2 , Humanos , Neoplasias Hepáticas/patologiaRESUMO
Hepatic nuclear factor 4α (HNF4A) is a nuclear transcription factor that regulates the expression of many genes involved in drug disposition. To identify additional molecular mechanisms that regulate HNF4A, we identified microRNAs (miRNAs) that target HNF4A expression. In silico analyses suggested that HNF4A is targeted by many miRNAs. We conducted in vitro studies to validate several of these predictions. With use of an HNF4A 3'-untranslated region (UTR) luciferase reporter assay, five of six miRNAs tested significantly down-regulated (â¼20-40%) the luciferase activity. In HepG2 cells, miR-34a and miR-449a also down-regulated the expression of both the HNF4A protein and an HNF4A target gene, PXR (â¼30-40%). This regulation appeared without reduction in HNF4A mRNA expression, suggesting that they must be blocking HNF4A translation. Using additional bioinformatic algorithms, we identified polymorphisms that are predicted to alter the miRNA targeting of HNF4A. Luciferase assays indicated that miR-34a and miR-449a were less effective in regulating a variant (rs11574744) than the wild-type HNF4A 3'-UTR. In vivo, subjects with the variant HNF4A had lower CYP2D6 enzyme activity, although this result was not statistically significant (p = 0.16). In conclusion, our findings demonstrate strong evidence for a role of miRNAs in the regulation of HNF4A.
Assuntos
Regulação da Expressão Gênica , Fator 4 Nuclear de Hepatócito/genética , MicroRNAs/fisiologia , Polimorfismo de Nucleotídeo Único , Regiões 3' não Traduzidas/genética , Citocromo P-450 CYP2D6/genética , Citocromo P-450 CYP2D6/metabolismo , Genes Reporter , Genótipo , Células HeLa , Células Hep G2 , Fator 4 Nuclear de Hepatócito/metabolismo , Hepatócitos/metabolismo , Humanos , Luciferases/genética , MicroRNAs/genética , Preparações Farmacêuticas/metabolismo , Plasmídeos , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , TransfecçãoRESUMO
The presystemic sulfate conjugation of the stereoisomers of 4'-methoxyfenoterol, (R,R')-MF, (S,S')-MF, (R,S')-MF, and (S,R')-MF, was investigated using commercially available human intestinal S9 fractions, a mixture of sulfotransferase (SULT) enzymes. The results indicate that the sulfation was stereospecific and that an S-configuration at the ß-OH carbon of the MF molecule enhanced the maximal formation rates with (S,R')-MF (S,S')-MF (R,S')-MF ≈ (R,R')-MF, and competition studies demonstrated that (S,R')-MF is an effective inhibitor of (R,R')-MF sulfation (IC(50) = 60 µM). In addition, the results from a cDNA-expressed human SULT isoform screen indicated that SULT1A1, SULT1A3, and SULT1E1 can mediate the sulfation of all four MF stereoisomers. Previously published molecular models of SULT1A3 and SULT1A1 were used in docking simulations of the MF stereoisomers using Molegro Virtual Docker. The models of the MF-SULT1A3 and MF-SULT1A1 complexes indicate that each of the two chiral centers of MF molecule plays a role in the observed relative stabilities. The observed stereoselectivity is the result of multiple hydrogen bonding interactions and induced conformational changes within the substrate-enzyme complex. In conclusion, the results suggest that a formulation developed from a mixture of (R,R')-MF and (S,R')-MF may increase the oral bioavailability of (R,R')-MF.
Assuntos
Fenoterol/análogos & derivados , Sulfatos/química , Sulfotransferases/metabolismo , Sítios de Ligação , Simulação por Computador , Fenoterol/química , Humanos , Modelos Moleculares , Isoformas de Proteínas/metabolismo , Estereoisomerismo , Sulfotransferases/genéticaRESUMO
The breast cancer resistance protein (BCRP), an ATP binding cassette (ABC) efflux transporter, plays a role in multiple drug resistance (MDR). Previous studies of the subcellular location of the ABC transporter P-glycoprotein indicated that this protein is expressed in nuclear membranes. This study examines the nuclear distribution of BCRP in seven human-derived glioblastoma (GBM) and astrocytoma cell lines. BCRP expression was observed in the nuclear extracts of 6/7 cell lines. Using the GBM LN229 cell line as a model, nuclear BCRP protein was detected by immunoblotting and confocal laser microscopy. Importantly, nuclear BCRP staining was found in a subpopulation of tumour cells in a human brain GBM biopsy. Mitoxantrone cytotoxicity in the LN229 cell line was determined with and without the BCRP inhibitor fumitremorgin C (FTC) and after downregulation of BCRP with small interfering RNA (siRNA). FTC inhibition of BCRP increased mitoxantrone cytotoxicity with a ~7-fold reduction in the IC50 and this effect was further potentiated in the siRNA-treated cells. In conclusion, BCRP is expressed in the nuclear extracts of select GBM and astrocytoma cell lines and in a human GBM tumour biopsy. Its presence in the nucleus of cancer cells suggests new role for BCRP in MDR.
Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Núcleo Celular/metabolismo , Glioblastoma/metabolismo , Glioblastoma/patologia , Proteínas de Neoplasias/metabolismo , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/antagonistas & inibidores , Astrocitoma/metabolismo , Astrocitoma/patologia , Biópsia , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Núcleo Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Cromatografia Líquida de Alta Pressão , Imunofluorescência , Técnicas de Silenciamento de Genes , Humanos , Immunoblotting , Indóis/análise , Indóis/farmacologia , Microscopia Confocal , Mitoxantrona/farmacologia , Proteínas de Neoplasias/antagonistas & inibidores , Transporte Proteico/efeitos dos fármacos , RNA Interferente Pequeno/metabolismo , Frações Subcelulares/efeitos dos fármacos , Frações Subcelulares/metabolismoRESUMO
The objective was to determine the cytochrome P450s (CYPs) responsible for the stereoselective and regiospecific hydroxylation of ketamine [(R,S)-Ket] to diastereomeric hydroxyketamines, (2S,6S;2R,6R)-HK (5a) and (2S,6R;2R,6S)-HK (5b) and norketamine [(R,S)-norKet] to hydroxynorketamines, (2S,6S;2R,6R)-HNK (4a), (2S,6R;2R,6S)-HNK (4b), (2S,5S;2R,5R)-HNK (4c), (2S,4S;2R,4R)-HNK (4d), (2S,4R;2R,4S)-HNK (4e), (2S,5R;2R,5S)-HNK (4f). The enantiomers of Ket and norKet were incubated with characterized human liver microsomes (HLMs) and expressed CYPs. Metabolites were identified and quantified using LC/MS/MS and apparent kinetic constants estimated using single-site Michaelis-Menten, Hill or substrate inhibition equation. 5a was predominantly formed from (S)-Ket by CYP2A6 and N-demethylated to 4a by CYP2B6. 5b was formed from (R)- and (S)-Ket by CYP3A4/3A5 and N-demethylated to 4b by multiple enzymes. norKet incubation produced 4a, 4c and 4f and minor amounts of 4d and 4e. CYP2A6 and CYP2B6 were the major enzymes responsible for the formation of 4a, 4d and 4f, and CYP3A4/3A5 for the formation of 4e. The 4b metabolite was not detected in the norKet incubates. 5a and 4b were detected in plasma samples from patients receiving (R,S)-Ket, indicating that 5a and 5b are significant Ket metabolites. Large variations in HNK concentrations were observed suggesting that pharmacogenetics and/or metabolic drug interactions may play a role in therapeutic response.
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Anestésicos Dissociativos/metabolismo , Sistema Enzimático do Citocromo P-450/metabolismo , Ketamina/análogos & derivados , Anestésicos Dissociativos/uso terapêutico , Síndromes da Dor Regional Complexa/tratamento farmacológico , Humanos , Hidroxilação , Ketamina/metabolismo , Ketamina/uso terapêuticoRESUMO
Race and ethnicity can contribute to differences in drug exposure and/or response. Here, we report that about 10% of the new molecular entities (NMEs) approved between 2014 and 2019 by the US Food and Drug Administration's Center for Drug Evaluation and Research showed differences in exposure and/or response based on race/ethnicity or pharmacogenetic factors known to vary in frequency across global populations. Fewer NMEs (10%) reported differences in the labeling in 2014 to 2019 when compared to about 21% of NMEs approved between 2008 and 2013 that had differences in pharmacokinetics, safety, response, and/or pharmacogenetics. Understanding the underlying mechanisms that lead to such differences and adequate enrollment of racial and ethnic subgroups is essential to obtain sufficient information on exposure and response. Though drug development is global, when heterogeneous populations are not adequately enrolled, the risk-benefit assessments can remain incomplete for certain subgroups. Consequently, this can result in regional differences in drug approval, population-specific prescribing recommendations, or need for additional postmarketing studies to address concerns related to exposure, response, or lack of representation that lead to gaps in information.
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Aprovação de Drogas , Etnicidade , Desenvolvimento de Medicamentos , Humanos , Preparações Farmacêuticas , Farmacogenética , Estados Unidos , United States Food and Drug AdministrationRESUMO
Clinical pharmacology is an integral discipline supporting the development, regulatory evaluation, and clinical use of drugs for the treatment of both common and rare diseases. Here, we evaluated the recommendations and information available from select clinical pharmacology studies in the therapeutic product labeling of new molecular entities (NMEs) approved from 2017 to 2019 for both common and rare diseases. A total of 151 NMEs, including 72 orphan and 79 non-orphan drugs, were analyzed for recommendations and information available related to food-drug interaction, drug-drug interaction, renal impairment, hepatic impairment, QT assessment, and human radiolabeled mass balance studies using data collected from the original labeling and other regulatory documents. The analysis showed no statistically significant difference in the recommendations between orphan and non-orphan drugs except for renal impairment related recommendations in section 8 of the labeling. Although not significant, fewer hepatic impairment labeling recommendations were available for orphan drugs when compared with non-orphan drugs. At the time of initial approval, 79 postmarketing requirements (PMRs) and postmarketing commitments (PMCs) for 33 orphan drugs and 39 PMRs and PMCs for 19 non-orphan drugs were established; with most difference observed for drug-drug interaction, hepatic impairment, and QT assessment. Overall, although there was a trend for more labeling recommendations and fewer postmarketing studies and clinical trials for non-orphan drugs, there appeared to be no substantial differences in how these select clinical pharmacology studies are leveraged during the development and approval of orphan and non-orphan drugs.
Assuntos
Aprovação de Drogas , Farmacologia Clínica , Estados Unidos , Humanos , United States Food and Drug Administration , Doenças Raras/tratamento farmacológico , Rotulagem de ProdutosRESUMO
The objective of this study was to assess how solubility and dissolution profile comparisons under different pH conditions can be used to predict gastric pH-mediated drug-drug interaction (DDI) potential. We collected information for new molecular entities (NMEs) approved from 2003 to 2019 by the U.S. Food and Drug Administration (FDA) that had dedicated clinical DDI studies with acid-reducing agents (ARAs). Among these, 67 NMEs with solubility under different pHs and dissolution profiles generated in pH 1.2, 4.5, and 6.8 aqueous media were included for analysis. Similarity factor (f2) was used to compare dissolution profiles at different pHs for pH-mediated DDI prediction (e.g., f2<50 predicts positive DDI). Prediction accuracy was calculated based on the outcome comparison between predicted and observed DDIs. Based on dissolution profile comparisons and observed DDI data, weak base drugs (WBDs) (n = 49) showed 72.5% prediction accuracy under the fasted conditions, and 66.7% prediction accuracy under fed conditions. While using solubility and clinical dose for prediction, the prediction accuracy was 80% under fasted conditions and 66.7% under fed conditions, respectively. Comparison of dissolution profiles generated at pH 1.2, 4.5, and 6.8 can be used to predict gastric pH-mediated DDI potential for WBDs. It demonstrated comparable prediction accuracy under both fasted and fed conditions when compared to the prediction using solubility and clinical dose. Furthermore, dissolution profile comparison could add an additional understanding of possible impact of pH change on the release behavior of the drug product. Graphical abstract.