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1.
J Pharmacol Toxicol Methods ; 101: 106654, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31730936

RESUMO

Any adverse event is reliant on three properties: the appropriate pharmacology to trigger the event, the appropriate exposure of compound, and intrinsic patient factors. Each alone is necessary but insufficient to predict the event. The Comprehensive in vitro Proarrhythmia Assessment (CiPA) initiative attempts to predict the risk of torsade de pointes (TdP) by focusing on an in-silico model with thresholds determined at modest multiples of the therapeutic exposure for the parent molecule. This emphasizes the pharmacologic properties necessary for TdP but does not account for situations where clinical exposure may be higher, or where hERG potassium channel active metabolites are involved. Could accounting for clinical worst-case scenarios and metabolites, as is already standard practice in thorough QTc studies, improve the prediction algorithm? Terfenadine, a drug classed as "Intermediate" risk by CiPA, was assessed differently in the in-silico model validation. The clinical concentration of terfenadine used for the model was the exposure in the presence of metabolic inhibition representing a 14 to 40-fold increase in exposure compared to the therapeutic plasma concentration. However, several other "Intermediate" risk compounds are also known to be sensitive to metabolic inhibition and/or to have therapeutically active major metabolites, some of which are known to block hERG. Risperidone and astemizole are relevant examples. If only parent exposure is used to calculate a therapeutic window, risperidone has a relatively large multiple between clinical exposure and the hERG potency. Using this exposure of risperidone, the drug borders the "Intermediate" and "Low/No" risk categories for the CiPA in-silico model's TdP metric. The desmethyl metabolite of astemizole likely contributes significantly to the effects on cardiac repolarization, being equipotent on hERG but circulating at much higher levels than parent. Recalculating the TdP metric and margin values for terfenadine, risperidone and astemizole using the unbound concentration normally associated with treatment and a clinical worst case changes the qNet metric to higher risk values and illustrates the potential benefit to the algorithm of consistently using a clinical high exposure scenario accounting for all "hERG-active species". This exercise suggests repeating the model qualification accounting for clinical exposures and metabolites under 'stressed' scenarios would improve prediction of the TdP risk.

2.
Clin Pharmacol Ther ; 107(1): 102-111, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31709525

RESUMO

This white paper presents principles for validating proarrhythmia risk prediction models for regulatory use as discussed at the In Silico Breakout Session of a Cardiac Safety Research Consortium/Health and Environmental Sciences Institute/US Food and Drug Administration-sponsored Think Tank Meeting on May 22, 2018. The meeting was convened to evaluate the progress in the development of a new cardiac safety paradigm, the Comprehensive in Vitro Proarrhythmia Assay (CiPA). The opinions regarding these principles reflect the collective views of those who participated in the discussion of this topic both at and after the breakout session. Although primarily discussed in the context of in silico models, these principles describe the interface between experimental input and model-based interpretation and are intended to be general enough to be applied to other types of nonclinical models for proarrhythmia assessment. This document was developed with the intention of providing a foundation for more consistency and harmonization in developing and validating different models for proarrhythmia risk prediction using the example of the CiPA paradigm.

3.
J Pharmacol Toxicol Methods ; 99: 106603, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31247306

RESUMO

INTRODUCTION: Scientists are increasingly in a position to ask whether or not to adopt new technologies. We present a visualization tool to help scientists swiftly evaluate the worth of new assays. METHODS: The parameters (prevalence, sensitivity and, specificity) relevant to use of a toxicity test have values between 0 and 1. The proper arrangement of the parameters can be used to define areas in a [0,1] × [0, 1] cross-space. Our analogy is a square plot of land subdivided into smaller lots. We call the resultant graphic a real estate diagram. RESULTS: We use the well studied example of predicting prolongation of the QT interval of the electrocardiogram to illustrate the diagrams. The experience in human clinical Thorough QT (TQT) studies has been described (Park et al., 2013). Within the data we chose two chronological sets: 2 blocks of two years (2005-2006 and 2011-2012). In the first block 13 of 29 (45%) submitted compounds had positive TQT studies; in the second block the prevalence was 4 of 42 (10%). In other studies, the hERG channel patch-clamp assay used in predicting TQT outcome had an expected sensitivity of 0.70 and expected specificity of 0.72. Real estate diagrams were constructed to yield insight into the positive and negative predictive value (PPV and NPV, respectively) of the TQT prediction. The structure of the real estate diagrams revealed that increasing assay sensitivity in the face of declining prevalence would have a trivial effect on PPV and NPV. DISCUSSION: Nonclinical safety scientists will be called upon to question whether a new technology has the potential to meaningfully increase the predictive value of testing regimens. The real-estate diagram is a useful tool in making that assessment.

4.
J Pharmacol Toxicol Methods ; 88(Pt 1): 85-91, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28797763

RESUMO

INTRODUCTION: Safety pharmacology is a growing discipline with scientists broadly distributed across international geographical regions. This electronic salary survey is the first to be distributed amongst the entire Safety Pharmacology Society (SPS) membership. An electronic survey was sent to all members of the Society. Categorical survey questions assessed membership employment types, annual incomes, and professional certifications, along with other associated career attributes. METHODS: This survey was distributed to the SPS membership that is comprised of safety pharmacologists, toxicologists and pharmacologists working globally in the pharmaceutical industry, at contract research organizations (CRO), regulatory agencies, and academia or within the technology provider industry. The survey was open for responses from December 2015 to March 2016. RESULTS: The survey response rate was 28% (129/453). North America (68%) was the region with the largest number of respondents followed by Europe (28%). A preponderance of respondents (77%) had 12years of industry experience or more. 52% of responders earned annually between $40,000 and $120,000. As expected, salary was generally positively correlated with the number of years of experience in the industry or the educational background but there was no correlation between salary and the number of employee's directly supervised. The median salary was higher for male vs female respondents, but so was median age, indicative of no gender 'salary gap'. DISCUSSION: Our 2016 SPS salary survey results showcased significant diversity regarding factors that can influence salary compensation within this discipline. These data provided insights into the complex global job market trends. They also revealed the level of scientific specialization embedded within the organization, presently uniquely positioned to support the dynamic career paths of current and future safety pharmacologists.


Assuntos
Farmacologia/economia , Salários e Benefícios/estatística & dados numéricos , Sociedades/economia , Toxicologia/economia , Adulto , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Inquéritos e Questionários
5.
J Pharmacol Toxicol Methods ; 86: 34-43, 2017 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-28223123

RESUMO

INTRODUCTION: The Safety Pharmacology Society (SPS) has conducted a survey of its membership to identify industry practices related to testing considered in the Comprehensive In vitro Proarrhythmia Assay (CiPA). METHODS: Survey topics included nonclinical approaches to address proarrhythmia issues, conduct of in silico studies, in vitro ion channel testing methods used, drugs used as positive controls during the conduct of cardiac ion channel studies, types of arrhythmias observed in non-clinical studies and use of the anticipated CiPA ion channel assay. RESULTS: In silico studies were used to evaluate effects on ventricular action potentials by only 15% of responders. In vitro assays were used mostly to assess QT prolongation (95%), cardiac Ca2+ and Na+ channel blockade (82%) and QT shortening or QRS prolongation (53%). For de-risking of candidate drugs for proarrhythmia, those assays most relevant to CiPA including cell lines stably expressing ion channels used to determine potency of drug block were most frequently used (89%) and human stem cell-derived or induced pluripotent stem cell cardiomyocytes (46%). Those in vivo assays related to general proarrhythmia derisking include ECG recording using implanted telemetry technology (88%), jacketed external telemetry (62%) and anesthetized animal models (53%). While the CiPA initiative was supported by 92% of responders, there may be some disconnect between current practice and future expectations, as explained. DISCUSSION: Proarrhythmia liability assessment in drug development presently includes study types consistent with CiPA. It is anticipated that CiPA will develop into a workable solution to the concern that proarrhythmia liability testing remains suboptimal.


Assuntos
Arritmias Cardíacas/induzido quimicamente , Animais , Bloqueadores dos Canais de Cálcio/farmacologia , Simulação por Computador , Indústria Farmacêutica , Eletrocardiografia/efeitos dos fármacos , Humanos , Células-Tronco Pluripotentes Induzidas , Canais Iônicos , Síndrome do QT Longo/induzido quimicamente , Miócitos Cardíacos/efeitos dos fármacos , Bloqueadores dos Canais de Sódio/farmacologia , Inquéritos e Questionários , Telemetria
6.
Handb Exp Pharmacol ; 229: 291-321, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26091645

RESUMO

Although the basic structure of the gastrointestinal tract (GIT) is similar across species, there are significant differences in the anatomy, physiology, and biochemistry between humans and laboratory animals, which should be taken into account when conducting a gastrointestinal (GI) assessment. Historically, the percentage of cases of drug attrition associated with GI-related adverse effects is small; however, this incidence has increased over the last few years. Drug-related GI effects are very diverse, usually functional in nature, and not limited to a single pharmacological class. The most common GI signs are nausea and vomiting, diarrhea, constipation, and gastric ulceration. Despite being generally not life-threatening, they can greatly affect patient compliance and quality of life. There is therefore a real need for improved and/or more extensive GI screening of candidate drugs in preclinical development, which may help to better predict clinical effects. Models to identify drug effects on GI function cover GI motility, nausea and emesis liability, secretory function (mainly gastric secretion), and absorption aspects. Both in vitro and in vivo assessments are described in this chapter. Drug-induced effects on GI function can be assessed in stand-alone safety pharmacology studies or as endpoints integrated into toxicology studies. In silico approaches are also being developed, such as the gut-on-a-chip model, but await further optimization and validation before routine use in drug development. GI injuries are still in their infancy with regard to biomarkers, probably due to their greater diversity. Nevertheless, several potential blood, stool, and breath biomarkers have been investigated. However, additional validation studies are necessary to assess the relevance of these biomarkers and their predictive value for GI injuries.


Assuntos
Descoberta de Drogas , Avaliação Pré-Clínica de Medicamentos/métodos , Trato Gastrointestinal/efeitos dos fármacos , Animais , Biomarcadores , Controle de Medicamentos e Entorpecentes , Esvaziamento Gástrico/efeitos dos fármacos , Motilidade Gastrointestinal/efeitos dos fármacos , Trato Gastrointestinal/anatomia & histologia , Trato Gastrointestinal/fisiologia , Humanos , Absorção Intestinal/efeitos dos fármacos
7.
Am Heart J ; 170(1): 23-35, 2015 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-26093861

RESUMO

Thorough QT studies conducted according to the International Council on Harmonisation E14 guideline are required for new nonantiarrhythmic drugs to assess the potential to prolong ventricular repolarization. Special considerations may be needed for conducting such studies with antidiabetes drugs as changes in blood glucose and other physiologic parameters affected by antidiabetes drugs may prolong the QT interval and thus confound QT/corrected QT assessments. This review discusses potential mechanisms for QT/corrected QT interval prolongation with antidiabetes drugs and offers practical considerations for assessing antidiabetes drugs in thorough QT studies. This article represents collaborative discussions among key stakeholders from academia, industry, and regulatory agencies participating in the Cardiac Safety Research Consortium. It does not represent regulatory policy.


Assuntos
Arritmias Cardíacas/induzido quimicamente , Sistema de Condução Cardíaco/anormalidades , Hipoglicemiantes/efeitos adversos , Síndrome do QT Longo/induzido quimicamente , Síndrome de Brugada , Doença do Sistema de Condução Cardíaco , Inibidores da Dipeptidil Peptidase IV/efeitos adversos , Eletrocardiografia , Receptor do Peptídeo Semelhante ao Glucagon 1 , Inibidores de Glicosídeo Hidrolases , Ventrículos do Coração , Humanos , Técnicas de Patch-Clamp , Receptores de Glucagon/agonistas , Inibidores do Transportador 2 de Sódio-Glicose , Compostos de Sulfonilureia/efeitos adversos , Tiazolidinedionas/efeitos adversos , Função Ventricular
8.
Artigo em Inglês | MEDLINE | ID: mdl-25959882

RESUMO

As with other professional disciplines there is a growing need from within industry as well as global regulatory authorities for implementation of a certification process in order to assure that appropriate expertise is developed and quality standards are identified for professionals involved in the practice of Safety Pharmacology (SP). In order to meet this need, the Safety Pharmacology Society (SPS) has developed the Diplomate in Safety Pharmacology (DSP) certification process. There are many benefits to certification including authentication of the discipline within the overall pharmaceutical community and with regulatory authorities. It also encourages participation in SPS activities by other professionals (toxicologists, clinicians, academics) who wish to broaden their professional expertise. It provides an opportunity for candidates to strengthen their fundamental scientific knowledge, and stimulates the sharing of data, methods and model development in the form of publications and presentations on relevant topics in SP. Accreditation in SP occurs after candidates successfully complete a written certification examination conducted at the annual SPS meeting. The DSP exam consists primarily of material pertinent to the conduct of SP vital function core battery studies (i.e., cardiovascular, respiratory and central nervous systems), supplemental SP studies (i.e., renal/urinary, gastrointestinal, immunology, and hematology), Regulatory Guidelines (ICH Guidelines) as well as relevant cross-functional knowledge (e.g., physiology, pharmacology, toxicology, biochemistry, pathology, pharmacokinetics, dosing formulation, analytical methods, and statistics). Maintenance of the DSP certification results from the accrual of credits which are gained from a range of educational and scientific contributions. Eligibility requirements include a combination of at least a bachelor degree in science and two years of relevant professional SP experience and one poster presentation on a SP topic as first author at a recognized major scientific meeting.


Assuntos
Certificação , Farmacologia/normas , Competência Profissional , Animais , Indústria Farmacêutica/normas , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos , Humanos , Sociedades Científicas/organização & administração
9.
J Pharmacol Toxicol Methods ; 62(2): 143-7, 2010.
Artigo em Inglês | MEDLINE | ID: mdl-20570744

RESUMO

Positive controls have often been used in nonclinical and clinical cardiovascular safety studies to evaluate the study's assay sensitivity with respect to drug-induced prolongation of the QT interval of the electrocardiogram (ECG). If the study is able to detect such QT prolongation by the control, then a finding of negative QT effect of comparable size for the test drug will constitute evidence that the test drug does not in fact prolong the QT interval by the amount of regulatory concern. Current regulatory guidance regarding QT interval prolongation includes ICH S7B (nonclinical) and ICH E14 (clinical). However, the underlying null hypothesis settings of the two documents are quite different. This paper quantitatively evaluates the utility of positive controls in nonclinical and clinical studies in which a test drug and a positive control are simultaneously assessed in a study. The results show that positive controls, when powered at 80%, can be beneficial in 58% of nonclinical QT studies with Prob(S)=0.8; when powered at 90%, positive controls can be beneficial 73% of clinical QT studies with Prob(S)=0.9, where Prob(S) represents the probability of success with no QT risk at the stage of development.


Assuntos
Drogas em Investigação/toxicidade , Eletrocardiografia/efeitos dos fármacos , Animais , Arritmias Cardíacas/induzido quimicamente , Avaliação Pré-Clínica de Medicamentos , Guias como Assunto , Humanos , Sensibilidade e Especificidade
10.
J Pharmacol Toxicol Methods ; 61(2): 178-91, 2010.
Artigo em Inglês | MEDLINE | ID: mdl-20172036

RESUMO

INTRODUCTION: Pharmacological inhibition of cardiac potassium channels encoded by hERG (human ether-à-go-go-related gene) is associated with QT interval prolongation and torsades de pointes arrhythmia. Electrophysiological assays of hERG channel inhibition are integral to the safety testing of novel drug candidates. This study was conducted to compare, for the high affinity hERG inhibitors dofetilide and cisapride, hERG blockade between action potential (AP) and conventional (step and step-ramp) screening waveforms. Furthermore, it evaluated dynamic (pulse-by-pulse) protocol-dependence of hERG channel inhibition by these drugs. METHODS: Whole-cell patch-clamp recordings were made at 37 degrees C from hERG-expressing HEK 293 cells. Half-maximal inhibitory concentrations (IC(50) values) for I(hERG) blockade were obtained using conventional voltage clamp and action potential clamp, using previously digitised ventricular and Purkinje fibre (PF) AP waveforms. RESULTS: A more marked variation in IC(50) values with different command waveforms was observed for cisapride (ranging from 7 to 72 nM) than for dofetilide (ranging from 4 to 15 nM), with higher IC(50)s obtained with AP than step or step-ramp commands. The two drugs differed little from one another in effects on voltage-dependent activation; however, I(hERG) blockade by each drug was initially voltage-dependent, but at steady-state was only voltage-dependent for cisapride. There was comparatively little difference between the two drugs in effects on I(hERG) availability or time constants of development of inactivation. Features of time-dependence of blockade and the use of protocols employing varying rest periods in drug or commands of alternating duration highlighted a pronounced ability of cisapride, but not dofetilide, to dissociate and reassociate from hERG on a pulse-by-pulse basis. DISCUSSION: Protocols described here that demonstrated dynamic variation (drug dissociation/reassociation) in hERG channel current blockade at 37 degrees C for cisapride may have future value for investigating drug interactions with the hERG channel. Downloadable digitised ventricular and PF AP waveforms that can be used in AP clamp experiments also accompany this article.


Assuntos
Cisaprida/farmacologia , Canais de Potássio Éter-A-Go-Go/antagonistas & inibidores , Fenetilaminas/farmacologia , Bloqueadores dos Canais de Potássio/farmacologia , Sulfonamidas/farmacologia , Potenciais de Ação/efeitos dos fármacos , Axônios/efeitos dos fármacos , Linhagem Celular , Cisaprida/metabolismo , Interpretação Estatística de Dados , Eletrofisiologia , Canais de Potássio Éter-A-Go-Go/metabolismo , Humanos , Técnicas de Patch-Clamp , Fenetilaminas/metabolismo , Bloqueadores dos Canais de Potássio/metabolismo , Sulfonamidas/metabolismo , Temperatura Ambiente
11.
J Pharmacol Toxicol Methods ; 58(2): 118-28, 2008.
Artigo em Inglês | MEDLINE | ID: mdl-18619862

RESUMO

INTRODUCTION: Adrenergic blockade as a treatment for chronic heart failure (CHF) has proved effective, but its pharmacological mechanism on CHF remains unclear. In the past two decades, studies on heart rate variability (HRV) have reported that CHF patients generally have a reduced temporal complexity in heart rate variability. On the other hand, adrenergic blockers have been shown to restore such complexity. Fractal analysis is a novel and efficient tool to explore the adrenergic blockade effect on HRV. This paper applies the detrended fluctuation analysis (DFA) and multifractal DFA (MF-DFA) methods in an attempt to understand the effect of adrenergic blockade on cardiac dynamics in conscious beagle dogs. METHODS: DFA and MF-DFA analysis are conducted on RR interval data generated from telemetry instrumented dogs receiving a combination of 15 mg/kg nadolol and 5 mg/kg phenoxybenzamine orally administered at the 22nd and 34th hour in a parallel design (n=12). All dogs had approximately 48 h of beat-to-beat heart rate measurements recorded in the left ventricle. Complexity measures for heartbeat series are compared between the blocker and vehicle group. We also compute traditional statistics for HRV and spectral parameters and examine their correlation with fractal analysis. RESULTS: When compared to the vehicle group, the adrenergic blocker group had: 1) longer RR intervals (p=0.02) and lower beat-to-beat variability (p=0.04); 2) decreased low frequency (LF) and high frequency (HF) power (p=0.03), and higher LF-to-HF ratio; 3) larger middle-range scaling exponents (p<0.01); 4) broader multifractal spectra (p=0.03) with higher dominant singularity indices (p=0.02). DISCUSSION: Our results show that 1) adrenergic blockade alters the sympathovagal balance; 2) adrenergic blockers enhance the complexity of the cardiac dynamics; 3) the adrenergic blockade effect on cardiac dynamics is primarily the attenuation of small fluctuations in RR intervals. Fractal analysis also has the potential to be applied to early QT diagnosis.


Assuntos
Antagonistas Adrenérgicos alfa/efeitos adversos , Antagonistas Adrenérgicos beta/efeitos adversos , Frequência Cardíaca/efeitos dos fármacos , Nadolol/efeitos adversos , Fenoxibenzamina/efeitos adversos , Administração Oral , Animais , Pressão Sanguínea , Cães , Nadolol/administração & dosagem , Fenoxibenzamina/administração & dosagem
12.
J Pharmacol Toxicol Methods ; 56(2): 194-202, 2007.
Artigo em Inglês | MEDLINE | ID: mdl-17583537

RESUMO

INTRODUCTION: Purkinje fibre repolarisation assays are valuable tools for identifying compounds which affect cardiac ion channels. The throughput of compound testing in this assay is low therefore we designed a novel recording system to improve screening and animal tissue usage efficiencies. METHODS: The system was used to evaluate compounds using standard sharp microelectrode techniques. Animal tissue usage efficiencies were quantified by adding up the total number of Purkinje fibres from which recordings were attempted and dividing this by the number of experimental data sets generated, to arrive at a 'fibres per data set' ratio. Test compounds were dofetilide (3 x 10(-10) to 10(-8) M), cisapride (10(-8) to 3 x 10(-7) M), terodiline (10(-6) to 3 x 10(-5) M) and verapamil (3 x 10(-7) to 10(-5) M). RESULTS: Using the novel modified system, 21 data sets were generated from 29 fibres, compared to 24 data sets from 41 fibres using the conventional manual recording system, demonstrating a 24% improvement in the efficiency of animal tissue usage. Comparing data from the manual and modified systems revealed differences in absolute values for all parameters including APD90 (308.73 +/- 9.97 ms, n = 24, compared to 275.27 +/- 8.25 ms, n = 21, respectively; P < 0.05). Differences in the magnitude of changes in action potential parameters between the systems were also evident for all compounds including terodiline (1 x 10(-5) M) which caused a -27.1 +/- 16.5% reduction in APD50 in the manual system, compared to a - 55.2 +/- 2.2% reduction in the modified system. DISCUSSION: Although the value of the present study is limited by the small sample sizes, it has demonstrated utility of the modified system in improving efficiency of animal tissue usage. It offers potential utility in a higher throughput screening environment for examining the electrophysiological properties of novel compounds in native cardiac tissues, particularly where functional patch clamp data are limited.


Assuntos
Técnicas Eletrofisiológicas Cardíacas/métodos , Coração/efeitos dos fármacos , Canais Iônicos/antagonistas & inibidores , Ramos Subendocárdicos/efeitos dos fármacos , Potenciais de Ação/efeitos dos fármacos , Potenciais de Ação/fisiologia , Animais , Antiarrítmicos/farmacologia , Butilaminas/farmacologia , Bloqueadores dos Canais de Cálcio/farmacologia , Cisaprida/farmacologia , Cães , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos/instrumentação , Avaliação Pré-Clínica de Medicamentos/métodos , Estimulação Elétrica , Técnicas Eletrofisiológicas Cardíacas/instrumentação , Feminino , Coração/fisiologia , Técnicas In Vitro , Canais Iônicos/fisiologia , Síndrome do QT Longo/diagnóstico , Síndrome do QT Longo/fisiopatologia , Masculino , Microeletrodos , Fenetilaminas/farmacologia , Ramos Subendocárdicos/fisiologia , Reprodutibilidade dos Testes , Sulfonamidas/farmacologia , Verapamil/farmacologia
13.
Biochem Biophys Res Commun ; 351(1): 273-80, 2006 Dec 08.
Artigo em Inglês | MEDLINE | ID: mdl-17056009

RESUMO

The phenothiazine antipsychotic agent thioridazine has been linked with prolongation of the QT interval on the electrocardiogram, ventricular arrhythmias, and sudden death. Although thioridazine is known to inhibit cardiac hERG K(+) channels there is little mechanistic information on this action. We have investigated in detail hERG K(+) channel current (I(hERG)) blockade by thioridazine and identified a key molecular determinant of blockade. Whole-cell I(hERG) measurements were made at 37 degrees C from human embryonic kidney (HEK-293) cells expressing wild-type and mutant hERG channels. Thioridazine inhibited I(hERG) tails at -40mV following a 2s depolarization to +20mV with an IC(50) value of 80nM. Comparable levels of I(hERG) inhibition were seen with physiological command waveforms (ventricular and Purkinje fibre action potentials). Thioridazine block of I(hERG) was only weakly voltage-dependent, though the time dependence of I(hERG) inhibition indicated contingency of blockade upon channel gating. The S6 helix point mutation F656A almost completely abolished, and the Y652A mutation partially attenuated, I(hERG) inhibition by thioridazine. In summary, thioridazine is one of the most potent hERG K(+) channel blockers amongst antipsychotics, exhibiting characteristics of a preferential open/activated channel blocker and binding at a high affinity site in the hERG channel pore.


Assuntos
Clorpromazina/administração & dosagem , Canais de Potássio Éter-A-Go-Go/fisiologia , Ativação do Canal Iônico/fisiologia , Rim/fisiologia , Potenciais da Membrana/fisiologia , Antipsicóticos/administração & dosagem , Linhagem Celular , Relação Dose-Resposta a Droga , Canal de Potássio ERG1 , Canais de Potássio Éter-A-Go-Go/química , Humanos , Rim/efeitos dos fármacos , Potenciais da Membrana/efeitos dos fármacos , Estrutura Terciária de Proteína , Relação Estrutura-Atividade
14.
J Pharmacol Toxicol Methods ; 54(2): 183-8, 2006.
Artigo em Inglês | MEDLINE | ID: mdl-16567113

RESUMO

INTRODUCTION: To account for heart rate-induced changes in the QT interval, correction formulas are generally applied to normalize the QT interval for heart rate. None of these formulas is entirely accurate because correction or normalization of any parameter in biology may introduce an additional source of variation in estimating the parameter. In this article, a one-step approach for the statistical analysis of the QT interval was proposed based on modeling the functional relationship between the QT interval and heart rate. METHODS: The QT-HR relationship was incorporated into the statistical analysis to provide a model-based correction. This was accomplished by including HR as a covariate in the QT interval analysis. The approach was demonstrated using data generated from Lilly Research Laboratories. We compared the false positive rate and statistical power of QT, QTcF, and the proposed one-step method. RESULTS: We found the one-step method demonstrated the greatest sensitivity in detecting a QT interval change without an increase in the false positive rate. It was shown that the one-step QT analysis could detect a 5%-6% increment of the QT interval. This is approximately equivalent to an increase of 11-13 ms in QT interval in beagle dogs. DISCUSSION: Several advantages and unique features of the one-step method are discussed. These include evaluating treatment effect on QT without applying a heart rate correction formula and estimating QT difference flexibly at any selected heart rate. In addition to the linear QT-HR relationship, other functional relationships can be easily implemented to this approach.


Assuntos
Síndrome do QT Longo/fisiopatologia , Telemetria , Algoritmos , Animais , Cães , Relação Dose-Resposta a Droga , Eletrocardiografia/efeitos dos fármacos , Reações Falso-Positivas , Frequência Cardíaca/efeitos dos fármacos , Síndrome do QT Longo/induzido quimicamente
15.
Biochem Biophys Res Commun ; 341(2): 500-6, 2006 Mar 10.
Artigo em Inglês | MEDLINE | ID: mdl-16446155

RESUMO

The HERG potassium channel might have a non-canonical drug binding site, distinct from the channel's inner cavity, that could be responsible for elements of closed-state pharmacological inhibition of the channel. The macrolide antibiotic erythromycin is a drug that may block unconventionally because of its size. Here we used whole-cell patch-clamp recording at 37 degrees C from heterologously expressed HERG channels in a mammalian cell line to show that erythromycin either produces a rapid open-state-dependent HERG channel inhibition, or components of both open-state-dependent and closed-state-dependent inhibition. Alanine-substitution of HERG's canonical determinants of blockade revealed that Y652 was not important as a molecular determinant of blockade, and that mutation of F656 resulted in only weak attenuation of inhibition. In computer models of the channel, erythromycin could make several direct contacts with F656, but not with Y652, in the open-state model, and erythromycin was unable to fit into a closed-state channel model.


Assuntos
Eritromicina/farmacologia , Canais de Potássio Éter-A-Go-Go/química , Linhagem Celular , Relação Dose-Resposta a Droga , Eletrofisiologia , Canais de Potássio Éter-A-Go-Go/metabolismo , Humanos , Concentração Inibidora 50 , Modelos Biológicos , Modelos Químicos , Modelos Moleculares , Mutação , Técnicas de Patch-Clamp , Canais de Potássio de Abertura Dependente da Tensão da Membrana/química , Conformação Proteica , Inibidores da Síntese de Proteínas/farmacologia , Software , Temperatura Ambiente , Fatores de Tempo
16.
Br J Pharmacol ; 147(8): 905-16, 2006 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-16474415

RESUMO

The fluoroquinolone antibiotic moxifloxacin has been associated with the acquired long QT syndrome and is used as a positive control in the evaluation of the QT-interval prolonging potential of new drugs. In common with other QT-prolonging agents, moxifloxacin is known to inhibit the hERG potassium K+ channel, but at present there is little mechanistic information available on this action. This study was conducted in order to characterise the inhibition of hERG current (I(hERG)) by moxifloxacin, and to determine the role in drug binding of the S6 aromatic amino-acid residues Tyr652 and Phe656. hERG currents were studied using whole-cell patch clamp (at room temperature and at 35-37 degrees C) in an HEK293 cell line stably expressing hERG channels. Moxifloxacin reversibly inhibited currents in a dose-dependent manner. We investigated the effects of different voltage commands to elicit hERG currents on moxifloxacin potency. Using a 'step-ramp' protocol, the IC50 was 65 microM at room temperature and 29 microM at 35 degrees C. When a ventricular action potential waveform was used to elicit currents, the IC50 was 114 microM. Block of hERG by moxifloxacin was found to be voltage-dependent, occurred rapidly and was independent of stimulation frequency. Mutagenesis of the S6 helix residue Phe656 to Ala failed to eliminate or reduce the moxifloxacin-mediated block whereas mutation of Tyr652 to Ala reduced moxifloxacin block by approximately 66%. Our data demonstrate that moxifloxacin blocks the hERG channel with a preference for the activated channel state. The Tyr652 but not Phe656 S6 residue is involved in moxifloxacin block of hERG, concordant with an interaction in the channel inner cavity.


Assuntos
Antibacterianos/farmacologia , Compostos Aza/farmacologia , Canais de Potássio Éter-A-Go-Go/antagonistas & inibidores , Quinolinas/farmacologia , Linhagem Celular , Relação Dose-Resposta a Droga , Canal de Potássio ERG1 , Fluoroquinolonas , Humanos , Moxifloxacina , Ofloxacino/farmacologia
17.
J Pharmacol Exp Ther ; 302(2): 828-33, 2002 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-12130750

RESUMO

QT interval prolongation of the electrocardiogram has been associated with the occurrence of life-threatening fatal ventricular arrhythmias. To understand the relationship between preclinical cardiac conduction assessment to clinical outcome, comparisons of free (unbound)-plasma drug concentrations and their associated effects in the conscious mongrel dog were made to the free plasma concentrations in humans reported to produce QT prolongation. E-4031 (an experimental class III antiarrhythmic), cisapride, terfenadine, terodiline, and verapamil all affect cardiac repolarization and can produce QT prolongation in humans. In the conscious dog, the QT interval was assessed on a beat-to-beat basis in relation to each preceding RR interval at concentrations approximating the same unbound human concentrations. E-4031, cisapride and terodiline statistically increased the QT(RR1000) interval [the QT interval at a 60 beats/min (bpm) heart rate] 23, 8, and 9 ms, respectively, at concentrations 0.3 to 15.8 times their relevant clinical level. Increases were not observed for terfenadine or verapamil (p > 0.05 at all doses). Inspection of individual dog QT versus RR interval relationships showed clear QT interval responses specific to each treatment but not readily apparent when data are averaged at a heart rate of 60 bpm. For specific rectifier K(+) current (IKr) blockers, robust effects on mean QT prolongation can be detected. However, for drugs that affect repolarization through multiple channels, the effect on the mean QT interval may be more difficult to detect. Inspection of the beat-to-beat QT-RR interval relationship in an individual animal can increase the sensitivity for more accurate clinical prediction.


Assuntos
Antiarrítmicos/toxicidade , Frequência Cardíaca/fisiologia , Síndrome do QT Longo/fisiopatologia , Animais , Butilaminas/toxicidade , Cisaprida/toxicidade , Modelos Animais de Doenças , Cães , Feminino , Frequência Cardíaca/efeitos dos fármacos , Síndrome do QT Longo/induzido quimicamente , Masculino , Piperidinas/toxicidade , Piridinas/toxicidade , Terfenadina/toxicidade , Verapamil/toxicidade
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