五、参考文献 1.ICH, ICH Guidance for Industry ICH S7A:Safety Pharmacology Studies for Human Pharmaceuticals, 2001. 2.ICH, ICH Guidance for Industry ICH S7B:Safety Pharmacology Studies for assessing the potential for delayed ventricular repolarization (QT interval prolongation)by Human Pharmaceuticals, 2005. 3.Liu T, Brown BS, Wu Y, Antzelevitch C, Kowey PR, Yan GX.Blinded validation of the isolated arterially perfused rabbit ventricular wedge in preclinical assessment of drug-induced proarrhythmias, Heart Rhythm 2006; 3: 948-956. 六、附录—试验方法 (一)体外试验 评价药物对离子电流的影响,主要包括IKr和其他几种参与心肌电活动的重要离子通道。体外IKr试验是采用原代心肌细胞或表达IKr通道蛋白(如由hERG编码的蛋白)的细胞系评价药物对离子电流的影响。相关研究方法见参考文献。检测中获得IC50的基本参数项目列于表2。 表1 致心律失常作用试验检测的心肌离子通道
通道种类
阳性对照(参考)
参考文献
Cav1.2 (L-type)
尼非地平(Nifedipine)
Shen J et al Comparison of L-type calcium channel blockade by nifedipine and/or cadmium in guinea pig ventricular myocytes JPET, 2000;294:562–70
Zahradnık I, Minarovic, I and Zahradnıkova A Inhibition of the cardiac L-typec calcium channel currentby antidepressant drugs JPET,2008;324:977–84
hERG
西沙必利(Cisapride)或特非那定(Terfenadine)
Helliwell R, Recording hERG potassium currents and assessing the effects of compounds using the whole-cell patch-clamp technique Jonathan D Lippiat (ed ), Methods in Molecular Biology, Potassium Channels, 2008;491: 279–95
Kamiya K et al Molecular determinants of hERG channel block by terfenadine and cisapride J Pharmacol Sci,2008;108:301-7
Gintant GA et al Utility of hERG assays as surrogate markers of delayed cardiac repolarization and QT safety ToxicolPathol, 2006;34:81-90
Kv1.5
4-氨基吡啶(4-Aminopyridine,4-AP)
De Biasi M et al Open channel block of human heart hKv1 5 by the beta-subunit hKv beta 1 2 Am J Physiol 1997 272: H2932-41 Lagrutta A et al Novel, potent inhibitors of human Kv1 5 K+ channels and ultrarapidly activating delayed rectifier potassium current JPET,2006;317:1054-63
Kv4.3
氟卡尼(又名哌氟酰胺,Flecainide)
或4-AP
Radicke S et al Effects of MiRP1 and DPP6 β-subunits on the blockade induced by flecainide of KV4 3/KchIP2 channels Br J Pharmacol 2008, 154: 774–86
Fischer F et al Inhibition of cardiac Kv1 5 and Kv4 3 potassium channels by the class Ia anti-arrhythmic ajmaline: mode of action Naunyn Schmiedebergs Arch Pharmacol, 2013, online, Jul
KvLQT1/minK
二乙酰醇293B (Chromanol-293B)
Yang WP et al KvLQT1, a voltage-gated potassium channel responsible forhuman cardiac arrhythmias PNAS, 1997; 94:4017- 4021
Seebohm G et al Molecular determinants of KCNQ1 channel block by a benzodiazepine Mol Pharmacol, 2003, 64:70-7
Nav1.5
利多卡因(Lidocaine)
Wu L et al Role of late sodium current in modulating the proarrhythmic and antiarrhythmic effects of quinidine Heart Rhythm, 2008;5: 1726-34
McNulty, MM andHanck ,DA State-dependent mibefradil block of Na+ channels Mol Pharmacol, 2004; 66:1652-61
Kir2.1
钡(Barium)或氯喹(Chloroquine)
Rodriguez-Menchaca AA et al , The molecular basis of chloroquine block of the inward rectifier Kir2 1 channel PNAS, 2008;105:1364-8
Schram G et al Barium block of Kir2 and human cardiac inward rectifier currents:evidence for subunit-heteromeric contribution to native currents Cardiovasc Res, 2003, 59: 328-38
Ollerstam, A et al Comparison of the QT interval response during sinus andpaced rhythm in conscious and anesthetized beagle dogs J PharmacolToxicol Methods, 2007;56: 131-44
Chiba K, et al In vivo experimental approach for the risk assessment of fluoroquinolone antibacterial agents-induced long QT syndrome Eur J Pharmacol, 2004;486:189-200
Ishizaka T et al Evaluation of drug-induced QT prolongation in a halothane-anesthetized monkey model: effects of sotalol J PharmacolToxicol Methods, 2009;59:86-93
猪
Authier S et al Cardiovascular and respiratory safety pharmacology in Göttingenminipigs: Pharmacological characterization JpharmacolToxicol Methods, 2011;64:53-9
遥
测
动
物
犬
Batey AJ, Doe PA A method for QT correction based on beat-to-beat analysis of the QT/RR interval relationship in conscious telemetred beagle dogs J PharmacolToxicol Methods,2002;48: 11-9
Fossa AA Assessing QT prolongation in conscious dogs: validation of a beat-to-beat method PharmacolTher, 2008;118:231-8
猴
Ando K et al QT PRODACT: In vivo QT assay with a consciousmonkey for assessment of the potential for drug-induced Qtinterval prolongation J Pharmacol Sci,2005;99: 487-500
Haushalter TM et al The cardiovascular and pharmacokineticprofile of dofetilide in conscious telemetered beagle dogs andcynomolgus monkeys Br J Pharmacology,2008;154: 1457-64
猪
Stubhan et al Evaluation of cardiovascular and ECG parameters in the normal, freely moving GöttingenMinipig J Pharmacol Toxicological Methods,2008, 57: 202–11
KanoM et al QT PRODACT: Usabilityof miniature pigs in safety pharmacology studies: Assessment for drug-inducedQT interval prolongation J Pharmacol Sci,2005, 99:501−11
麻
醉
动
物
犬
Takahara A et al Effects of mexiletine on the canine model of sparfloxacin-induced long QT syndrome Eur J Pharmacol,2003,476:115-22
Chiba K et al Proarrhythmic effects of fluoroquinolone antibacterial agents: in vivo effects as physiologic substrate for Torsades ToxicolApplPharmacol, 2000, 169:8-16
Kimura K et al Hemodynamic and electrophysiological effects of mitemcinal (GM-611), a novel prokinetic agent derived from erythromycin in a halothane-anesthetized canine model J ToxicolSci, 2007,32:231-9
Terrar, DA et al Comparison of guinea-pig ventricular myocytes and dog Purkinje fibres for in vitro assessment of drug-induced delayed repolarization J Pharmacol Toxicol Methods, 2007; 56: 171-85
Lightbown ID et al Towards automation of a valuable preclinical cardiac safety pharmacology assay: Evaluation of the effects of cardiac ion channel blockers on cardiac repolarisation in vitro J Pharmacol Toxicol Methods, 2007;56:194-202
Limberis JT et al The effects of plasma proteins on delayed repolarization in vitro with cisapride, risperidone, and D, L-sotalol J Pharmacol Toxicol Methods, 2007; 56:11-7
兔蒲肯野纤维
Himmel HM Suitability of commonly used excipients for electrophysiological in-vitrosafety pharmacology assessment of effects on hERG potassiumcurrent and on rabbit Purkinje fiber action potential J Pharmacol Toxicol Methods, 2007; 56: 145-58
Ducroq J et al Action potential experiments complete hERG assay and QT-interval measurements in cardiac preclinical studies J Pharmacol Toxicol Methods, 2007; 159-70
豚鼠心室肌细胞
Hagiwara T et al A comparative study of the fluoroquinolone antibacterial agents on the action potential duration in guinea pig ventricular myocardia Jpn J Pharmacol, 2001; 87: 231–34
Shuba LM et al Action potentials, contraction, and membrane currents in guinea pig ventricular preparations treated with the antispasmodic agent terodiline JPET, 1999; 290:1417-26
豚鼠心室乳头肌
Hayashi S et al QT PRODACT: a multi-site study of in vitro action potential assays on 21 compounds in isolated guinea-pig papillary muscles J PharmacolSci,2005; 99:423-37
Langendorff离体心脏
Milberg P et al Proarrhythmia as a class effect of quinolones: increased dispersion of repolarization and triangulation of action potential predict torsades de pointes J Cardiovasc Electrophysiol, 2007; 18:647-54
Clements-Jewery H et al Actions of flecainide on susceptibility to phase-2 ventricular arrhythmias during infarct evolution in rat isolated perfused hearts British J Pharmacol, 2006; 147: 468–75
ChengHC and Incardona J Models of torsades de pointes: effects of FPL64176, DPI201106, dofetilide, and chromanol 293B in isolated rabbit and guinea pig hearts J Pharmacol Toxicol Methods, 2009; 60: 174-84
冠状动脉灌注兔左心室肌楔形标本
ChenX et al Use of arterially perfused rabbit ventricular wedge inpredicting arrhythmogenic potentials of drugs J Pharmacol Toxicol Methods, 2006;54: 261-72
Liu T, Brown BS, Wu Y, Antzelevitch C, Kowey PR, Yan GX.Blinded validation of the isolated arterially perfused rabbit ventricular wedge in preclinical assessment of drug-induced proarrhythmias Heart Rhythm 2006; 3: 948-956
上述研究相关试验建议最大可能执行GLP
表5 心肌动作电位试验中的重要参数项目
三浓度检测参数项目
测试
测试浓度数
3
检测每一个受试物所需的蒲肯野纤维数或心肌细胞数
4
用于对照品的蒲肯野纤维数或心肌细胞数
4
累积曲线
+
在生理温度 (37º±1º C) 下进行试验
+
统计学分析
+
如果系GLP实验室,收集样本的分析
+
报告
+
如果是GLP实验室,要求有质量保证部门(Quality Assurence Unit, QAU)的检查,包括同步、数据和报告的检查