An evaluation of hERG current assay performance: Translating preclinical safety studies to clinical QT prolongation
Introduction
Delayed cardiac repolarization is a well-accepted clinical surrogate marker of Torsades-de-Pointes arrhythmia, a rare but potentially lethal ventricular arrhythmia. Numerous studies of acquired long QT syndrome have implicated blockade of the delayed rectifier current IKr, a small but influential cardiac K+ repolarizing current, as a primary factor in promoting pharmacologically-induced delayed repolarization in humans that enhances proarrhythmic risk (see, for example, De Bruin et al. (2005) and Roden (2008)). The human ether-a-go-go related gene (hERG) encodes the pore-forming alpha-subunit of the IKr channel in humans (Sanguinetti et al., 1995). Thus, reduction of IKr/hERG current is considered a surrogate marker of delayed cardiac repolarization, and functional assays measuring drug-induced block of hERG current expressed in heterologous systems are routinely used to characterize the potency of hERG blockade (Gintant et al., 2006, Hancox et al., 2008). Indeed, the value of the hERG assay is recognized by its key role in the integrated risk assessment within the ICH S7b Guidance Document for the Industry (S7b reference) as the primary in vitro assay to consider when evaluating delayed repolarization risk. Despite the generally accepted link between hERG block and delayed repolarization, the relationship between hERG block and clinical QTc prolongation is poorly understood, and reliance on the hERG assay likely results in unwarranted attrition of otherwise promising drug candidates.
In drug discovery, potency of hERG block is typically used to as an early screen for evolving preclinical drug candidates to avoid the risk of delayed cardiac repolarization. The IC50 value for hERG block is often compared to predicted or measured maximal plasma drug concentrations (Cmax) to define a predictive value (IC50/Cmax) referred to as the hERG safety margin. Earlier studies relating hERG safety margins with QTc prolongation and proarrhythmic risk relied on case reports of suspected adverse drug reactions and diverse sets of preclinical and clinical study conditions and impressions (Redfern et al., 2003, De Bruin et al., 2005). While highlighting the utility of the hERG assay and providing provisional safety margins, these studies did not provide a rigorous assessment of the performance of the hERG assay in predicting the extent of small delays in QTc prolongation. Such validation is recognized as an unmet need of safety pharmacology when assessing the risk of untoward clinical effects based on preclinical surrogate markers (Pugsley et al., 2008).
In clinical development, drug-induced delayed cardiac repolarization is rigorously evaluated in “thorough QT” (TQT) studies. A TQT study essentially represents a standardized clinical trial to assess the potential for a compound to affect small changes in ventricular repolarization (manifest as QTc prolongation). A positive control (typically moxifloxacin) is employed to demonstrate study sensitivity (see ICH E-14 document, also Darpo et al. (2006) and Morganroth (2007)). A mean increase in baseline-corrected, time-matched, placebo-controlled, QTc interval of 5 ms is recognized as one indication of concern.
The purpose of this retrospective study was to define the performance of the preclinical hERG current assay in assessing the potential risk of clinical QTc prolongation. This goal was accomplished by comparing TQT study results (prolongation ≥ 5 ms, a level of regulatory concern) and hERG safety margins (calculated based on IC50 values for hERG block and mean maximal drug concentrations (Cmax) measured during TQT studies) for 39 drugs. As drug binding to plasma proteins modulates drug distribution, total and free drug concentrations were both considered when defining safety margins. Assay performance was evaluated based on measures of sensitivity, specificity, receiver–operator characteristics, and likelihood ratios (for all approaches for evaluating the utility of diagnostic tests, see Sackett et al. (1991), Jaeschke et al. (1994), Altman and Bland (1994) and Deeks and Altman (2004)). An overview of TQT study results (and moxifloxacin responses from these studies) was also included as part of this review, along with a comparison of hERG block potency to product labeling. Results demonstrate that lower hERG safety margins (values less than 30 based on calculated free drug concentrations) provide moderate confidence in identifying risk of QTc prolongation, with an optimal decision threshold achieved with a safety margin ≤ 45. Likelihood ratios (providing another measure of test accuracy) demonstrate that QTc prolonging drugs are 7 times as likely to demonstrate hERG safety margin values below 30 compared to drugs that do not prolong QTc. Paradoxically, higher safety margins (greater than 1000) provide lesser confidence of reduced repolarization risk, an effect likely due to QTc prolongation arising from non hERG-dependent mechanisms and higher drug exposures. While lower safety margins are most useful in identifying the risk of delayed repolarization clinically, neither the potency of hERG block nor hERG safety margins alone are adequate to confidently avoid QTc prolongation. These results provide a strategy for supplementing hERG current assay results with additional preclinical assays early in drug discovery to avoid unwarranted attrition of novel compounds while mitigating delayed repolarization risk.
Section snippets
Methods and background
Information on 39 drugs was obtained from published reports describing hERG and TQT studies, product labels, FDA drug approval packages and EMA approval documents; all information was available in the public domain. No biologics were included in this study, as data regarding potency of hERG current block is often not obtained (as recently supported by a review by Vargas et al. (2008)). For inclusion in this analysis, the following information was required: a) from in vitro studies: IC50 values
Relation of hERG block potency to product labeling
Fig. 1 examines hERG block potency and its relation to product labeling regarding delayed cardiac repolarization. The distribution of IC50 values (based on a 5 μM bin size) for 29 drugs with definitive IC50 values (the standard dataset) is shown in Panel A. The majority of the drugs (72%) were characterized with IC50 values less than 50 μM. Panel B compares block potency for the 39 drug extended datasets with product labeling. Block potency spanned a wide concentration range (~7 log units).
Discussion
This retrospective study of 39 drugs represents the first rigorous quantitative evaluation of the relationship between hERG safety margins (calculated based on hERG block potency in relation to drug exposures during TQT studies) and QTc prolongation assessed during the same TQT studies. As expected, hERG potency alone is inadequate to define either product labeling for QT prolongation or the extent of QT prolongation in thorough QT studies. A general trend relating QTc prolongation with
Acknowledgments
Special thanks to Drs. C. Thomas Lin, Lei Shu and Damian Wandler for statistical discussions, Dr. Anton Safer for early comments, and Dr. Brian F. Hoffman for providing insights still relevant today.
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