Human engineered heart tissue as a model system for drug testing☆
Graphical abstract
Introduction
Drug development is a long and costly process with high attrition rates. With costs rising exponentially particularly in late stages of development involving clinical studies, the failure of drugs in late development or after approval is a worst-case scenario. Many pharmaceutical companies therefore follow a “fail early, fail cheap” approach, bearing the risk of wasting potentially valuable drugs. Cardiac side effects such as arrhythmias are the most common reason for the withdrawal of drugs [1], [2]. An estimated ~ 45% of all withdrawals and ~ 30% of restrictions to drug application are due to unwanted cardiovascular effects [2]. Estimates suggest that 70% of the toxicity seen in clinical trials could have been detected in the preclinical phase [3] and that a 10% improvement of predictive value of preclinical tests could save averaged development costs of 100 million dollars per drug [4]. This situation has prompted regulatory agencies in the US and the EU (Federal Drug and Food Administration, FDA, and European Medicines Agency, EMA) a decade ago to establish guidance for industry, e.g., for the preclinical evaluation of proarrhythmic drug effects [5]. Though the currently approved test systems increased the sensitivity for the detection of certain side effects, they still have major shortcomings, including the lack of consideration of species differences and complex interactions between more than one ion channel [6], [7]. A reliable human-based cost-effective preclinical screening tool may circumvent some of them.
The ideal test-bed for the prediction of cardiac side effects would be the human heart, but the limited availability of these preparations (e.g., atrial appendages as a by-product of cardiac surgery or ventricular muscle strips from explanted failing human hearts in the course of heart transplantation) precludes this possibility. The prospects of hiPSC-derived cardiomyocytes (hiPSC-CM) readily available (e.g., from commercial sources such as Cellular Dynamics International (CDI), Axiogenesis, Pluriomics) and devoid of the ethical concerns related to embryonic stem cells, have raised hopes in the field. HiPSC-CM express most, if not all, structural and regulatory elements present in a human CM [8], [9], [10], [11], display a cross-striated ultrastructure, contract spontaneously and show basic functional properties of the human heart, including typical responses to drugs (Table 1). This unambiguously establishes hiPSC-CM as bona fide CM. Yet, most parameters differ quantitatively between hiPSC-CM and adult CM. HiPSC-CM are smaller, beat spontaneously (which is a clear sign of immaturity), show a lower degree of ultrastructural organization (which is almost crystalline in adult CM), no t-tubules, less negative diastolic membrane potential, slower action potential upstroke velocity and lower contractile force, and smaller responses to β-adrenergic stimulation (Table 2) or the Ca2 +-channel agonist Bay K 8644 [12]. Their maturation state has been estimated to resemble 16-week old human fetal CM [13], raising the question whether testing in hiPSC-CM has indeed a higher validity than classical animal experiments. Thus, much effort is currently directed towards means to maturate CM and improve the functional readout. This article reviews the potential of hydrogel-based engineered heart tissue (EHT) for this purpose.
Section snippets
Established systems (FDA and EMA guidelines)
Cardiac side effects such as arrhythmias are common and potentially life threatening [2]. Many drugs causing arrhythmias interact with the human ether-á-go-go (hERG) related potassium channel [14], [15]. The hERG-dependent current IKr plays a major role in human cardiac repolarization and its inhibition is a common cause of severe ventricular arrhythmias, such as “Torsades-des-Pointes” (TdP) arrhythmias [16]. Thus, screening for hERG-interaction is obligatory in the preclinical phase of drug
General aspects of EHTs
Engineered heart tissues (EHTs) are three-dimensional, hydrogel-based muscle constructs that can be generated from isolated heart cells of chicken, rat, mouse, hESC and hiPSC [41], [42], [43], [44]. The method for the generation of EHTs was introduced in 1997 [41] and has not been principally changed since then. It requires (i) heart cells, (ii) a liquid hydrogel that solidifies and promotes tissue formation, (iii) a casting mold that determines the 3D shape of the developing tissue and (iv) a
Disease modeling
Besides drug screening, EHTs may prove valuable as a robust, simple and high content assay for hiPSC-mediated disease modeling with patient-specific hiPSC-CM (review in [81]). The principal idea to detect consequences of individual mutations in patient-derived hiPSC-CM has been validated in several studies for cardiac diseases such as long QT syndrome type 1 (LQT1, [82]), long QT syndrome type 2 (LQT2, [83]), Timothy syndrome [84], catecholaminergic polymorphic ventricular tachycardia (CPVT,
Conclusion & outlook
Preclinical drug testing with EHTs offers a number of advantages compared to standard 2D culture formats, including a more physiological 3D muscle environment, longitudinal alignment, increased maturity, directed contractions and simple access to measurements of force, the most important parameter of heart function. Though drug testing has already been formulated as the major goal 20 years ago [41], the EHT technology just now starts to enter wider practice. The two major reasons are the design
Funding
This work is supported by grants from the European Union (Biodesign, FP7-NMP-2010-LARGE-4 #262948), the European Research Council (ERC-AG IndivuHeart erca.c(2014)1813543), the German Research Foundation (DFG Es 88/12-1, DFG Ha 3/1), the British National Centre for the Replacement Refinement & Reduction of Animals in Research (NC3Rs, CRACK IT challenge), the German Centre for Cardiovascular Research (DZHK) and the German Ministry of Research (BMBF), the German Heart Foundation and the Freie and
Conflict of interest
The authors have founded a company (EHT Technologies GmbH) based on a patent TE is holding for the method (patent number: PCT/EP0100856).
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This review is part of the Advanced Drug Delivery Reviews theme issue on “Tissue engineering of the heart: from in vitro models to regenerative solutions.”
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