Stem cell derived cardiomyocytes as models of pharmacology, physiology, and toxicology

Drug discovery and development requires preclinical models to eliminate flawed compounds from development pipelines. Unfortunately, current models have limitations, occasionally resulting in toxic or ineffective compounds progressing to the clinic at great cost and patient risk. Utilising stem cell technology, it is now possible to generate sophisticated models with human biology and physiological context, potentially overcoming the limitations of more established preclinical models. In this thesis I have investigated the use of embryonic stem cell derived cardiac cells for use as models in pharmacology, physiology and toxicology studies. Mouse embryonic stem cells were differentiated to generate cardiomyocytes in multicellular aggregates containing not only myocytes, but also pacemaker cells, fibroblasts and endothelium. These aggregates were used for pharmacology studies, where the signaling resulting from β-adrenoceptor and adenosine receptor stimulation was explored. Using the same differentiation method, in combination with a pan-cardiac reporter for cell enrichment, the function of individual cardiac cells was measured using calcium imaging. Following extensive method development, multiple phenotypes were identified in the enriched population based on spontaneous calcium oscillations. These distinct phenotypes were characterised based on calcium oscillation kinetics, pharmacology and immunocytochemistry. Using human stem cell derived cardiomyocyte aggregates I studied the effects of doxorubicin (a known cardio-toxin) and Trastuzumab, a humanised antibody with disputed cardio-toxicity. Following extensive method development, toxicity was observed for both doxorubicin and Trastuzumab. Furthermore, mechanistic studies implicate multiple cell types mediating Trastuzumab toxicity via a complicated signaling pathway. Based on my results, as models of pharmacology stem cell derived cardiomyocytes provide access to a physiologically heterogeneous model that may be useful for the screening of compounds for non-specific cardiac activity. Unfortunately, the complexity of multicellular aggregates limits their use in characterizing less established, or complicated receptor signaling pathways. Results from calcium imaging studies indicate that at a single cell level, there is considerable heterogeneity of stem cell derived cardiac cells. Focusing on cells with spontaneous calcium oscillations, presumably pacemaker cells, it may be possible to gain greater insight into the mechanisms required to maintain spontaneous cardiac activity, and identify drugs that disrupt it. The results of the Trastuzumab toxicity study provide evidence of a novel mechanism of Trastuzumab cardio-toxicity. More importantly, these results support the use of stem cell derived models for toxicology screening, particularly of humanised antibodies whose toxicity may be missed in classical models. The work presented in this thesis identified novel pacemaker phenotypes previously unreported, and a novel mechanism for Trastuzumab toxicity. Furthermore, this thesis highlights the strengths and weaknesses of stem cell derived models for use in pharmacology, physiology and toxicology assays.