Bioengineering human embryonic stem cell-derived ventricular-like cardiomyocytes for preclinical translation, cardiomyopathy modelling and drug screening

Abstract

Human cardiac disease modelling and therapeutic discovery are facing challenges in translating biomedical research from preclinical to clinical stages. Despite the valuable insight of the complex human heart physiology and pathophysiology gained from animal models, extrapolations of these findings are limited by the species-related differences between humans and other animals. However, an authenticated and robust human cardiac model system remains elusive. Advances in stem cell biology have opened the possibilities of generating an unlimited supply of human cardiomyocytes (CMs) from human pluripotent stem cells (hPSC). Noting that hPSC-derived CMs (hPSC-CMs) recapitulates some of the cellular phenotypes of typical native human heart cells, the utilisation of these cells in bench research and preclinical studies has therefore been pursued. In this dissertation, human ventricular-like CMs (vCM) were produced from human embryonic stem cells (hESCs) through directed cardiac differentiation. Three potential applications of these hESC-vCMs were critically evaluated: 1) The bioengineering of hESC-vCMs into nodal-like CMs as a preclinical trial for the creation of bioartificial pacemaker. 2) The modelling of doxorubicin-induced cardiomyopathy in hESC-vCMs for the identification of pathological pathways and novel cardioprotective compounds. 3) The fabrication and verification of human ventricular cardiac tissue strips (hvCTS) for drug screening.

Firstly, hESC-vCMs were functionally reprogrammed into pacemaker-like CMs by overexpressing engineered hyperpolarisation-activated cyclic-nucleotide-gated channel 1 (HCN1) via the recombinant adeno-associated virus. The resulting HCN1-transformed cells were functional and exhibited automaticity as well as electrophysiological parameters that resembled those of native nodal CMs, which sustained for over one month. This work demonstrated the preclinical translation capacity of using hESC-vCMs as a tool for creating bioartificial pacemaker.

Secondary, to examine the capacity of hESC-vCMs in modelling cardiac diseases, hESC-vCMs were treated with doxorubicin, a chemotherapeutic agent that have been associated with acute and chronic cardiomyopathy in cancer patients. Doxorubicin significantly reduced the cell viability in a dose-dependent manner, and caused remarkable morphological changes and sarcomeric damages. The secretion of cardiac injury marker growth/differentiation factor 15 was also induced. In addition, when the hESC-vCMs were pre-treated with several compounds derived from traditional Chinese medicine, a paradoxically more intense cytotoxic effect was observed, further suggesting the potential use of hESC-vCMs for identifying novel cardioactive compounds.

Lastly, hESC-vCMs were used as building blocks for fabricating 3-dimensional hvCTS which allowed a direct measurement of contractility. hvCTS exhibited biologically relevant response to an array of cardioactive compounds of different classes, and demonstrated excellent sensitivity and accuracy in identifying negative inotropes in a blinded assay. As such, hvCTS could serve as an effective drug screening platform for preliminary safety and efficacy assessment in the drug development pipeline.

In summary, the present study comprehensively evaluated the capacity and limits of hESC-vCMs in preclinical translation, disease modelling and drug screening. A better understanding on the applications of hESC-vCMs shall enable a more efficient drug and therapy development process, and provides a better model for future cardiac research.