Advances in homology directed genetic engineering of human pluripotent and adult stem cells

Abstract

The ability to introduce precise genomic modifications in human cells has profound implications for both basic and applied research in stem cells, ranging from identification of genes regulating stem cell self-renewal and multilineage differentiation to therapeutic gene correction and creation of in vitro models of human diseases. However, the overall efficiency of this process is challenged by several factors including inefficient gene delivery into stem cells and low rates of homology directed site-specific targeting. Recent studies report the development of novel techniques to improve gene targeting efficiencies in human stem cells; these methods include molecular engineering of viral vectors to efficiently deliver episomal genetic sequences that can participate in homology directed targeting, as well as the design of synthetic proteins that can introduce double-stranded breaks in DNA to initiate such recombination events. This review focuses on the potential of these new technologies to precisely alter the human stem cell genome and also highlights the possibilities offered by the combination of these complementary strategies.

Keywords: Human stem cells, Genetic engineering, Engineered viruses, Synthetic restriction endonucleases

Core tip: This manuscript focuses on the development of novel technologies to precisely alter the human stem cell genome and highlights their implications for both basic and applied stem cell research. Specifically, we discuss the development of two main technologies: molecular engineering of viral vectors and design of artificial endonucleases. We also discuss the merits of combining these complementary approaches and suggest other possible strategies that could be explored to further improve genetic engineering of human stem cells.