Generation of FOXO3 engineered human stem cells with enhanced efficacy and safety

FOXO3 is an evolutionally conserved transcription factor that has been linked to longevity. Here we asked whether human stem cells could be functionally enhanced by engineering them to express an activated form of FOXO3. This was accomplished via HDAdV-mediated gene editing of human embryonic stem cells. These cells were then differentiated into a range of vascular cell types, as FOXO3 has been shown to play a protective role in the cardiovascular system. FOXO3-enhanced vascular cells exhibited delayed senescence and increased resistance to oxidative injuries, compared with wild type cells. When tested in a therapeutic context, FOXO3-enhanced human mesenchymal stem cells promoted vasculature regeneration in a mouse model of ischemic injury, and were resistant to tumorigenic transformation, both in vitro and in vivo. Mechanistically, constitutively active FOXO3 conferred cytoprotection by transcriptionally downregulating CSRP1. Taken together, our findings provide mechanistic insights into FOXO3-mediated vascular protection, and indicate that FOXO3 activation may provide a means for generating more effective and safe biomaterials for cell replacement therapies. In this study, we investigated the functions of FOXO3-enhanced human embryonic stem cells (hESCs), vascular endothelial cells (hVECs), vascular smooth muscle cell (hVSMCs), mesenchymal stem cells (hMSCs) and transformed mesenchymal stem cells (transMSCs) through multi-omics analysis.