Generation of virus-free induced pluripotent stem cells using fusogenic lipid vesicles to deliver reprogramming proteins to adult neural stem cells

• Applicant: Dr. Juliane Nguyen
• Subject Area: Pharmacy
• Term: from 2010 to 2013
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 188950407

The objective of this research is to use liposomes composed of novel fusogenic lipids to increase the low reprogramming efficiency of induced pluripotent stem cells (iPSCs) by delivery of protein transcription factors (reprogramming proteins) to adult neural stem cells (NSCs). iPSCs are adult cells which are reprogrammed to undifferentiated embryonic stem cell-like states. They eliminate the need for human embryos, and potentially provide a safe cell source for applications in transplantation medicine. This research is enabled by a novel, efficient and simple liposome production method I devised that results in high levels of protein encapsulation into small diameter liposomes. It also relies upon novel lipids developed within our liposome group. Delivery of reprogramming proteins, instead of viral or plasmid DNA encoding the protein sequence, will avoid potential insertional mutagenesis and reduce the risk of adverse phenotypic changes in the iPSCs. I will first optimize the protein delivery system using a transcription factor-induced luciferase reporter gene. After optimization, the recombinant pluripotency-associated protein Oct3/4 will be delivered to reprogram NSCs. To validate reprogramming of NSCs, I will examine their gene expression profile, DNA methylation status, and ability to differentiate into three germ layers after forming embryoid bodies in suspension culture. Finally, the ability of these iPSCs for germline competence after chimera formation will be investigated. Success in this research will provide a better understanding of the pharmacology of reprogramming factors delivered as proteins and might open the way for the use of a patient¿s own iPSCs in regenerative medicine.

• DFG Programme: Research Fellowships
• International Connection: USA
• Host: Professor Dr. Francis Szoka