Understanding iPSC reprogramming using single cycle Measles vector

Abstract. In the last 10 years, the field of induced pluripotent stem cells (iPSCs) has come far, but the reprogramming process is still sub-inefficient; around 1-3% of the transduced cells reprogram, and the iPSCs are just starting to be translated into a clinical trial. Therefore, there is a need for an efficient reprogramming system that would allow the in-depth analysis of the different phases of the reprogramming process and the role of the innate immune response. We have developed a new vector system, based on a human Paramyxovirus, measles virus (MeV). From this virus, we have shown that the measles virus can be modified into a “one cycle” reprogramming vector expressing the four reprogramming factors (RFs, OCT4, KLF4, SOX2 and cMYC) in one single genome and successfully generate genomic modification-free iPSCs from human fibroblasts. We showed that the MeV vector is equivalent to or superior to the Sendai (SeV) vector and that the MeV-derived iPSC clones present all the characteristics of iPSC clones. The long-term goal is to understand the process of reprogramming by MeV vectors to produce better human iPSCs for the treatment of degenerative diseases. The central hypothesis is that because the MeV reprogramming vector expresses the four reprogramming factors in one single vector, it can be used as a tool to understand the underlying process of iPSC reprogramming. The objectives of this particular application are to (1) identify the difference in the state of the innate immune response activation during MeV vector reprogramming, (2) understand how the control of the innate immunity by MeV affects reprogramming efficiency, and (3) understand the role of exogenous RFs expression in the reprogramming process. The proposed work is innovative because it capitalizes on a new MeV vector expressing the four RFs for the reprogramming of somatic cells into iPSC, and our group developed this technology. Also, the RFs expression can be further modulated in an innovative way by either post-transcriptional regulation, using miRNA that are naturally up-regulated or down-regulated during the reprogramming process or by changing their position in the genome. The proposed work is significant because it will collectively validate MV vector as a platform to produce clinically relevant iPSCs, but also as a new tool to study the reprogramming process. This work will lead to the identification of pathways important in MeV reprogramming, but these pathways could be investigated in the context of the related SeV vector and other reprogramming technologies.