The induced pluripotent stem cells (iPSCs) technology enables derivation of patient-specific pluripotent stem cells from adult somatic cells. We have demonstrated reproducible derivation of genomic modification-free iPSCs from patients using Sendai viral vectors. However, the Sendai vector system, which is based on a murine Paramyxovirus has never been approved by the FDA for the use in human clinical trial. Here, we propose to develop a new vector system, based on a human Paramyxovirus, measles virus (MV). We have developed a reverse genetic system allowing the production of recombinant virus equivalent to the Moraten vaccine strain, which is currently used for children vaccination in the US. Additionally, clinical grade genetically modified MV is being produced to treat cancer patients and is currently being evaluated in human clinical trials.
In aim 1, we will test the hypothesis that measles virus can be modified into "one cycle" reprogramming vectors expressing the four reprogramming genes (OCT4, KFL4, SOX2 and cMYC). We will produce four "one cycle" MV vectors expressing one reprogramming factor (RF) and test their ability to reprogram adult somatic cells in iPSCs.
In aim 2, we will test the hypothesis that we can improve the efficacy and safety of the "one-cycle" MV vectors to reprogram adult somatic cells in iPSCs. We will increase efficacy by producing a single MV vector expressing the four RFs from a single genome. We will take advantage of the measles gradient of transcription, to optimize the expression levels of the four factors for the highest reprogramming efficiency. In order to increase the safety of the vector, we will arm the vector with a suicide mechanism, which will allow extinction of the RF expression as well as the elimination of the residual genome and viral protein expressions as soon as differentiation of the iPSCs starts. We will insert the target sequence of a miRNA specifically expressed in the definitive endoderm. This will increase the efficiency of differentiation and safety of our vectors. The result of these studes will lead to a new vector system allowing the production of genomic modification-free iPSCs, enabling rapid iPSCs translation into the clinic.
The proposed research will advance the field of the stem cells research, as it will develop a new safe way of producing pluripotent stem cells that can be differentiated into various types of cells. This research has relevance to public health and will have an important impact in the development of new regenerative medicine approaches for diabetes, cardiac, respiratory and other diseases.