An induced pluripotent stem cell-mediated and integration-free factor VIII expression system.

Human artificial chromosome (HAC) has several advantages as a gene therapy vector, including stable episomal maintenance and the ability to carry large gene inserts. Induced pluripotent stem (iPS) cells also have a great potential for gene therapy, which can be generated from an individual's own tissues and contribute to any tissues when reintroduced. A Sendai virus (SeV) vector with reprogramming factors is a powerful tool for generating iPS cells because of the high infection efficiency without the risk of integration into host chromosomes. In this study, we developed an iPS cell-mediated and integration-free coagulation factor VIII (FVIII) expression system using non-integrating SeV- and HAC-vectors. Multiple human FVIII genes, which were under the control of the megakaryocyte-specific platelet factor-4 (PF4) promoter for development of a treatment for hemophilia A, were inserted into a HAC vector (PF4-FVIII-HAC). The PF4-FVIII-HAC was introduced into SeV vector-mediated iPS cells derived from a mouse model of hemophilia A. After in vitro differentiation of iPS cells with the PF4-FVIII-HAC into megakaryocytes/platelets, the PF4-FVIII-HAC resulted in expression of FVIII. This study has developed the iPS cell-mediated PF4-driven FVIII expression system using two non-integrating vectors; therefore, this system may be a promising tool for safer gene- and cell-therapy of hemophilia A.