Development of a CENH3-based and non-viral vector system

New vectors for successful transgene delivery in patients are more than needed. Current gene therapeutic vectors are mostly based on virus genomes. A main reason is the very efficient administration of the vector into the target cells. The viral background of these systems also has severe side effects like e.g. immunological reactions of the patient or the activation of oncogenes due to insertional mutagenesis, which can result in cancer development. A possibility to overcome these problems is the establishment of extrachromosomally maintained, autonomously replicating and non-viral vectors. The goal of this work was to establish such a novel system with the help of the CENH3 protein. The centromere-specific histone 3 variant CENH3 (H.sapiens: CENH3CENP-A, D.melanogaster: CENH3CID) is sufficient for centromere formation. It confers a stable and epigenetically heritable mark at the centromeric region and does this also on plasmids. With the help of CENH3 I changed the passive piggyback segregation mechanism of Epstein-Barr virus-derived vectors into an active segregation mechanism for plasmid DNA (pDNA) vectors. I demonstrated that the new and active pDNA vector segregation mechanism prolongs pDNA vector retention in cells. The information gained from basic research might have great impact in the field of gene-therapeutic research, as this mechanism might be a helpful tool in future gene therapeutic vectors.