In our ongoing work within the DFG-funded priority program (SPP1230) we have identified important variables that regulate the transformation of hematopoietic cells by integrating gene vectors. We showed that the potential development of serious adverse events following insertional mutagenesis strongly depends upon the biological properties of the target cells and the configuration of the gene vector. Preliminary data also supported a role for extrinsic environmental factors, especially cell culture conditions used prior to transplantation of gene-modified cells, in the regulation of clonal competition in vivo. The present proposal continues this work in murine models of bone marrow transplantation and cell-based assays. We hypothesize that intrinsic cell features and/or engraftment conditions regulate the fate of hematopoietic cells following insertional mutagenesis, and will perform molecular and functional studies to address the underlying mechanisms. In the course of this work we will attempt to define the actual frequency of proto-oncogene upregulation in hematopoietic stem cells by insertional mutagenesis when using conventional and safety-improved vectors. This project opens a mechanistic understanding of cell fate in response to insertional mutagenesis, establishes platforms for the functional evaluation, and may provide a rationale for the improved design of clinical trials in gene therapy.

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