AAV is a promising candidate for gene therapy since it provides prolonged transgene expression without the risk of insertional mutagenesis. This can be used in the context of bone regeneration by packing genes into the AAV vector that promote the formation of new bone. In order to improve gene expression, enhancers which are compatible with in vivo and potential clinical use were developed and tested in vitro.

Different newly developed proprietary poloxamers were combined with AAV vectors and were tested for reporter gene expression of cell lines and primary cells in vitro. The mixes included thermo-responsive hydrogels. The polymer and serotype with the best results were selected for subsequent experiments using ovine bone marrow-derived mesenchymal progenitor cells. AAVs either coded for reporter gene eGFP or growth factor genes BMP-2 or VEGF. Transduction efficiency was quantitated by fluorescence measurements (eGFP) or ELISA (growth factors). Cell viability was determined by XTT assay.

The experiments revealed a cationic poloxamer, which not only increased transduction efficiency in a dose dependent manner, but also promoted cell survival. Enhancement was seen with AAV2, AAV6 and AAV8 capsids, albeit at different rates. Cationic poloxamer additionally protected AAV from drying, potentially allowing for lyophylization. Thus, a cationic poloxamer with potential for in vivo application and clinical use was identified, which enhanced AAV-mediated gene transfer in general and in cells relevant to bone regeneration.

 

SBR has received funding from the European Union’s Horizon 2020 research and innovation program under grant agreement No. 874896.