Project description
A novel way to improve gene delivery
Gene therapy and gene editing necessitate the delivery of nucleic acids into cells. However, there are various cellular bottlenecks that hamper the effectiveness of delivery. These include endosomal sequestration, poor nuclear translocation and, most importantly, retention of introduced nucleic acids in membrane cages following recognition by the barrier-to-autointegration factor (BAF) protein. Scientists of the EU-funded IBAF project will address this issue and increase the cytosolic availability and mobility of transfected DNA through the transient suppression of the BAF. The project's findings will advance our understanding of this unexplored DNA retention mechanism, and provide transfection enhancers that can boost already-existing gene delivery platforms.
Objective
Safe and efficient delivery of nucleic acids to tissues and cells is a shared challenge in the clinical translation of gene therapy and gene editing. At the intracellular level, DNA delivery is hindered by endo/lysosomal sequestration, inefficient transport into the nucleus and a retention mechanism mediated by the barrier-to-autointegration factor (BAF) protein that detects, clusters and locks away intruding double-stranded DNA in membrane cages. The first two of these intracellular obstacles are well-known bottlenecks and are being addressed by many laboratories. However, preliminary in vitro data suggest the detrimental impact of BAF’s mechanism on transgene expression is underestimated. We hypothesize that transiently suppressing BAF or one of its regulating factors will increase the cytosolic availability and mobility of transfected DNA, facilitating its transport to the nucleus and ultimately boosting transfection efficiency. We will tackle this rather uninvestigated mechanism through two parallel strategies: i) identifying novel small-molecule inhibitors of BAF or its regulators via high-throughput screening of chemical libraries, and ii) producing a recombinant kinase that phosphorylates BAF in situ and thus supresses its DNA-clustering function. The BAF suppressors will be co-delivered to cells with rationally-designed nucleoprotein nanoparticles consisting of a reporter plasmid and a dual-function fusion protein to facilitate endocytosis and endosomal escape. The combined transfection-enhancing effect of the BAF suppressors and the nanoparticles will be extensively characterized in vitro, and in vivo proof-of-concept will be obtained in mice. Besides expanding our fundamental knowledge on this unexplored DNA retention mechanism, the project will provide powerful transfection enhancers that can boost already-existing gene delivery platforms (viral and non-viral), so that these can reach their full potential for gene therapy and gene-editing applications.
Fields of science
- natural sciencesbiological sciencesbiochemistrybiomoleculesnucleic acids
- medical and health sciencesmedical biotechnologygenetic engineeringgene therapy
- natural sciencesbiological sciencesgeneticsDNA
- natural sciencesbiological sciencesbiochemistrybiomoleculesproteins
- engineering and technologynanotechnologynano-materials
Programme(s)
Topic(s)
Funding Scheme
ERC-ADG - Advanced GrantHost institution
8092 Zuerich
Switzerland