Therapeutic gene delivery with Sleeping Beauty transposon vectors: Assessment of preclinical efficacy and safety in a mouse model of Gaucher disease

• Applicants: Professor Dr. Zoltan Ivics; Professor Dr. Thomas Georg Ohm
• Subject Area: Human Genetics
• Term: from 2015 to 2020
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 283749119

Gaucher disease (GD) is the most common lysosomal storage disease caused by inherited mutations that result in a deficiency of the enzyme beta-glucorebrosidase (GCase). Clinically, three different subtypes can be distinguished: GD type I (with visceral symptoms), and II/III (with symptoms affecting the central nervous system). While enzyme replacement therapy offers at least some improvement of symptoms of GD type I, some patients are unresponsive and lack a suitable bone marrow donor. There are no treatment options available for the neuronopathic subforms of GD. A gene therapy approach potentially offers a unique possibility to provide life-long correction of disease by tackling their actual genetic cause. In this project, efficacy and biosafety of the non-viral Sleeping Beauty (SB) transposon system will be assessed for stable genetic modification of GCase-deficient hematopoietic stem cells (HSCs) in a non-neuronopathic GD mouse model (GD1 mice), which exhibits normal lifespan and relatively slow disease progression. We aim at improving our non-viral therapeutic gene delivery into HSCs by combining the SB transposon system with minicircle technology, which allows for a significant reduction in vector size by deleting bacterial backbone sequences present in conventional plasmid DNA vectors. This approach is expected not only to enhance the efficacy of genetic correction, but it also serves to further improve the biosafety of non-viral gene delivery in the context of a gene therapy application. In this proof-of-concept study, we will also address the feasibility of enzyme delivery into the central nervous system, which is a prerequisite for a successful application of the strategy for the severe neuronopathic forms of the disease. For that purpose, we are going to challenge our preclinical gene therapy approach by an advanced vector design, in which a secretion signal coupled to GCase is expected to enable its systemic distribution after being expressed from genetically corrected HSCs engrafted in the bone marrow. In addition, apolipoprotein B- and apolipoprotein E domains attached to the C-terminus of the enzyme are expected to promote its transfer through the blood-brain-barrier and hence its brain-specific delivery. GD1 mice transplanted with genetically corrected HSCs will be thoroughly characterized with respect to engraftment, gene marking and vector insertion site analysis, transgene expression and biodistribution and visceral organ phenotyping. This project is expected to significantly accelerate the development and clinical translation of safe preclinical protocols for GD gene therapy.

• DFG Programme: Research Grants