Aitchison, KL; (2015) Lentiviral vectors for gene therapy of Gaucher disease. Doctoral thesis , UCL (University College London).

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Abstract

Gaucher disease (GD), a recessive disorder characterised by hepatosplenomegaly, pancytopenia and skeletal complications, is caused by deficiency of the enzyme glucocerebrosidase (GC). GD leads to the accumulation of glucocerebrosides within macrophages, particularly in the liver and spleen. Current treatment is limited to enzyme replacement therapy (ERT) which is effective for most symptoms however skeletal problems are slow to respond. Treatment also has significant cost and impact on quality of life as infusions must be administered every two weeks. GD is a candidate for gene therapy as bone marrow transplantation has been shown to be curative which serves as a proof-of-concept that correction of haematopoietic stem cells (HSCs) can alleviate disease. This project produced lentiviral vectors carrying a range of constructs. GC was modified to contain a protein transduction domain (PTD) which could facilitate cross-correction of untreated cells in vivo. Recombinant vectors carrying PTD-GBA cDNA corrected the metabolic defect in patient-derived fibroblasts with levels of enzyme activity restored to within the healthy range. Transduced cells secreted active protein, uptake of which by untransduced cells was mediated by fusion of a PTD to the C- but not the N-terminus of the enzyme. The skeletal complications of GD are likely to be caused by enzyme deficiency in the osteoclast, a cell of haematopoietic origin. Therefore it is possible that by transducing HSCs we will be able to alleviate skeletal symptoms. To this end it is shown that modification of HSCs does not affect their ability to generate osteoclasts. It is also demonstrated that osteoclasts derived in vitro from the neuronopathic GD mouse model have increased activity and this could be a useful model for osteoclast correction when treating GD. In conclusion, this project generated lentiviral vectors for use in treating Gaucher disease. Further work should include correction of the osteoclast phenotype and further investigation of the potential for cross-correction in vivo.

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