Demonstration of Feasibility of In Vivo Gene Therapy for Gaucher Disease Using a Chemically Induced Mouse Model

Abstract

Progress towards developing gene therapy for Gaucher disease has been hindered by the lack of an animal model. Here we describe a mouse model of Gaucher disease which has a chemically induced deficiency of glucocerebrosidase and that accumulates elevated levels of glucosylceramide (GL-1) in the lysosomes of Kupffer cells. Administration of mannose-terminated glucocerebrosidase (Cerezyme) resulted in the reduction of GL-1 levels in the livers of these animals. Gene transduction of hepatocytes with a plasmid DNA vector encoding human glucocerebrosidase (pGZB-GC) generated high-level expression and secretion of the enzyme into systemic circulation with consequent normalization of Kupffer cell GL-1 levels. This suggested that the de novo synthesized and unmodified enzyme produced by hepatocyte transduction was also capable of being delivered to the cells that are primarily affected in Gaucher disease. Immunolocalization studies also revealed that preferential transduction and expression of human glucocerebrosidase in the Kupffer cells with subsequent reduction in the GL-1 levels could be attained with a low dose of a recombinant adenoviral vector encoding the human enzyme (Ad2/CMV-GC). This observation raises the possibility of gene therapy for Gaucher disease that involves directly transducing the affected histiocytes using recombinant adenoviral vectors. Together, these data demonstrate the potential for use of in vivo gene therapy vectors for treating Gaucher disease.

Expression and secretion of human glucocerebrosidase mediated by recombinant lentivirus vectors in vitro and in vivo: implications for gene therapy of Gaucher disease

“…It is important to determine whether unmodified enzyme also localized to the Kupffer cells. Marshall et al [15] showed that unmodified GC has the ability to localize to Kupffer cells and exhibits a longer half life than the modified enzyme. The unmodified GC expressed by gene transfer could be a useful therapeutic for Gaucher patients.…”

“…However, low transduction efficiency and poor long-term expression in these cells have hampered their clinical use [13,14]. Liver-targeted GC gene transfer using an adenovirus vector has demonstrated that enzyme secreted from transduced murine hepatocytes can reach tissues that are primarily affected [15]. This treatment, however, has immunological limitations.…”

Feasibility of gene therapy in Gaucher disease using an adeno-associated virus vector

“…Marshall et al showed that GC produced by adenovirus vector-mediated gene transfer has the ability to localize to Kupffer cells and exhibits a longer half-life than the modified enzyme (Marshall et al 2002). The GC expressed by rAAVmediated gene transfer could be a useful therapeutic approach for patients with Gaucher disease.…”

Dexamethasone-Mediated Up-Regulation of the Mannose Receptor Improves the Delivery of Recombinant Glucocerebrosidase to Gaucher Macrophages

“…After incubation with Cerezyme, the wells were washed twice with PBS containing 1 mg/ml yeast mannan and twice more with PBS and then lysed in 1 ml of KP buffer as for the macrophages. Lysates were assayed for glucocerebrosidase activity using the artificial fluorogenic substrate 4-methylumbelliferyl-␤-D-glucopyranoside as described previously (Marshall et al, 2002). Protein levels were quantified using the MicroBCA protein assay kit (Pierce Chemical, Rockford, IL).…”