EXPRESSION OF THE HUMAN GLUCOCEREBROSIDASE AND ARYLSULFATASE-A GENES IN MURINE AND PATIENT PRIMARY FIBROBLASTS TRANSDUCED BY AN ADENOASSOCIATED VIRUS VECTOR

We have constructed two recombinant adeno-associated virus (AAV) vecto rs (pJJ-3GC and pJJ-3ASA) which contained either the human glucocerebr osidase (GC) or arylsulfatase A (ASA) cDNA under the control of an SV4 0 promoter. These plasmids were co-transfected to 293 cells with a hel per plasmid containing trans-acting AAV genes required for packaging t he vectors. The two recombinant vectors successfully infected murine a nd patient fibroblasts. The human glucocerebrosidase and arylsulfatase A genes were expressed at high levels in the cells as determined by W estern blot analysis, enzyme assay and immunochemical staining. GC enz yme activity in Gaucher patient fibroblasts (GM-0877) infected by AAV- GC was 15-fold higher than in non-infected cells. ASA enzyme activity in MLD 577 g cells infected by AAV-ASA was up to 500-fold higher than in the metachromatic leukodystrophy (MLD) control cells. Southern blot ting results showed that the vector integrated 1-2 copies of pJJ-3GC a nd ASA in the targeted cell genome. These two vectors will be useful i n studying AAV-mediated transfer of the GC and ASA genes in cells and animals.