Distribution of a Lysosomal Enzyme in the Adult Brain by Axonal Transport and by Cells of the Rostral Migratory Stream

Abstract

A portion of the lysosomal enzymes produced by cells is secreted, diffuses through extracellular spaces, and can be taken up by distal cells via mannose-6-phosphate receptor-mediated endocytosis. This provides the basis for treating lysosomal storage diseases, many of which affect the CNS. Normal enzyme secreted from a cluster of genetically corrected cells has been shown to reverse storage lesions in a zone of surrounding brain tissue in mouse disease models. However, low levels of enzyme activity and reduction of storage lesions also have been observed at sites in the brain that may not be explained by a contiguous gradient of secreted enzyme diffusing away from the genetically corrected cells. No direct evidence for alternative mechanisms of enzyme transport has been shown, and little is understood about the intracellular movement of lysosomal enzymes in neurons. We investigated whether axonal transport could occur, by expressing an eukaryotic lysosomal enzyme that can be visualized in tissue sections (beta-glucuronidase) in brain structures that have defined axonal connections to other structures. This resulted in the transfer of enzyme to, and a reversal of storage lesions in, neurons that project to the gene expression site, but not in nearby structures that would have been corrected if the effect had been mediated by diffusion. In addition, transduction of cells in the subventricular zone resulted in the uptake of beta-glucuronidase by cells entering the rostral migratory stream. Gene transfer to specific neuronal circuits or cells in migratory pathways may facilitate delivery to the global brain lesions found in these disorders.

AAV2-mediated CLN2 gene transfer to rodent and non-human primate brain results in long-term TPP-I expression compatible with therapy for LINCL

“…As has been observed with AAV2 vectors expressing other transgenes, 28,29 we observed a high level of TPP-I expression in cell populations not directly at the site of injection, but whose axonal projections were located in the site of the injection. As widespread delivery of the gene product throughout the brain is a goal of the current study, we examined other brain regions that project to the striatum to determine if expression of TPP-I could be detected in projection neuronal populations following single intrastriatal administration of AAV2 CU hCLN2.…”

Adeno-associated virus vector-mediated transduction in the cat brain

“…We have also observed this in mice where it has been shown that episomal forms of vector DNA persist in brain tissue, therefore, small amounts of sense riboprobe may have hybridized to the nontranscribed strand of the episomal DNA. 3,42 No evidence of hybridization was seen in regions of uninjected brain using either the sense or antisense riboprobe.…”

“…21 No evidence of retro-or anterograde transport in the cat brain was found with any of the three serotypes used. Although axonal transport of a lysosomal enzyme has been reported in the mouse, 42 there are several possible explanations for this apparent difference. First, the regions of the brain in which transport was examined in the mouse, the intrahippocampal and septohippocampal pathways, were not tested in the cat so a direct comparison is not possible.…”

“…In the mouse brain, AAV2 transduces primarily neurons. 3,8,17,42,43 In the cat brain, the fact that AAV2/2 transduces only the gray matter, in addition to the neuronal-like shape of the transduced cells (Figure 2), suggests that neuronal transduction is also occurring. In the cat, AAV2/1 transduced both gray and white matter in the cat brain, suggesting that AAV2/1 is capable of also 17,35 In the cat experiments, the expression cassette was flanked by AAV2 ITRs and a eucaryotic promoter was used.…”

Adeno-associated virus type 4 (AAV4) targets ependyma and astrocytes in the subventricular zone and RMS

“…While prior work with AAV type 2 (AAV2) vectors showed poor NPC transduction following direct SVZ injection, AAV2 did successfully transduce human NPCs in vitro. 9,10 We previously showed that AAVs possessing different capsid structures have altered tropism when introduced into the mammalian brain. 11 For example, AAV4 transduced ependyma with high efficiency when injected into the striata or lateral ventricles of adult mice.…”