Expression of human β–hexosaminidase α–subunit gene (the gene defect of Tay–Sachs disease) in mouse brains upon engraftment of transduced progenitor cells

Abstract

In humans, beta-hexosaminidase alpha-subunit deficiency prevents the formation of a functional beta-hexosaminidase A heterodimer resulting in the severe neurodegenerative disorder, Tay-Sachs disease. To explore the feasibility of using ex vivo gene transfer in this lysosomal storage disease, we produced ecotropic retroviruses encoding the human beta-hexosaminidase alpha-subunit cDNA and transduced multipotent neural cell lines. Transduced progenitors stably expressed and secreted high levels of biologically active beta-hexosaminidase A in vitro and cross-corrected the metabolic defect in a human Tay-Sachs fibroblasts cell line in vitro. These genetically engineered CNS progenitors were transplanted into the brains of both normal fetal and newborn mice. Engrafted brains, analyzed at various ages after transplant, produced substantial amounts of human beta-hexosaminidase alpha-subunit transcript and protein, which was enzymatically active throughout the brain at a level reported to be therapeutic in Tay-Sachs disease. These results have implications for treating neurologic diseases characterized by inherited single gene mutations.

Global gene and cell replacement strategies via stem cells

“…These gene products can be delivered to the host CNS in a direct, immediate, and stable manner. 5,8,40,42 While NSCs can migrate and integrate widely throughout the brain particularly well when implanted into germinal zones, allowing reconstitution of enzyme or cellular deficiencies in a global manner, 5,8,42 this extensive migratory ability is present even in the parenchyma of the diseased adult 5,40 and aged 43 brain. Despite their extensive plasticity, NSCs never give rise to cell types inappropriate to the brain (eg muscle, bone, teeth) or yield neoplasms.…”

“…Injecting the progenitors into the cerebral ventricles presumably allowed them to gain access to most of the subventricular germinal zone (SVZ), as well as to networks of cerebral vasculature, along the surface of which they would also migrate. This approach worked equally well in the fetus where donor NSCs gained access to the ventricular germinal zone (VZ), 42 migrating into the parenchyma within 24-48 h. This engraftment technique, exploiting many of the inherent properties of NSCs, permitted missing gene products to be delivered without disturbing other neurobiological processes and suggested a strategy for gene therapy of a class of neurogenetic diseases that, heretofore, had not been adequately treated ( Figure 1a). While MPS VII may be regarded as 'uncommon', the broad category of diseases that it models (neurogenetic conditions) afflicts as many as 1 in 1500 children and serves as a model for many adult neurodegenerative processes of genetic origin (AD, for example, could broadly fall into this category).…”

Syngeneic central nervous system transplantation of genetically transduced mature, adult astrocytes

“…5,6 However, the widespread use of fetal-derived tissues in human studies is problematic due to ethical considerations and legislative regulations 7,8 and relative concerns regarding allogeneic transplantation and widespread dissemination throughout the CNS. Other cell sources have included CNS progenitor cells [9][10][11][12] and immortalized derived cell lines. 13,14 The latter is limited by its proclivity toward unregulated cell growth and the former by difficulties in producing well-characterized cell populations and in isolating these cells from human donors.…”

Absence of Metabolic Cross-correction in Tay-Sachs Cells

“…Moreover, we are investigating whether the increase in ␤-subunit level is related to the ␣-subunit overexpression. Lacorazza et al (40) showed similar findings in C17 neuroblastoma clone transduced with a different retroviral vector expressing the ␣-subunit gene.…”