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Background: GM1-gangliosidosis is a glycosphingolipid (GSL) lysosomal storage disease caused by a genetic deficiency of acid β-galactosidase (βgal), which results in the accumulation of GM1-ganglioside and its asialo-form (GA1) primarily in the CNS. Age of onset ranges from infancy to adulthood, and excessive ganglioside accumulation produces progressive neurodegeneration and psychomotor retardation in humans. Currently, there are no effective therapies for the treatment of GM1-gangliosidosis., Methodology/principal Findings: In this study we examined the effect of thalamic infusion of AAV2/1-βgal vector in adult GM1 mice on enzyme distribution, activity, and GSL content in the CNS, motor behavior, and survival. Six to eight week-old GM1 mice received bilateral injections of AAV vector in the thalamus, or thalamus and deep cerebellar nuclei (DCN) with pre-determined endpoints at 1 and 4 months post-injection, and the humane endpoint, or 52 weeks of age. Enzyme activity was elevated throughout the CNS of AAV-treated GM1 mice and GSL storage nearly normalized in most structures analyzed, except in the spinal cord which showed ∼50% reduction compared to age-matched untreated GM1 mice spinal cord. Survival was significantly longer in AAV-treated GM1 mice (52 wks) than in untreated mice. However the motor performance of AAV-treated GM1 mice declined over time at a rate similar to that observed in untreated GM1 mice., Conclusions/significance: Our studies show that the AAV-modified thalamus can be used as a 'built-in' central node network for widespread distribution of lysosomal enzymes in the mouse cerebrum. In addition, this study indicates that thalamic delivery of AAV vectors should be combined with additional targets to supply the cerebellum and spinal cord with therapeutic levels of enzyme necessary to achieve complete correction of the neurological phenotype in GM1 mice.

Glioblastoma multiforme (GBM) is a devastating form of brain cancer for which there is no effective treatment. Here, we report a novel approach to brain tumor therapy through genetic modification of normal brain cells to block tumor growth and effect tumor regression. Previous studies have focused on the use of vector-based gene therapy for GBM by direct intratumoral injection with expression of therapeutic proteins by tumor cells themselves. However, as antitumor proteins are generally lethal to tumor cells, the therapeutic reservoir is rapidly depleted, allowing escape of residual tumor cells. Moreover, it has been difficult to achieve consistent transduction of these highly heterogeneous tumors. In our studies, we found that transduction of normal cells in the brain with an adeno-associated virus (AAV) vector encoding interferon-beta (IFN-beta) was sufficient to completely prevent tumor growth in orthotopic xenograft models of GBM, even in the contralateral hemisphere. In addition, complete eradication of established tumors was achieved through expression of IFN-beta by neurons using a neuronal-restricted promoter. To our knowledge this is the first direct demonstration of the efficacy of targeting gene delivery exclusively to normal brain cells for brain tumor therapy.

GM1-gangliosidosis is a glycosphingolipid (GSL) lysosomal storage disease caused by autosomal recessive deficiency of lysosomal acid beta-galactosidase (betagal), and characterized by accumulation of GM1-ganglioside and GA1 in the brain. Here we examined the effect of neonatal intracerebroventricular (i.c.v.) injection of an adeno-associated virus (AAV) vector encoding mouse betagal on enzyme activity and brain GSL content in GM1-gangliosidosis (betagal(-/-)) mice. Histological analysis of betagal distribution in 3-month-old AAV-treated betagal(-/-) mice showed that enzyme was present at high levels throughout the brain. Biochemical quantification showed that betagal activity in AAV-treated brains was 7- to 65-fold higher than in wild-type controls and that brain GSL levels were normalized. Cerebrosides and sulfatides, which were reduced in untreated betagal(-/-) mice, were restored to normal levels by AAV treatment. In untreated betagal(-/-) brains, cholesterol was present at normal levels but showed abnormal cellular distribution consistent with endosomal/lysosomal localization. This feature was also corrected in AAV-treated mice. The biochemical and histological parameters analyzed in this study showed that normal brain neurochemistry was achieved in AAV-treated betagal(-/-) mice. Therefore we show for the first time that neonatal AAV-mediated gene delivery of lysosomal betagal to the brain may be an effective approach for treatment of GM1-gangliosidosis.

Adeno-associated virus (AAV) vectors have gained a preeminent position in the field of gene delivery to the normal brain through their ability to achieve extensive transduction of neurons and to mediate long-term gene expression with no apparent toxicity. In adult animals direct infusion of AAV vectors into the brain parenchyma results in highly efficient transduction of target structures. However AAV-mediated global delivery to the adult brain has been an elusive goal. In contrast, widespread global gene delivery has been obtained by i.c.v. injection of AAV1 or AAV2 in neonates. Among the novel AAV serotypes cloned and engineered for production of recombinant vectors, AAV8 has shown a tremendous potential for in vivo gene delivery with nearly complete transduction of many tissues in rodents after intravascular infusion. Here we compare the efficiency of an AAV8 serotyped vector with that of AAV1 and AAV2 serotyped vectors for the extent of gene delivery to the brain after neonatal injection into the lateral ventricles. The vectors all encoded green fluorescent protein (GFP) under control of a hybrid CMV enhancer/chicken beta-actin promoter with AAV2 inverted terminal repeats, but differed from each other with respect to the capsid type. A total of 6.8 x 10(10) genome copies were injected into the lateral ventricles of postnatal day 0 mice. Mice were killed at postnatal day 30 and brains analyzed for distribution of GFP-positive cells. AAV8 proved to be more efficient than AAV1 or AAV2 vectors for gene delivery to all of the structures analyzed, including the cerebral cortex, hippocampus, olfactory bulb, and cerebellum. Moreover the intensity of gene expression, assessed using a microarray reader, was considerably higher for AAV8 in all structures analyzed. In conclusion, the enhanced transduction achieved by AAV8 compared with AAV1 and AAV2 indicates that AAV8 is the superior serotype for gene delivery to the CNS.