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ILAR Journal V36(3/4) 1994 [FORMERLY ILAR NEWS]
Advances in Gene Therapy

Most inherited neurological disorders are not amenable to existing treatments, however they are potential candidates for gene therapy, an approach that combines recent developments in molecular diagnosis with novel gene transfer techniques. Genes may be transferred directly to neural tissue using viral vectors or by physical methods, or indirectly by injection of cell vehicles that express therapeutic genes. The application of gene therapy to the treatment of neurological disease is at an early stage, and inherent problems, such as accessing sites of injury or degeneration and stimulating neuronal regeneration, have yet to be resolved. Promising gene transfer techniques have many potential clinical applications for brain diseases, so it is imperative that they be validated prior to use in humans. For this purpose, there are many suitable animal models of neurological disease that can be used to provide a scientific rationale for safe and effective treatment. This review focuses on inherited neurological disorders and gene therapy approaches to their treatment.

CHALLENGES TO THE TREATMENT OF NEUROLOGICAL DISEASE

Compared to other organs, the brain is particularly vulnerable to degenerative disease, and it is a difficult organ to treat. Fragile neural tissue has a limited capacity for repair, and its sophisticated functions depend on the maintenance of its structural integrity. Neurons do not usually divide in adult life, except in a few locations such as the olfactory bulb, dentate gyrus (Gould and McEwen, 1993) and subventricular zone (Lois and Alvarez-Buylla, 1993), although neurotrophic factors can stimulate replication in vitro (Reynolds and Weiss, 1992; Richards et al., 1992). Instead the brain is protected from injury by the bony skull and cerebrospinal fluid cushion, and from injurious molecules in the circulation by the tight junctions of the cerebrovascular endothelium, which form the blood-brain barrier. Some of the unique obstacles to treatment of diseases of the brain are listed in Table 1. Diagnosis, treatment, and cure of many disorders is hampered by our imperfect understanding of vital aspects of neural function, including memory, learning, and consciousness.

RATIONALE FOR GENE THERAPY OF NEUROLOGICAL DISEASE

There are many potential applications for therapies that promote neural regeneration and repair and, considering the limited success of conventional therapy, the incentives to vigorously pursue new approaches to treatment are compelling. The major degenerative disorders of the human central nervous system (CNS) such as Parkinson's disease and Alzheimer's disease, are currently incurable. They cause prolonged suffering with progressive disability and often consume considerable resources in supportive care. Brain disorders with a multifactorial etiology as well as those disorders caused by a single gene defect, such as Huntington's disease and amyotrophic lateral sclerosis (MoreIl, 1993; Rosen et al., 1993), are potential candidates for genetic intervention.

Neurotrophic factors, which promote survival and repair of neural cells in vivo, have considerable therapeutic potential for many neurodegenerative diseases (Friden et al., 1993) if they can be delivered for prolonged periods (Kawaja et al., 1992). For example, an animal model of Parkinson's disease induced by unilateral nigrostriatal injury responded to treatment with neurotransmitter precursors or neurotrophic factors (Gage et al., 1990a, b).

The neurological lysosomal storage diseases are caused by single gene defects and are promising candidates for gene therapy. In these diseases cells are deficient in lysosomal enzymes and are able to endocytose enzymes from the extracellular fluid. Replacement enzyme can be supplied by direct injection or from enzyme-producing cells, such as host cells corrected with a retroviral vector (Moullier et al., 1993). There is convincing evidence from animal studies that neurological storage disease can respond to enzyme replacement. For example, mice with mucopolysaccharidosis (MPS) type VII had reduced CNS storage lesions following weekly enzyme infusions beginning at birth (before the blood-brain barrier is complete) (Sands et al., in press), dogs with fucosidosis had high levels of CNS enzyme after marrow transplantation (Taylor et al., 1992), and amannosidosis cats had enzyme in neurons and reduced storage lesions after bone marrow transplantation (Walkley et al., 1994). Lysosomal enzymes do not cross the adult blood-brain barrier, but following bone marrow engraftment, they are delivered to the brain by donor monocytes that cross the blood-brain barrier and differentiate into microglia (Unger et al., 1993). These studies demonstrate that concentrations of gene products, which are within the range that can be provided by current gene therapy technology, can have potent effects when delivered to the site of neurological damage. Moreover, once the current difficulties with longevity of gene expression are overcome, gene therapy could provide a permanent supply of therapeutic proteins, thereby simplifying treatment.

GENE DELIVERY TO NEURAL TISSUE

The structure of the brain is complex and the diseases that affect it are diverse, so a multiplicity of approaches to treatment may be required. Two major approaches have emerged: direct treatment of cells in situ, or indirect treatment, by injection of cells that deliver the gene product (Table 2). Both approaches use viral vectors or physical methods of DNA transfer to provide the therapeutic product.

Direct (in situ) Treatment of the Brain

Direct delivery of genes to neurons in situ has many advantages. The need for surgical removal and in vitro manipulation of host tissue is eliminated, and it permits localized treatment of affected areas of the brain.

Viral Vectors

The major advantage of viral vectors is their gene transfer efficiency, which exceeds the efficiency of direct DNA transfer by several orders of magnitude. Three viruses have been used as vectors: herpes, adeno-, and retroviruses.

Herpes Virus. Herpes virus vectors have been developed to directly transfer genes to neural cells in situ (Breakefield and DeLuca, 1991; Kennedy and Steiner, 1993). Herpes simplex virus (HSV) type I has natural tropism for cells of the nervous system. HSV infects neurons and other nonreplicating cells both in situ and in culture. The virus can be injected directly into the brain, or administered to the peripheral nerve terminals for retrograde transport to central neurons. Acute infection may cause cytotoxicity, but is followed by enduring latency, and during this time latency associated transcript genes (LAT) are expressed. Two types of vectors, plasmid ampiicons and replication defective viruses, have been developed to exploit the unique neurotropism of HSV. The utility of these vectors in vivo has been hampered by toxicity in some models of neurological disease and difficulties in achieving an adequate level of gene expression in more than a few cells. Nevertheless, stable, long-term gene expression was seen in the CNS of mice with MPS type VII, which were infected with an HSV vector containing the b-glucuronidase gene driven by the LAT promoter (Wolfe et al., 1992). Specific, low-level gene expression in neurons was achieved with the neuron-specific enolase promoter (Andersen et al., 1993). The virus genome of 152kb can carry a large insert of up to 30kb DNA, but also contains many genes with poorly defined functions. HSV I vectors have not been tipproved for clinical use, but have potential for treatment of neuroanatomically localized disorders.

Adenovirus. An adenovirus vector (Ad type 5) has emerged as the vector of choice for diffuse, transient gene delivery in many tissues (Kozarsky and Wilson, 1993). It has a wide host range and is not associated with tumor formation. The 30kb vector has deletions in Ela, Elb, and E3 regions of the viral genome and can take up to a 7kb insert. The vector is produced by homologous recombination and is packaged by 293 cells, which provide missing El functions (viral transcription for replication) in trans.The vector efficiently infects a wide range of nonreplicating cells, including mature neurons, glia, and ependymal cells following direct injection into the brain, yielding high level reporter gene expression (Davidson et al., 1993; Salle et al., 1993). It has low pathogenicity when injected at low titer (3 x 10⁵) in the CNS, but causes toxicity and gliosis at higher concentrations (>10⁷) (Akli et al., 1993). Ependymal cells infected with an adenovirus vector secreted a-l-anti-trypsin into the cerebrospinal fluid for one week, an approach that may be useful for widespread delivery of gene products to the brain parenchyma (Bajocchi et al., 1993). Clinical trials using adenovirus vectors for cystic fibrosis have been hampered by rapid loss of gene expression and by immune responses to the vector. Immune responses are more attenuated in the brain, due to its protected status, and gene expression persisted for at least 2 months in the .CNS (Salle et al., 1993) compared to loss within a few weeks in other tissues. New generation adenovirus vectors with modifications in E3 and E4 regions are being developed to reduce immunogenicity and increase longevity of expression.

Retrovirus. Retroviral vectors permanently deliver genes to cells and their progeny, but require mitosis for integration of a proviral DNA copy of the viral RNA into the host cell genome. This makes these vectors unsuitable for gene delivery to terminally differentiated neurons in the adult brain. Crippled retroviral vectors are created by deletions in the viral gag, pol, and env genes. These functions are supplied in transby packaging cells, providing space in the vector for an insert up to 8-10 kb (McLachlin et al., 1990). Additional mutations have been created in the vectors and packaging cells to limit the possibility of the generation of wild type virus (Miller and Rosman, 1989).

Retroviral vectors are useful as cell lineage markers in the developing nervous system because they are replication defective, so the integrated proviral DNA and reporter gene expression provide a permanent marker of cell lineage (Walsh and Cepko, 1993). Direct injection of retroviral vectors is currently not useful for therapy because the virions are very fragile and label few cells. Efficiency of infection with high titer (e.g. 10⁶) retroviral vectors is high in rapidly dividing cells in vitro, but can be improved. For example, if the vector is packaged in vesicular stomatitis virus envelope the virions can be concentrated to very high titer (10⁹) without loss of activity (Burns et al., 1993); alternatively, the inclusion of replication-defective adenovirus can increase the effective titer (Wang et al., 1994).

Provirus integration in the cell's genome requires cell division, and this restriction has been used to facilitate treatment of brain tumors. Retroviral packaging cells are transplanted into the tumor and release virions that contain a "suicide'' gene, herpes simplex thymidine kinase (tk). This vector infects rapidly dividing tumor cells which become sensitive to the antiviral drug, gancilovir. Infected tumor cells and bystander tumor cells are selectively killed without damage to the surrounding brain tissue (Culver et al., 1992). This approach has been approved for clinical use.

Other Potential Vectors. Other viruses are being developed as vectors, but have not reached an equivalent degree of safety and efficacy. Adeno-associated virus infects nondividing cells and permanently integrates at a specific chromosomal site in some cells. This very simple vector can carry a gene of 4.5kb and requires adenovirus helper functions for replication (Flotte et al., 1993). Its use in neural cells has not yet been reported. Viruses with some potential as vectors include vaccinia, papilloma virus, and neurotropic human JC virus (Tomatore et al., 1994) and these are in the early stages of investigation. Pseudorabies is a neurotropic herpes virus and, like HSV 1, is transported from peripheral injection sites transneuronally to the CNS (Card et al., 1990; Levine et al., 1994). Modified virus encoding [3-galactosidase (the LacZ reporter gene) has been used as a neurotracer, but it causes progressive neurotoxicity, so it is currently unsuitable as a therapeutic vector.

Direct Delivery of Genes

The main advantages of direct gene delivery are: (1) its simplicity (only plasmid constructs are required), (2) the absence of constraints on the size or number of genes transferred, and (3) the option to use only mammalian genes and regulatory elements. Down regulation of vector-encoded mammalian genes is a common finding in gene therapy studies (Palmer et al., 1991; Scharfmann et al., 1991) and occurs in the brain (Schinstine and Gage, 1993). In vivo loss of gene expression is associated with methylation of viral sequences (Challita and Kohn, 1994) but is not well understood.

DNA/RNA Injection. DNA directly injected into muscle is taken up, remains in a stable extra-chromosomal location, and is expressed in vivo (Wolff et al., 1990). The process is very inefficient and has not been reported in neural tissue. Direct DNA delivery by other physical methods such as shock wave, gene gun, and particle bombardment (Cheng et al., 1993; Yang, 1992) is also very inefficient. RNA is surprisingly stable in the brain, and when vasopressin mRNA was injected into hypothalamic axons of Brattleboro (di) rats, a model for diabetes insipidus, vasopressin was produced and corrected the diabetes for 5 days (Jirikowski et al., 1992).

DNA-Liposome. Brain cells, both neuronal and glial, have been transfected with liposome-DNA mixtures in culture. Cells take up and retain DNA episomally for extended periods, and, following transplantation, the transferred gene is expressed for up to 2 months (Jiao et al., 1992). However, liposome-mediated DNA transfer is inefficient, and in situ treatment of neurons has not been reported. Fusogenic liposomes created with inactivated Sendai virus fuse non-specifically with cell membranes at neutral pH (Kaneda et al., 1993), increasing the target cell range and efficiency of liposome delivery.

DNA-receptor Complexes. Specific binding of a ligand to its receptor can improve the delivery of DNA (and other products) into target cells (Ferkol et al., 1994). This approach can increase transport of molecules across the blood-brain barrier. For example, an antibody to the transferrin receptor was linked to nerve growth factor and facilitated its transport into the brain (Friden et al., 1993). Unfortunately much of the internalized DNA is degraded in cell endosomes. The inclusion of replication-incompetent adenovirus causes endosomolysis and enables DNA to bypass this degradation path, thereby increasing delivery to the nucleus (Cristiano et al., 1993).

Indirect (ex vivo) Cell Treatment and Injection

Cells may be injected into the brain to perform various functions, including: (I) supply a gene product of therapeutic value (such as enzyme or neurotrophic factor), (2) directly replace lost neurons, or (3) provide glial cell functions (such as remyelination)

Fibroblasts. Fibroblasts transduced with retroviral vectors were the first genetically engineered cells used to deliver neurotransmitters and growth factors to the brain (Rosenberg et al., 1988). The aim of this approach was to supply the product long-term, without local damage in the area of treatment. Immortalized fibroblasts secrete proviral gene products in vivo, but may be tumorigenic. Primary autologous fibroblasts are an attractive alternative cell vehicle for their availability, histocompatibility to the host, ease of culture, rapid expansion in vitro, and quiescence after transplantation (Gage et al., 1991; Suhr and Gage, 1993). Unfortunately, the stability of these cells after injection is accompanied by loss of expression of proviral sequences after a few weeks in vivo (down regulation) (Schinstine and Gage, 1993). In experimental animals primary fibroblasts formed stable grafts that survived for long periods without inducing neurological signs (Kawaja et al., 1991, 1992). Fibroblasts transfected by a retroviral vector containing the nerve growth factor gene promoted neuron survival and axonal regrowth in rats with experimental lesions (Kawaja et al., 1992). In a spontaneous murine model of lysosomal storage disease, MPS type VII, engineered fibroblasts injected into the brain secreted the enzyme b-glucuronidase and reduced the storage lesions nearby, but the proviral gene expression fell to a low level after 3 weeks, even though the transplanted cells survived for at least 11 months (Taylor and Wolfe, 1994). Cells were lost from intracerebral fibroblast grafts in rats in long-term quantitative studies (Kawaja et al., 1991) compounding the problem of loss of gene expression.

Myoblasts. Muscle cells have considerable potential for gene delivery and secretion of gene products as a treatment for non-myogenic disease. Muscle can be readily collected by biopsy, expanded, transfected by physical means, or retrovirally infected. Following gene transfer, myoblasts and differentiated myotubes secrete gene products in vitro (Smith et al., 1990) and in vivo (Dai et al., 1992). Plasmid-transfected primary myoblasts and myotubes survived long-term in the rat brain after stereotaxic injection. They expressed the transfected tyrosine hydroxylase (th) gene and secreted dopamine, which reduced the rotational behavior in this experimental model of Parkinson's disease (Jiao et al., 1993).

Neural Cells. Cells of neural origin have the dual capacity to supply a genetically engineered protein and participate directly in cellular repair. Transplants of normal fetal neural tissue have met with some success in Parkinson's disease patients, but have raised ethical controversy (Fisher and Gage, 1993). There are two major difficulties with primary neural cells as vehicles for gene transfer: (1) lack of a suitable supply of nonessential neural cells in the patient for removal and ex vivo manipulation, and (2) growth, expansion, and survival of cells in vitro is poor and deteriorates with donor age, so they are difficult to infect with retroviral vectors. Recently, it has been shown that neurons from adult CNS tissue can be induced to limited proliferation in vitro with epidermal growth factor (Reynolds and Weiss, 1992), but currently they cannot be produced in sufficient numbers from a biopsy for gene therapy. Mixed glial cell cultures divide and are efficiently infected with retroviral vectors (unpublished observations), but Tsuda et al. (1990), reported the chloramphenicol acetyltransferase (cat) gene was only expressed for 3 weeks in glial cells after injection.

O2A Cells. Transplants of glial ceils, particularly bi-potential O2A glial progenitors, may be useful in treating acquired demyelinating disease like multiple sclerosis, and inherited disorders with oligodendrocyte dysfunction, for example metachromatic leukodystrophy and Krabbe's disease. O2A progenitors are primary cells expanded from neural tissue with platelet-derived growth factor and basic fibroblastic growth factor. They proliferate in vitro and can be labelled with retroviral vectors. Following injection they differentiate into oligodendrocytes (Espinosa et al., 1993) and remyelinate experimental spinal cord lesions (Groves et al., 1993). Labelled oligodendrocytes (from transgenic mice), engrafted in the CNS of Twitcher (twi) mice after injection and increased the life span of these animals (Huppes et al., 1992).

Schwann cells. Schwann cells also remyelinate CNS lesions and secrete growth factors that prevent neuronal atrophy (Pizzorusso et al., 1994). They have potential as a gene transfer vehicle in treating demyelinating disease, especially because they are more easily collected than oligodendrocytes, can be expanded in vitro, and can be infected with retroviral vectors (Langford and Owens, 1990).

Neural Progenitor. The ultimate goal of treatment in many neurological disorders is to replace lost and damaged neurons. Research into neuropoiesis and developmental plasticity in the brain has identified multipotent neuroglial progenitor cells that have remarkable plasticity (Marvin and McKay, 1992; Snyder, 1992). C17-2 progenitor cells were derived from mouse cerebellum by infection with retroviral vectors encoding the oncogene v-myc and the LacZ reporter gene (Ryder et al., 1989). These progenitors respond to environmental cues in vitro and in vivo and differentiate into astrocytes, oligodendrocytes, and neurons accordingly (Snyder et al., 1992). These cells are nontumorigenic, appear to be well tolerated by histoincompatible strains of mice and can be transfected in vitro by physical or viral methods. C 17-2 cells express the transferred LacZ gene long-term in vivo and were used to deliver endogenous b-glucuronidase in mice with MPS VII by neonatal intraventricular injection. The neuroglial progenitor cells engrafted, provided a permanent supply of enzyme, and reduced the lysosomal storage lesions (Snyder et al., in press).

Gene Delivery to the CNS by Hematopoietic Cells. Transplantation of normal hematopoietic cells has been used as a treatment of lysosomal disorders with beneficial effects on brain lesions in some disorders (Taylor et al., 1992; Walkley et al., 1994) but not others (Haskins et al., 1991). If donor origin monocytes (from a bone marrow, fetal liver cell, or cord blood transplant) migrate to the brain and differentiate into microglial cells as has been suggested, then genetically modified hematopoietic cells might provide a long-term source of gene products in the brain, particularly if treatment occurs early in life, during the period of ameboid microglial migration in the brain.

CONCLUSIONS

Gene therapy holds great promise, even for a tissue as complex and difficult to treat as the brain. Many approaches are being developed to overcome the unique barriers to treating the brain, which will increase our understanding of neural regeneration and repair and may improve the treatment of disease. The wider problems facing gene therapy, particularly stability and level of expression, are the subject of intense investigation, and may be circumvented by the development of more efficient methods of receptor-mediated gene transfer or by replacement of neurons and glial cells in adult life with multipotent neuropoietic progenitor cells.

ACKNOWLEDGEMENTS

I am grateful to John Wolfe, in whose laboratory I performed the MPS VII mouse studies and to our collaborator in these studies, Evan Snyder. I thank Peter Williamson and Graeme Stewart for their helpful discussions on the manuscript. I was supported by a Kleberg Fellowship and a Sandoz Fellowship from the Transplantation Society of Australia and New Zealand.

REFERENCES

Akli, S.. C. Calllaud, E. Vigne, L. D. Stratford-Perricaudet, L. Poenaru, M. Perricaudet. A. Kahn, and M. R. Peschanski. 1993. Transfer of a foreign gene into the brain using adenovirus vectors. Nature Cmnet. 3:224-228.

Andersen, J. K., D. M. Frim, and X. O. Breakefield. 1993. Herpesvirus-mediated gene delivery into the rat brain: Specificity and efficiency of the neuron-specific enolase promoter. Cell. Mol. Neurobiol. 13:503-515.

Bajocchi, G., S. H. Feldman, R. G. Crystal, and A. Mastrangeli. 1993. Direct in vivo gene transfer to ependymal cells in the central nervous system using recombinant adenovirus vectors. Nature Genet. 3:229-234.

Breakefield, X. O., and N. A. DeLuca. 1991. Herpes simplex virus as a vector for neurons. Pp. 287-319 in Treatment of Genetic Diseases, R. J. Desnick, eds. New York: Churchill Livingstone.

Bums, J. C., T. Friedmann, W. Driever, M. Burrascano, and J. K. Yee. 1993. Vesicular stomatitis virus G glycoprotein in pseudotyped retroviral vectors: Concentration to very high titer and efficient gene transfer into mammalian and nonmammalian cells. Proc. Natl. Acad. Sci. USA. 90:8033-8037.

Card. J. P., L. Rinaman, J. S. Schwaber, R. R. Miselas, M. E. Whealy, A. K. Robbins. and L. W. Enquist. 1990. Neurotropic properties of pseudorabies virus: Uptake and transneuronal passage in the rat central nervous system. J. Neurosci. 10:1974-1994.

Challita. P.-M., and D. B. Kohn. 1994. Lack of expression from a retroviral vector after transduction of murine hematopoietic stem cells is associated with methylation in vivo. Proc. Natl. Acad. Sci. USA. 91:2567-2571.

Cheng, L., P. R. Ziegelhoffer, and N. S. Yang. 1993. In vivo promoter activity and transgene expression in mammalian somatic tissues evaluated by using particle bombardment. Proc. Natl. Acad. Sci. USA. 90:4455-4459.

Cristiano, R. J., L. C. Smith, M. A. Kay, B. R. Brinkley, and S. L. Woo. 1993. Hepatic gene therapy: Efficient gene delivery and expression in primary hepatocytes utilizing a conjugated adenovirus-DNA complex. Proc. Natl. Acad. Sci. USA. 90:11548-11552.

Culver, K. W., Z. Ram, S. Wallbridge, H. Ishii, E. H. Oldfield, and R. M. B laese. 1992. In vivo gene transfer with retroviral vector-producer cells for treatment of experimental brain tumors. Science 256:1550-1552.

Dai, Y.. M. Roman, R. K. Naviaux, and I. M. Verma. 1992. Gene therapy via primary myoblasts: Long-term expression of factor IX protein following transplantation in vivo. Proc. Natl. Acad. Sci. USA. 89:10892-10895.

Davidson, B. L., E. D. Allen, K. F. Kozarsky, J. M. Wilson, and B. J. Roessler. 1993. A model system for in vivo gene transfer into the central nervous system using an adenoviral vector. Nature Genet. 3:219-223.

Dobson, A.T., F. Sedarati, G. Devi-Rao, W.M. Flanagan, M.J. Fartell, J.G> Stevens, E. K. Wagner, and L.T. Feldman. 1989. Identification of the latency-associated transcript promoter by expression of rabbit betaglobin mRNA in mouse sensory nerve ganglia latently infected with a recombinant herpes simplex virus. J. Virol. 63:3844-3851.

During, M. J., A. Freese, A. Y. Deutch, P. G. Kibat, B. A. Sabel, R. Langet, and R. H. Roth. 1992. Chemical and behavioural recovery in a rodent model of Parkinson's disease following stereotactic implantation of dopamine-containing liposomes. Exp. Neurol. 115:193-199.

Espinosa, A., D. L. Momeros, M. Zhang, and J. de Veilis. 1993. O2A progenitor cells transplanted into the neonatal rat brain develop into oligodendrocytes but not astrocytes. Proc. Natl. Acad. Sci. USA. 90:50-54.

Federoff, H.J.. M.D. Geschwind, A.L. Geller, and J.A. Kessler. 1992. Expression of nerve growth factor in vivo from a defective herpes simplex virus 1 vector presents effects of axotomy on sympathetic ganglia. Proc. Natl. Acad. Sci. USA 89:1636-1640.

Ferkol, T., C. S. Kaetzel, and P. B. Davis. 1994. Gene transfer into respiratory epithelial cells by targeting the polymeric immunoglobulin receptor. J. Clin. Invest. 92:2394-2400.

Fisher, L. J., and F. H. Gage. 1993. Grafting in the mammalian central nervous system. Physiol. Rev.73:583-616.

Fisher, L. J., M. Schinstine, P. Salvaterra, A. J. Dekker, L. Thai, and F. H Gage. 1993. In vivo production and release of acetylcholine from primary fibroblasts genetically modified to express choline acetyltransferase. J. Neurochem. 61:1323-1332.

Flotte, T. R., S. A. Aflone, C. Conrad, S. A. McGrath, R. Solow, H. Oka, P.L. Zeitlin, W. B. Guggino, and B. J. Carter. 1993. Stable in vivo expression of the cystic fibrosis transmembrane conductance regulator with an adeno-associated virus vector. Proc. Natl. Acad. Sci. USA. 90:10613-10617.

Friden, P. M., L. R. Walus, P. Watson, S. R. Doctrow, J. W. Kozarich, C. Backman, H. Bergman, B. Hoffer, F. Bloom, and A. C. Granholm. 1993. Blood-brain barrier penetration and in vivo activity of an NGF conjugate. Science 259:373-377.

Gage, F. H., L. J. Fisher, H. A. Jinnah, M. B. Rosenberg, M. H. Tuszynski and T. Friedmann. 1990a. Grafting genetically modified cells to the brain: Conceptual and technical issues. Prog. Brain Res. 82:1-10.

Gage, F. H.. M. B. Rosenberg, M. H. Tuszynski, K. Yoshida, D. M. Arrestton. R. C. Hayes, and T. Friedmann. 1990b. Gene therapy in the CNS: Intracerebral grafting of genetically modified cells. Prog. Brain Res. 86:205-217.

Gage, F. H., M. D. Kawaja, and L. J. Fisher. 1991. Genetically modified cells: Applications for intracerebral grafting. Trends Neurosci. 14:328-333

Gould, E., and B. S. McEwen. 1993. Neuronal birth and death. Cure. Opin. Neurobiol. 3:676-682

Groves, A. K., S. C. Barnett, R. J. M. Franklin, A. J. Crang, M. Mayer, W. F. Blakemore, and M. Noble. 1993. Repair ofdemyelinated lesions by transplantation of purified O-2A progenitor cells. Nature 362:453-455.

Haskins, M. E., H. J. Raker, E. Birkenmeier. P. M. Hoogerbrugge, B. J. H. M. Poorthuis, T. Sakiyama, R. M. Shull, R. M. Taylor, M. A. Thrall, and S. U. Walkley. 1991. Transplantation in animal model systems. Pp. 183-201 in Treatment of Genetic Diseases, R. J. Desnick, ed. New York: Churchill Livingstone.

Huppes, W., C. J. A. d. Groot, R. H. Ostendorf, J. G. J. Bauman, J. A. Gossen, V. Smit, J. Vijg, and C. D. Dijkstra. 1992. Detection of migrated allogeneic oligodendrocytes throughout the central nervous system of the galactocerebrosidase-deficient twitcher mouse. J. Neurocyt. 21:129-136.

Jiao, S., G. Acsadi, A. Jani, P. L. Feigner, and J. A. Wolff. 1992. Persistence of plasmid DNA and expression in rat brain cells in vivo. Exp. Neurol. 115:400-413.

Jiao, S., V. Gurevich, and J. Wolff. 1993. Long-term correction of rat model of Parkinson's disease by gene therapy. Nature 362:450-453.

Jirikowski, G. F., P. P. Sanna, D. Maciejewski-Lenoir, and F. E. Bloom. 1992. Reversal of diabetes insipidus in Brattleboro rats: Intrahypothalamic injection of vasopressin mRNA. Science 255:996-998.

Kaneda, Y., K. Kato, M. Nakanishi, and T. Uchida. 1993. Introduction of plasmid DNA and nuclear protein into cells using erythrocyte ghosts, liposomes, and Sendal virus. Meth. Enzymol. 221:317-327.

Kawaja, M. D., A. M. Fagan, B. L. Firestein, and F. H. Gage. 1991. Intracerebral grafting of cultured autologous skin fibroblasts into the rat striaturn: An assessment of graft size and ultrastructure. J. Comp. Neurol. 307:695-706.

Kawaja. M. D., M. B. Rosenberg, K. Yoshida, and F. H. Gage. 1992. Somatic gene transfer of nerve growth factor promotes the survival of axotomized septal neurons and the regeneration of their axons in adult rats. J. Neurosci. 12:2849-2864.

Kennedy. P. G. E., and 1. Steiner. 1993. The use of herpes simplex virus vectors for gene therapy in neurological diseases. Quart. J. Med. 86:697-702.

Kozarsky. K. F., and J. M. Wilson. 1993. Gene therapy: Adenovirus vectors. Curt. Opin. Genet. Devel. 3:499-503.

Langford, L. A., and G. C. Owens. 1990. Resolution of the pathway taken by implanted Schwann cells to a spinal cord lesion by prior infection with a retrovirus encoding betagalactosidase. Acta Neuropath. 80:514-520.

Levine, J. D., X. S. Xhao, and R. R. Miselis. 1994. Direct and indirect retinohypothalamic projections to the supraoptic nucleus in the female albino rat. J. Comp. Neurol. 341:214-224.

Lois, C., and A. Alvarez-Buylla. 1993. Proliferating subventricular zone cells in the adult mammalian forebrain can differentiate into neurons and glia. Proc. Natl. Acad. Sci. USA. 90:2074-2077.

Marvin. M., and R. McKay. 1992. Mulipotential stem cells in the vertebrate CNS. Semin. Cell Biol. 3:401-411

McLachlin, J. R., K. Cometm, M. A. Eglitis, and W. F. Anderson. 1990. Retroviral-mediated gene transfer. Prog. Nucleic Acid Res. 38:91-135.

Miller, A. D., and G. J. Rosman. 1989. Improved retroviral vectors for gene transfer and expression. BioTechniques 7:980-989.

Morell, V. 1993. Huntington's gene finally found. Science 260:28-30.

Moullier, P., D. Bohl, J.-M. Heard, and O. Danos. 1993. Correction of lysosomal storage in the liver and spleen of MPS VII mice by implantation of genetically-modified skin fibroblasts. Nature Genet. 4:154-159.

Palmer, T. D., G. J. Rosman, W. R. A. Osborne, and A. D. Miller. 1991. Genetically modified skin fibroblasts persist long after transplantation but gradually inactivate introduced genes. Proc. Natl. Acad. Sci. USA. 88:1330-1334.

Pizzorusso, T., M. Fagiolini, M. Fabtis, G. Ferrari, and L. Maffei. 1994. Schwann cells transplanted in the lateral ventricles prevent the functional and anatomical effects of mononuclear deprivation in the rat. Proc. Natl. Acad. Sci. USA. 91:2572-2576.

Reynolds, B. A., and S. Weiss. 1992. Generation of neurons and astrocytes from isolated cells of the adult mammalian central nervous system. Science 255:1707-1710.

Richards, L. J., T. J. Kilpatrick, and P. F. Bartlett. 1992. De novo generation of neuronal cells from the adult mouse brain. Proc. Natl. Acad. Sci. USA. 89:8591-8595.

Rosen, D. R., T. Siddique, D. Patterson, D. A. Figlewicz. P. Sapp, A. Hentati, D. Donaldson, J. Goto, J. P. O'Regan, H. X. Deng, et al.. 1993. Mutations in Cu/Zn superoxide dismutase gene are associated with familial amyotrophic lateral sclerosis. Nature 362:59-62.

Rosenberg, M. B., T. Friedmann, R. C. Robertson, M. Tuszynski, J. A. Wolff, X. O. Breakefield, and F. H. Gage. 1988. Grafting genetically modified cells to the damaged brain: Restorative effects of NGF expression. Science 242:1575-1577.

Ryder, E. F., E. Y. Snyder, and C. L. Cepko. 1989. Establishment and characterization of multipotent neural cell lines using retrovirus vector-mediated oncogene transfer. J. Neurobiol. 21:356-375.

Salle, G. L., J. J. Robert, S. Berrard, V. Ridoux, L. D. Stratford-Perricaudet, M. Perricaudet, and J. Mallet. 1993. An adenovirus vector for gene transfer into neurons and gila in the brain. Science 259:988-993.

Sands, M. A., C. Vogler, J. W. Kyle, J. H. Grubb, B. Levy. N. Galvin. W. S. Sly. and E. Birkenmeier. In press. Enzyme replacement therapy for murine mucopolysaccharidosis type VII. J. Clin. Invest.

Scharfmann, R., J. H. Axelrod, and I. M. Verma. 1991. Long-term in vivo expression of retrovirus-mediated gene transfer in mouse fibroblast implants. Proc. Natl. Acad. Sci. USA. 88:4626-4630.

Schinstine, M., and F. H. Gage. 1993. Factors affecting proviral expression in primary cells grafted into the CNS. Pp. 311-323 in Molecular and Cellular Approaches to the Treatment of Neurological Disease, S. G. Waxman, ed. New York: Raven Press. Ltd.

Smith, B. F., R. K. Hoffman, U. Giger, and J. H. Wolfe. 1990. Genes transferred by retroviral vectors into normal and mutant myoblasts in primary cultures are expressed in myotubes. Mol. Cell. Biol. 10:3268-3271.

Snyder, E.Y. 1992. Neural transplantation: An approach to cellular plasticity in the developing central nervous system. Sem. Perinat. 16:106-121.

Snyder, E. Y., D. L. Deitcher, C. Walsh, S. Amold-Aidea, E. A. Hartwieg, and C. L. Cepko. 1992. Multipotent neural cell lines can engraft and participate in development of mouse cerebellum. Cell 68:33-51.

Snyder, E. Y., R. M. Taylor, and J. H. Wolfe. In press. Neural progenitor cell engraftment corrects lysosomal storage throughout the MPS VII mouse brain. Nature.

Suhr, S. T., and F. H. Gage. 1993. Gene therapy for neurologic disease. Arch. Neurol. 50:1252-1268.

Taylor, R. M., and J. H. Wolfe. 1994. Enzyme replacement in MPS VII brain following transplantation of genetically modified fibroblasts. J. Cell. Biochem. 18A Supp:247.

Taylor, R. M., B. R. H. Farrow, and G. J. Stewart. 1992. Bone marrow transplantation in fucosidase deficient dogs. Am. J. Med. Genet. 42:628-632.

Tomatore, C., K. Meyers, and G. Major. 1994. Can JC viruses be used as a vector for gene delivery to oligodendrocytes and astrocytes of the CNS white matter? J. Cell. Biochem. 18A supp:247.

Tsuda, M., S. Yuasa, Y. Fujino, M. Sekikawa, K. Ono, T. Tsuchiya, and K. Kawamura. 1990. Retrovirus-mediated gene transfer into mouse cerebellar primary culture and its application to the neural transplantation. Brain Res. Bull. 24:787-792.

Unger, E.R., J. H. Sung, J. C. Manivel, M. L. Chenggis, B. R. Blazar, and W. Krivit. 1993. Male donor-derived cells in the brains of female sex-mismatched bone marrow transplant recipients: A Y-chromosome specific in situ hybridization study. J. Neuropath. Exp. Neurol. 53:460-470.

Walkley, S. U., M. A. Thrall, K. Dobrenis. M. Huang, P. A. March, D. A. Siegel, and S. Wurzelmann. 1994. Bone marrow transplantation corrects the enzyme defect in neurons of the central nervous system in a lysosomal storage disease. Proc. Natl. Acad. Sci. USA. 91:2970-2974.

Walsh, C., and C. L. Cepko. 1993. Clonal dispersion in proliferative layers in developing cerebral cortex. Nature 362:632-635.

Wang, M., D. Steffen, and F. D. Ledley. 1994. Replication defective adenovirus enables transduction by retroviral vectors of cells outside their host range. J. Cell. Biochem. 18A Supp:222.

Wolfe, J. H., S. L. Deshmane, and N. W. Fraser. 1992. Herpesvirus vector gene transfer and expression of betaglucuronidase in the central nervous system of MPS VII mice. Nature Genet. 1:379-384.

Wolff, J. A., R. W. Malone, P. Williams, W. Chond, G. Acsadi, A. Jani, and P.L. Feiger. 1990. Direct gene transfer into mouse muscle in vivo. Science 247:1465-1468.

Yang, N. S. 1992. Gene transfer into mammalian somatic cells in vivo. Crit. Rev. Biotech. 12:335-356.

TABLE 1 Overcoming obstacles in treatment of neurological disease

TABLE 2 Animal Studies of CNS Gene Therapy

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