Parkinson's Disease: A Neurodegenerative Disease Particularly Amenable to Gene Therapy
Authors: Martha C. Bohn
Journal: Molecular Therapy (2000)

Most neurodegenerative diseases, such as Alzheimer's disease, amyotrophic lateral sclerosis (ALS), lipid storage diseases, and leukodystrophies, globally affect the nervous system and involve many types of cells. Thus, the application of gene therapy in a global manner to these diseases raises the problem of how to distribute a gene therapy vector to the nervous system. In contrast, Parkinson's disease is confined primarily to a well-defined, compact group of neurons. This fact, along with the availability of well-defined animal models, has helped bring Parkinson's disease to the forefront in studies of gene therapy for neurodegenerative diseases. The neurons primarily affected in Parkinson's disease are located in the substantia nigra (SN) of the brainstem. These cells synthesize dopamine and innervate large areas of the forebrain that are essential to the control of movement. Supplying therapeutic genes to these neurons by injection of viral vectors into the forebrain would appear to be straightforward and relatively safe. The therapeutic gene would then be expected to be retrogradely transported to the dopamine neurons in the brainstem via transport of the vector itself. Alternately, the gene product would provide a therapeutic effect by acting on the dopamine terminals or their target neurons at the site of injection. A second advantage of Parkinson's disease over other neurodegenerative diseases is the variety of excellent rodent and primate models that have long been available. Neurotoxins, such as 6-hydroxydopamine (6-OHDA) and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), selectively kill dopamine neurons in rodents and monkeys. These have provided models for testing effects of putative therapeutic genes not only on neuronal protection, but also on behaviors that result from a deficiency in dopamine. The behavioral consequences of administering MPTP to monkeys closely mimic the clinical symptoms in Parkinsonian patients. Finally, the molecular mechanisms involved in the development, survival, and function of dopamine neurons are very well understood. This knowledge has led to the pursuit of two strategies of gene therapy for Parkinson's. One is aimed at replacing enzymes involved in dopamine synthesis. The second strategy, which is the focus of this commentary, is to provide genes to enhance the survival of dopamine neurons and to rejuvenate their function. Although therapies that slow or reverse the degeneration of dopamine neurons have long been sought, none is currently available. Thus, gene therapy with neurotrophic factors is one strategy that holds high promise for Parkinson's disease in this regard. Neurotrophic factors are small glycoproteins with potent effects on the survival and differentiation of specific classes of neurons. Dissociation constants for neurotrophic factors with their receptors are in the nanomolar to picomolar range; thus, very low levels of these factors would be needed for biological effects. However, direct protein therapies are ineffective because these proteins do not efficiently cross the blood–brain barrier. Even direct injection into the ventricles does not result in sufficient amounts of the factors reaching affected neurons (1Kordower J.H. Palfi S. Chen E.Y. Ma S.Y. Sendera T. Cochran E.J. Cochran E.J. Mufson E.J. Penn R. Goetz C.G. Comella C.D. Clinicopathological findings following intraventricular glial-derived neurotrophic factor treatment in a patient with Parkinson's disease.Ann. Neurol. 1999; 46: 419-424Crossref PubMed Scopus (361) Google Scholar). Moreover, Parkinson's disease is a slowly progressive disease and if neurotrophic factor therapies are to be effective, chronic administration will likely be required for long-term neuronal protection. Therefore, strategic delivery of a neurotrophic gene to diseased dopamine neurons may be the most judicious way to achieve a chronic therapy. Glial cell line-derived neurotrophic factor (GDNF), a survival factor for dopamine neurons, is one strong candidate gene for therapeutic intervention. GDNF is a remarkable molecule. First isolated in 1993 from a glial cell line, it was shown to have extremely potent effects in the picomolar range on the survival and growth of dopamine neurons cultured from the embryonic rat brain (2Engele J. Schubert D. Bohn M.C. Conditioned media derived from glial cell lines promote survival and differentiation of dopaminergic neurons in vitro:Role of mesencephalic glia.J. Neurosci. Res. 1991; 30: 359-371Crossref PubMed Scopus (134) Google Scholar, 3Lin L.F. Doherty D.H.H. Lile J.D. Bektesh S. Collins F. GDNF: A glial cell line-derived neurotrophic factor for midbrain dopaminergic neurons.Science. 1993; 260: 1130-1132Crossref PubMed Scopus (2875) Google Scholar). Over the next several years, numerous studies in rat, mouse, and monkey models of Parkinson's disease demonstrated the effectiveness of GDNF in protecting dopamine neurons against the damaging effects of neurotoxins [for review, see Ref. (4Bohn M.C. Choi-Lundberg D.L. Gene therapies for Parkinson's disease.in: Chiocca E.A. Breakefield X.O. In Gene Therapy for Neurological Disorders and Brain Tumors. Humana Press, Towata, NJ1998: 377-395Crossref Google Scholar)]. Subsequently, several groups showed that GDNF gene delivery via injection of adenoviral (Ad) or adeno-associated viral vectors (AAV) prior to a neurotoxic insult could protect dopamine neurons from dying in a 6-OHDA rat model (5Choi-Lundberg D.L. Lin Q. Chang Y.N. Chiang Y.L. Hay C.M. Mohajeri H. Davidson B.L. Bohn M.C. Dopaminergic neurons protected from degeneration by GDNF gene therapy.Science. 1997; 275: 838-841Crossref PubMed Scopus (573) Google Scholar, 6Choi-Lundberg D.L. Lin Q. Schallert T. Crippens D. Davidson B.L. Chang Y.N. Chiang Y.L. Qian J. Bardwaj L. Bohn M.C. Behavioral and cellular protection of rat dopaminergic neurons elicited by an adenoviral vector encoding glial cell line-derived neurotrophic factor.Exp. Neurol. 1998; 154: 261-275Crossref PubMed Scopus (170) Google Scholar, 7Bilang-Bleuel A. Revah F. Colin P. Locquet I. Robert J.J. Mallet J. Horellou P. In trastriatal injection of an adenoviral vector expressing glial-cell-line-derived neurotrophic factor prevents dopaminergic neuron degeneration and behavioral impairment in a rat model of Parkinson disease.Proc. Natl. Acad. Sci. USA. 1997; 94: 8818-8823Crossref PubMed Scopus (274) Google Scholar, 8Mandel R.J. Spratt S.K. Snyder R.O. Leff S.E. Midbrain injection of recombinant adeno-associated virus encoding rat glial cell line-derived neurotrophic factor protects nigral neurons in a progressive 6-hydroxydopamine-induced degeneration model of Parkinson's disease in rats.Proc. Natl. Acad. Sci. USA. 1997; 94: 14083-14088Crossref PubMed Scopus (299) Google Scholar). The first of these studies demonstrated that only 10–15% of dopamine neurons degenerated if AdGDNF was injected near the dopamine perikarya in the SN prior to inducing a 6-OHDA lesion (5Choi-Lundberg D.L. Lin Q. Chang Y.N. Chiang Y.L. Hay C.M. Mohajeri H. Davidson B.L. Bohn M.C. Dopaminergic neurons protected from degeneration by GDNF gene therapy.Science. 1997; 275: 838-841Crossref PubMed Scopus (573) Google Scholar). This vector provided nanogram levels of GDNF in the vicinity of the dopamine neurons. Because it is a secreted protein, GDNF acted at a distance from the vector injection site and did not require direct infection of dopamine neurons. Subsequent studies showed that injection of GDNF vectors even near the terminals of dopamine neurons in the striatum provided significant protection of dopamine perikarya in the SN (6Choi-Lundberg D.L. Lin Q. Schallert T. Crippens D. Davidson B.L. Chang Y.N. Chiang Y.L. Qian J. Bardwaj L. Bohn M.C. Behavioral and cellular protection of rat dopaminergic neurons elicited by an adenoviral vector encoding glial cell line-derived neurotrophic factor.Exp. Neurol. 1998; 154: 261-275Crossref PubMed Scopus (170) Google Scholar, 7Bilang-Bleuel A. Revah F. Colin P. Locquet I. Robert J.J. Mallet J. Horellou P. In trastriatal injection of an adenoviral vector expressing glial-cell-line-derived neurotrophic factor prevents dopaminergic neuron degeneration and behavioral impairment in a rat model of Parkinson disease.Proc. Natl. Acad. Sci. USA. 1997; 94: 8818-8823Crossref PubMed Scopus (274) Google Scholar). This effect could be explained by retrograde transport by axons of the dopamine neurons, either of the vector itself or of GDNF protein secreted from infected cells in the striatum, or both. Studies in rat clearly show that GDNF gene delivery prior to causing a lesion of dopamine neurons significantly inhibits the death of these neurons. This protective effect is observed whether the vector is injected near the dopamine perikarya or near their terminals. However, these two sites are not equivalent if one looks at the effects of delivering the GDNF gene after damage has already commenced. In a "rescue" strategy, we have observed that AdGDNF GDNF gene delivery can rescue these neurons from death if the vector is injected in the SN, but not in the striatum (9Kozlowski D.A. Connor B. Tillerson J.L. Schallert T. Bohn M.C. Delivery of a GDNF gene into the substantia nigra after a progressive 6-OHDA lesion maintains functional nigrostriatal connections.Exp. Neurol. 2000; (in press)Google Scholar). In this case, the GDNF vector was injected 1 week following 6-OHDA induction. Injection of the vector near the terminals after damage had commenced was ineffective probably because enough terminals were damaged to the degree that insufficient amounts of either GDNF protein or the GDNF vector could be transported back to the SN. However, if AdGDNF was injected into the SN, twice as many dopamine neurons that projected to the lesion site survived compared to those in control rats, and dopamine-dependent function was protected. Behavioral studies in rat models of Parkinson's disease also suggest that the forebrain and brainstem sites are not equivalent sites for GDNF gene therapy (9Kozlowski D.A. Connor B. Tillerson J.L. Schallert T. Bohn M.C. Delivery of a GDNF gene into the substantia nigra after a progressive 6-OHDA lesion maintains functional nigrostriatal connections.Exp. Neurol. 2000; (in press)Google Scholar, 10Connor B. Kozlowski D.A. Schallert T. Tillerson J.L. Davidson B.L. Bohn M.C. Differential effects of glial cell line-derived neurotrophic factor (GDNF) in the striatum and substantia nigra of the aged Parkinsonian rat.Gene Ther. 1999; 6: 1936-1951Crossref PubMed Scopus (126) Google Scholar). To maintain dopamine-dependent behaviors, intact dopamine terminals in the forebrain are required. In Parkinson's patients, there is already an 80–90% loss of dopamine in the forebrain when clinical symptoms appear. Prior to this, dopamine levels are apparently maintained by the dopamine system's remarkable ability for compensation. Although it would be safer to inject vectors into the human forebrain rather than into the brainstem where the dopamine perikarya are located, there will not be a substrate for GDNF action if most of the dopamine terminals have degenerated prior to GDNF gene therapy. On the other hand, if the vector is injected into the SN alone, it may protect the neurons from dying, but not restore dopamine levels in the forebrain. It is likely that the most effective approach will be to inject GDNF vectors into both the brainstem and forebrain. Furthermore, as means become available to diagnosis this devastating disease prior to the degeneration of the majority of the dopamine neurons, GDNF gene therapy could be applied to early Parkinson's patients with promise of a more remarkable therapeutic effect. Although Parkinson's disease is not immediately life threatening, it is nevertheless a very debilitating disease that seriously compromises the patient's quality of life for many years. It also has a relatively high incidence, affecting about 1% of individuals over age 50. During early stages of the disease, pharmacological therapies with dopaminergic agonists or L-DOPA, the precursor for dopamine, are very effective in ameliorating clinical symptoms. However, patients usually become refractory to these treatments as the disease progresses. If GDNF gene expression were to elicit only a small increase in dopamine levels, this could lead to a prolonged masking of clinical symptoms, as the onset of clinical symptoms does not occur until there is about an 80% depletion in dopamine. Of course, earlier gene therapy intervention would make it more likely that there would still be enough dopamine neurons to protect and rejuvenate. What is required to bring GDNF gene therapy for Parkinson's disease to the clinic? First, a vector needs to be identified that has the potential of providing high levels of long-term transgene expression in the human brain in the absence of pathology. Such vector testing is in progress, and new vectors such as lentivirus, helper-free AAV, and internally deleted Ad are all promising. GDNF gene therapy also needs to be shown to be effective in the MPTP monkey model of Parkinson's disease. Again, such studies are in progress in several laboratories, and the results should soon be known. Finally, it will be important to carefully correlate neurochemical, morphological, and biochemical effects of GDNF gene delivery in the MPTP monkey model to the levels of GDNF transgene expression in different brain regions. The effects of expressing high levels of neurotrophic factors ectopically in the human brain are essentially unknown. A potent molecule such as GDNF, if chronically expressed at high levels in ectopic locations in the brain, may eventually lead to unexpected and deleterious side effects. Also unknown are the possible consequences of providing a factor that may continually upregulate growth and function in dopamine neurons. Motor control is under the fine control of competing neuronal systems. There could, therefore, be a threshold level of GDNF expression below which clinical symptoms in Parkinson's patients might initially be ameliorated and above which symptoms might worsen. Thus, a vector with the GDNF gene under a regulatable promoter should be considered for clinical trials. In summary, the recognition and excitement over the past decade that neurotrophic factors hold therapeutic potentials for neurodegenerative diseases might first be realized in gene therapy trials for Parkinson's disease. However, it remains to be determined whether animal models in which dopamine neurons are chemically lesioned, although closely representative of this disease, faithfully reflect what will happen to diseased dopamine neurons in humans when exposed to GDNF. The author thanks Elizabeth Crown for editorial assistance and NIH (Grants NS31957 and NS39267) for support.