The introduction of mutant, disease-causing genes into the germline of rodents and other animals has been a great boost to neurodegenerative disease research, but there are some limitations to the method, including the time and expense it takes to raise transgenic animals and the possibility that the introduced genes fundamentally alter development. One of the most promising alternatives is the introduction of transgenes into older animals via viruses, but early models were beset by problems with viral toxicity and efficient gene expression. Recent versions have overcome these limitations, and several groups have now used the recombinant adeno-associated virus (rAAV) vector to create or treat models of Parkinson's disease.
Killing Dopaminergic Cells with α-Synuclein
Two groups working independently--Ron Klein and colleagues at the University of Florida in Gainesville and Deniz Kirik, Anders Bjorklund and colleagues at Lund University in Sweden--have produced parkinsonian pathology by using rAAV vectors to overexpress α-synuclein in rats. Bjorklund's group was also able to show Parkinson-like behavioral deficits.
α-Synuclein is a major constituent of Lewy bodies and dystrophic neurites, both pathological hallmarks of PD. Mutations in the gene for α-synuclein can cause familial forms of the disease. Germline transgenic mouse models expressing these mutated genes have mimicked damage to axons and motor impairments, but have failed to produce the characteristic selective loss of dopaminergic cells in the substantia nigra (SN).
Klein's group, reporting in the 20 March Human Gene Therapy, used rAAV to express a mutant human α-synuclein gene. They showed accumulation of α-synuclein in SN neurons, Lewy-like dystrophic neurites in the SN and its target area, the striatum, and a 53 percent loss of dopaminergic neurons.
Bjorklund's group, reporting in the April 1 Journal of Neuroscience, expressed a different human mutant gene, as well as wildtype human α-synuclein. They demonstrated Lewy-like α-synuclein accumulation in the dopaminergic cells, as well as dystrophic neurites, accompanied by large-scale loss of dopaminergic cells and depletion of dopamine in the striatum. They also found significant motor impairment in animals whose cell loss exceeded 50-60 percent. Interestingly, both the human wildtype and mutant α-synuclein genes produced these effects.
Saving Dopaminergic Cells With L-Dopa
Kirik, Bjorklund and colleagues also demonstrate a more complex, therapeutic use of the rAAV vector approach in yesterday's Proceedings of the National Academy of Sciences (USA). They delivered a constant supply of the dopamine precursor levodopa (L-dopa) to the striatum in a mouse model of Parkinson's and thereby reduced, or eliminated, motor impairments.
The intermittent, pulsatile bioavailability of current oral L-dopa treatment in humans is thought to be partly responsible for the drug's diminishing effectiveness and increasing side effects when given for long periods of time. Viral vectors containing the gene for the enzyme tyrosine hydroxylase (TH), which synthesizes L-dopa, have been deployed to the striatum in an attempt to boost dopamine levels, alas without success. The Swedish group used the rAAV vector to deliver genes for TH and a needed cofactor to the striatum of animals with partial or total lesions of the nigrostriatal pathway.
Even in animals with total lesions, this restored striatal L-dopa production to levels sufficient to partially reverse the behavioral effects of the lesions. In animals where some of the nigrostriatal pathway was left intact (a state that perhaps more accurately reflects the situation in PD), the transgenes completely reversed the behavioral deficits of the lesions.
"These data suggest that local intrastriatal L-dopa delivery may be a viable therapeutic strategy in PD not only for treatment of underlying motor deficits but also for control of adverse side effects associated with oral L-dopa therapy such as on-off fluctuations and drug-induced dyskinesias," the authors conclude.—Hakon Heimer
- Klein RL, King MA, Hamby ME, Meyer EM. Dopaminergic cell loss induced by human A30P alpha-synuclein gene transfer to the rat substantia nigra. Hum Gene Ther. 2002 Mar 20;13(5):605-12. PubMed.
- Kirik D, Rosenblad C, Burger C, Lundberg C, Johansen TE, Muzyczka N, Mandel RJ, Björklund A. Parkinson-like neurodegeneration induced by targeted overexpression of alpha-synuclein in the nigrostriatal system. J Neurosci. 2002 Apr 1;22(7):2780-91. PubMed.
- Kirik D, Georgievska B, Burger C, Winkler C, Muzyczka N, Mandel RJ, Bjorklund A. Reversal of motor impairments in parkinsonian rats by continuous intrastriatal delivery of L-dopa using rAAV-mediated gene transfer. Proc Natl Acad Sci U S A. 2002 Apr 2;99(7):4708-13. PubMed.