1. ¹Division of Neurology, Department of Medicine, Center for Molecular Medicine, Jichi Medical School, Tochigi 329-0498, Japan
2. ²Divison of Genetic Therapeutics, Center for Molecular Medicine, Jichi Medical School, Tochigi 329-0498, Japan
3. ³Department of Life Science, Tokyo Institute of Technology, Kanagawa 226-8501, Japan
4. ⁴Institut de Génétique et de Biologie Moléculaire et Cellulaire, Centre National de la Recherche Scientifique, Institut National de la Santé et de la Recherche Médicale, Université Louis Pasteur, Collège de France, and Institut Clinique de la Souris, 67404 Illkirch Cedex, France

Correspondence: Keiya Ozawa, Division of Genetic Therapeutics, Center for Molecular Medicine, Jichi Medical School, 3311-1 Yakushiji, Minamikawachi, Tochigi 329-0498, Japan. Fax: +81 285 44 8675.. E-mail: kozawa@ms.jichi.ac.jp; Shin-ichi Muramatsu, Division of Neurology, Department of Medicine, Jichi Medical School, 3311-1 Yakushiji, Minamikawachi, Tochigi 329-0498, Japan. Fax: +81 285 44 5118. E-mail: muramats@ms.jichi.ac.jp

Received 14 May 2005; Revised 26 July 2005; Accepted 1 August 2005.

Abstract

Regulation of gene expression is necessary to avoid possible adverse effects of gene therapy due to excess synthesis of transgene products. To reduce transgene expression, we developed a viral vector-mediated somatic regulation system using inducible Cre recombinase. A recombinant adeno-associated virus (AAV) vector expressing Cre recombinase fused to a mutated ligand-binding domain of the estrogen receptor (CreER^T2) was delivered along with AAV vectors expressing dopamine-synthesizing enzymes to rats of a Parkinson disease model. Treatment with 4-hydroxytamoxifen, a synthetic estrogen receptor modulator, activated Cre recombinase within the transduced neurons and induced selective excision of the tyrosine hydroxylase (TH) coding sequence flanked by loxP sites, leading to a reduction in transgene-mediated dopamine synthesis. Using this strategy, aromatic L-amino acid decarboxylase (AADC) activity was retained so that L-3,4-dihydroxyphenylalanine (L-dopa), a substrate for AADC, could be converted to dopamine in the striatum and the therapeutic effects of L-dopa preserved, even after reduction of TH expression in the case of dopamine overproduction. Our data demonstrate that viral vector-mediated inducible Cre recombinase can serve as an in vivo molecular switch, allowing spatial and temporal control of transgene expression, thereby potentially increasing the safety of gene therapy.