Dopamine Transporter (DAT) Inhibitors Alleviate Specific Parkinsonian Deficits in Monkeys: Association with DAT Occupancy in Vivo Bertha K. Madras, Michele A. Fahey, Martin Goulet, 1 Zhicheng Lin, Jacob Bendor, 2 Claudia Goodrich, Peter C. Meltzer, David R. Elmaleh, Eli Livni, Ali A. Bonab, and Alan J. Fischman Department of Psychiatry, Harvard Medical School and New England Primate Research Center, Southborough, Massachusetts (B.K.M., M.A.F., M.G., Z.L., J.B., C.G.); Organix, Inc., Woburn, Massachusetts (P.C.M.); and Department of Nuclear Medicine, Massachusetts General Hospital, Boston, Massachusetts (D.R.E., E.L., A.A.B., A.J.F.) Received March 27, 2006; accepted August 1, 2006 ABSTRACT Viable dopamine neurons in Parkinson’s disease express the do- pamine transporter (DAT) and release dopamine (DA). We postu- lated that potent DAT inhibitors, with low affinity for the serotonin transporter (SERT), may elevate endogenously released extracel- lular dopamine levels to provide therapeutic benefit. The thera- peutic potential of eight DAT inhibitors was investigated in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-treated cynomol- gus monkeys (Macaca fascicularis), with efficacy correlated with DAT occupancy as determined by positron emission tomography imaging in striatum. Four potent DAT inhibitors, with relatively high norepinephrine transporter, but low SERT affinities, that occupied the DAT improved activity in parkinsonian monkeys, whereas three high-affinity DAT inhibitors with low DAT occupancy did not. 2-Carbomethoxy-3-(3,4-dichlorophenyl)-7-hydroxy-8-meth- yl-8-azabicyclo[3.2.1.]octane (O-1163) occupied the DAT but had short-lived pharmacological effects. The benztropine analog dif- luoropine increased general activity, improved posture, reduced body freeze, and produced sleep disturbances at high doses. (1R)-2-(1-Propanoyl)-3-(4-fluorophenyl)tropane (O-1369) allevi- ated parkinsonian signs in advanced parkinsonian monkeys, by increasing general activity, improving posture, reducing body freeze, and sedation, but not significantly reducing bradykinesia or increasing locomotor activity. In comparison with the D 2 -D 3 DA receptor agonist quinelorane, O-1369 elicited oral/facial dyskine- sias, whereas quinelorane did not improve posture or reduce balance and promoted stereotypy. In conclusion, DAT inhibitors with therapeutic potential combine high DAT affinity in vitro and high DAT occupancy of brain striatum in vivo with enduring day- time effects that do not extend into the nighttime. Advanced parkinsonian monkeys (80% DAT loss) respond more effectively to DAT inhibitors than mild parkinsonian monkeys (46% DAT loss). The therapeutic potential of dopamine transport inhibitors for Par- kinson’s disease warrants preclinical investigation. Parkinson’s disease (PD) is the most common of the neu- rodegenerative movement disorders, with approximately 1% of the population older than 65 years presenting with this progressive disease. Disease symptoms are caused by degen- eration of dopamine neurons in the substantia nigra, with significant losses of dopamine and the dopamine transporter (DAT) in the basal ganglia (e.g., Kish et al., 1988; Kaufman and Madras, 1991). The dopamine precursor L-dopa alone or L-dopa combined with D 2 -D 3 dopamine receptor agonists are drug therapies of choice for treating idiopathic Parkinson’s disease. In symptomatic Parkinson’s disease, exogenous L- dopa replenishes the endogenous dopamine precursor and This study was supported by National Institutes of Health Grants NS30556 (NINDS), DA11558 and DA06303 (NIDA), and RR00168 (NCRR) and Boston Life Sciences, Inc. (Hopkinton, MA). The receptor-screening program was supported in part by the National Institute of Mental Health Psychoactive Drug Screening Program Grant NO1 MH80005. This study was presented in abstract form. Madras BK (2001) Occupancy of the dopamine transporter by a transport inhibitor, as measured by PET imaging, is predictive of therapeutic efficacy for parkinsonism. J Nucl Med 42 (Suppl):210P; Madras BK (2002) The therapeutic potential of a dopamine transport (DAT) inhibitor for Parkinson’s disease: comparison with a D 2 dopamine agonist in MPTP-treated monkeys. Society for Neuroscience, Washington, DC. 1 Current affiliation: EnVivo Pharmaceuticals, Watertown, Massachusetts. 2 Current affiliation: Laboratory of Cellular Molecular Neurosciences, Rock- efeller University, New York, New York. Article, publication date, and citation information can be found at http://jpet.aspetjournals.org. doi:10.1124/jpet.106.105312. ABBREVIATIONS: PD, Parkinson’s disease; DAT, dopamine transporter; DA, dopamine; GBR 12909, 1-{2-[bis-(4-fluorophenyl)methoxy]ethyl}- 4-(3-phenylpropyl)piperazine; MPTP, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine; CFT (WIN 35,428), 2-carbomethoxy-3-4-(fluorophenyl)- tropane; SERT, serotonin transporter; NET, norepinephrine transporter; difluoropine or O-620, (S)-(+)-2-carbomethoxy-3-(di-4-fluorophenyl- methoxy)tropane; O-1369, (1R)-2-(1-propanoyl)-3-(4-fluorophenyl)tropane; O-1014, 2-carbomethoxy-3-(3,4-dichlorophenyl)-8-oxabi- cyclo[3.2.1]oct-2-ene; O-1163, 2-carbomethoxy-3-(3,4-dichlorophenyl)-7-hydroxy-8-methyl-8-azabicyclo[3.2.1.]octane; O-1231, 2-carbome- thoxy-3-(3,4-dichlorophenyl)bicyclo[3.2.1]oct-2-ene; O-1973, 2-(1-propanoyl)-3-(4-chlorophenyl)-8-oxabicyclo[3.2.1]octane; O-2099, 2- (1-propanoyl)-3-(3,4-dichlorophenyl)-7-hydroxy-8-methyl-8-azabicyclo[3.2.1]octane; O-2240, 2-(1-propanoyl)-3-(3,4-dichlorophenyl)-7- hydroxy-8-methyl-8-azabicyclo[3.2.1]octane laurate; PET, positron emission tomography; Ctrl, control; veh, vehicle; BTS 74 398, (1-[1-(3,4- dichlorophenyl)cyclobutyl]-2-(3-diaminethylaminopropylthio)ethanone monocitrate; NS 2330, tesofensine. 0022-3565/06/3192-570–585$20.00 THE JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS Vol. 319, No. 2 Copyright © 2006 by The American Society for Pharmacology and Experimental Therapeutics 105312/3145387 JPET 319:570–585, 2006 Printed in U.S.A. 570 at Harvard University on May 27, 2009 jpet.aspetjournals.org Downloaded from