The Hunt for a Cure for Parkinson’s Diseasesageke.sciencemag.org/cgi/reprint/2001/1/re1.pdf · affected with the disease with unaffected siblings (sib pair analysis). This would

sageke.sciencemag.org/cgi/content/full/sageke;2001/1/re1 Page 1

Several exciting new scientific advances have beenmade in the past decade toward both understandingthe causes of and finding a cure for Parkinson’s dis-ease. Heartened by an acceleration in research find-ings in the past several years, the government hasrecently called for an infusion of funds from both theNational Institutes of Health and private foundationsinto this burgeoning area of biomedical research.Most currently available conventional treatments forthe disease only temporarily delay symptom presen-tation while doing nothing to halt disease progres-sion. However, the rapidly accelerating pace of re-search in this field has left researchers hopeful thatParkinson’s will be the first major age-related neu-rodegenerative disease for which we have a viablecure. In this article, advances in various areas ofParkinson’s disease research are reviewed.

How Soon Will We Have a Cure?Fueled by a renewed sense of optimism both within the researchcommunity and among clinicians and patient advocates,Congress recently issued the Morris K. Udall Parkinson’s Re-search Act. Encouraged by the rapid pace of scientific advancesin the past several years, this directive called for a 5-year agendaoutlining efforts to expand combined National Institutes ofHealth (NIH)- and foundation-funded research toward the realis-tic goal of a cure for and/or prevention of Parkinson’s disease(PD) (a complete copy of the agenda is available atwww.ninds.nih.gov/about_ninds/nihparkinsons_agenda.htm).PD is a chronic, progressive, and disabling disorder that afflictsapproximately 1% of the U.S. population over the age of 50;those numbers rise to over 5% by the age of 85 (1). PD is present-ly second only to Alzheimer’s disease (AD) as the most commonage-related neurodegenerative disorder (see http://sageke.sciencemag.org/cgi/content/full/sageke;2001/1/oa2 and http://sageke.sciencemag.org/cgi/content/full/sageke;2001/1/dn2). As thepopulation of the United States continues to age, PD will becomemore and more prevalent. This devastating illness is characterizedby a decreased ability to initiate voluntary motor movement, calledbradykinesia (an excellent video depicting these symptoms can befound at http://medweb.bham.ac.uk/http/depts/clin_neuro/teach-ing/tutorials/parkinsons/parkinsons2.html).

Bradykinesia results primarily from the gradual loss ofneurons containing the neurotransmitter dopamine in a tinyarea of the brain located near the brain stem called the sub-stantia nigra (“black substance”) or SN (2, 3). The presence ofneuromelanin in the dopaminergic cells in this brain regiongives them their characteristic darkened appearance. These

neurons normally make connections to a part of the midbraincalled the striatum (ST). The nigrostriatal connection is part ofa larger system of neuronal interconnections that help convertabstract thoughts of movement into voluntary action (Fig. 1).The release of dopamine by the SN neurons into the striatumnormally acts to modulate release of the neurotransmitter γ-aminobutyric acid (GABA) from the striatum. GABA releasefrom the striatum results in inhibition of motor movement. In-hibition of GABA release by dopamine normally is balancedby release of the excitatory neurotransmitter acetylcholinefrom the motor cortex into the striatum, which acts to stimu-late striatal GABA release. Therefore, loss of dopaminergicmodulation of GABA release results in muscle rigidity, restingtremor, and gait and balance disturbances. The cause ofdopaminergic cell loss in PD is still a mystery to researchers;however, accelerated research discoveries in the past decade

have yielded some tantalizing clues that may help halt the pro-gression, restore function, or even prevent the disease.

Unfortunately, most current therapies for PD, while oftendrastically halting or suppressing the symptoms at early stagesin the disease, do nothing to prevent disease progression. Themost common treatment for the disease is oral administrationof the dopamine precursor L-3,4-dihydroxyphenylalanine (L-DOPA). L-DOPA acts to up-regulate the production ofdopamine in the remaining cells in the SN, but does nothing toprevent the eventual death of these cells (3). As more and moreof the cells are lost, increased doses of the drug must be ad-ministered, which can result in side effects that might be as dis-tressing to the patients as the disease itself. These includedyskinesia (involuntary motor movements), psychoses, and

The Hunt for a Cure for Parkinson’s Disease

Julie K. Andersen, Jyothi Kumar, Bharath Srinivas, Deepinder Kaur, Michael Hsu, and Subramanian Rajagopalan

(Published 3 October 2001)

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The authors are in the Buck Institute for Age Research, Novato, CA94945, USA. E-mail: [email protected] (J.K.A.)

Fig. 1. Schematic representation of the important neuronal con-nections within the midbrain relevant to Parkinson’s disease. (A)Sagittal section of the human brain showing connections betweenthe SN (blue) and basal ganglia of the striatum (dark pink). (B)The connections described in (A) are shown in a horizontal brainsection as seen from the inferior side of the human brain (thecerebellum has been excluded). The SN is in blue and the basalganglia of the striatum are in dark pink. The top of the diagram in-dicates the forebrain.

SAGE KESCIENCE OF AGING KNOWLEDGE ENVIRONMENT


fluctuations in motorcontrol. As a result ofcontinued cell loss, thedrug eventually loses itseff icacy. The ultimategoal in the PD researchfield is the ability to di-agnose the condition atan early stage and to pre-vent subsequent cell lossfrom occurring in sus-ceptible individuals.

What Genetic StudiesHave Told UsLike AD, PD does notappear to be a primarilygenetic disorder butrather to arise from othercauses (such as exposureto environmental toxins,head trauma, viruses,etc.), perhaps coupledwith genetic susceptibili-ty (4, 5). However, muchis likely to be gleanedfrom understanding themolecular pathways thatare disrupted in rare fa-milial cases of PD, asthese are likely to be thesame pathways affectedin the more commonsporadic forms of thedisorder (6). For example, although mutations in the gene forthe presynaptic protein alpha synuclein are present only in afew PD pedigrees, the discovery of this rare autosomal reces-sive gene mutation led to the realization that aggregates of theprotein are a major component of intraneuronal occlusionscalled Lewy bodies, a pathological hallmark associated withsporadic PD (7-12). Aggregation of alpha synuclein and otherproteins within these Lewy bodies may play a role in the ensu-ing cell death. Aberrant protein aggregation is indeed emergingas a common feature of many different neurodegenerative dis-eases, including AD, Huntington’s disease, spinocellular ataxi-as, and amyotrophic lateral sclerosis (ALS) (13-15). Mutationsin the so-called parkin gene were discovered in another subsetof families with a rare juvenile form of PD, and subsequentstudies identified this gene as an E3 ubiquitin protein ligase, animportant component of the ubiquitin-proteosome proteindegradation pathway. This finding has further fueled the notionthat the lack of appropriate disposal of proteins may lead totheir aggregation and contribute to disease pathology (16, 17).In yet another early-onset PD pedigree, a missense mutationwas reported in the gene encoding ubiquitin carboxyl-terminalhydrolase L1 (UCH-L1), another enzyme involved in the ubiq-uitin-proteosome protein degradation pathway (18). Proteo-some function itself recently has been reported to be impairedin the SN of patients with sporadic forms of PD (19). Polymor-phisms on other loci on chromosomes 1p, 2p, and 4p have beenreported in other rare inherited forms of PD, but the corre-

sponding genes have yet to be identified (20-22). The identification of inherited gene mutations resulting in

PD phenotypes has spurred researchers into attempting to cre-ate genetic models of the disease via expression of the corre-sponding genes in either cultured cells or whole animals. Thehope is that expression of these genes will result in models thatmimic at least some aspects of the disease. For example, trans-genic Drosophila and mouse models that overexpress either thewild-type or mutant forms of the alpha synuclein gene recentlyhave been created (23-26). Alpha synuclein transgenic flies dis-play pathological changes similar to those observed in the PDbrain, including the selective age-dependent loss of midbraindopaminergic neurons and the development of intranuclear in-clusions resembling Lewy bodies. The flies also display an age-related loss in locomotor function, although it is not knownwhether this can specifically be attributed to dopaminergic cellloss or whether dopamine agonists reverse this behavioral phe-notype. Mice that express wild-type alpha synuclein in neuronsdisplay age-related Lewy body-like formation in the SN, cortex,and hippocampus. However, some of these occlusions appear tobe nuclear rather than cytoplasmic and not to be of the normalfibrillary type observed in autopsied brains from PD patients(24). Although there is no actual loss of midbrain dopaminergicneurons by 1 year of age in the alpha synuclein transgenic mice,there is a slight reduction in the number of striatal dopamineterminals as measured by immunocytochemistry performedwith antibodies to tyrosine hydroxylase (TH). In addition, a

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Fig. 2. Schematic diagram of electron transfer in the mitochondria and the sources of reactive oxygenspecies (ROS) resulting in oxidative stress within the organelle. The numbers I, II, III , IV, and V in thecross section of the mitochondria refer to the multisubunit protein complexes present along the inner mi-tochondrial membrane. These complexes play key roles in the transfer of electrons necessary as an ener-gy source for ATP production. The red and yellow arrows that connect the complexes indicate the direc-tion of electron transport. The blue arrows show oxygen consumption and the generation of ROS withinthe organelle. UQ, ubiquinone. Cyt, cytochrome c.



drop in TH activity was observed in the highest-expression line(the neurons of which express alpha synuclein in the cortex at alevel of 80% compared to the corresponding neurons in the nor-mal human brain), suggesting an initiation of the disease. Thisline also displayed impaired motor ability as measured by therotorod test, although as with the transgenic flies, it was not de-termined whether this could be reversed with dopamine replace-ment therapy (23, 24). Even if these animals do not completelyrecapitulate all of the PD pathology, transgenic animal modelsthat express genes mutated in rare familial forms of PD con-ceivably could be important tools for understanding more aboutdisease symptomology and could aid in diagnosing the condi-tion (27). They would also be useful for testing the efficacy ofvarious pharmacologicaltreatments (as well as safeand effective gene thera-pies) before moving thesetherapies into clinical trialsin humans. Finally, suchmodel systems may alsoassist in the identificationof novel therapeutic targetsfor the disease. Suppressormutant screens of alphasynuclein flies, for in-stance, could be used toidentify candidate protec-tive gene products.

Although most casesof PD don’t appear to beprimarily genetic in na-ture, there still might ex-ist genetic susceptibilitiesthat might render an indi-vidual more prone tomanifesting the disease inthe presence of certainenvironmental factors.Cardiovascular disease isan illustration of anotherage-related disorder thatappears to result from acombination of both ge-netic risk and environ-mental influences. Possi-ble methods for identify-ing “risk factor” genes include genetic comparison of thoseaffected with the disease with unaffected siblings (sib pairanalysis). This would allow the identification of candidategenes involved in susceptibility to the disease. Past twin anal-ysis studies in which disease frequency was compared be-tween cohorts of identical versus fraternal twins found nosignificant difference between the two, suggesting no under-lying genetic cause for sporadic PD (28, 29). However, recentreanalysis of the data suggests that low penetrance could ex-plain these results. Furthermore, subsequent positron emis-sion tomography (PET) analysis suggests that the asymp-tomatic identical twin sibs of affected patients displayedpresymptomatic signs of the disease (30). In PET, fluorodopais administered intravenously and is allowed to be convertedinto dopamine within the nigrostriatal system, where it can

then be noninvasively imaged at high resolution to assessdopamine synthesis capacity in this brain region as a measureof neuronal cell function. Although these results suggest thatthere might be some genetic involvement in PD, shared envi-ronmental influences cannot be ruled out (31).

The analysis of complex genetic traits at a population level iscurrently being revolutionized through the large influx of infor-mation from the human genome project and the use of noveltechnologies such as gene microarray and proteomic analyses. Inthese types of analyses, patterns of expression of tens of thou-sands of genes or proteins can be assessed simultaneously to lookfor a “fingerprint” of gene or protein expression associated withthe disease (32-34). This fingerprint can then be used both as a

diagnostic tool and toidentify novel genes andproteins that are differen-tially expressed during var-ious stages of the diseaseprocess. Such genes andproteins may representnovel targets for therapy orbiomarkers for the disease.

What Studies of Envi-ronmental Interac-tions Have Told Us PD appears to be mostprevalent in industrial-ized countries. In addi-tion, people moving fromnonindustrial countrieswith low disease inci-dence to industrial coun-tries with higher diseaseincidence show an in-creased occurrence of thedisease (4). This suggeststhat industrial toxinsmight play a role in PD.On the basis of these ob-servations, numerousepidemiological studieshave been performed toidentify environmentalfactors that might predis-pose individuals for the

disease (35-41). The most common of these types of studiesinvolve comparisons between people with and without PD(case-control studies). Results from several epidemiologicalstudies, for example, have suggested that populations thatdrink well water have a higher incidence of sporadic forms ofthe disease (42). This observation gives credence to the no-tion that exposure to environmental industrial toxins such ascertain herbicides and pesticides may increase the risk for PD(43). Identification of such potential toxins allows scientiststo follow people who have experienced exposure to a particu-lar putative toxin to see if they develop symptoms of the dis-ease (long-term studies). These symptoms can be correlatedwith gene/protein profiles to assess the interplay betweentoxin exposure and genetic susceptibility. The profiles mayalso aid in identifying preclinical characteristics of disease.

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Fig. 3. How MPTP causes selective death of dopaminergic SN neurons inthe brain. After systemic injection, MPTP crosses the blood-brain barrierinto the brain, where it is believed to diffuse into all cell types. It is convert-ed into MPDP+, an intermediate product, by the enzyme MAO-B withinbrain astrocytes. MPDP+ can then spontaneously form MPP+ either withinthe astrocyte itself or after diffusion into the extracellular space. MPP+ isthen specifically taken up into dopaminergic neurons via the dopaminetransporter (DAT). Once inside the dopaminergic neuron, MPP+ is takenup into the mitochondria via an energy-dependent transport process,where it acts as a specific inhibitor of mitochondrial complex I.



In PD, as is the case with many other multifactorial diseases,interpretation of these experiments may be difficult becauseof the complex nature of the illness. In order to expedite suchwork, large pedigrees with familial PD need to be identifiedand carefully characterized for use in testing the relation be-tween environment and genetic predisposition and to assessthe efficacy of preventative treatments. In addition, the roleof age, gender (PD is reported to be higher in men than wom-en), race (PD is reported to be higher in Caucasian than inother racial populations), and geographic distribution needsto be assessed (4). These analyses would help to address is-sues such as whether noted epidemiological trends are due toactual disease incidence or rather to the underreporting of PDas the cause of death in minority or other populations. Studiesin animal models of PD appear to support the noted epidemi-ological male-female differences in PD susceptibility in hu-mans. This raises the possibility of a protective role for fe-male hormones such as estrogen (44-46).

What Basic Scientific Research Has Told UsAdvances over the past couple of decades in the basic neuro-sciences have increased our understanding of the fundamentalmechanisms involved in neuronal cell death, which may helpus to know more about the dopaminergic cell loss that occursin PD. Both postmortem data from human PD patients and datafrom research using various cellular and animal models of thedisease strongly suggest that mitochondrial dysfunction is amajor cause of neuronal cell death (47-49). Mitochondria arethe power plants of the cell. Compromise of their function re-sults in the cell’s inability to produce adequate energy suppliesin the form of adenosine 5′-triphosphate (ATP) to sustain itself.ATP is synthesized via an intricate series of electron transferreactions involving five sequential mitochondrial protein com-plexes (I through V) (Fig. 2) (for review, see http://sageke.sciencemag.org/cgi/content/full/sageke;2001/1/oa5). Mito-chondria also generate reactive oxygen species (ROS), whichare toxic byproducts generated during conversion of high-ener-

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Fig. 4. Aberrant metabolic events associated with PD at the cellular level and possible therapeutic interventions. The flow chart depictsvarious interacting elements that, acting in concert, may lead to the dopaminergic SN cell death associated with PD. Italicized wordsshown in blue depict current or potential therapeutic measures to prevent or alleviate the particular metabolic events indicated by theblack arrows. ROS, Reactive oxygen species; DA, dopamine; GSH, glutathione; MAO-B, monoamine oxidase B; L-DOPA, L-3,4-Dihy-droxyphenylalanine; PD, Parkinson’s disease; gliosis, nonneuronal proliferative response to injury.



gy oxygen to low-energy water as fuel for ATP production.ROS production may also contribute to cellular degeneration.

The link between neuronal cell demise in PD and mito-chondrial dysfunction was originally discovered as a conse-quence of the manufacturing of an illegal drug gone awry. In1982, drug addicts in the San Francisco Bay area, while at-tempting to make an illegal heroin analog, overcooked theconcoction, which resulted in the formation of a compoundknown as 1-methyl-4-phenyl 1,2,3,6-tetrahydropyridine(MPTP). Injection of MPTP resulted in acute “end stage”Parkinsonism in these individuals, due to the rapid loss of thesame set of neurons that are destroyed in PD (50). Adminis-tration of MPTP to experimental animals results in similar ef-fects (51, 52). Animal studies have demonstrated that MPTPacts as an inhibitor of mitochondrial complex I, one of theprinciple entry points for electrons into the electron transportchain. Interestingly, the activity of this enzyme complex isdecreased selectively in the SN of PD patients (Fig. 3) (53-55). In addition, when rodents are treated with chronic lowdoses of rotenone, a mitochondrial complex I inhibitor thatwas once used both as an insecticide and as a means to cullfish populations, they develop PD-like pathology, includingprogressive nigrostriatal degeneration, formation of Lewybody-like occlusions, and motor deficits. These pathologieshave been shown to be reversed upon treatment with adopamine receptor agonist (56).

Aberrant mitochondrial function that results in cell deathcan take forms other than energetic failure or production ofROS, including dysregulation of the control of intracellularcalcium levels or induction of a form of genetically regulatedcell suicide known as apoptosis (57). Indices of all of thesetypes of damage are found to various degrees in the brains ofPD patients and in various animal and cellular models of thedisease [for examples, see (2, 5, 25-27, 49, 55, 58-62)].These various mechanisms of cell death can act singly or inconcert to elicit the cell’s demise. In our laboratory, for ex-ample, we recently demonstrated that reduction of the an-tioxidant glutathione (GSH) in dopamine-containing cells invitro results in the selective inhibition of mitochondrial com-plex I activity, suggesting there is a causative link betweenthe two (63). GSH depletion is the earliest reported de-tectable biomarker for PD, and the degree of loss closely cor-relates with the severity of the disease (64-66). Loss of GSHcoupled with the presence of dopamine, which can be oxi-dized to form toxic oxidant metabolites, and a high iron con-tent in this brain region, which can exacerbate this oxidationprocess, may lead to increased ROS in dopaminergic SNneurons, making them more vulnerable to degeneration (67-69). L-DOPA treatment itself has been proposed to result inthe generation of toxic oxidative species (“oxidative stress”)that may contribute to cell loss, although this hypothesis iscontroversial (70-72). Oxidative stress and subsequent dam-age to cellular proteins may secondarily lead to protein ag-gregation, which can contribute to the pathology of PD. Inaddition, recent unpublished results from our laboratory sug-gest that GSH depletion can directly affect the ubiquitinpathway by allowing increased thiol oxidation of the E1 andE2 ligases, which are crucial enzymes involved in the trans-fer of ubiquitin to the appropriate protein. This in turn couldaffect the ability of proteins to be properly degraded and leadto aggregation.

From Scientific Findings to Pharmacological InterventionBecause mitochondrial impairment appears to play a pivotalrole in the demise of dopaminergic SN neurons in PD, nutrition-al agents that act to enhance bioenergetic function may be ther-apeutic (73). Oral administration of CoQ, a cofactor involved inthe transport of electrons between complexes I and II and a po-tential antioxidant, and nicotinamide adenine dinucleotide(NADH), a substrate for complex I, have both been reported toprevent cell death in various animal models of PD and to im-prove symptoms in PD patients themselves (74-78). CoQ subse-quently was found to be ineffective in a short clinical trial in hu-mans, although there was a trend toward increased mitochondri-al complex I activity (79). On the basis of these results, the peri-od of the trial has been extended. NADH given either intra-venously or orally has been reported to improve disability in PDpatients. Creatine, a substrate for creatine kinase, which is in-volved in ATP production, has been reported to be protective inthe MPTP model of PD (80). These results taken in total sug-gest that bioenergetic supplementation may hold promise as atherapeutic for PD.

ROS production has emerged as another important con-tributing factor in PD-related neuronal cell death (58, 62, 81-83). On the basis of earlier epidemiological reports, a largeclinical trial called DATATOP was conducted in the early1990s to assess the efficacy of oral administration of the an-tioxidant vitamin E with respect to PD symptoms. Although vi-tamin E was found to be ineffectual in this study, the chemicaldeprenyl was found to delay the need for L-DOPA therapy andprovided at least some temporary symptomatic relief to pa-tients (84, 85). Deprenyl is an inhibitor of monoamine oxidaseB (MAO-B), which catalyzes the oxidation of dopamine. It isimportant to note that the levels of vitamin E used in theDATATOP study were less than those found to be successful ina previous, smaller clinical study, so dosage may be an issue(86). Another small clinical trial in which GSH was adminis-tered intravenously to PD patients over a 1-month period re-vealed that this treatment showed some efficacy in terms ofimproved motor ability (87). GSH does not easily cross theblood-brain barrier (BBB), which may limit its usefulness as atherapy on its own. However, alternatives to GSH therapy havebeen suggested for testing in clinical trials, including the GSHprecursor N-acetylcysteine or the GSH substitute lipoic acid, oresterized forms of the compound that can more easily cross theBBB (88). In addition, extremely potent antioxidant enzymemimetics called EUKs have been developed recently and wereshown to prevent MPTP-induced cell death in PD models (89).These compounds have the added important benefit of beingable to cross the BBB easily. They also have been shown to in-crease life-span in Caenorhabditis elegans, presumablythrough reduction of age-related oxidative stress (90).

Although initially controversial, evidence is building thatapoptosis may play an important role in the neuronal cell deathassociated with PD (91, 92). Induction of apoptosis in PD as inother neuronal cell death scenarios probably involves the activa-tion of a cascade of cysteine proteases called caspases. Twomembers of this cascade, upstream caspase 8 and downstreamcaspase 3, have been reported to be present in dying SN neu-rons in the brains of autopsied tissue from PD patients (93). Un-published data from our own laboratory suggest that mice thatexpress the general caspase inhibitor protein baculoviral p35 inneurons are more resistant to MPTP toxicity, and Beal and col-

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leagues have demonstrated that mice expressing a dominant-negative form of caspase 1 also display decreased MPTP-in-duced SN cell death (94). These observations suggest that phar-macological caspase inhibitors may constitute another possibleavenue of therapy for PD. Such compounds are already beingtested clinically for their efficacy in other neurological condi-tions in which apoptosis has been suggested to play a role (forexample, stroke) (95).

How various interrelated mechanisms of cell death such asmitochondrial dysfunction, oxidative stress, apoptosis, and pro-tein aggregation result in the selective death of dopaminergicSN neurons in PD is still not completely understood. However,by understanding the basic molecular processes involved inneuron loss, we might be able to delay or even halt dopaminer-gic SN cell death through a combination of therapeutic ap-proaches (for example, bioenergetic supplements, antioxidants,metal chelators, caspase inhibitors, etc.) (96). In addition, withhigh-throughput drug screening assays that measure the func-tions of molecular players in the process of selective cell death,it is possible to test the effects of large numbers of smallmolecules on these molecular targets for drug therapy. This, ofcourse, requires adequate cellular models of the disease, butcould include, for instance, assessment of the ability of drugs toinhibit toxin-induced apoptosis or alpha synuclein aggregationin dopaminergic neurons in culture.

Possible Roads to TherapeuticsProlonged exposure of neurons to the excitatory neurotransmit-ter glutamate can result in excitotoxic cell death. This is espe-cially the case for vulnerable cells with compromised energymetabolism. Dopaminergic neurons of the SN express gluta-mate receptors, and these cells degenerate when exposed to ex-citoxins. In vivo treatment with various excitotoxin agonists hasbeen demonstrated to attenuate MPTP-induced cell death, espe-cially with agonists specific for the NMDA-type glutamate re-ceptor [for review, see (97-99)]. Attentuation of glutamate trans-mission by other means, including both pharmacological andsurgical interventions (see below), also appears to attenuate thesymptoms of PD. It is not clear whether removing excess gluta-mate input into the nigrostriatal system, however, really protectsor only delays dopaminergic cell death.

Inflammation, initiated most likely in response to nerve celldamage, also appears to be involved in PD pathogenesis. Mi-croglial activation has been demonstrated both in the MPTP an-imal model and in PD patients [for example, see (100-105)]. Ithas been suggested that microglia elicit their toxic effects byproduction of nitric oxide (NO) through induction of the en-zyme nitric oxide synthase (NOS) in these cells. NO can in turnreact with superoxide to form peroxynitrite, which can elicit ni-tration of tyrosine residues on proteins such as TH, the rate-lim-iting enzyme in dopamine synthesis. Nitration of tyrosineresidues can thus affect the function of these target proteins.Evidence suggesting the presence of nitrotyrosine has been ob-served both in PD animal models and in patients (106-108).NOS inhibitors might be another possible therapy for the dis-ease and indeed have been reported to be effective in attenuat-ing disease progression in various animal models (109). Neu-roimmunophilins have also been reported to elicit both protec-tive and restorative effects, although interestingly not due totheir immunosuppressive actions. Rather, these effects appear toresult from inhibition of calcineurin, which can induce NOS ac-

tivity (110-112). Neuronally derived NOS may be another im-portant component of cell death in PD, as NOS inhibitors alsoappear to be protective in animal models (113, 114).

Delivery of protective neurotrophic growth factors todopaminergic SN neurons and their terminals in the striatum isanother hotly pursued line of research in the PD field. This ap-proach was previously hindered by the lack of effective meth-ods of delivery for clinical usage (115). Most neurotrophicagents are large molecules that don’t easily cross the BBB. Mi-croinjection has worked well in animal models but less well inpatients. Recently, Kordower and colleagues demonstrated thatdelivery of glial-derived growth factor (GDNF), a neurotrophicfactor relatively selective for dopaminergic neurons, usinglentivirus as a vector for its expression, resulted in sustained ex-pression of the protein over a several-month period and func-tional recovery from MPTP-mediated Parkinsonism in the ma-jority of rhesus monkeys treated with the agent (116). The de-gree of recovery correlated well with prevention of loss ofdopaminergic terminals in the striatum. This produced much ex-citement in the Parkinson’s research community. However, sev-eral parameters still need to be carefully assessed before humantrials can be contemplated. For example, how safe for humanuse is lentivirus, an HIV-related virus? Concern was raised overthe fact that a small number of the injected animals died aftertreatment with the virus. Is the virus capable of expressing thetransgene for years as opposed to only months? Sustained ex-pression may be necessary to maintain the function of thedopamine SN neurons over the life-span of the affected individ-ual. Also, not all monkeys that displayed effective delivery andexpression of the transgene showed recovery—what needs to bedone to optimize the procedure?

Toxic models such as MPTP or rotenone administration,along with novel genetic models of PD, now allow scientists toexamine the progression of PD as it occurs. This provides re-searchers with the opportunity to identify which molecular pro-cesses are important at each stage of the disease and to pinpointplaces at which therapeutic interventions can be administered.Such model systems also allow scientists to assess the types ofcompensatory changes that occur in the cells or circuitry as aresult of the disease. For example, researchers can determinewhether loss of dopamine in the SN neurons leads to chronic al-terations in related synapses, neurons, and circuits, such asmolecular changes in the concentrations of neurotransmitter re-ceptors or signal transduction molecules that can affect the pre-sentation of disease symptoms. If these physiological changeshelp to delay the course of the disease, it may be possible to em-ulate these changes using pharmacological agents. By under-standing dopaminergic SN neuronal function and how it fits in-to the larger neural circuitry of the brain, it also might be possi-ble to restore the lost function via protein (for example, neu-rotrophin) or dopaminergic cell replacement. Understanding thecomplexities of the affected circuitry has already yielded infor-mation leading to surgical manipulations that control some ofthe symptoms of PD. An example is neuronal ablation of path-ways from other brain regions that send inhibitory signals to thestriatum, such as from the globus pallidus. A depiction of thisso-called “pallidotomy” can be seen at http://medweb.bham.ac.uk/http/depts/clin_neuro/teaching/tutorials/parkinsons/sur-gery/parkinsons-surgery.html or www.pallidotomy.com/parkin-son_s_disease.html). Another procedure developed more re-cently is deep brain stimulation of the subthalamus nuclei (117-

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119). In this procedure, a device is implanted into the brain thatallows chronic electrical stimulation of neurons in an attempt torestore normal neuronal circuitry; this relieves many of thesymptoms of the disease and the affects appear to be long-last-ing. However, the problem with current surgical interventions isthat, in general, they only deal with the symptoms of the diseaseand not the ongoing progressive cell loss.

Cell Replacement Therapy: How Feasible Is It?Another exciting area of current PD research involves the re-placement via transplantation of the dopaminergic SN neuronslost in the disease. A plus for the use of cell transplantationtherapy in PD is that the nigrostriatal system is fairly easily ac-cessible and well-defined structurally. Transplantation is usuallydone directly into the striatum, so that the transplanteddopaminergic cells can easily make connections with the striatalneurons (120). Studies in this area of research suggest thattransplantation works best when the recipient is young (that is,during fetal development), although grafted cells are capable ofsurviving and making functional connections even in the adultbrain. Numerous studies in various animal models of PD havesuggested that transplanted cells can undergo normal dopaminesynthesis, turnover, and release and that their presence can actto attenuate behavioral symptoms (120). However, it is impor-tant to note in this context that the vast majority of PD patientsare over the age of 65, and the older the host, the less efficientthe transplant is likely to be. In addition, in humans we knowlittle about how to regulate cell growth and function once cellsare transplanted into the brain. On the basis of some promisingpreliminary results from animal studies, surgeries were per-formed on several PD patients involving the implantation oftheir own dopamine-producing carotid bodies into the striatumbefore it was realized that this method of dopaminergic cell de-livery was ineffective because of poor graft survival (121).There also have been reports of several scattered clinical casestouting the efficacy of fetal tissue grafts as a therapeutic for thedisease; however, a recent large double-blind fetal transplantstudy produced disappointing results with less-than-desired lev-els of benefits and side effects (122). These results preclude forthe moment the widespread usage of fetal transplantation in PDpatients until more clinical trials are completed [for review, see(123-125)]. The published study was done without immunosup-pressants, although PET scan analysis suggests that the graftssurvived even without the use of these agents. Other importantfactors to be assessed in terms of fetal transplant include meth-ods and timing of tissue storage before transplant, amount oftissue to be transplanted, and which growth factors allow thebest maintenance of the cells in vivo. Such parameters mayhave major impacts on the number of neurons that ultimatelysurvive the grafting procedure. It is estimated that approximate-ly 80,000 functional dopaminergic neurons, or roughly one-fifthof the normal SN complement of neurons, are needed for thetreatment to succeed (120). There are many moral and social is-sues connected with the use of fetal tissue, even though thus farthese types of cells appear to work best in grafting experiments.And only a limited amount of such tissue is available.

Another source of tissue for transplantation is embryonicstem cells. Stem cells are undifferentiated cells that can renewthemselves by cell division and can give rise to many types ofmore specialized cells (126). Stem cells from the embryo arepluripotent; that is, they differentiate into all known cell types. In

contrast, adult stem cells may give rise only to certain definedlineages. The dream among researchers is that easily accessiblecells might someday be used to treat diseases such as PD. In-deed, cells from the bone marrow recently were shown to be ca-pable of migrating into the brain and becoming neuronal-likecells (127). The ability to use cells such as those from the bonemarrow has two benefits. One is that it would eliminate the ethi-cal concerns associated with the use of fetal tissue. The second isthe eradication of immune rejection, as the patient’s own cellscould be used. Multipotent stem cells potentially can be an un-limited source of transplantation materials. First, however, theymust be expanded into sufficient numbers for transplantation (asthey are rare) and predifferentiated in vitro toward a dopaminer-gic cell fate. What controls the proliferation and differentiationof stem cells is still poorly understood. It is particularly impor-tant to ascertain whether they retain their pluripotent capacity af-ter transplantation and differentiate into fully functionaldopaminergic neurons. It is also unknown whether regional-specific glial support cells will be required in order to get properdopaminergic cell induction and function in vivo.

Many exciting new avenues of potential PD therapy havebeen opened up thanks to major advances in various researchareas over the past several years (F. 4). With the influx of newcombined government and foundation spending to bolster re-search efforts toward finding its cause and treatment, hopes arehigh that PD will be the first major neurodegenerative diseasefor which we find a cure.

Further ReadingNIH: www.ncbi.nlm.nih.gov/disease/Parkinson.htmlParkinson’s Disease Foundation: www.pdf.orgMichael J. Fox Foundation for Parkinson’s Disease Research: www.michaeljfox.orgAmerican Parkinson’s Disease Association: www.apdaparkinson.orgNational Parkinson Foundation: www.parkinson.orgParkinson’s Action Network: www.parkinsonaction.orgThe Parkinson’s Alliance: www.parkinsonalliance.netThe Parkinson’s Institute: www.parkinsonsinstitute.org

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The Hunt for a Cure for Parkinson’s Diseasesageke.sciencemag.org/cgi/reprint/2001/1/re1.pdf · affected with the disease with unaffected siblings (sib pair analysis). This would