Genomic and proteomic biomarkers for Parkinson disease.pdf

Genomic and proteomic biomarkers forParkinson diseaseThomas Gasser,MDABSTRACTBiomarkers are objective,accessible,and easily measurable biologic parameters that correlateeither with the presence(trait)or the severity(state)of a disease.As the major neurodegenerativediseases,such as Alzheimer disease or Parkinson disease,are likely to be etiologically heteroge-neous disorders,sensitive and reliable biomarkers that reflect the underlying disease process areurgently needed,and are a prerequisite for a more refined diagnosis and the development of noveldisease-modifying therapeutic strategies.“Genetic biomarkers,”in the form of disease genes orrisk-modifying variants,are able to define the risk of an individual developing a disease,and allowstratification of patient populations according to the underlying molecular defect.Alterations ofthe transcriptome or the proteome,on the other hand,may provide a means to monitor diseaseprogression or severity.However,because of the complex relationship of genotypes and pheno-types in neurodegenerative disorders,the development of useful biomarkers is still in an earlyphase.NEUROLOGY 2009;72(Suppl 2):S27S31Neurodegenerative diseases,such as Alzheimer dis-ease(AD)and Parkinson disease(PD),are a growingchallenge to social and healthcare systems worldwide.Because of the demographic development in westerncountries,a growing number of patients will seektreatment and care.Current treatment strategies forneurodegenerative diseases are mostly aimed at re-lieving major symptoms,not at the disease-processitself.1However,because of the complexity of thepathologic changes and pathogenetic mechanisms un-derlyingclinicallyobservedsymptoms,itisunlikelythatsubstantially better symptomatic treatments will beavailable any time soon.The development of disease-modifying or preventive treatment strategies is,there-fore,of utmost importance.These urgently needed new treatment strategies are,of course,hampered by the fact that the etiology andpathogenesis of neurodegenerative diseases are,in mostcases,still unknown,at least at the molecular level.Asdefined by clinicopathologic criteria,neurodegenerativediseases are,in reality,most likely a group of many dis-eases,each caused by interplay of a number of differentgenetic and nongenetic causes.Therefore,patient co-horts selected according to clinical criteria and used instudies of etiology,disease-modifying treatment or pre-vention,probably represent an etiologically heterogeneousgroup,resultinginlessstatisticalpowerinthesestudies.Furthermore,neurodegenerative diseases usuallyfollow a slowly chronic progressive course,and firstsymptoms appear only when the degenerative processhas progressed for a long time,in most cases probably5 to 10 years.Because of the slow progression,a reli-able estimation of disease onset and disease activity isdifficult,particularly since the affected tissue is notdirectly available to be examined,for example by bi-opsy,for obvious reasons.In addition,the long“pre-symptomatic”(and probably also“prodromal”)phase,results in a diagnosis that occurs many yearsafter the onset of the neurodegenerative process.Forall these reasons,we can at present only look at thepathobiologic underpinnings of PD and other neu-rodegenerative disorders indirectly.Biomarkers thatare objective,accessible,and easily measurable bio-logic parameters,and that correlate either with thepresence(trait)or the severity(state)of a neurode-generative disease,are therefore a major prerequisitefor a more refined diagnosis and the development ofnovel therapeutic strategies.Currently,there is no reliable biomarker for PD,except in a small minority of patients with a mono-From the Hertie-Institute for Clinical Brain Research,Department of Neurodegenerative Diseases,Tu bingen,Germany.Disclosure:This article was written following an expert meeting supported by Teva and Lundbeck.The author has no other financial or nonfinancialconflicts to report.Neurologysupplementsarenotpeer-reviewed.InformationcontainedinNeurologysupplementsrepresentstheopinionsoftheauthors.Theseopinionsare not endorsed by nor do they reflect the views of the American Academy of Neurology,Editor-in-Chief,or Associate Editors of Neurology.Address correspondence and reprintrequests to Thomas Gasser,MD,Hertie-Institute for Clinical BrainResearch,Department ofNeurodegenerative Diseases,Tu bingen,Germanythomas.gasseruni-tuebingen.deCopyright 2009 by AAN Enterprises,Inc.S27genetic form of the disorder,in whom at least theunderlying disease trait can be unequivocally deter-mined by mutational analysis.However,even in thesepatients,the relationship between the biomarker(inthis case the mutation)and disease development iscomplex because of the often reduced and age-dependent penetrance and the variable expressivity.No markers exist to objectively measure the severityand the rate of progression of PD-related cellulardysfunction and neurodegeneration in the substantianigra,let alone in the many other affected brain ar-eas.Despite being clinically very useful,even themost advanced imaging methods,including magnetic resonance spectroscopy and PET,are surrogatemarkers that reflect only certain consequences of thedisease process.Nevertheless,recent progress in our knowledge ofthe genetic and molecular etiology of PD has pro-vided the basis to develop genomic and proteomicbiomarkers that may eventually be instrumental inthe development of new treatment strategies.State and trait biomarkers.A biomarker is a substanceused as an indicator of a biologic state.It should beeasily and objectively measurable and evaluated as anindicator of normal biologic processes,pathogenicprocesses,or responses to a therapeutic intervention.A mutation or other genetic variant can be con-sidered a biomarker for an inherited trait if it is asso-ciated with a specific phenotype,i.e.,a disease.Thisbiomarker can,in principle,be determined easily andunequivocally in the laboratory by sequencing or byanother molecular genetic method.In clinical prac-tice,it has to be kept in mind that the feasibility ofdetermining a genetic mutation depends on the sizeof the gene and the type of mutation.Mutationalanalysis in diseases that are caused by many differentmutations in a very large gene as for exampleleucine-rich repeat kinase 2(LRRK2)-related PD,seelater are still time-consuming and expensive.As the genetic information does not change dur-ing life,a genetic biomarker does not reflect a condi-tion of an organism at a given point in time of its life,but rather an invariable feature or trait,for example,a statistical risk for developing a disease at a certainage.Genetic variants should,therefore,be considered“trait biomarkers.”In contrast,alterations in the proteome,the sumof all proteins of a cell or an organism,reflect itsactual condition at a given moment.If specific de-tectable changes in the concentration or distributionof a protein,or a set of proteins,are associated with adisease,they often follow the course of disease pro-gression and can therefore be called“state biomark-ers.”They may be particularly useful if they are notonly specific for a disease but also show a correlationwith the severity of the disease process.Genomic(trait)and proteomic(state)biomarkers,therefore,reflect different aspects of a biologic process.Genetic and genomic biomarkers for PD.The discov-ery of mutations that cause monogenetic forms ofPD has allowed clinical investigators to determinethe cause of the disease,at least in a small minority ofpatients,namely in mutation carrying individuals,andto define the probability of a mutation carrier develop-ing the disease.Point mutations,duplications,and trip-lications in the?-synuclein gene(SNCA),have beenidentified as a rare but well recognized cause of autoso-mal dominantly inherited PD.24The fact that the en-coded protein,?-synuclein(?SYN),is the majorfibrillary component of the Lewy body,underlinesthe close link between these mutations,the proteinand the pathogenic process.Point mutations,dupli-cations,and triplications of SNCA cause PD withhigh penetrance.This means that carrying the muta-tion confers a high probability of developing the dis-ease.Age at onset is,however,highly variable,ranging from the mid-30s to the late 80s and de-pends,to some extent,on the type of mutation.TheA53T-mutation is often associated with an early on-set(mid-40s),a severe phenotype,rapid progressionand early development of dementia.However,rareindividuals who carry the mutation but have not de-veloped the disease until their 70s or even later,havebeen found in these families.Early onset is also therule for patients with SNCA triplications,while du-plications usually cause a late-onset disease.That thisis,in fact,due to the SNCA“dose”has been shownclearly in a family in whom duplication and triplica-tion of the identical genomic fragment bearing theSNCA gene was found in different branches,5lead-ing to late-and early-onset disease,respectively.However,age at onset of each mutation type is asso-ciated with a fairly high variability.Therefore,evenin the relatively simple case of monogenetic causa-tion of a disease with high penetrance,prediction ofonset and course in individual cases is not possiblewith a high degree of confidence.A second autosomal dominant gene causing PD isthe gene for LRRK2.6,7Mutations in the LRRK2gene are much more common than mutations inSNCA,being responsible for about 1015%of fa-milial cases,6and 12%of sporadic cases,in whitepopulations.7In some genetically isolated popula-tions such as the Ashkenazi Jews8and the North Af-rican Arabs,9the prevalence of a particular LRRK2mutation,G2019S,is much higher and reaches 3040%in familial cases.Age at onset is usually in the60s.Although the causative role of this mutation isnot in question,it is clear that penetrance is age-dependent and incomplete:healthy 90-year-old mu-S28Neurology 72(Suppl 2)February 17,2009tation carriers have been described.Based onpopulation studies,penetrance of the G2019S muta-tion has been estimated to be only 3040%.10Al-though this might be an underestimate,it is clearthat carrying the G2019S mutation does not un-equivocally predict development of PD during a life-time.The value of a genetic biomarker in predictingan individuals risk of developing the disease is,therefore,questionable.Other mutations in theLRRK2 gene may have a higher penetrance,as theyhave been identified in large multigenerational fami-lies only.These considerations are even more importantwith respect to genetic variants in the SNCA andLRRK2 genes,which have been identified as modifi-ers of disease risk in sporadic patients.It has nowbeen clearly shown in several studies that geneticvariability in SNCA modifies the risk of developingsporadic PD.Single nucleotide polymorphisms,i.e.,variations in the DNA sequence that occur com-monly in the human genome,in multiple regions ofthe gene including the promotor,intronic regionsand the 3?UTR(the“tail end”of the gene)havebeen found to be associated with PD in differentpopulations.1113The mechanism by which these ge-netic variants influence disease susceptibility is notclear,but may include altered gene expression by dif-ferential binding of transcription factors to the pro-moter region,14as well as other mechanisms such asalternative splicing or altered RNA processing.15Another common risk allele appears to be theG2385R variant of the LRRK2 gene.This variant isvery rare in Europeans,but occurs in about 35%ofall individuals of the Han Chinese population.Prev-alence of this variant in Chinese PD patients isclearly increased to about 810%,as shown by sev-eral studies of G2385R,16,17and was recently shownto extend to the Japanese population.18Becauseoftherelatively high frequency of these risk alleles,they ac-count for a substantial proportion of the population-attributable risk.However,the frequency of the riskalleles can be as high as 5%in controls for the G2385Rvariant or even approximately 30%for certain SNCArisk variants,again rendering this biomarker useless forindividual cases of disease prediction.Use of genetic and genomic biomarkers.As has beendemonstrated above,genetic mutations and variantsfulfill the criteria for biomarkers as they are objec-tively measurable traits that reflect the risk of devel-oping a specific disease.However,genetic orgenomic biomarkers are probably not so valuable interms of individual risk determination.Rather,theywill serve to stratify patient populations according tocertain risk profiles.This will be useful for epidemio-logic studies including those that address questionsof geneenvironment interaction.They will also al-low identification of relatively homogeneous patientpopulations for neuroprotection studies.For exam-ple,if a neuroprotective agent targets increasedSNCA expression as its mode of action,it may beadvantageous to be able to select those patients inwhom?SYN over-expression is an important factorin disease pathogenesis.If,on the other hand,LRRK2 kinase signaling is the target for pharmaco-logical intervention,it may be reasonable to selectpatients with risk variants associated with an increasein LRRK2 kinase activity.If the neuroprotective po-tential of a drug can,in fact,be shown by this ap-proach,geneticriskprofilesmayevenbeusedtostratifycohorts for preventive or disease-modifying treatmentin clinical practice.Proteomic biomarkers.Ideally,a biomarker that is ex-pected to accurately reflect a disease process shouldbe accessible for examination in the affected tissue,i.e.,in the case of PD in the ailing dopaminergicneurons.The fact that this is not possible is certainlyone of the major obstacles to developing causative ordisease-modifying treatments in PD.The value ofthis approach is obvious,for example,in modern tu-mor treatment that can be tailored according to theindividual hormone receptor status of the malignantcells.As this approach cannot be taken in PD,pro-teomic disease-associated alterations must be searchedfor in accessible body fluids such as blood plasma orCSF,orinperipheraltissues.Although it seems to be along shot at first glance,there are indications thatdetected changes may,in fact,reflect at least certainaspects of the disease process in the brain.Many metabolic pathways have been implicatedin the pathogenesis of PD,including mitochondrialand proteasomal dysfunction,but measurements ofthe concentration of their protein components ortheir enzyme activities in peripheral tissue have,sofar,produced contradicting results.A detailed discus-sion of these studies is beyond the scope of this re-view.As a consequence of the genetic findingsdiscussed above,?SYN has also become a major fo-cus of research at the proteomic level.?SYN is one of the most abundant proteins ex-pressed in the brain.It is found in many brain areas,particularly in neurons.In brain homogenates of pa-tients with PD,decreased?SYN concentrations havebeen reported.19These findings need to be inter-preted with caution,however