Abstract
Background: Background The leucine-rich repeat kinase 2 gene (LRRK2) harbours highly penetrant mutations that are linked to familial parkinsonism. However, the extent of its polymorphic variability in relation to risk of Parkinson's disease (PD) has not been assessed systematically. We therefore assessed the frequency of LRRK2 exonic variants in individuals with and without PD, to investigate the role of the variants in PD susceptibility.
Methods: LRRK2 was genotyped in patients with PD and controls from three series (white, Asian, and Arab-Berber) from sites participating in the Genetic Epidemiology of Parkinson's Disease Consortium. Genotyping was done for exonic variants of LRRK2 that were identified through searches of literature and the personal communications of consortium members. Associations with PD were assessed by use of logistic regression models. For variants that had a minor allele frequency of 0·5% or greater, single variant associations were assessed, whereas for rarer variants information was collapsed across variants.
Findings: 121 exonic LRRK2 variants were assessed in 15 540
individuals: 6995 white patients with PD and 5595 controls, 1376 Asian patients and 962 controls, and 240 Arab-Berber patients and 372 controls. After exclusion of carriers of known pathogenic mutations, new independent risk associations were identified for polymorphic variants in white individuals (M1646T, odds ratio 1·43, 95% CI 1·15-1·78; p=0·0012) and Asian individuals (A419V, 2·27, 1·35-3·83; p=0·0011). A protective haplotype (N551K-R1398H-K1423K) was noted at a frequency greater than 5% in the white and Asian series, with a similar finding in the Arab-Berber series (combined odds ratio 0·82, 0·72-0·94; p=0·0043). Of the two previously reported Asian risk variants, G2385R was associated with disease (1·73, 1·20-2·49; p=0·0026), but no association was noted for R1628P (0·62, 0·36-1·07; p=0·087). In the Arab-Berber series, Y2189C showed potential evidence of risk association with PD (4·48, 1·33-15·09; p=0·012).
Interpretation: The results for LRRK2 show that several rare and common genetic variants in the same gene can have independent effects on disease risk. LRRK2, and the pathway in which it functions, is important in the cause and pathogenesis of PD in a greater proportion of patients with this disease than previously believed. These results will help discriminate those patients who will benefit most from therapies targeted at LRRK2 pathogenic activity.
Funding: Michael J Fox Foundation and National Institutes of Health.
| Original language | English |
|---|---|
| Journal | The Lancet Neurology |
| Volume | 10 |
| Issue number | 10 |
| Pages (from-to) | 898-908 |
| Number of pages | 11 |
| ISSN | 1474-4422 |
| DOIs | |
| Publication status | Published - 01.10.2011 |
Funding
The results of our study, one of the largest so far of the genetics of PD, show that a single gene, ). Although population stratification is an inherent caveat of this type of large-scale collaborative effort (and a potential limitation of the present study in the absence of genome-wide population control markers), these findings exemplify the confluence and independent effects of rare and common variations on gene loci that have a major effect in shaping both familial and sporadic disease. LRRK2 , harbours many rare and common variants that confer susceptibility to PD in diverse populations ( panel About a third of variants we assessed were not identified in any study participant. These included four previously documented pathogenic mutations (LRRK2 N1437H, R1441G, Y1699C, and I2020T), showing that they are rare mutations in the population samples we assessed. 26 variants were recorded at a frequency greater than 0·5% in any of the three series, and only 13 were noted at a frequency greater than 0·5% in all three series. This finding draws attention to the importance of studying genetic variability in large samples and in different ethnic groups, because frequencies and genetic effects might vary substantially. 26 The newly identified associations warrant further discussion. M1646T in the COR (C-terminal of Ras) domain of LRRK2 was identified in the white series, and the effect was consistent in many countries ( figure 1 ). This variant was not identified in participants of Asian descent and was rare in the series of Arab–Berber participants. LRRK2 A419V was consistently more common in patients than in controls in Asian sites ( figure 2 ). Although we cannot exclude the possibility of a non-coding element in linkage disequilibrium, the N-terminal region of the protein seems functionally relevant to disease development. LRRK2 M1646T is the first common-risk factor to have been identified in white populations, whereas A419V is now the third risk factor reported to be specific to individuals of Asian ancestry, along with R1628P and G2385R. 12,14,15 LRRK2 R1628P was not significantly associated with risk in our Asian series. This variant was common only in the Taiwanese series, in which a non-significant protective effect was noted. Our inability to replicate the previously reported risk effect of R1628P is likely to be due to a combination of the low frequency of this variant, natural sampling variation, and population heterogeneity, in view of the results of previous studies of ethnic Han Chinese populations (of note, G2385R did show association). 14,15 The identification of a common three-variant haplotype (N551K-R1398H-K1423K) that seems to act in a protective manner ( figure 3 ) is also important. It suggests that the reduced penetrance that is noted in patients with LRRK2 -associated parkinsonism might be due to variants acting in cis or trans with the pathogenic variant and that LRRK2 activity can be exploited to modify symptom onset in patients. Any future therapeutic strategies that lower risk in LRRK2 -associated parkinsonism might protect against symptomatic onset in idiopathic PD. 14,27 The previous report 14 of a protective effect with N551K and R1398H showed a reduced kinase activity for the R1398H variant, suggesting this Roc domain substitution might be the most likely functional allele on the haplotype. Although the results of our study have identified an association of PD only with common variants, they also draw attention to the many rare variants in LRRK2 that could contribute to disease risk. Genetic loci that contribute to disease risk might do so through variants that span the whole range of minor allele frequencies, from rare mutations to frequent single nucleotide polymorphisms. 28 Despite the very large sample size, we noted only three of seven previously described pathogenic LRRK2 mutations. Hence, the search for mutations contributing to familial PD should include an analysis of single pedigrees, with further assessment in very large population studies. Single pedigrees might result in some false-positive results, which can be filtered out with large population samples. For example, two variants (I1371V and T2356I) have been proposed as pathogenic and to account for the clinical and functional features of LRRK2- associated parkinsonism. 29,30 However, in our study, both variants were noted in patients and controls at the same frequency ( table 4 ). Conversely, we noted other possible rare risk (E334K, R1325Q, and T1410M) and protective (A211V and A1151T) variants; however, because of their low frequency, large meta-analytical approaches are necessary to define their roles fully. In this study, we focused on exonic variants because all pathogenic variants identified in LRRK2 so far have been single nucleotide missense changes. However, silent, synonymous variants were also included because they can result in alternative splicing and, since protein translation is a function of codon use and transfer RNA abundance, could affect the rate of protein domain folding and secondary modifications. 31 Neither copy number variants nor other risk factors in non-coding regions that regulate LRRK2 expression or alter splicing were assessed in our study. As new loci for susceptibility to diverse diseases are continuously being discovered in genome-wide association and whole-genome sequencing studies, the results of our study show the importance of revisiting loci at which rare or common variants have been identified, since they could harbour many more independent signals of genetic risk in different populations. 25,32,33 Furthermore, LRRK2 sequencing studies in under-represented populations (eg, from South America, sub-Saharan Africa, Middle East, and western Asia) will undoubtedly show novel ethnic-group-specific risk variants and could clarify the role of variants that were rare or absent in our study. LRRK2 variants, including novel exonic variants, were reported as part of the 1000 Genome Project, lending support to this hypothesis. 34 Large-scale parallel resequencing (targeted genomic capture of the specific regions—eg, gene-specific, exome, transcriptome, and whole-genome sequencing) is likely to identify many more variants in candidate genes that might predispose to PD. Characterisation of each variant will require this type of collaborative international effort to define their pathogenicity, frequency in different populations, and contribution to disease pathogenesis through genotype–phenotype assessment. This online publication has been corrected. The corrected version first appeared at thelancet.com/neurology on September 19, 2011 Contributors OAR and MJF were the principal investigators and were responsible for the concept and design of the study. AIS-O, JAB, OAR, and CVG were responsible for the technical aspects of the study. MGH and NND were responsible for all the analyses; OAR and MJF were responsible for drafting the report. All authors participated in study design and approach, sample collection, data acquisition, and critical revision and final approval of the report. Conflicts of interest JOA, MJF, and ZKW report holding a patent on LRRK2 genetic variability and MJF has received royalties for licensing of genetically modified LRRK2 mouse models. DMM declares a patent pending entitled Methods to treat PD . CK and RK declare receiving payment in their role as consultants for Centogene and Takeda Pharmaceutical, respectively. All other authors declare that they have no conflicts of interest. Acknowledgments This report is dedicated to the memory of J Mark Gibson (1953–2010). The work in this study was supported by a grant from the Michael J Fox Foundation for Parkinson's Research (OAR and MJF). Original funding for GEO-PD was supported by a grant from the Michael J Fox Foundation for Parkinson's Research Edmond J Safra Global Genetics Consortia programme. The Mayo Clinic is a Morris K Udall Center of Excellence in Parkinson's Disease Research (P50 NS072187) and was supported by a gift from the family of Carl Edward Bolch Jr and Susan Bass Bolch (DWD, RJU, ZKW, and OAR). This research was undertaken, in part, thanks to funding from the Canada Excellence Research Chairs programme (MJF and CV-G). Leading Edge Endowment Funds, provided by the Province of British Columbia, LifeLabs, and Genome BC, support the Dr Donald Rix BC Leadership Chair (MJF). Studies at individual sites were supported by different funding agencies worldwide—the Italian Ministry of Health (Ricerca Corrente 2010, Ricerca Finalizzata 2006); Fondazione Livio Patrizi; Swedish Parkinson Academy; the Swedish Parkinson Foundation; Lund University Research Fund, American Fidelity Assurance Insurance and the Royal Physiographic Society, Lund (AP and CN); Federal Ministry for Education and Research (BMBF, NGFNplus; 01GS08134; RK); NGFNplus (Neuron-Parkinson-subproject 7; SG); South African Medical Research Council and the University of Stellenbosch (SB, JC); Centre Hospitalier Régional Universitaire (CHRU) de Lille, University Lille 2 INSERM; French Ministry Programme Hospitalier de Recherche Clinique (1994/2002/1918, 2005/1914); Association France Parkinson (2005); Fondation de France 2004-013306; Fondation de la Recherche Médicale (2006); Le Programme Pluri-Formations (synucléothèque 2005–2009); Centres de Ressources Biologiques (L'Institut Pasteur de Lille, CHRU-Lille) and their scientific committee; the Agence Nationale de la Recherche (ANR-05-NEUR-019 and ANR-08-MNP-012; AB, SL); grant ES10758 from the National Institutes of Health; Swedish Research Council; Swedish Society for Medical Research; Swedish Society of Medicine; funds from the Karolinska Institutet and the Parkinson Foundation in Sweden (KW); Special Research Fund of the University of Antwerp; Research Foundation Flanders (Fonds Wetenschappelijk Onderzoek–Vlaanderen [FWO]); the Agency for Innovation by Science and Technology in Flanders (IWT); Interuniversity Attraction Poles Program P6/43 of the Belgian Federal Science Policy Office; Methusalem Excellence Grant of the Flanders Government and the Medical Research Foundation Antwerp and Neurosearch, Belgium; National Institutes of Health and National Institute of Neurological Disorders and Stroke 1RC2NS070276, NS057567, P50NS072187; Mayo Clinic Research Committee Clinical Research programmes (MCF and ZKW); Geriatric Medical Foundation of Queensland (GDM); a career development award from the Volkswagen Foundation and from the Hermann and Lilly Schilling Foundation (CK); Research Committee of University of Thessaly (code 2845); and Institute of Biomedical Research and Technology, CERETETH (code 01-04-207; GH and ED); and GlaxoSmithKline for past sponsorship of research into familial parkinsonism in Tunisia (RG and FH). DC is a holder of an FWO PhD fellowship and JT receives an FWO postdoctoral fellowship. For their contributions to make this work possible, we acknowledge Ferdinanda Annesi, Patrizia Tarantino (Institute of Neurological Sciences, National Research Council, Piano Lago di Mangone, Cosenza, Italy); Chiara Riva (Department of Neuroscience and Biomedical Technologies, University of Milano-Bicocca, Monza, Italy); Roberto Piolti (Department of Neurology, Ospedale San Gerardo, Monza, Italy); Magdalena Boczarska-Jedynak (Department of Neurology, Medical University of Silesia, Katowice, Poland); Aurélie Duflot, (UMR837 INSERM-University Lille 2, CHRU de Lille); Jean-Philippe Legendre, Nawal Waucquier (Neurologie et Pathologie du Mouvement, Clinique de Neurologie du CHU de Lille); Anna Rita Bentivoglio, Tamara Ialongo, Arianna Guidubaldi, Carla Piano (Institute of Neurology, Catholic University, Rome, Italy); Karen Nuytemans (Neurodegenerative Brain Diseases Group, Department of Molecular Genetics, Vlaams Instituut voor Biotechnologie; Laboratory of Neurogenetics, Institute Born-Bunge and University of Antwerp, Belgium); Sebastiaan Engelborghs; Peter De Deyn (Department of Neurology, ZiekenhuisNetwerk Antwerpen Middelheim and Laboratory of Neurochemistry and Behaviour, Institute Born-Bunge and University of Antwerp); David Crosiers, Patrick Cras (Department of Neurology, University Hospital Antwerp and Laboratory of Neurobiology, Institute Born-Bunge and University of Antwerp, Belgium); Phil Hyu Lee (Department of Neurology, Yonsei University College of Medicine, Seoul, South Korea); Susanne Lindskov (Department of Geriatrics and Neurology, Central Hospital Kristianstad, Northeast Skåne Health Care District, Kristianstad, Sweden); Karin Nilsson (Department of Clinical Science, Section of Geriatric Psychiatry, Lund University, Sweden); Jan Reimer (Department of Neurology, Skåne University Hospital, Sweden); Manabu Funayama, Yuanzhe Li, Hiroyo Yoshino (Juntendo University School of Medicine, Tokyo, Japan); and we acknowledge all the patients and controls who kindly donated DNA to make collaborative studies like these possible. A full list of GEO-PD consortia is provided in webappendix pp 27–30 .
