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Association of the DRD2 CAn-STR and DRD3 Ser9Gly polymorphisms with Parkinson's disease and response to dopamine agonists
Association of the DRD2 CAn -STR and DRD3 Ser9Gly polymorphisms with Parkinson’s disease and response to dopamine agonists Shaoqing Xu, Jiujiang Liu, Xiaodong Yang, Yiwei Qian, Qin Xiao PII: DOI: Reference:
S0022-510X(16)30484-1 doi: 10.1016/j.jns.2016.08.005 JNS 14735
To appear in:
Journal of the Neurological Sciences
Received date: Revised date: Accepted date:
22 February 2016 6 July 2016 1 August 2016
Please cite this article as: Shaoqing Xu, Jiujiang Liu, Xiaodong Yang, Yiwei Qian, Qin Xiao, Association of the DRD2 CAn -STR and DRD3 Ser9Gly polymorphisms with Parkinson’s disease and response to dopamine agonists, Journal of the Neurological Sciences (2016), doi: 10.1016/j.jns.2016.08.005
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ACCEPTED MANUSCRIPT Association of the DRD2 CAn-STR and DRD3 Ser9Gly polymorphisms with Parkinson's disease and response to dopamine
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agonists
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Shaoqing Xu, Jiujiang Liu, Xiaodong Yang, Yiwei Qian, Qin Xiao*
University School of Medicine, Shanghai 200025, China
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Department of Neurology & Institute of Neurology, Ruijin Hospital affiliated to Shanghai JiaoTong
*Correspondence to: Qin Xiao, email: [email protected] Abstract
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Dopamine agonists (DAs) play important roles in the treatment of Parkinson's disease (PD). Currently, it is thought that genetic variations in the genes encoding dopamine receptors (DR) are important factors in determining inter-individual variability in drug responses. To investigate the association
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between Dopamine receptor D type 2 (DRD2) dinucleotide short tandem repeat (CAn-STR) and Dopamine receptor D type 3 (DRD3) Ser9Gly polymorphisms and the risk of PD, as well as the possible reasons for PD patients using different doses of DAs, we recruited 168 idiopathic PD patients
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and 182 controls. There were no significant differences in DRD2 CAn-STR and DRD3 Ser9Gly
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genotypes (p=0.184, p=0.196) or in allele frequencies (p=0.239, p=0.290) between PD patients and controls. There was no association between DRD2 CAn-STR polymorphism and doses of DAs. Among three different DRD3 Ser9Gly genotypes (Ser/Ser, Ser/Gly, Gly/Gly), patients carrying Gly/Gly
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genotype used higher doses of DAs than patients with Ser/Gly and Ser/Ser genotypes (p=0.001). In pramipexole subgroup, the Gly/Gly group took more pramipexole than the other genotype groups (p<0.001), whereas the doses of piribedil were not significantly different among three genotypes
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(p=0.735). Our results suggest that genotype in DRD3 Ser9Gly was the main factor determining different doses of DAs and PD patients carrying Gly/Gly genotype require higher doses of pramipexole for effective treatment. This study may provide insights into understanding possible reasons for different responses to DAs in Chinese PD patients. Key words: DRD2 CAn-STR, DRD3 Ser9Gly, Parkinson's disease, dopamine agonists 1 Introduction Parkinson's disease (PD) is a common neurodegenerative disorder, affecting 1%-3% of people above 65 years of age and with increased prevalence in advancing age [1]. The pathological changes of PD manifest as a progressive loss of dopaminergic neurons in the substantia nigra pars compacta (SNc) and an abnormal aggregation of Lewy bodies. It is commonly recognized that the interaction between heredity and environment contributes to the pathogenesis of the disease. In clinical practice, dopamine replacement therapy is still the principle way to treat PD. However, the dopamine precursor, L-3,4-dihydroxyphenylalanine (L-dopa), is mainly effective in the restitution of most components of motor dysfunction but poorly remedies postural instability and freezing of gait or co-morbid symptoms, such as depression, cognitive impairment, perturbed sleep and sensory dysfunction [2]. Further, 1
ACCEPTED MANUSCRIPT long-term use of L-dopa often leads to motor complications such as wearing-off effects and dyskinesias [3]. As a result, dopamine agonists (DAs), which have a longer half-life compared with L-dopa, are increasingly employed either in conjunction with L-dopa or as monotherapy. Currently, the most investigated DAs are mainly divided into two categories: ergot-derived and
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non-ergot-derived DAs. Nevertheless, there is growing evidence that the ergot-derived dopamine
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agonists, for instance, bromocriptine, cabergoline and pergolide, can give rise to cardiac valvular regurgitation, pulmonary changes and even liver cancer [4-6], and as a result, they are being gradually
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phased out. The non-ergot-derived DAs, pramipexole and piribedil, are two key DAs prescribed in China, and pramipexole is more widely used because of its better efficacy and fewer side effects. Inter-individual differences in response to dopamine agonist treatment remain a critical problem in the
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management of PD patients, and it is thought that genetic variations in the genes encoding the dopamine receptors (DRs) may be causal of the variability in drug responses. The natural targets for DAs consist of five DR subtypes, of which DRD2, DRD3, and DRD4 are composed of D2-like
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receptors, while DRD1 and DRD5 consist of D1-like receptors [7]. When activated, the D1-like receptor subtypes stimulate adenylate cyclase (AC) activity; whereas the D2-like receptor subtypes generally inhibit adenylate cyclase activity [8]. Nearly all of the dopamine agonists mainly act through
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D2-like receptors, rather than the D1-like receptors [9].
Among all DR genes, DRD2 (chromosome 11q22-q23) and DRD3 (chromosome 3q13) have been
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the most investigated [7]. Several polymorphisms in the DRD2 gene have been identified to be associated with drug efficacy in PD. Wang and colleagues found that the DRD2 Taq1A (rs1800497)
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A1/A1 genotype was associated with an increased risk of motor fluctuation in PD [10]. PD patients carrying 13 and 14 repeat alleles of DRD2 CAn-STR was associated with less occurrences of L-dopa induced dyskinesia [11]. Interestingly, another group reported that this protective effect of DRD2 CAn-STR 13 and 14 repeat alleles on dyskinesia risk was restricted to men [12]. Besides, the absence
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of the DRD2 CAn-STR 15 repeat allele was found to be associated with decreased rates of discontinuation of non-ergoline DAs [13]. The most widely investigated polymorphic site in DRD3 was DRD3 Ser9Gly, which had three different genotypes (Ser/Ser, Ser/Gly and Gly/Gly). A serine to glycine substitution in the N-terminal extracellular part of the receptor was reported to be associated with altered dopamine binding affinity [14]. A prospective study including 30 PD patients conducted in China found that the response rates were significantly higher in the Ser/Ser group (60%) than in the group containing the Gly allele (13%) after treatment with pramipexole for 2 months [15]. However, the dosage of pramipexole used was only 0.375 mg/day, which was much smaller than the normally prescribed doses. In our study, we investigated the association between the DRD2 CAn-STR and DRD3 Ser9Gly polymorphisms and the risk of PD. We also examined possible reasons for the variable responses to DAs among PD patients, especially as it relates to the variation in daily DAs dosages between individuals. 2. Materials and methods 2.1. Subjects 2
ACCEPTED MANUSCRIPT A total of 168 idiopathic PD patients and 182 age and gender matched controls were included in the study. The PD patients were recruited from the Movement Disorder Clinic at the Department of Neurology of Ruijin Hospital between December 2013 and April 2016. Inclusion criteria were (1) clinical diagnosis of PD according to the United Kingdom Parkinson's Disease Society Brain Bank
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criteria [16], (2) no history of stroke or significant neurological disorders, and (3) no family history of
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PD extended to the first-degree relatives. The study was approved by the ethics committee of Ruijin Hospital. All participants signed an informed consent.
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All PD patients were examined by at least two movement disorder specialists, and the basic clinical features were recorded including the age of onset, duration of the disease, the Hoehn and Yahr stage (H&Y stage), levodopa equivalent dose (LED), and doses of DAs, including pramipexole and piribedil.
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Additional factors recorded included the Unified Parkinson's Disease Rating Scale (UPDRS) scores and Non-Motor Symptoms questionnaire for Parkinson's disease (NMS). Patients were also evaluated by the Hamilton Anxiety Scale (HAMA), Hamilton Depression Scale (HAMD), and Mini-Mental State
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Examination (MMSE). The LEDs of patients were calculated according to a previous study [17], where 1 mg of pramipexole is equivalent to 100 mg of piribedil and 100 mg L-dopa. 2.2. Genotyping for the DRD2 CAn-STR and DRD3 Ser9Gly polymorphisms
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Genomic DNA was isolated from peripheral blood leucocytes with phenol–chloroform extraction followed by ethanol precipitation. Analysis of CAn-STR polymorphism in the DRD2 gene was by
PCR
amplification
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performed
using
the
following
set
of
primers:
F:
5′-
TGAAGACTCGCCATGAAA-3′ and R: 5′-GGTAGAAGCCAAAGGTGC-3′. PCR products were then
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fluorescently labeled at the 5'-end by 6′-FAM dideoxynucleotides (GENEWIZ, China) and analyzed using the ABI Prism 3730 DNA Sequencer. Allele size was established using Genemapper V4.0 software. Sequence analysis was used to confirm differentiation of alleles by size. PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism) method was performed to test the
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genotype of Ser9Gly polymorphism. A 174-bp product was amplified with an initial 5-min denaturation at 94˚C, followed by 35 cycles of 94 ˚C for 30 s, 56˚C for 30 s, 72 ˚C for 30 s, and a final extension period at 72 ˚C for 5 min, using the upstream primer, 5′-CCAGGCCATTGCCGAAGA-3′ and the downstream primer, 5′-TGGCATCACGCACCTCCCT-3′. The PCR products were digested by MscI enzyme and specific fragments were generated. If it was the Ser allele, then a 174-bp product was detected, and 122-bp and 52-bp product were detected if it was the Gly allele. Genotyping was performed by individuals who were blinded to the clinical status of the subjects. 2.3. Statistical analysis Statistical analyses were performed with SPSS software (ver. 13.0 for Windows, Chicago, IL).The Hardy–Weinberg equilibrium chi-square test was used to assess the data’s goodness of fit. All analyses were 2-tailed, and the level of statistical significance was set at p<0.05. Categorical data were analyzed with the chi-square test or Fisher exact test to establish whether there was an association between PD and the genotypes. The clinical variables were compared between the groups by the 2-sample, unpaired t-test for DRD2 CAn-STR polymorphism continuous variables with approximate normal distribution, 3
ACCEPTED MANUSCRIPT and the chi-square test or Fisher's exact test for discrete variables. For DRD3 Ser9Gly genotype continuous variables, we performed the homogeneity test for variances and then one-way ANOVA. The multiple comparisons were performed by Bonfferoni test and either Pearson's chi-square test or Fisher’s exact test for discrete variables. After revealing the differential variables among genotypes, we carried
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out single-factor linear regression and multiple stepwise linear regression to assess the precise
3. Results
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3.1. Demographic features of PD patients and healthy controls
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association between the differential variables and genotypes with a criterion for entry of p<0.05.
A total of 168 idiopathic PD patients and 182 controls participated in this cross-sectional study, and there were no significant differences in the age (p=0.679) and gender (p=0.452) distributions between
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the patients with PD and the control groups (PD patients: age 66.0±8.9 years, 99males/69 females; controls: age 65.0±8.5 years, 100males/82 females).
3.2. Genotype and allele distributions of DRD2 CAn-STR and DRD3 Ser9Gly in PD patients and
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controls
The genotype and allele distributions of DRD2 CAn-STR and DRD3 Ser9Gly are shown in Table 1. All the observed genotype or allele frequencies did not differ from the expected frequencies according to
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the Hardy-Weinberg equilibrium. Previous studies showed that carrying the 13 and 14 repeat of DRD2 CAn-STR was associated with less risk of L-dopa induced dyskinesia, indicating that 13 and 14 carriers
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might provide some protection [11, 12]. Due to the low frequencies of some genotypes in the DRD2 CAn-STR, subjects were categorized as 13, 14+ or 13, 14− (subjects carrying at least 1 of allele 13 or 14
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in their genotype or carrying neither allele 13 nor 14, respectively) in our study according to Oliveri’s and Zappia’s categorization [11, 12]. The genotype or allele frequencies in the PD patients did not differ from the controls (p=0.184, p=0.239). In addition, there were no significant differences in the DRD3 Ser9Gly genotype distributions when comparing between PD patients and controls (p=0.196) nor in the
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allele distributions (p=0.290).
3.3. Demographic and clinical features of patients taking dopamine agonists There were 102 patients using dopamine agonists in our study. As shown in Table 2, for DRD2 CAn-STR polymorphism, there were no differences in any of the clinical features; however, for DRD3 Ser9Gly genotypes (Ser/Ser, Ser/Gly, Gly/Gly), no significant differences were found in clinical features such as age of onset, duration of the disease, H-Y stage, LED, or the scores of UPDRS total, NMS, HAMA, HAMD or MMSE, except for doses of DAs (p=0.001). Patients with the Gly/Gly genotype used higher doses of DAs compared with those from the Ser/Gly and Ser/Ser genotype groups (141.0±49.6 mg vs. 109.0±33.2 mg and 100.4±36.2 mg, respectively). Doses of DAs in the Gly/Gly subjects significantly differed from Ser/Ser (p=0.001) and Ser/Gly groups (p=0.011) by multiple comparison. After adjusting for gender, age, and other clinical features, by carrying out single-factor linear regression and multiple stepwise linear regression, the doses of DAs were significantly correlated with DRD3 Ser9Gly polymorphisms (p=0.001, p=0.001 respectively). Moreover, we further analyzed pramipexole (n=62) and piribedil (n=40) subgroups, and data were 4
ACCEPTED MANUSCRIPT shown in Table 3 and 4. We found that in DRD2 CAn-STR polymorphisms, there were not any significant differences in the pramipexole (p=0.840) subgroup, while in the piribedil group, the doses of DAs were obviously different between 1314+ and 1314- genotype subjects (120.8±33.5mg vs. 102.5±18.4mg, respevtively, p=0.039). However, single-factor linear regression showed that there was
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no significant association between DRD2 CAn-STR genotypes and daily doses of piribedil. For DRD3
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Ser9Gly polymorphisms, the daily doses of pramipexole in the Gly/Gly group were significantly higher
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than the Ser/Gly and Ser/Ser groups (153.4±54.2 mg vs. 102.2±36.1 mg and 93.5±36.1 mg, respectively, p<0.001), whereas in patients using piribedil, the dosages were not significantly different among the three genotypes (p=0.735). In the pramipexole subgroup, significant differences were seen between the
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Gly/Gly and Ser/Ser groups (p<0.001) as well as the Gly/Gly and Ser/Gly groups (p=0.003) by Benferroni test. Single-factor linear regression and multiple stepwise linear regression showed that after
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adjustment for other factors, a relationship still existed (p=<0.001 in single-factor linear regression, p=0.001 in multiple stepwise linear regression). Results of single-factor linear regression were shown in Table 5. Besides, age of onset was shown to be negatively associated with doses of DAs (B= -1.075,
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p=0.016 in in single-factor linear regression, B= -0.899, p=0.032 in multiple stepwise linear regression).
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4. Discussion
PD is the second most common neurodegenerative disorder, affecting more than 2 million people in China. Though there are several ways to treat PD, such as deep brain stimulation (DBS) [18] and gene
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therapy [19], drug therapy is still the primary approach. L-dopa can be prescribed for dopamine replacement therapy as it enters the brain via the blood brain barrier and is transformed to dopamine by dopa decarboxylase. The efficacy of L-dopa treatment is affected by dopamine metabolism enzymes,
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such as catechol-O-methyltransferase (COMT) and monoamine oxidase B (MAOB), as well as the activity of DRs and the dopamine transporter (DAT). The role of dopamine can also be imitated by DAs that directly act on the DRs in the brain. Pramipexole and piribedil are two new types of non-ergot-derived DAs and mainly act on D2-like dopamine receptors. The heritable variations of dopamine receptor genes could lead to structural and functional abnormalities, potentially resulting in a large inter-individual variability in response to DA drugs. Several studies conducted on PD patients have focused on the variable efficacy of L-dopa treatment as well as the risk of associated motor complications, which may have genetic origins. Bialecka et al. investigated the most common COMT gene haplotypes (formed by single nucleotide polymorphisms (SNPs): rs6269: A>G; rs4633C>T; rs4818: C>G; and rs4680: A>G) in 322 PD patients and 357 controls and reported that the doses of L-dopa administered to G_C_G_G (high activity) haplotype carriers were significantly higher than those administered to non-carriers during the fifth year of treatment [20]. Lee and colleagues observed patients with peak dose dyskinesia and diphasic dyskinesia in a longitudinal study in 503 Korean PD patients and found that DRD3 Ser9Gly polymorphism was associated with diphasic dyskinesia [21]. 5
ACCEPTED MANUSCRIPT Our data showed that DRD2 CAn-STR and DRD3 Ser9Gly were not significantly different in the distribution of genotypes and alleles among PD patients and healthy controls, indicating that neither DRD2 CAn-STR nor DRD3 Ser9Gly was associated with increased susceptibility to PD. A meta-analysis containing 4,279 cases and 5,661 controls indicated that rs6280 (DRD3 Ser9Gly) was not
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associated with PD [22], which is consistent with our results. Though previous study had shown that the
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15 allele distribution in DRD2 CAn-STR was higher in controls than PD patients (p=0.04) [11], we found no association between DRD2 CAn-STR and PD susceptibility herein. Multiple environmental
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and genetic factors are thought to play important roles in the etiology and pathogenesis of PD. Single gene polymorphism may not be sufficient to induce the disease. Moreover, gene polymorphisms exist not only in ethnicity but also in regions. Multi-center, large-scale trials are needed to confirm our
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findings.
We did not find an association between the DRD2 CAn-STR polymorphism and doses of DAs, indicating that the DRD2 CAn-STR genotypes might not be associated with the inter-individual
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responses to DAs, neither pramipexole nor piribedil. The DRD2 gene is mainly expressed in the dorsal regions of the striatum and the globus pallidus and is involved in locomotion as well as drug abuse [15]. The DRD2 CAn-STR polymorphism is located in the non-coding region of chromosome 11q22-q23 and
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its function has not been completely elucidated. Previous studies indicated that the absence of a 15× DRD2 CA repeat allele was significantly correlated with a decreased discontinuation of non-ergoline
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treatment (ropinirole and pramipexole). The aim of our study is to find an association between DRD2 CAn-STR polymorphism and daily doses of DAs, which has not been explored to date. Since a DRD2
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CAn-STR is located in a non-coding region, and might not be a functional polymorphism, this might partially explain why there is no association with DAs dosage. More studies are needed to understand the function of CAn-STR and its relationship with the response to DAs. Herein, when the PD patients were grouped by the DRD3 Ser9Gly polymorphism, we found that the
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Gly/Gly subgroup took higher doses of DAs compared with the Ser/Ser and Ser/Gly subgroups. Furthermore, we found that PD patients with the Gly/Gly genotype took more pramipexole than patients with the Ser/Ser and Ser/Gly genotypes. However, the dosages of piribedil in the three genotypes were not significantly different. DRD3 receptors are most abundant in the islands of Calleja of the ventral striatum/nucleus accumbens region as well as the dentate gyrus and striate cortex and may play important roles in limbic-related functions, such as emotion and cognition, and non-limbic functions, such as processing of motor and sensory information [23]. The Ser9Gly polymorphism located in the DRD3 gene is a functional polymorphic site, and the substitution of glycine for serine may lead to the structural and functional changes of the gene, with increased affinity for dopamine and that the function of the D3 receptor was attenuated [14，15], thus potentially contributing to the variable responses to DAs. This may partly account for why the Gly/Gly groups use more DAs than Ser/Ser and Ser/Gly groups. Pramipexole and piribedil are both non-ergot-derived DAs and partially activate DRD2 and DRD3. In PD patients, pramipexole exerts therapeutic action on motor deficits mainly via the post-synapses of D2 and D3 receptors by the indirect and direct pathways. In the pathogenesis of PD, the direct pathway is 6
ACCEPTED MANUSCRIPT restrained, and the inhibiting effect of the indirect pathway is alleviated. Therefore, pramipexole helps to correct the increased inhibitory impact of the direct pathway on motor activity by enhancing the direct transmission (through D3 receptors) and reducing the indirect transmission (through D2 receptors) [24]. Piribedil is a mixed D2/D3 receptor partial agonist, and the binding affinity for DRD2 and DRD3 is
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similar. However, pramipexole shows a seven-to ten-fold higher affinity to D3 than to D2 receptors [25].
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Pramipexole also has a higher affinity for DRD3 than piribedil [26]. This may explain why the DRD3 Ser9Gly genotypes are related to dosage of pramipexole rather than piribedil. Besides, we found that age
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of onset was negatively associated with DAs, whereas, in the pramipexole or piribedil subgroups, no association was found, might due to the reduced number of patients.
While there were no significant differences in clinical features or therapeutic effects among different
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DRD3 Ser9Gly genotypes; the Gly/Gly group took more DAs and pramipexole than Ser/Gly and Ser/Ser groups, indicating that patients carrying the Gly/Gly genotype might not respond as well as the other two subgroups to DAs if using the same dosage. Our finding was partly consistent with another group in
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China who found that the Ser/Ser group responded better to pramipexole than the group containing the Gly allele [15]; however, considerable differences existed. Firstly, the daily dose of pramipexole used was only 0.375 mg/day, which was insufficient to achieve desirable effect in the previous study. Patients
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in our study used much higher dose of pramipexole (approximately 1.1±0.4 mg/day), which was much
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closer to the clinical practice. Secondly, duration for patients using pramipexle was much longer in our
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research (3.9±1.6 years vs. 2 months), which was more compelling in the long-term treatment. Furthermore, they investigated different drug responses among the DRD3 Ser9Gly genotypes using same dose of pramipexole, whereas we explored various doses of DAs in different genotypes. They found the Gly carriers responded less to pramipexole, while only patients with Gly/Gly homozygote
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took more pramipexole in our study. Whether the heterozygous mutation (Ser/Gly genotype) of DRD3 Ser9Gly may influence the response to pramipexole needs to be further confirmed in other countries and races.
5. Conclusions
In this cross-sectional study, we investigated the association of DRD2 CAn-STR as well as DRD3 Ser9Gly and daily doses of DAs, especially pramipexole. Our data showed that patients carrying the DRD3 Ser9Gly Gly/Gly genotype took more pramipexole than patients with the Ser/Ser or Ser/Gly genotypes, indicating that to obtain the best therapeutic results, higher doses of pramipexole should be used in patients with the Gly/Gly genotype. Furthermore, these results provide some insights into the pharmacogenomics of PD treatment and may help us better understand individual variability in response to DAs. Large-scale, multi-center, and even prospective studies are needed to further evaluate the impact of genetic variation on the different responses of individuals to DAs. Conflict of interest The authors declare no conflict of interest. Acknowledgments 7
ACCEPTED MANUSCRIPT This work was supported by the National Natural Science Foundation of China (Grant No. 81071023) and the Natural Science Foundation of Shanghai (Grant No. 14ZR1425700). References
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[1] de Lau LM, Breteler MM,Epidemiology of Parkinson's disease, Lancet Neurol. 5 (2006) 525-535.
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http://dx.doi.org/ 10.1016/S1474-4422(06)70471-9
[2] Sethi K, Levodopa unresponsive symptoms in Parkinson disease, Mov Disord. 23 (2008) S521-533.
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http://dx.doi.org/10.1002/mds.22049
[3] Fahn S, Oakes D, Shoulson I, Kieburtz K, Rudolph A, Lang A, Olanow CW, Tanner C, Marek K, Parkinson Study Group, Levodopa and the progression of Parkinson's disease, N Engl J Med. 351 (2004) 2498-2508. http://dx.doi.org/10.1056/NEJMoa033447
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[4] Andersohn F, Garbe E, Cardiac and noncardiac fibrotic reactions caused by ergot-and nonergot-derived dopamine agonists, Mov Disord. 24 (2009) 129-133.
MA
http://dx.doi.org/10.1002/mds.22385
[5] Trifirò G, Mokhles MM, Dieleman JP, van Soest EM, Verhamme K, Mazzaglia G, Herings R, de Luise C, Ross D, Brusselle G, Colao A, Haverkamp W, Schade R, van Camp G, Zanettini R, Sturkenboom MC, Risk of cardiac valve regurgitation with dopamine agonist use in Parkinson's
D
disease and hyperprolactinaemia: a multi-country, nested case-control study, Drug Saf. 35 (2012)
TE
159-171. http://dx.doi.org/10.2165/11594940-000000000-00000 [6] Wang V, Chao TH, Hsieh CC, Lin CC, Kao CH, Cancer risks among the users of ergot-derived
CE P
dopamine agonists for Parkinson's disease, a nationwide population-based survey, Parkinsonism Relat Disord. 21 (2015) 18-22. http://dx.doi.org/10.1016/j.parkreldis.2014.10.015 [7] Schumacher-Schuh AF, Rieder CR, Hutz MH, Parkinson's disease pharmacogenomics:new findings and perspectives, Pharmacogenomics. 15 (2014) 1253-1271. http://dx.doi.org/10.2217/pgs.14.93
AC
[8] Emilien G, Maloteaux JM, Geurts M, Hoogenberg K, Cragg S, Dopamine receptors--physiological understanding to therapeutic intervention potential, Pharmacol Ther. 84 (1999) 133-156. http://dx.doi.org/10.1016/S0163-7258 (99)00029-7 [9] Jenner P, Pharmacology of dopamine agonists in the treatment of Parkinson’s disease, Neurology. 58 (2002) S1-S8. http://dx.doi.org/10.1212/WNL.58.suppl_1.S1 [10] Wang J, Liu ZL, Chen B, Association study of dopamine D2, D3 receptor gene polymorphisms with motor fluctuations in PD, Neurology. 56 (2001) 1757-1759. http://dx.doi.org/10.1212/WNL.56.12.1757 [11] Oliveri RL, Annesi G, Zappia M, Civitelli D, Montesanti R, Branca D, Nicoletti G, Spadafora P, Pasqua AA, Cittadella R, Andreoli V, Gambardella A, Aguglia U, Quattrone A, Dopamine D2 receptor gene polymorphism and the risk of levodopa-induced dyskinesias in PD, Neurology. 53 (1999) 1425-1430. http://dx.doi.org/10.1212/WNL.53.7.1425 [12] Zappia M, Annesi G, Nicoletti G, Arabia G, Annesi F, Messina D, Pugliese P, Spadafora P, Tarantino P, Carrideo S, Civitelli D, De Marco EV, Cirò-Candiano IC, Gambardella A, Quattrone A, Sex differences in clinical and genetic determinants of levodopa peak-dose dyskinesias in Parkinson 8
ACCEPTED MANUSCRIPT disease: an exploratory study, Arch Neurol. 62 (2005) 601-605. http://dx.doi.org/10.1001/archneur.62.4.601 [13] Arbouw ME, Movig KL, Egberts TC, Poels PJ, van Vugt JP, Wessels JA, van der Straaten RJ, Neef C, Guchelaar HJ, Clinical and pharmacogenetic determinants for the discontinuation of
T
non-ergoline dopamine agonists in Parkinson's disease, Eur J Clin Pharmacol 65 (2009) 1245-1251.
IP
http://dx.doi.org/ 10.1007/s00228-009-0708-6
[14] Jeanneteau F, Funalot B, Jankovic J, Deng H, Lagarde JP, Lucotte G, Sokoloff P, A functional
SC R
variant of the dopamine D3 receptor is associated with risk and age-at-onset of essential tremor, Proc Natl Acad Sci U S A. 103 (2006) 10753-10758. http://dx.doi.org/10.1073/pnas.0508189103 [15] Liu YZ, Tang BS, Yan XX, Liu J, Ouyang DS, Nie LN, Fan L, Li Z, Ji W, Hu DL, Wang D, Zhou
NU
HH, Association of the DRD2 and DRD3 polymorphisms with response to pramipexole in Parkinson’s disease patients, Eur J Clin Pharmacol, 65 (2009) 679-683. http://dx.doi.org/10.1007/s00228-009-0658-z
MA
[16] Hughes AJ, Daniel SE, Kilford L, Lees AJ, Accuracy of clinical diagnosis of idiopathic Parkinson's disease: a clinico-pathological study of 100 cases, J Neurol Neurosurg Psychiatry. 55 (1992) 181-184. http://dx.doi.org/ 10.1136/jnnp.55.3.181
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[17]Tomlinson CL, Stowe R, Patel S, Rick C, Gray R, Clarke CE, Systematic review of levodopa dose
10.1002/mds.23429
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equivalency reporting in Parkinson's disease, Mov Disord. 25 (2010) 2649-2653. http://dx.doi.org/
[18] Hacker ML, Tonascia J, Turchan M, Currie A, Heusinkveld L, Konrad PE, Davis TL, Neimat JS,
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Phibbs FT, Hedera P, Wang L, Shi Y, Shade DM, Sternberg AL, Drye LT, Charles D, Deep brain stimulation may reduce the relative risk of clinically important worsening in early stage Parkinson's disease, Parkinsonism Relat Disord. 21 (2015) 1177-1183. http://dx.doi.org/10.1016/j.parkreldis.2015.08.008
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[19] Bartus RT, Gene therapy for Parkinson's disease: a decade of progress supported by posthumous contributions from volunteer subjects, Neural Regen Res. 10 (2015) 1586-1588. http://dx.doi.org/10.4103/1673-5374.167783 [20] Bialecka M, Kurzawski M, Klodowska-Duda G, Opala G, Tan EK, Drozdzik M, The association of functional catechol-O-methyltransferase haplotypes with risk of Parkinson’s disease, levodopa treatment response, and complications, Pharmacogenet Genomics. 18 (2008) 815-821. http://dx.doi.org/10.1097/FPC.0b013e328306c2f2 [21] Lee JY, Cho J, Lee EK, Park SS, Jeon BS, Differential genetic susceptibility in diphasic and peak-dose dyskinesias in Parkinson's disease, Mov Disord. 26 (2011) 73-79. http://dx.doi.org/10.1002/mds.23400 [22] Dai D, Wang Y, Wang L, Li J, Ma Q, Tao J, Zhou X, Zhou H, Jiang Y, Pan G, Xu L, Ru P, Lin D4, Pan J, Xu L, Ye M, Duan S, Polymorphisms of and genes and Parkinson's disease: A meta-analysis, Biomed Rep. 2 (2014) 275-281. http://dx.doi.org/10.3892/br.2014.220 [23] Suzuki M, Hurd YL, Sokoloff P, Schwartz JC, Sedvall G, D3 dopamine receptor mRNA is widely expressed in the human brain, Brain Res. 779 (1998) 58-74. 9
ACCEPTED MANUSCRIPT http://dx.doi.org/10.1016/S0006-8993(97)01078-0 [24] Mierau J, Schneider FJ, Ensinger HA, Chio CL, Lajiness ME, Huff RM, Pramipexole binding and activation of cloned and expressed dopamine D2, D3 and D4 receptors, Eur J Pharmacol. 290 (1995) 29-36. http://dx.doi.org/ 10.1016/0922-4106(95)90013-6
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[25] Piercey MF, Pharmacology of pramipexole, a dopamine D3-preferring agonist useful in treating
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Parkinson's disease, Clin Neuropharmacol. 21 (1998) 141-151. [Pubmed: 9617505] [26] Deleu D, Northway MG, Hanssens Y, Clinical pharmacokinetic and pharmacodynamic properties
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http://dx.doi.org/10.2165/00003088-200241040-00003
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of drugs used in the treatment of Parkinson's disease, Clin Pharmacokinet. 41 (2002) 261-309.
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ACCEPTED MANUSCRIPT Table 1 Genotype and allele distributions of DRD2 CAn-STR and DRD3 Ser9Gly in PD patients and controls Genotypes
p
Alleles
p
value value 1314+
1314-
13
14
15
41(24.4%
126(69.2
=182)
%)
Ser9Gly PD
(n=182)
56(3
9(2
0.8%)
Ser/S
Ser/
.5%)
Gly/
er
Gly
Gly
77(45.8%)
66
25
(39.3%)
(14.9%)
86
80
16
(47.3%)
(44.0%)
(8.7%)
(n=168) Controls
18(5.4%) 154(45.8%) 19(5.7%) 143(42.6%) 2(0.6%)
155(4
2.6%)
23(
6.3%)
174(47.8 %)
0.196
0.239
3( 0.8%)
Ser
Gly
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Controls(n
DRD3
0.184
)
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127(75.6%)
220(65.5%)
116(34.5%)
252 (69.2%)
0.290
112 (30.8%)
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PD ( n=168)
17
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CAn-STR
16
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DRD2
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1314+: subjects carrying at least 1 of allele 13 or 14 in DRD2 CAn-STR; 1314-: subjects carrying neither 13 nor 14 allele in CAn-STR Data is expressed as the number of subjects in each category (frequency). All p values refer to the chi-square (2) test.
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Table 2 Demographic and clinical features of patients using DAs (n=102)
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DRD2 CAn-STR 1314+(7
Age
Male
p
Ser/Ser
Ser/Gly
Gly/Gly
9)
)
value a
(45)
(39)
(18)
65.1±9.4
65.3±8.1
0.946
65.9±8.5
65.4±10.0
62.8±8.2
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Gender
1314-(23
DRD3 Ser9Gly p value b
0.465
0.093
0.582
53(67.1%)
11 (47.8%)
30(66.7%)
22 (56.4%)
12(66.7%)
26(32.9%)
12(52.2%)
15(33.3%)
17(43.6%)
6(33.3%)
59.3±9.2
58.3±8.1
0.644
59.2±8.3
60.0±9.7
57.2±9.2
0.604
Duration of disease
6.0±4.6
6.9±4.1
0.391
6.7±4.6
5.7±4.0
6.2±5.1
0.581
H&Y stage
2.1±0.8
2.0±0.6
0.670
2.2±0.7
2.1±0.9
2.1±0.5
0.646
LED (mg/d)
419.8±247.3
386.3±231.3
0.563
395.0±232.8
406.4±255.0
468.1±247.2
0.554
Dosage of DAs (=LED, mg/d)
112.1±43.3
106.5±27.1
0.458
100.4±36.2
109.0±33.2
141.0±49.6
0.001**
UPDRS-Total
37.5±15.5
37.3±13.3
0.968
40.1±15.1
33.7±14.3
38.6±14.8
0.157
NMS
6.9±4.0
6.6±4.7
0.778
7.3±4.1
6.4±4.0
6.6±4.6
0.648
HAMA
6.1±5.2
7.0±6.4
0.479
6.1±4.6
6.2±6.3
7.0±5.9
0.831
HAMD
5.1±5.6
5.1±5.2
0.987
6.0±6.2
4.0±4.0
5.0±6.1
0.299
MMSE
27.8±2.6
27.4±3.0
0.552
Female Age of onset
27.4±2.7
27.8±2.8
28.4±1.4
0.40 1
Values are expressed as the mean ± SD except for the gender category. : p value between patients with different DRD2 CAn-STR genotypes. The 2 test or Fisher’s exact test for discrete variables and the unpaired t-test for a
11
ACCEPTED MANUSCRIPT
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continuous variables were used. b : p value among patients with different DRD3 Ser9Gly genotypes. The 2 test or Fisher’s exact test for discrete variables and the one-way ANOVA for continuous variables were used. ** : p0.01
Table 3
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Demographic and clinical features of patients using Pramipexole (n=62) DRD2 CAn-STR 1314-(13)
66.0±8.2
66.3±8.0
D
Age
1314+(49)
Gender
Female
21(42.9%)
Age of onset
60.1±8.5
Duration of disease
0.891
DRD3 Ser9Gly Ser/Ser (28)
Ser/Gly(23)
Gly/Gly(11)
p value b
66.1±7.1
66.9±8.7
64.1±9.4
0.643
0.091
0.899
4(30.8%)
15(51.6%)
11(47.8%)
6(54.5%)
9(69.2%)
13(48.4%)
12(52.2%)
5(45.5%)
0.972
59.9±7.2
61.5±8.6
57.7±10.2
0.479
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28(57.1%)
60.2±7.8
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Male
p value
a
6.0±4.6
6.2±4.3
0.913
6.1±4.2
5.7±4.3
6.4±5.7
0.916
2.2±0.8
2.0±0.7
0.508
2.3±0.7
2.1±0.9
2.1±0.6
0.682
422.5±249.4
329.6±225.1
0.228
396.5±258.3
377.6±215.3
472.7±281.4
0.345
106.8±47.8
109.6±32.7
0.840
93.5±36.1
102.2±36.1
153.4±54.2
<0.001***
38.8±15.9
35.7±12.6
0.525
38.7±14.7
35.8±15.1
41.0±17.4
0.641
7.0±4.3
6.2±3.9
0.561
7.0±3.9
6.4±4.2
7.4±4.9
0.803
6.2±5.9
6.8±6.8
0.727
6.0±4.8
6.5±7.1
6.7±7.1
0.930
HAMD
5.3±6.2
3.5±3.9
0.356
5.8±6.6
3.8±4.2
4.8±6.6
0.483
MMSE
28.0±2.4
27.6±3.0
0.651
27.6±3.1
28.2±2.1
28.0±1.7
0.736
H&Y stage LED (mg/d)
UPDRS-Total NMS HAMA
AC
Dosage of DAs (=LED, mg/d)
Values are expressed as the mean ± SD except for the gender category. a: p value between patients with different DRD2 CAn-STR genotypes. The 2 test or Fisher’s exact test for discrete variables and the unpaired t-test for continuous variables were used. b: p value among patients with different DRD3Ser9Gly genotypes. The 2 test or Fisher’s exact test for discrete variables and the one-way ANOVA for continuous variables were used. : p 0.001
***
Table 4 Demographic and clinical features of patients using Piribedil (n=40)
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ACCEPTED MANUSCRIPT DRD2 CAn-STR 1314+(
Age
1314-(1
DRD3 Ser9Gly p
30)
0)
value
63.7±11.0
63.9±8.4
0.965
Ser/Gl
Gly/Gl
p
(17)
y(16)
y(7)
value b
65.5±10.6
63.3±11.6
60.7±6.0
0.576 0.345
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0.361
Male
25(83.3%)
7(70.0%)
15(88.2%)
11(68.8%)
6(85.7%)
Female
5(16.7%)
3(30.0%)
2(11.8%)
5(31.2%)
1(14.3%)
Age of onset
56.2±14.7
56.0±8.4
0.968
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Gender
Ser/Ser
a
57.7±10.9
48.3±22.4
0.230
Duration of disease
6.0±4.7
7.9±3.7
0.254
H&Y stage
2.0±0.7
2.1±0.6
0.795
LED (mg/d)
415.5±247.9
460.0±229.2
0.620
Dosage of DAs
120.8±33.5
102.5±18.4
0.039*
UPDRS-Total
35.3±14.7
39.5±14.5
0.447
NMS
6.7±3.5
7.1±5.8
HAMA
5.9±3.6
7.2±6.1
HAMD
4.7±4.5
7.0±6.0
MMSE
27.5±2.9
27.2±3.0
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5.6±3.8
5.8±4.2
0.412
2.1±0.6
1.9±0.9
2.0±0.3
0.760
392.6±191.0
447.8±306.1
460.7±202.5
0.751
111.8±34.4
118.8±26.6
121.4±36.6
0.735
47.1±17.3
30.1±14.5
41.3±11.5
0.071
0.801
7.8±4.4
6.4±4.0
5.4±4.2
0.440
0.537
6.3±4.4
5.6±4.8
7.4±3.6
0.688
0.226
6.2±5.7
4.4±3.7
5.3±5.7
0.638
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7.6±5.2
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(=LED, mg/d)
57.9±9.9
27.0±2.
27.2±3.
29.0±1.
5
7
4
0. 290
D
0.758
Values are expressed as the mean ± SD except for the gender category. : p value between patients with different DRD2 CAn-STR genotypes. The 2 test or Fisher’s exact test for discrete variables and
the unpaired t-test for continuous variables were used.
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a
one-way ANOVA for
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: p value among patients with different DRD3Ser9Gly genotypes. The 2 test or Fisher’s exact test for discrete variables and the
b
continuous variables were used. : p 0.05
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*
Table 5 Single-factor linear regression of doses of DAs with demographic and clinical features of patients PD patients taking DAs (n=102) p value Gen
95% CI
PD patients taking pramipexole (n=62) p value
95% CI
PD patients taking piribedil (n=40) p value
95% CI
0.592
-20.81-11.93
0.953
-22.30-23.66
0.494
-33.80-16.62
Age
0.059
-1.70-0.03
0.209
-2.31-0.52
0.164
-1.65-0.29
Age
0.016*
-1.95--0.20
0.127
-2.46-0.31
0.057
-1.99--0.93
0.198
-0.62-2.96
0.503
-1.73-3.49
0.184
-0.75-3.77
0.659
-13.47-8.56
0.507
-19.38-9.67
0.648
-11.50-18.26
der
a
a
a
of onset
Dur ation of disease a H& Y stage
a
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ACCEPTED MANUSCRIPT UPDRS-Total a
0.681
-0.44-0.67
0.675
-0.62-0.10
0.865
-0.68-0.80
NM
0.721
-2.36-1.64
0.669
-3.51-2.27
0.962
-2.52-2.64
0.969
-1.51-1.57
0.952
-2.05-1.93
0.788
-2.18-2.86
0.265
-2.39-0.67
a
HA
a
MD
-2.58-1.61
0.99
-4.83-4.88
4
MM SE
0.64
a
0.8 83
DR
-3.41-2.9 4
0.560
3
-24.51-13.35
0.840
D2 CAn-STR
DR D3
01
-28.20--
<0.0
8.02
01
a
***
-39.92--12.14
0.093
-3.92-0.32
0.868
-4.12-3.50
0.110
-40.98-4.3 1
0.445
-18.97-8.5 0
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Ser9Gly a
0.0 **
-2.35-31.06
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a
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HA
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a
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S
：independent variable
Dependent variable: doses of DAs, pramipexole or piribedil 95% CI: 95% confidence interval
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: p0.05; **: p<0.01; ***: p<0.001
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*
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ACCEPTED MANUSCRIPT Highlights •
We investigated the association between DRD2 CAn-STR as well as DRD3 Ser9Gly polymorphisms and doses of DAs, especially pramipexole
•
We found that PD patients carrying DRD3 Ser9Gly Gly/Gly genotype took more pramipexole Our study indicated that to obtain better therapeutic results, higher doses of pramipexole should be used in patients with Gly/Gly genotype
Our results provide some insights into the pharmacogenomics of PD treatment and may help
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to better understand individual variability in response to DAs
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•
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•
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than patients with Ser/Ser and Ser/Gly genotypes
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S0022-510X(16)30484-1 doi: 10.1016/j.jns.2016.08.005 JNS 14735
To appear in:
Journal of the Neurological Sciences
Received date: Revised date: Accepted date:
22 February 2016 6 July 2016 1 August 2016
Please cite this article as: Shaoqing Xu, Jiujiang Liu, Xiaodong Yang, Yiwei Qian, Qin Xiao, Association of the DRD2 CAn -STR and DRD3 Ser9Gly polymorphisms with Parkinson’s disease and response to dopamine agonists, Journal of the Neurological Sciences (2016), doi: 10.1016/j.jns.2016.08.005
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ACCEPTED MANUSCRIPT Association of the DRD2 CAn-STR and DRD3 Ser9Gly polymorphisms with Parkinson's disease and response to dopamine
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agonists
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Shaoqing Xu, Jiujiang Liu, Xiaodong Yang, Yiwei Qian, Qin Xiao*
University School of Medicine, Shanghai 200025, China
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Department of Neurology & Institute of Neurology, Ruijin Hospital affiliated to Shanghai JiaoTong
*Correspondence to: Qin Xiao, email: [email protected] Abstract
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Dopamine agonists (DAs) play important roles in the treatment of Parkinson's disease (PD). Currently, it is thought that genetic variations in the genes encoding dopamine receptors (DR) are important factors in determining inter-individual variability in drug responses. To investigate the association
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between Dopamine receptor D type 2 (DRD2) dinucleotide short tandem repeat (CAn-STR) and Dopamine receptor D type 3 (DRD3) Ser9Gly polymorphisms and the risk of PD, as well as the possible reasons for PD patients using different doses of DAs, we recruited 168 idiopathic PD patients
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and 182 controls. There were no significant differences in DRD2 CAn-STR and DRD3 Ser9Gly
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genotypes (p=0.184, p=0.196) or in allele frequencies (p=0.239, p=0.290) between PD patients and controls. There was no association between DRD2 CAn-STR polymorphism and doses of DAs. Among three different DRD3 Ser9Gly genotypes (Ser/Ser, Ser/Gly, Gly/Gly), patients carrying Gly/Gly
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genotype used higher doses of DAs than patients with Ser/Gly and Ser/Ser genotypes (p=0.001). In pramipexole subgroup, the Gly/Gly group took more pramipexole than the other genotype groups (p<0.001), whereas the doses of piribedil were not significantly different among three genotypes
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(p=0.735). Our results suggest that genotype in DRD3 Ser9Gly was the main factor determining different doses of DAs and PD patients carrying Gly/Gly genotype require higher doses of pramipexole for effective treatment. This study may provide insights into understanding possible reasons for different responses to DAs in Chinese PD patients. Key words: DRD2 CAn-STR, DRD3 Ser9Gly, Parkinson's disease, dopamine agonists 1 Introduction Parkinson's disease (PD) is a common neurodegenerative disorder, affecting 1%-3% of people above 65 years of age and with increased prevalence in advancing age [1]. The pathological changes of PD manifest as a progressive loss of dopaminergic neurons in the substantia nigra pars compacta (SNc) and an abnormal aggregation of Lewy bodies. It is commonly recognized that the interaction between heredity and environment contributes to the pathogenesis of the disease. In clinical practice, dopamine replacement therapy is still the principle way to treat PD. However, the dopamine precursor, L-3,4-dihydroxyphenylalanine (L-dopa), is mainly effective in the restitution of most components of motor dysfunction but poorly remedies postural instability and freezing of gait or co-morbid symptoms, such as depression, cognitive impairment, perturbed sleep and sensory dysfunction [2]. Further, 1
ACCEPTED MANUSCRIPT long-term use of L-dopa often leads to motor complications such as wearing-off effects and dyskinesias [3]. As a result, dopamine agonists (DAs), which have a longer half-life compared with L-dopa, are increasingly employed either in conjunction with L-dopa or as monotherapy. Currently, the most investigated DAs are mainly divided into two categories: ergot-derived and
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non-ergot-derived DAs. Nevertheless, there is growing evidence that the ergot-derived dopamine
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agonists, for instance, bromocriptine, cabergoline and pergolide, can give rise to cardiac valvular regurgitation, pulmonary changes and even liver cancer [4-6], and as a result, they are being gradually
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phased out. The non-ergot-derived DAs, pramipexole and piribedil, are two key DAs prescribed in China, and pramipexole is more widely used because of its better efficacy and fewer side effects. Inter-individual differences in response to dopamine agonist treatment remain a critical problem in the
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management of PD patients, and it is thought that genetic variations in the genes encoding the dopamine receptors (DRs) may be causal of the variability in drug responses. The natural targets for DAs consist of five DR subtypes, of which DRD2, DRD3, and DRD4 are composed of D2-like
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receptors, while DRD1 and DRD5 consist of D1-like receptors [7]. When activated, the D1-like receptor subtypes stimulate adenylate cyclase (AC) activity; whereas the D2-like receptor subtypes generally inhibit adenylate cyclase activity [8]. Nearly all of the dopamine agonists mainly act through
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D2-like receptors, rather than the D1-like receptors [9].
Among all DR genes, DRD2 (chromosome 11q22-q23) and DRD3 (chromosome 3q13) have been
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the most investigated [7]. Several polymorphisms in the DRD2 gene have been identified to be associated with drug efficacy in PD. Wang and colleagues found that the DRD2 Taq1A (rs1800497)
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A1/A1 genotype was associated with an increased risk of motor fluctuation in PD [10]. PD patients carrying 13 and 14 repeat alleles of DRD2 CAn-STR was associated with less occurrences of L-dopa induced dyskinesia [11]. Interestingly, another group reported that this protective effect of DRD2 CAn-STR 13 and 14 repeat alleles on dyskinesia risk was restricted to men [12]. Besides, the absence
AC
of the DRD2 CAn-STR 15 repeat allele was found to be associated with decreased rates of discontinuation of non-ergoline DAs [13]. The most widely investigated polymorphic site in DRD3 was DRD3 Ser9Gly, which had three different genotypes (Ser/Ser, Ser/Gly and Gly/Gly). A serine to glycine substitution in the N-terminal extracellular part of the receptor was reported to be associated with altered dopamine binding affinity [14]. A prospective study including 30 PD patients conducted in China found that the response rates were significantly higher in the Ser/Ser group (60%) than in the group containing the Gly allele (13%) after treatment with pramipexole for 2 months [15]. However, the dosage of pramipexole used was only 0.375 mg/day, which was much smaller than the normally prescribed doses. In our study, we investigated the association between the DRD2 CAn-STR and DRD3 Ser9Gly polymorphisms and the risk of PD. We also examined possible reasons for the variable responses to DAs among PD patients, especially as it relates to the variation in daily DAs dosages between individuals. 2. Materials and methods 2.1. Subjects 2
ACCEPTED MANUSCRIPT A total of 168 idiopathic PD patients and 182 age and gender matched controls were included in the study. The PD patients were recruited from the Movement Disorder Clinic at the Department of Neurology of Ruijin Hospital between December 2013 and April 2016. Inclusion criteria were (1) clinical diagnosis of PD according to the United Kingdom Parkinson's Disease Society Brain Bank
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criteria [16], (2) no history of stroke or significant neurological disorders, and (3) no family history of
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PD extended to the first-degree relatives. The study was approved by the ethics committee of Ruijin Hospital. All participants signed an informed consent.
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All PD patients were examined by at least two movement disorder specialists, and the basic clinical features were recorded including the age of onset, duration of the disease, the Hoehn and Yahr stage (H&Y stage), levodopa equivalent dose (LED), and doses of DAs, including pramipexole and piribedil.
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Additional factors recorded included the Unified Parkinson's Disease Rating Scale (UPDRS) scores and Non-Motor Symptoms questionnaire for Parkinson's disease (NMS). Patients were also evaluated by the Hamilton Anxiety Scale (HAMA), Hamilton Depression Scale (HAMD), and Mini-Mental State
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Examination (MMSE). The LEDs of patients were calculated according to a previous study [17], where 1 mg of pramipexole is equivalent to 100 mg of piribedil and 100 mg L-dopa. 2.2. Genotyping for the DRD2 CAn-STR and DRD3 Ser9Gly polymorphisms
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Genomic DNA was isolated from peripheral blood leucocytes with phenol–chloroform extraction followed by ethanol precipitation. Analysis of CAn-STR polymorphism in the DRD2 gene was by
PCR
amplification
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performed
using
the
following
set
of
primers:
F:
5′-
TGAAGACTCGCCATGAAA-3′ and R: 5′-GGTAGAAGCCAAAGGTGC-3′. PCR products were then
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fluorescently labeled at the 5'-end by 6′-FAM dideoxynucleotides (GENEWIZ, China) and analyzed using the ABI Prism 3730 DNA Sequencer. Allele size was established using Genemapper V4.0 software. Sequence analysis was used to confirm differentiation of alleles by size. PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism) method was performed to test the
AC
genotype of Ser9Gly polymorphism. A 174-bp product was amplified with an initial 5-min denaturation at 94˚C, followed by 35 cycles of 94 ˚C for 30 s, 56˚C for 30 s, 72 ˚C for 30 s, and a final extension period at 72 ˚C for 5 min, using the upstream primer, 5′-CCAGGCCATTGCCGAAGA-3′ and the downstream primer, 5′-TGGCATCACGCACCTCCCT-3′. The PCR products were digested by MscI enzyme and specific fragments were generated. If it was the Ser allele, then a 174-bp product was detected, and 122-bp and 52-bp product were detected if it was the Gly allele. Genotyping was performed by individuals who were blinded to the clinical status of the subjects. 2.3. Statistical analysis Statistical analyses were performed with SPSS software (ver. 13.0 for Windows, Chicago, IL).The Hardy–Weinberg equilibrium chi-square test was used to assess the data’s goodness of fit. All analyses were 2-tailed, and the level of statistical significance was set at p<0.05. Categorical data were analyzed with the chi-square test or Fisher exact test to establish whether there was an association between PD and the genotypes. The clinical variables were compared between the groups by the 2-sample, unpaired t-test for DRD2 CAn-STR polymorphism continuous variables with approximate normal distribution, 3
ACCEPTED MANUSCRIPT and the chi-square test or Fisher's exact test for discrete variables. For DRD3 Ser9Gly genotype continuous variables, we performed the homogeneity test for variances and then one-way ANOVA. The multiple comparisons were performed by Bonfferoni test and either Pearson's chi-square test or Fisher’s exact test for discrete variables. After revealing the differential variables among genotypes, we carried
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out single-factor linear regression and multiple stepwise linear regression to assess the precise
3. Results
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3.1. Demographic features of PD patients and healthy controls
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association between the differential variables and genotypes with a criterion for entry of p<0.05.
A total of 168 idiopathic PD patients and 182 controls participated in this cross-sectional study, and there were no significant differences in the age (p=0.679) and gender (p=0.452) distributions between
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the patients with PD and the control groups (PD patients: age 66.0±8.9 years, 99males/69 females; controls: age 65.0±8.5 years, 100males/82 females).
3.2. Genotype and allele distributions of DRD2 CAn-STR and DRD3 Ser9Gly in PD patients and
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controls
The genotype and allele distributions of DRD2 CAn-STR and DRD3 Ser9Gly are shown in Table 1. All the observed genotype or allele frequencies did not differ from the expected frequencies according to
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the Hardy-Weinberg equilibrium. Previous studies showed that carrying the 13 and 14 repeat of DRD2 CAn-STR was associated with less risk of L-dopa induced dyskinesia, indicating that 13 and 14 carriers
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might provide some protection [11, 12]. Due to the low frequencies of some genotypes in the DRD2 CAn-STR, subjects were categorized as 13, 14+ or 13, 14− (subjects carrying at least 1 of allele 13 or 14
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in their genotype or carrying neither allele 13 nor 14, respectively) in our study according to Oliveri’s and Zappia’s categorization [11, 12]. The genotype or allele frequencies in the PD patients did not differ from the controls (p=0.184, p=0.239). In addition, there were no significant differences in the DRD3 Ser9Gly genotype distributions when comparing between PD patients and controls (p=0.196) nor in the
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allele distributions (p=0.290).
3.3. Demographic and clinical features of patients taking dopamine agonists There were 102 patients using dopamine agonists in our study. As shown in Table 2, for DRD2 CAn-STR polymorphism, there were no differences in any of the clinical features; however, for DRD3 Ser9Gly genotypes (Ser/Ser, Ser/Gly, Gly/Gly), no significant differences were found in clinical features such as age of onset, duration of the disease, H-Y stage, LED, or the scores of UPDRS total, NMS, HAMA, HAMD or MMSE, except for doses of DAs (p=0.001). Patients with the Gly/Gly genotype used higher doses of DAs compared with those from the Ser/Gly and Ser/Ser genotype groups (141.0±49.6 mg vs. 109.0±33.2 mg and 100.4±36.2 mg, respectively). Doses of DAs in the Gly/Gly subjects significantly differed from Ser/Ser (p=0.001) and Ser/Gly groups (p=0.011) by multiple comparison. After adjusting for gender, age, and other clinical features, by carrying out single-factor linear regression and multiple stepwise linear regression, the doses of DAs were significantly correlated with DRD3 Ser9Gly polymorphisms (p=0.001, p=0.001 respectively). Moreover, we further analyzed pramipexole (n=62) and piribedil (n=40) subgroups, and data were 4
ACCEPTED MANUSCRIPT shown in Table 3 and 4. We found that in DRD2 CAn-STR polymorphisms, there were not any significant differences in the pramipexole (p=0.840) subgroup, while in the piribedil group, the doses of DAs were obviously different between 1314+ and 1314- genotype subjects (120.8±33.5mg vs. 102.5±18.4mg, respevtively, p=0.039). However, single-factor linear regression showed that there was
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no significant association between DRD2 CAn-STR genotypes and daily doses of piribedil. For DRD3
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Ser9Gly polymorphisms, the daily doses of pramipexole in the Gly/Gly group were significantly higher
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than the Ser/Gly and Ser/Ser groups (153.4±54.2 mg vs. 102.2±36.1 mg and 93.5±36.1 mg, respectively, p<0.001), whereas in patients using piribedil, the dosages were not significantly different among the three genotypes (p=0.735). In the pramipexole subgroup, significant differences were seen between the
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Gly/Gly and Ser/Ser groups (p<0.001) as well as the Gly/Gly and Ser/Gly groups (p=0.003) by Benferroni test. Single-factor linear regression and multiple stepwise linear regression showed that after
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adjustment for other factors, a relationship still existed (p=<0.001 in single-factor linear regression, p=0.001 in multiple stepwise linear regression). Results of single-factor linear regression were shown in Table 5. Besides, age of onset was shown to be negatively associated with doses of DAs (B= -1.075,
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p=0.016 in in single-factor linear regression, B= -0.899, p=0.032 in multiple stepwise linear regression).
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4. Discussion
PD is the second most common neurodegenerative disorder, affecting more than 2 million people in China. Though there are several ways to treat PD, such as deep brain stimulation (DBS) [18] and gene
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therapy [19], drug therapy is still the primary approach. L-dopa can be prescribed for dopamine replacement therapy as it enters the brain via the blood brain barrier and is transformed to dopamine by dopa decarboxylase. The efficacy of L-dopa treatment is affected by dopamine metabolism enzymes,
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such as catechol-O-methyltransferase (COMT) and monoamine oxidase B (MAOB), as well as the activity of DRs and the dopamine transporter (DAT). The role of dopamine can also be imitated by DAs that directly act on the DRs in the brain. Pramipexole and piribedil are two new types of non-ergot-derived DAs and mainly act on D2-like dopamine receptors. The heritable variations of dopamine receptor genes could lead to structural and functional abnormalities, potentially resulting in a large inter-individual variability in response to DA drugs. Several studies conducted on PD patients have focused on the variable efficacy of L-dopa treatment as well as the risk of associated motor complications, which may have genetic origins. Bialecka et al. investigated the most common COMT gene haplotypes (formed by single nucleotide polymorphisms (SNPs): rs6269: A>G; rs4633C>T; rs4818: C>G; and rs4680: A>G) in 322 PD patients and 357 controls and reported that the doses of L-dopa administered to G_C_G_G (high activity) haplotype carriers were significantly higher than those administered to non-carriers during the fifth year of treatment [20]. Lee and colleagues observed patients with peak dose dyskinesia and diphasic dyskinesia in a longitudinal study in 503 Korean PD patients and found that DRD3 Ser9Gly polymorphism was associated with diphasic dyskinesia [21]. 5
ACCEPTED MANUSCRIPT Our data showed that DRD2 CAn-STR and DRD3 Ser9Gly were not significantly different in the distribution of genotypes and alleles among PD patients and healthy controls, indicating that neither DRD2 CAn-STR nor DRD3 Ser9Gly was associated with increased susceptibility to PD. A meta-analysis containing 4,279 cases and 5,661 controls indicated that rs6280 (DRD3 Ser9Gly) was not
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associated with PD [22], which is consistent with our results. Though previous study had shown that the
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15 allele distribution in DRD2 CAn-STR was higher in controls than PD patients (p=0.04) [11], we found no association between DRD2 CAn-STR and PD susceptibility herein. Multiple environmental
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and genetic factors are thought to play important roles in the etiology and pathogenesis of PD. Single gene polymorphism may not be sufficient to induce the disease. Moreover, gene polymorphisms exist not only in ethnicity but also in regions. Multi-center, large-scale trials are needed to confirm our
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findings.
We did not find an association between the DRD2 CAn-STR polymorphism and doses of DAs, indicating that the DRD2 CAn-STR genotypes might not be associated with the inter-individual
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responses to DAs, neither pramipexole nor piribedil. The DRD2 gene is mainly expressed in the dorsal regions of the striatum and the globus pallidus and is involved in locomotion as well as drug abuse [15]. The DRD2 CAn-STR polymorphism is located in the non-coding region of chromosome 11q22-q23 and
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its function has not been completely elucidated. Previous studies indicated that the absence of a 15× DRD2 CA repeat allele was significantly correlated with a decreased discontinuation of non-ergoline
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treatment (ropinirole and pramipexole). The aim of our study is to find an association between DRD2 CAn-STR polymorphism and daily doses of DAs, which has not been explored to date. Since a DRD2
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CAn-STR is located in a non-coding region, and might not be a functional polymorphism, this might partially explain why there is no association with DAs dosage. More studies are needed to understand the function of CAn-STR and its relationship with the response to DAs. Herein, when the PD patients were grouped by the DRD3 Ser9Gly polymorphism, we found that the
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Gly/Gly subgroup took higher doses of DAs compared with the Ser/Ser and Ser/Gly subgroups. Furthermore, we found that PD patients with the Gly/Gly genotype took more pramipexole than patients with the Ser/Ser and Ser/Gly genotypes. However, the dosages of piribedil in the three genotypes were not significantly different. DRD3 receptors are most abundant in the islands of Calleja of the ventral striatum/nucleus accumbens region as well as the dentate gyrus and striate cortex and may play important roles in limbic-related functions, such as emotion and cognition, and non-limbic functions, such as processing of motor and sensory information [23]. The Ser9Gly polymorphism located in the DRD3 gene is a functional polymorphic site, and the substitution of glycine for serine may lead to the structural and functional changes of the gene, with increased affinity for dopamine and that the function of the D3 receptor was attenuated [14，15], thus potentially contributing to the variable responses to DAs. This may partly account for why the Gly/Gly groups use more DAs than Ser/Ser and Ser/Gly groups. Pramipexole and piribedil are both non-ergot-derived DAs and partially activate DRD2 and DRD3. In PD patients, pramipexole exerts therapeutic action on motor deficits mainly via the post-synapses of D2 and D3 receptors by the indirect and direct pathways. In the pathogenesis of PD, the direct pathway is 6
ACCEPTED MANUSCRIPT restrained, and the inhibiting effect of the indirect pathway is alleviated. Therefore, pramipexole helps to correct the increased inhibitory impact of the direct pathway on motor activity by enhancing the direct transmission (through D3 receptors) and reducing the indirect transmission (through D2 receptors) [24]. Piribedil is a mixed D2/D3 receptor partial agonist, and the binding affinity for DRD2 and DRD3 is
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similar. However, pramipexole shows a seven-to ten-fold higher affinity to D3 than to D2 receptors [25].
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Pramipexole also has a higher affinity for DRD3 than piribedil [26]. This may explain why the DRD3 Ser9Gly genotypes are related to dosage of pramipexole rather than piribedil. Besides, we found that age
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of onset was negatively associated with DAs, whereas, in the pramipexole or piribedil subgroups, no association was found, might due to the reduced number of patients.
While there were no significant differences in clinical features or therapeutic effects among different
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DRD3 Ser9Gly genotypes; the Gly/Gly group took more DAs and pramipexole than Ser/Gly and Ser/Ser groups, indicating that patients carrying the Gly/Gly genotype might not respond as well as the other two subgroups to DAs if using the same dosage. Our finding was partly consistent with another group in
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China who found that the Ser/Ser group responded better to pramipexole than the group containing the Gly allele [15]; however, considerable differences existed. Firstly, the daily dose of pramipexole used was only 0.375 mg/day, which was insufficient to achieve desirable effect in the previous study. Patients
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in our study used much higher dose of pramipexole (approximately 1.1±0.4 mg/day), which was much
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closer to the clinical practice. Secondly, duration for patients using pramipexle was much longer in our
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research (3.9±1.6 years vs. 2 months), which was more compelling in the long-term treatment. Furthermore, they investigated different drug responses among the DRD3 Ser9Gly genotypes using same dose of pramipexole, whereas we explored various doses of DAs in different genotypes. They found the Gly carriers responded less to pramipexole, while only patients with Gly/Gly homozygote
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took more pramipexole in our study. Whether the heterozygous mutation (Ser/Gly genotype) of DRD3 Ser9Gly may influence the response to pramipexole needs to be further confirmed in other countries and races.
5. Conclusions
In this cross-sectional study, we investigated the association of DRD2 CAn-STR as well as DRD3 Ser9Gly and daily doses of DAs, especially pramipexole. Our data showed that patients carrying the DRD3 Ser9Gly Gly/Gly genotype took more pramipexole than patients with the Ser/Ser or Ser/Gly genotypes, indicating that to obtain the best therapeutic results, higher doses of pramipexole should be used in patients with the Gly/Gly genotype. Furthermore, these results provide some insights into the pharmacogenomics of PD treatment and may help us better understand individual variability in response to DAs. Large-scale, multi-center, and even prospective studies are needed to further evaluate the impact of genetic variation on the different responses of individuals to DAs. Conflict of interest The authors declare no conflict of interest. Acknowledgments 7
ACCEPTED MANUSCRIPT This work was supported by the National Natural Science Foundation of China (Grant No. 81071023) and the Natural Science Foundation of Shanghai (Grant No. 14ZR1425700). References
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[1] de Lau LM, Breteler MM,Epidemiology of Parkinson's disease, Lancet Neurol. 5 (2006) 525-535.
IP
http://dx.doi.org/ 10.1016/S1474-4422(06)70471-9
[2] Sethi K, Levodopa unresponsive symptoms in Parkinson disease, Mov Disord. 23 (2008) S521-533.
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http://dx.doi.org/10.1002/mds.22049
[3] Fahn S, Oakes D, Shoulson I, Kieburtz K, Rudolph A, Lang A, Olanow CW, Tanner C, Marek K, Parkinson Study Group, Levodopa and the progression of Parkinson's disease, N Engl J Med. 351 (2004) 2498-2508. http://dx.doi.org/10.1056/NEJMoa033447
NU
[4] Andersohn F, Garbe E, Cardiac and noncardiac fibrotic reactions caused by ergot-and nonergot-derived dopamine agonists, Mov Disord. 24 (2009) 129-133.
MA
http://dx.doi.org/10.1002/mds.22385
[5] Trifirò G, Mokhles MM, Dieleman JP, van Soest EM, Verhamme K, Mazzaglia G, Herings R, de Luise C, Ross D, Brusselle G, Colao A, Haverkamp W, Schade R, van Camp G, Zanettini R, Sturkenboom MC, Risk of cardiac valve regurgitation with dopamine agonist use in Parkinson's
D
disease and hyperprolactinaemia: a multi-country, nested case-control study, Drug Saf. 35 (2012)
TE
159-171. http://dx.doi.org/10.2165/11594940-000000000-00000 [6] Wang V, Chao TH, Hsieh CC, Lin CC, Kao CH, Cancer risks among the users of ergot-derived
CE P
dopamine agonists for Parkinson's disease, a nationwide population-based survey, Parkinsonism Relat Disord. 21 (2015) 18-22. http://dx.doi.org/10.1016/j.parkreldis.2014.10.015 [7] Schumacher-Schuh AF, Rieder CR, Hutz MH, Parkinson's disease pharmacogenomics:new findings and perspectives, Pharmacogenomics. 15 (2014) 1253-1271. http://dx.doi.org/10.2217/pgs.14.93
AC
[8] Emilien G, Maloteaux JM, Geurts M, Hoogenberg K, Cragg S, Dopamine receptors--physiological understanding to therapeutic intervention potential, Pharmacol Ther. 84 (1999) 133-156. http://dx.doi.org/10.1016/S0163-7258 (99)00029-7 [9] Jenner P, Pharmacology of dopamine agonists in the treatment of Parkinson’s disease, Neurology. 58 (2002) S1-S8. http://dx.doi.org/10.1212/WNL.58.suppl_1.S1 [10] Wang J, Liu ZL, Chen B, Association study of dopamine D2, D3 receptor gene polymorphisms with motor fluctuations in PD, Neurology. 56 (2001) 1757-1759. http://dx.doi.org/10.1212/WNL.56.12.1757 [11] Oliveri RL, Annesi G, Zappia M, Civitelli D, Montesanti R, Branca D, Nicoletti G, Spadafora P, Pasqua AA, Cittadella R, Andreoli V, Gambardella A, Aguglia U, Quattrone A, Dopamine D2 receptor gene polymorphism and the risk of levodopa-induced dyskinesias in PD, Neurology. 53 (1999) 1425-1430. http://dx.doi.org/10.1212/WNL.53.7.1425 [12] Zappia M, Annesi G, Nicoletti G, Arabia G, Annesi F, Messina D, Pugliese P, Spadafora P, Tarantino P, Carrideo S, Civitelli D, De Marco EV, Cirò-Candiano IC, Gambardella A, Quattrone A, Sex differences in clinical and genetic determinants of levodopa peak-dose dyskinesias in Parkinson 8
ACCEPTED MANUSCRIPT disease: an exploratory study, Arch Neurol. 62 (2005) 601-605. http://dx.doi.org/10.1001/archneur.62.4.601 [13] Arbouw ME, Movig KL, Egberts TC, Poels PJ, van Vugt JP, Wessels JA, van der Straaten RJ, Neef C, Guchelaar HJ, Clinical and pharmacogenetic determinants for the discontinuation of
T
non-ergoline dopamine agonists in Parkinson's disease, Eur J Clin Pharmacol 65 (2009) 1245-1251.
IP
http://dx.doi.org/ 10.1007/s00228-009-0708-6
[14] Jeanneteau F, Funalot B, Jankovic J, Deng H, Lagarde JP, Lucotte G, Sokoloff P, A functional
SC R
variant of the dopamine D3 receptor is associated with risk and age-at-onset of essential tremor, Proc Natl Acad Sci U S A. 103 (2006) 10753-10758. http://dx.doi.org/10.1073/pnas.0508189103 [15] Liu YZ, Tang BS, Yan XX, Liu J, Ouyang DS, Nie LN, Fan L, Li Z, Ji W, Hu DL, Wang D, Zhou
NU
HH, Association of the DRD2 and DRD3 polymorphisms with response to pramipexole in Parkinson’s disease patients, Eur J Clin Pharmacol, 65 (2009) 679-683. http://dx.doi.org/10.1007/s00228-009-0658-z
MA
[16] Hughes AJ, Daniel SE, Kilford L, Lees AJ, Accuracy of clinical diagnosis of idiopathic Parkinson's disease: a clinico-pathological study of 100 cases, J Neurol Neurosurg Psychiatry. 55 (1992) 181-184. http://dx.doi.org/ 10.1136/jnnp.55.3.181
D
[17]Tomlinson CL, Stowe R, Patel S, Rick C, Gray R, Clarke CE, Systematic review of levodopa dose
10.1002/mds.23429
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equivalency reporting in Parkinson's disease, Mov Disord. 25 (2010) 2649-2653. http://dx.doi.org/
[18] Hacker ML, Tonascia J, Turchan M, Currie A, Heusinkveld L, Konrad PE, Davis TL, Neimat JS,
CE P
Phibbs FT, Hedera P, Wang L, Shi Y, Shade DM, Sternberg AL, Drye LT, Charles D, Deep brain stimulation may reduce the relative risk of clinically important worsening in early stage Parkinson's disease, Parkinsonism Relat Disord. 21 (2015) 1177-1183. http://dx.doi.org/10.1016/j.parkreldis.2015.08.008
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[19] Bartus RT, Gene therapy for Parkinson's disease: a decade of progress supported by posthumous contributions from volunteer subjects, Neural Regen Res. 10 (2015) 1586-1588. http://dx.doi.org/10.4103/1673-5374.167783 [20] Bialecka M, Kurzawski M, Klodowska-Duda G, Opala G, Tan EK, Drozdzik M, The association of functional catechol-O-methyltransferase haplotypes with risk of Parkinson’s disease, levodopa treatment response, and complications, Pharmacogenet Genomics. 18 (2008) 815-821. http://dx.doi.org/10.1097/FPC.0b013e328306c2f2 [21] Lee JY, Cho J, Lee EK, Park SS, Jeon BS, Differential genetic susceptibility in diphasic and peak-dose dyskinesias in Parkinson's disease, Mov Disord. 26 (2011) 73-79. http://dx.doi.org/10.1002/mds.23400 [22] Dai D, Wang Y, Wang L, Li J, Ma Q, Tao J, Zhou X, Zhou H, Jiang Y, Pan G, Xu L, Ru P, Lin D4, Pan J, Xu L, Ye M, Duan S, Polymorphisms of and genes and Parkinson's disease: A meta-analysis, Biomed Rep. 2 (2014) 275-281. http://dx.doi.org/10.3892/br.2014.220 [23] Suzuki M, Hurd YL, Sokoloff P, Schwartz JC, Sedvall G, D3 dopamine receptor mRNA is widely expressed in the human brain, Brain Res. 779 (1998) 58-74. 9
ACCEPTED MANUSCRIPT http://dx.doi.org/10.1016/S0006-8993(97)01078-0 [24] Mierau J, Schneider FJ, Ensinger HA, Chio CL, Lajiness ME, Huff RM, Pramipexole binding and activation of cloned and expressed dopamine D2, D3 and D4 receptors, Eur J Pharmacol. 290 (1995) 29-36. http://dx.doi.org/ 10.1016/0922-4106(95)90013-6
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[25] Piercey MF, Pharmacology of pramipexole, a dopamine D3-preferring agonist useful in treating
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Parkinson's disease, Clin Neuropharmacol. 21 (1998) 141-151. [Pubmed: 9617505] [26] Deleu D, Northway MG, Hanssens Y, Clinical pharmacokinetic and pharmacodynamic properties
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http://dx.doi.org/10.2165/00003088-200241040-00003
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of drugs used in the treatment of Parkinson's disease, Clin Pharmacokinet. 41 (2002) 261-309.
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ACCEPTED MANUSCRIPT Table 1 Genotype and allele distributions of DRD2 CAn-STR and DRD3 Ser9Gly in PD patients and controls Genotypes
p
Alleles
p
value value 1314+
1314-
13
14
15
41(24.4%
126(69.2
=182)
%)
Ser9Gly PD
(n=182)
56(3
9(2
0.8%)
Ser/S
Ser/
.5%)
Gly/
er
Gly
Gly
77(45.8%)
66
25
(39.3%)
(14.9%)
86
80
16
(47.3%)
(44.0%)
(8.7%)
(n=168) Controls
18(5.4%) 154(45.8%) 19(5.7%) 143(42.6%) 2(0.6%)
155(4
2.6%)
23(
6.3%)
174(47.8 %)
0.196
0.239
3( 0.8%)
Ser
Gly
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Controls(n
DRD3
0.184
)
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127(75.6%)
220(65.5%)
116(34.5%)
252 (69.2%)
0.290
112 (30.8%)
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PD ( n=168)
17
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CAn-STR
16
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DRD2
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1314+: subjects carrying at least 1 of allele 13 or 14 in DRD2 CAn-STR; 1314-: subjects carrying neither 13 nor 14 allele in CAn-STR Data is expressed as the number of subjects in each category (frequency). All p values refer to the chi-square (2) test.
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Table 2 Demographic and clinical features of patients using DAs (n=102)
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DRD2 CAn-STR 1314+(7
Age
Male
p
Ser/Ser
Ser/Gly
Gly/Gly
9)
)
value a
(45)
(39)
(18)
65.1±9.4
65.3±8.1
0.946
65.9±8.5
65.4±10.0
62.8±8.2
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Gender
1314-(23
DRD3 Ser9Gly p value b
0.465
0.093
0.582
53(67.1%)
11 (47.8%)
30(66.7%)
22 (56.4%)
12(66.7%)
26(32.9%)
12(52.2%)
15(33.3%)
17(43.6%)
6(33.3%)
59.3±9.2
58.3±8.1
0.644
59.2±8.3
60.0±9.7
57.2±9.2
0.604
Duration of disease
6.0±4.6
6.9±4.1
0.391
6.7±4.6
5.7±4.0
6.2±5.1
0.581
H&Y stage
2.1±0.8
2.0±0.6
0.670
2.2±0.7
2.1±0.9
2.1±0.5
0.646
LED (mg/d)
419.8±247.3
386.3±231.3
0.563
395.0±232.8
406.4±255.0
468.1±247.2
0.554
Dosage of DAs (=LED, mg/d)
112.1±43.3
106.5±27.1
0.458
100.4±36.2
109.0±33.2
141.0±49.6
0.001**
UPDRS-Total
37.5±15.5
37.3±13.3
0.968
40.1±15.1
33.7±14.3
38.6±14.8
0.157
NMS
6.9±4.0
6.6±4.7
0.778
7.3±4.1
6.4±4.0
6.6±4.6
0.648
HAMA
6.1±5.2
7.0±6.4
0.479
6.1±4.6
6.2±6.3
7.0±5.9
0.831
HAMD
5.1±5.6
5.1±5.2
0.987
6.0±6.2
4.0±4.0
5.0±6.1
0.299
MMSE
27.8±2.6
27.4±3.0
0.552
Female Age of onset
27.4±2.7
27.8±2.8
28.4±1.4
0.40 1
Values are expressed as the mean ± SD except for the gender category. : p value between patients with different DRD2 CAn-STR genotypes. The 2 test or Fisher’s exact test for discrete variables and the unpaired t-test for a
11
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continuous variables were used. b : p value among patients with different DRD3 Ser9Gly genotypes. The 2 test or Fisher’s exact test for discrete variables and the one-way ANOVA for continuous variables were used. ** : p0.01
Table 3
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Demographic and clinical features of patients using Pramipexole (n=62) DRD2 CAn-STR 1314-(13)
66.0±8.2
66.3±8.0
D
Age
1314+(49)
Gender
Female
21(42.9%)
Age of onset
60.1±8.5
Duration of disease
0.891
DRD3 Ser9Gly Ser/Ser (28)
Ser/Gly(23)
Gly/Gly(11)
p value b
66.1±7.1
66.9±8.7
64.1±9.4
0.643
0.091
0.899
4(30.8%)
15(51.6%)
11(47.8%)
6(54.5%)
9(69.2%)
13(48.4%)
12(52.2%)
5(45.5%)
0.972
59.9±7.2
61.5±8.6
57.7±10.2
0.479
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28(57.1%)
60.2±7.8
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Male
p value
a
6.0±4.6
6.2±4.3
0.913
6.1±4.2
5.7±4.3
6.4±5.7
0.916
2.2±0.8
2.0±0.7
0.508
2.3±0.7
2.1±0.9
2.1±0.6
0.682
422.5±249.4
329.6±225.1
0.228
396.5±258.3
377.6±215.3
472.7±281.4
0.345
106.8±47.8
109.6±32.7
0.840
93.5±36.1
102.2±36.1
153.4±54.2
<0.001***
38.8±15.9
35.7±12.6
0.525
38.7±14.7
35.8±15.1
41.0±17.4
0.641
7.0±4.3
6.2±3.9
0.561
7.0±3.9
6.4±4.2
7.4±4.9
0.803
6.2±5.9
6.8±6.8
0.727
6.0±4.8
6.5±7.1
6.7±7.1
0.930
HAMD
5.3±6.2
3.5±3.9
0.356
5.8±6.6
3.8±4.2
4.8±6.6
0.483
MMSE
28.0±2.4
27.6±3.0
0.651
27.6±3.1
28.2±2.1
28.0±1.7
0.736
H&Y stage LED (mg/d)
UPDRS-Total NMS HAMA
AC
Dosage of DAs (=LED, mg/d)
Values are expressed as the mean ± SD except for the gender category. a: p value between patients with different DRD2 CAn-STR genotypes. The 2 test or Fisher’s exact test for discrete variables and the unpaired t-test for continuous variables were used. b: p value among patients with different DRD3Ser9Gly genotypes. The 2 test or Fisher’s exact test for discrete variables and the one-way ANOVA for continuous variables were used. : p 0.001
***
Table 4 Demographic and clinical features of patients using Piribedil (n=40)
12
ACCEPTED MANUSCRIPT DRD2 CAn-STR 1314+(
Age
1314-(1
DRD3 Ser9Gly p
30)
0)
value
63.7±11.0
63.9±8.4
0.965
Ser/Gl
Gly/Gl
p
(17)
y(16)
y(7)
value b
65.5±10.6
63.3±11.6
60.7±6.0
0.576 0.345
T
0.361
Male
25(83.3%)
7(70.0%)
15(88.2%)
11(68.8%)
6(85.7%)
Female
5(16.7%)
3(30.0%)
2(11.8%)
5(31.2%)
1(14.3%)
Age of onset
56.2±14.7
56.0±8.4
0.968
IP
Gender
Ser/Ser
a
57.7±10.9
48.3±22.4
0.230
Duration of disease
6.0±4.7
7.9±3.7
0.254
H&Y stage
2.0±0.7
2.1±0.6
0.795
LED (mg/d)
415.5±247.9
460.0±229.2
0.620
Dosage of DAs
120.8±33.5
102.5±18.4
0.039*
UPDRS-Total
35.3±14.7
39.5±14.5
0.447
NMS
6.7±3.5
7.1±5.8
HAMA
5.9±3.6
7.2±6.1
HAMD
4.7±4.5
7.0±6.0
MMSE
27.5±2.9
27.2±3.0
SC R
5.6±3.8
5.8±4.2
0.412
2.1±0.6
1.9±0.9
2.0±0.3
0.760
392.6±191.0
447.8±306.1
460.7±202.5
0.751
111.8±34.4
118.8±26.6
121.4±36.6
0.735
47.1±17.3
30.1±14.5
41.3±11.5
0.071
0.801
7.8±4.4
6.4±4.0
5.4±4.2
0.440
0.537
6.3±4.4
5.6±4.8
7.4±3.6
0.688
0.226
6.2±5.7
4.4±3.7
5.3±5.7
0.638
NU
7.6±5.2
MA
(=LED, mg/d)
57.9±9.9
27.0±2.
27.2±3.
29.0±1.
5
7
4
0. 290
D
0.758
Values are expressed as the mean ± SD except for the gender category. : p value between patients with different DRD2 CAn-STR genotypes. The 2 test or Fisher’s exact test for discrete variables and
the unpaired t-test for continuous variables were used.
TE
a
one-way ANOVA for
CE P
: p value among patients with different DRD3Ser9Gly genotypes. The 2 test or Fisher’s exact test for discrete variables and the
b
continuous variables were used. : p 0.05
AC
*
Table 5 Single-factor linear regression of doses of DAs with demographic and clinical features of patients PD patients taking DAs (n=102) p value Gen
95% CI
PD patients taking pramipexole (n=62) p value
95% CI
PD patients taking piribedil (n=40) p value
95% CI
0.592
-20.81-11.93
0.953
-22.30-23.66
0.494
-33.80-16.62
Age
0.059
-1.70-0.03
0.209
-2.31-0.52
0.164
-1.65-0.29
Age
0.016*
-1.95--0.20
0.127
-2.46-0.31
0.057
-1.99--0.93
0.198
-0.62-2.96
0.503
-1.73-3.49
0.184
-0.75-3.77
0.659
-13.47-8.56
0.507
-19.38-9.67
0.648
-11.50-18.26
der
a
a
a
of onset
Dur ation of disease a H& Y stage
a
13
ACCEPTED MANUSCRIPT UPDRS-Total a
0.681
-0.44-0.67
0.675
-0.62-0.10
0.865
-0.68-0.80
NM
0.721
-2.36-1.64
0.669
-3.51-2.27
0.962
-2.52-2.64
0.969
-1.51-1.57
0.952
-2.05-1.93
0.788
-2.18-2.86
0.265
-2.39-0.67
a
HA
a
MD
-2.58-1.61
0.99
-4.83-4.88
4
MM SE
0.64
a
0.8 83
DR
-3.41-2.9 4
0.560
3
-24.51-13.35
0.840
D2 CAn-STR
DR D3
01
-28.20--
<0.0
8.02
01
a
***
-39.92--12.14
0.093
-3.92-0.32
0.868
-4.12-3.50
0.110
-40.98-4.3 1
0.445
-18.97-8.5 0
MA
Ser9Gly a
0.0 **
-2.35-31.06
NU
a
T
HA
IP
MA
a
SC R
S
：independent variable
Dependent variable: doses of DAs, pramipexole or piribedil 95% CI: 95% confidence interval
CE P
TE
D
: p0.05; **: p<0.01; ***: p<0.001
AC
*
14
ACCEPTED MANUSCRIPT Highlights •
We investigated the association between DRD2 CAn-STR as well as DRD3 Ser9Gly polymorphisms and doses of DAs, especially pramipexole
•
We found that PD patients carrying DRD3 Ser9Gly Gly/Gly genotype took more pramipexole Our study indicated that to obtain better therapeutic results, higher doses of pramipexole should be used in patients with Gly/Gly genotype
Our results provide some insights into the pharmacogenomics of PD treatment and may help
CE P
TE
D
MA
NU
SC R
to better understand individual variability in response to DAs
AC
•
IP
•
T
than patients with Ser/Ser and Ser/Gly genotypes
15