N-acetyltransferase-2 polymorphism in Parkinson’s disease: the Rotterdam study
- B Sanjay Harhangia,
- Ben A Oostrab,
- Peter Heutinkb,
- Cornelia M van Duijna,
- Albert Hofmana,
- Monique M B Bretelera
- aDepartment of Epidemiology and Biostatistics, bDepartment of Clinical Genetics, Erasmus University Medical School, PO Box 1738, 3000 DR Rotterdam, The Netherlands
- Dr M M B Breteler, Department of Epidemiology and Biostatistics, Erasmus University Medical School, PO Box 1738, 3000 DR, Rotterdam, The Netherlands. Telephone 0031 104087489; fax 0031 104089382; email
- Received 25 January 1999
- Revised 10 May 1999
- Accepted 21 May 1999
The N-acetyltransferase-2 gene (NAT-2) has been associated with Parkinson’s disease. The genotype associated with slow acetylation has been reported to be increased in patients with Parkinson’s disease. Three mutant alleles M1, M2, and M3 of NAT-2 were investigated in 80 patients with idiopathic Parkinson’s disease and 161 age matched randomly selected controls from a prospective population based cohort study. The allelic frequencies and genotypic distributions in patients were very similar to those found in controls. In controls the frequency of the wild type allele increased significantly with age suggesting that the mutant alleles are associated with an increased risk of mortality. These findings suggest that NAT-2 polymorphism is not a major genetic determinant of idiopathic Parkinson’s disease, but may be a determinant of mortality in the general population.
Parkinson’s disease is a common neurodegenerative disorder among elderly people.1 2 The aetiology of Parkinson’s disease still remains largely unknown but most likely results from interaction between genetic and environmental factors.3-5 Several functionally relevant polymorphisms in xenobiotic metabolism have been studied in relation to patients with Parkinson’s disease, but with no consistent results.6 7 Explanations for these inconsistencies include different methodologies, different diagnostic criteria, poor selection of control groups, and small sample sizes. Recently, an association of slow acetylators for NAT-2 with Parkinson’s disease was reported.8 NAT-2, which maps to chromosome 8p22,9 10 is associated with speed of acetylation of certain drugs and xenobiotics.9 11 Slow acetylators are homozygous for any of the mutant alleles and may be more susceptible to low level environmental exposure to neurotoxins.8 The finding of an increased frequency of patients homozygous for the NAT-2 mutations is compatible with the view that patients with Parkinson’s disease may be less capable of handling certain endogenous or exogenous toxins.12 However, the findings on NAT-2 were not confirmed.13 The aim of this study was to investigate the possible association of NAT-2 polymorphism in idiopathic Parkinson’s disease.
The study formed part of the Rotterdam study, a prospective population based cohort study on the frequency, aetiology, and prognosis of chronic diseases. The cohort exists of 7983 independently living or institutionalised inhabitants from a suburb of Rotterdam, The Netherlands, aged 55 years or older. The study started in June 1990 and has been described extensively elsewhere.14 Informed consent was obtained from each participant and the study was approved by the medical ethics committee of Erasmus University, Rotterdam. Participants were screened at baseline (1990–3) and at follow up (1993–4) for symptoms of parkinsonism by study physicians. All screen positives had a diagnostic investigation by a neurologist. Parkinson’s disease was diagnosed in persons with at least two out of four cardinal signs (resting tremor, bradykinesia, rigidity, and postural disturbances) and no other apparent cause of parkinsonism.2 In the Rotterdam study 97 prevalent and 35 incident patients with Parkinson’s disease were identified until 1994. Blood samples for DNA extraction and genotyping were available for 80 patients (68 prevalent patients and 12 incident patients, mean age 77.3 (SD 8.3); range 57.5–99.2; 27 men, 53 women). For each patient we randomly selected two age matched ( within 5 years) controls (mean age 76.8 (SD 8.3); range 57.9–98.8, 63 men and 98 women) from the same study population who did not have Parkinson’s disease and of whom baseline data regarding smoking history as well as blood samples were available. A polymerase chain reaction (PCR) was conducted using specific PCR primers for NAT-2.15 The amplification products were digested using restriction enzymes, separated on an agarose gel and visualised with ultraviolet light. We investigated the genotype of NAT-2 using KpnI, TaqI, and BamHI for the three mutant alleles M1, M2, and M3 and the wild type allele of NAT-2.8 Genotyping was performed on coded samples without knowledge of the clinical diagnosis of the patients. The mutations M1, M2, and M3 account for most of the slow acetylators in white patients.16 Slow acetylators were defined as carrying any two of the mutant alleles M1, M2, and M3. Allele frequencies were determined by counting alleles and calculating sample proportions. Allele frequencies and genotype frequencies were compared using χ2 statistics.
Genotype distributions were in Hardy-Weinberg equilibrium. Frequencies of mutations in patients and controls and stratified on age are listed in the table. Overall, the mutation frequencies were similarly distributed among patients and controls.
The proportion of slow acetylators in the youngest age category, 55–64 years, was significantly lower in patients than controls. An interesting finding was that the frequency of the wild type allele in controls increased significantly with age (p trend<0.001).
Our results do not confirm the association between the slow acetylator genotype for NAT-2 and Parkinson’s disease as was found previously.8 Sample size calculations with a 5% significance level showed that we had a power of 90% in our study to detect differences in genotype frequencies at an odds ratio level of 2.5 for slow acetylators as was found in the study of Bandmannet al.8 This study was based on pathologically proved Parkinson’s disease, whereas we used clinical diagnostic criteria to assess Parkinson’s disease. It might be considered that we misclassified some patients in our study and that this has biased our estimates. It is highly unlikely, however, that the resulting bias, if any, would be big enough to explain the discrepant findings. Moreover, the overall frequencies of slow acetylators in patients and controls found in our study are similar to that found in other studies.13 17
There are several other possible explanations for the discrepancy between our findings and those of Bandmann et al.8 Firstly, they reported a significant association between NAT-2 polymorphism and familial Parkinson’s disease, but not with sporadic Parkinson’s disease. However, the frequency of the slow acetylator genotype in sporadic Parkinson’s disease in their initial analysis was significantly higher than controls (odds ratio=2.45; p=0.003) and after correction for multiple comparison the point estimate of the association remained the same but only became of borderline significance (odds ratio=2.45; p=0.06). Secondly, the association between NAT-2 and Parkinson’s disease may not be due to a causal relation but rather to the NAT-2 gene being in linkage disequilibrium with a neighbouring gene that is involved in the aetiology of Parkinson’s disease. This explanation is in line with the finding of a lower proportion of slow acetylators in the early onset group in the present study whereas Bandmann et al found a higher proportion of slow acetylators in patients with Parkinson’s disease compared with controls. Thirdly, their controls came from a heterogeneous population submitted to the United Kingdom Parkinson’s Disease Brain Bank and the brain bank at the institute of Psychiatry, London, UK and were not matched for ethnic background. This could explain the relatively low frequency of slow acetylators found in their control population compared with our study and others,13 17 and may have introduced some bias. Fourthly, we found that the frequency of the wild type allele in controls increased significantly with age suggesting that the mutant alleles are associated with an increased risk of mortality, possibly because of the increased risk of cancer for mutation carriers.7 18 This suggests that bias may be introduced if cases and controls are not matched for age which was the case in the series of Bandmann et al,8 in which the mean age of the control population (77.1 years) was almost 9 years higher than the mean age from familial patients (68.4 years). A final explanation could be that slow acetylators are not only more susceptible to neurotoxins which are inactivated by an acetylation reaction,8 but are simultaneously less susceptible to potential neurotoxins which are activated by acetylation. This dual activation-detoxification of the NAT-2 polymorphism makes the interpretation of any association debatable.19
The findings of this population based study suggest that NAT-2 polymorphism is not a major genetic determinant of idiopathic Parkinson’s disease, but may be a determinant of mortality in the general population.
We thank Erwin Wauters for the technical assistance. This study was supported in part by the Prinses Beatrix Fund and Biomed II, grant PL95–0664.