A genetic predisposition to the development of neuroleptic malignant syndrome (NMS) has been suggested by clinical studies. Although the molecular basis of NMS is unclear, a dopaminergic blockade mechanism has been considered the main cause. We therefore investigated the association between NMS and three functional polymorphisms of the dopamine D2 receptor (DRD2) gene: TaqI A, −141C Ins/Del, and Ser311Cys. Subjects included 32 Japanese patients, previously diagnosed with NMS, and 132 schizophrenic patients treated with neuroleptics without occurrence of NMS. Polymerase chain reaction and restriction fragment length polymorphism analyses were performed to determine each genotype. We found significant differences in genotypic and allelic frequencies of the –141C Ins/Del polymorphism between patients with and without NMS. The −141C Del allele was significantly more frequent in the NMS group (23.4 vs 11.7%, P=0.026). Similarly, the proportion of −141C Del allele carriers was significantly higher in the NMS group (40.6 vs 20.5%, P=0.022). No significant differences between the two groups were seen for allelic and genotypic frequencies of the TaqI A and Ser311Cys polymorphisms. This result suggests that the −141C Ins/Del polymorphism is likely to predispose toward the development of NMS, probably together with other unidentified factors.
Neuroleptic malignant syndrome (NMS) is a severe adverse effect of psychotropic drugs characterized by hyperthermia, extrapyramidal signs, altered consciousness, and autonomic dysfunction. In addition to acquired physical risk factors, previous case reports including descriptions of familial occurrence have suggested a genetic predisposition toward NMS,1,2 and a molecular basis for NMS has been pursued.
A dopaminergic blockade mechanism has been considered to play a major role in NMS because most neuroleptics with a blocking effect at the dopamine D2 receptor (DRD2) can cause NMS, while discontinuation of neuroleptic medication usually improves symptoms of NMS. Therefore, central dopaminergic dysfunction, especially at DRD2, is believed to contribute to the development of NMS. Accordingly, several genetic studies with respect to DRD2 have been performed.
A number of functional polymorphisms have been identified in the DRD2 gene to date. Among these, the Ser311Cys polymorphism in exon 7 was suggested to affect the interaction between DRD2 and its G protein.3 The −141C Ins/Del polymorphism situated in the promoter region was suggested to affect promoter activity by an expression study using human cells.4 The Del allele was found to increase the DRD2 density in the striatum of the human brain, while the TaqI A1 allele in the 3′-untranslated region was demonstrated to reduce receptor density.5,6 Recent studies have shown a possible association of these polymorphisms with interindividual differences in both early therapeutic effects and adverse effects of neuroleptics.7,8,9 In terms of NMS, Suzuki et al10 suggested a possible association between NMS in Japanese patients and the TaqI A polymorphism, but we could not replicate their result in larger patient samples.11
In the present study, we performed systematic screening for these DRD2 polymorphisms and assessed genetic associations with the occurrence of NMS in Japanese patients.
We studied 32 patients (17 men and 15 women; mean age, 49.2 years; SD, 18.2) who had experienced NMS (27 patients with schizophrenia, one with schizoaffective disorder, three with mood disorder, and one with a psychotic disorder followed after viral encephalitis). Our control subjects were 132 patients with schizophrenia who had taken neuroleptics for more than 1 year and had never developed NMS (70 men and 62 women; mean age, 46.8 years; SD, 15.7). Psychiatric diagnoses were made according to the diagnostic criteria of the Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition (DSM-IV). NMS was diagnosed retrospectively according to the criteria of Pope et al,12 with all NMS cases confirmed as definite by these criteria. All subjects were Japanese, and no subject had a diagnosis of substance abuse. This study was approved by the Ethics Committee of Yokohama City University School of Medicine, and written informed consent was obtained from all subjects.
Genomic DNA was extracted from peripheral white blood cells from all patients using standard techniques. Polymerase chain reaction and restriction fragment length polymorphism analyses were performed to determine TaqI A genotypes according to Grandy et al;13 −141C Ins/Del genotypes according to Hori et al;14 and Ser311Cys genotypes according to Arinami et al,15 respectively. Genetic analyses were performed by an investigator kept unaware of which subjects had developed NMS.
Statistical analyses were performed using χ2 tests with the Monte Carlo method or Fisher's exact test to determine the presence of allelic and/or genotypic associations. In addition, the logistic regression analysis was performed to evaluate simultaneously the possible associations between NMS and independent variables (DRD2 genotypes, gender, and age). For the regression model, the allelic effect was assumed to be additive, that is, scores of 0, 1, and 2 were assigned for genotype wild homozygotes, heterozygotes, and mutant homozygotes of each mutation, respectively.
The differences in demographic characteristics between NMS patients with and without polymorphisms were evaluated by Fisher's exact test for categorical variables (gender distribution and NMS recurrence) and the Mann–Whitney U-test for continuous variables (age of onset of NMS, duration of illness, and neuroleptic dose). The neuroleptic dose before NMS onset was calculated in terms of chlorpromazine equivalents.16,17 The presence of Hardy–Weinberg equilibrium was determined using the χ2 test. These analyses were performed using SPSS 10.0 for Windows (SPSS Japan, Tokyo). Pairwise linkage disequilibrium and estimated haplotypes were analyzed using ARLEQUIN 2.000.18
Table 1 shows genotypic and allelic frequencies of three polymorphisms investigated in NMS patients and in schizophrenic control patients. No deviation from the Hardy–Weinberg equilibrium was seen in either the NMS or control group. There were no significant differences in age (t=0.769, df=154, P=0.443, Student's t-test) or gender (P=1.00, Fisher's exact test) distribution between the two groups. We found a significant difference in a genotypic distribution associated with the –141C Ins/Del polymorphism between patients with and without NMS (χ2=5.685, df=2, P=0.044, χ2 test with the Monte Carlo method). Frequencies of the −141C Del allele and proportion of Del carriers (ie, the patients with either Ins/Del or Del/Del) were significantly higher in the NMS group (allelic distribution, P=0.026; proportion of Del carriers, P=0.022, Fisher's exact test). The odds ratio for the development of NMS associated with the Del allele was 2.30 (95% CI, 1.16–4.59). Similarly, as shown in Table 2, the logistic regression analysis showed a significant association between NMS and −141C Ins/Del genotypes. (P=0.018; odds ratio, 2.30; 95% CI, 1.15–4.57).
On the other hand, no significant differences were noted in genotypic and allelic frequencies of the TaqI A polymorphism (genotypic distribution, χ2=1.474, df=2, P=0.484, χ2 test with the Monte Carlo method; allelic distribution, P=0.475, Fisher’s exact test), or of the Ser311Cys polymorphism (genotypic distribution, P=0.228; allelic distribution, P=0.235, Fisher’s exact test) between the two groups.
Table 3 shows the demographic characteristics of NMS patients with and without polymorphisms at the A1, Del, and Cys alleles. In any polymorphisms, no significant differences were noted for gender, age at NMS onset, duration of psychiatric illness, neuroleptic dose at NMS onset, or proportion of patients with recurrence of NMS among A1, Del, or Cys carriers and noncarriers.
Haplotype analysis revealed a marginally significant linkage disequilibrium between the TaqI A and Ser311Cys polymorphisms (D′=1.000, r2=0.022, P=0.007). On the other hand, no significant linkage disequilibrium was detected between the –141C Ins/Del and Ser311Cys polymorphisms (D′=0.352, r2=0.001, P=0.632) or the TaqI A and –141C Ins/Del polymorphisms (D′=0.002, r2=0.000, P=0.987). As shown in Table 4, a significant difference in an estimated haplotype distribution between patients with and without NMS was detected (χ2=16.3, df=4, P=0.03, χ2 test with the Monte Carlo method). Moreover, a frequency of the A2-Del-Ser haplotype was estimated to be significantly higher in NMS than that in controls, whereas a frequency of A2-Ins-Ser haplotype was estimated to be lower in NMS (P=0.01, Fisher’s exact test; odds ratio, 3.25; 95% confidence interval, 1.71–6.16).
Using SPSS Sample Power for Windows, we estimated that our sample had a power of 0.95 to detect a medium effect size (w=0.25), and a power of 0.30 to detect a small effect size (w=0.10) at a P<0.05 level for allelic comparison.
A dopaminergic blockade mechanism in the central nervous system is presumed to be the main cause of NMS, and this hypothesis has been supported by several reports. Nishijima et al19 showed that the cerebrospinal fluid concentration of homovanilic acid (HVA), a metabolite of dopamine, was low in the active phase of NMS and also after recovery. Jauss et al20 demonstrated using single photon emission computed tomography (SPECT) that DRD2 in the basal ganglia had been completely occupied during the acute state of NMS. Naturally, the DRD2 gene is of great interest in investigating the vulnerability to NMS. Furthermore, recent pharmacogenetic studies have shown that the DRD2 gene polymorphisms confer interindividual differences in responses to neuroleptic treatment. In the present study, we performed systematic screening for three functional DRD2 polymorphisms, and assessed the relationship between NMS and these polymorphisms. As a result, we found significant differences in genotypic and allelic frequencies of the –141C Ins/Del polymorphism between patients with and without NMS. Frequencies of the −141C Del allele and Del carriers were significantly higher in the NMS group. These results suggest that the Del allele is likely to be associated with a predisposition to the development of NMS.
The Del allele frequency, 11.7%, in our schizophrenic controls was lower than those previously reported in Japanese healthy controls (23%, Ishigro et al;21 25%, Katsuragi et al22), but consistent with those previously reported in Japanese schizophrenics (14%, Arinami et al;4 10%, Ohara et al23). Although Arinami et al4 and Ohara et al23 have reported a possible association between the −141 Ins/Del polymorphism and schizophrenia, these results have not been replicated.14
Recently, Suzuki et al8 suggested the clinical importance of −141 Ins/Del polymorphism, reporting that patients without the Del allele were particularly likely to show an improvement in anxiety and depressive symptoms when treated with DRD2 antagonists. Jönsson et al5 demonstrated by positron emission tomography (PET) that striatal DRD2 density in healthy subjects with the Del allele was higher than in others without the Del allele, although Pohjalainen et al24 could not replicate these results. Arinami et al3 examined the transient expression of luciferase enzymatic activity for reporter plasmids containing the Del or Ins allele, noting less efficient expression with the Del allele. These various observations suggested that this polymorphism could be an important factor affecting clinical phenotypes concerning the dopaminergic system. Furthermore, the unreplicated result concerning the human striatal DRD2 density remains of great interest. Relatively decreased DRD2 expression might predispose NMS occurrence. However, the results of receptor density studies in vivo are inconsistent so far, and we could not elucidate the pathogenetic mechanism of NMS in our present study.
In the present study, no significant differences in allelic or genotypic frequencies of the TaqI A or Ser311Cys polymorphisms were seen between the two groups. In previous studies, Otani and colleagues reported an association between the A1 allele of the TaqI A polymorphism and NMS,10,25 whereas the –141C Ins/Del polymorphism was not associated with NMS.25 We suspect that a difference in a sample size can explain the discrepancy in the results between the previous and present studies. While we examined 32 NMS cases, the previous investigators compared 15–17 NMS cases with 138–163 control cases, which might have influenced the results. On the other hand, we found a significant difference in a haplotype distribution between patients with and without NMS. Especially, a frequency of A2-Del-Ser haplotype was estimated to be significantly higher in NMS. Combinations of these three polymorphisms could be a marker for the prediction of NMS, and further haplotype studies based on larger samples are needed.
In terms of the demographic profiles of NMS patients, we noted no significant differences in the clinical background, including the neuroleptic dose between NMS patients with and without the −141C Del allele. Although, in general, the incidence of NMS is fairly low, some patients in our study had recurrent NMS. We know that some NMS patients experience recurrence even without known acquired physical risk factors, and we hypothesized that at least some recurrences would involve constitutional factors regulated by genetics. We could not confirm this hypothesis, finding no significant differences in genotypic distribution in patients with NMS recurrence. However, the numbers of cases with NMS recurrence appeared to be too small to resolve the issue.
In conclusion, our findings suggest that the −141C Ins/Del polymorphism is likely to affect vulnerability toward development of NMS. However, the sample size for our NMS patients was relatively small, and further large case–control studies will be needed to confirm population frequencies and the status of the Del allele with respect to NMS. In addition, we believe that by itself the Del allele cannot explain all NMS occurrences. Other genetic association studies concerning NMS have been performed from both pharmacokinetic and pharmacodynamic points of view,26,27 and additional genetic studies considering effects of factors in combination are needed to elucidate the nature of genetic predisposition toward NMS.
Deuschl G, Oepen G, Hermle L . Neuroleptic malignant syndrome: observations on altered consciousness. Pharmacopsychiatry 1987; 20: 168–170.
Otani K, Horiuchi M, Kondo T, Kaneko S, Fukushima Y . Is the predisposition to neuroleptic malignant syndrome genetically transmitted? Br J Psychiatry 1991; 158: 850–853.
Cravchik A, Sibley DR, Gejman PV . Functional analysis of the human D2 dopamine receptor missense variants. J Biol Chem 1996; 271: 26013–26017.
Arinami T, Gao M, Hamaguchi H, Toru M . A functional polymorphism in the promoter region of the dopamine D2 receptor gene is associated with schizophrenia. Hum Mol Genet 1997; 6: 577–582.
Jönsson EG, Nöthen MM, Grünhage F, Farde L, Nakashima Y, Propping P et al. Polymorphisms in the dopamine D2 receptor gene and their relationships to striatal dopamine receptor density of healthy volunteers. Mol Psychiatry 1999; 4: 290–296.
Pohjalainen T, Rinne JO, Någren K, Lehikoinen P, Anttila K, Syvälahti EKG et al. The A1 allele of the human D2 dopamine receptor gene predicts low D2 receptor availability in healthy volunteers. Mol Psychiatry 1998; 3: 256–260.
Schäfer M, Rujescu D, Giegling I, Guntermann A, Erfurth A, Bondy B et al. Association of short-term response to haloperidol treatment with a polymorphism in the dopamine D2 receptor gene. Am J Psychiatry 2001; 158: 802–804.
Suzuki A, kondo T, Mihara K, Yasui-Furukori N, Ishida M, Furukori H et al. The −141C Ins/Del polymorphism in the dopamine D2 receptor gene promoter region is associated with anxiolytic and antidepressive effects during treatment with dopamine antagonists in schizophrenic patients. Pharmacogenetics 2001; 11: 545–550.
Mihara K, Kondo T, Suzuki A, Yasui N, Nagashima U, Ono S et al. Prolactin response to nemonapride, a selective antagonist for D2 like dopamine receptors, in schizophrenic patients in relation to Taq1 A polymorphism of DRD2 gene. Psychopharmacology 2000; 149: 246–250.
Suzuki A, Kondo T, Otani K, Mihara K, Yasui-Furukori N, Sano A et al. Association of the TaqI A polymorphism of the dopamine D2 receptor gene with predisposition to neuroleptic malignant syndrome. Am J Psychiatry 2001; 158: 1714–1716.
Kishida I, Kawanishi C, Furuno T, Matsumura T, Hasegawa H, Sugiyama N et al. Lack of association in Japanese patients between neuroleptic malignant syndrome and the Taq I A polymorphism of the dopamine D2 receptor gene. Psychiatric Genet 2003; 13: 55–57.
Pope GH, Keck PE, McElroy SL . Frequency and presentation of neuroleptic malignant syndrome in a large psychiatric hospital. Am J Psychiatry 1986; 143: 1227–1233.
Grandy DK, Litt M, Allen L, Bunzow JR, Marchionni M, Makam H et al. The human dopamine D2 receptor gene is located on chromosome 11 at q22–q23 and identifies a TaqI RFLP. Am J Hum Genet 1989; 45: 778–785.
Hori H, Ohmori O, Shinkai T, Kojima H, Nakamura J . Association between three functional polymorphisms of dopamine D2 receptor gene and tardive dyskinesia in schizophrenia. Am J Med Genet 2001; 105: 774–778.
Arinami T, Itokawa M, Enguchi H, Tagaya H, Yano S, Shimizu H, Hamaguchi H et al. Association of dopamine D2 receptor molecular variant with schizophrenia. Lancet 1994; 343: 703–704.
Inagaki A, Inada T, Fujii Y, Yagi G . Dose equivalence orally administered neuroleptics. In: Dose Equivalence of Psychotropic Drugs. Seiwa Shoten: Tokyo, 1999; 11–60, (in Japanese).
American Psychiatry Association. Practice guideline for the treatment of patients with schizophrenia. III. Treatment principles and alternatives. Am J Psychiatry 1997; 154: 7–34.
Schneider S, Roessli D, Excoffier L . ARLEQUIN: a Software for Population Genetic Data Analysis. University of Geneva: Geneva, 2000.
Nishijima K, Ishiguro T . Neuroleptic malignant syndrome: a study of CSF monoamine metabolism. Biol Psychiatry 1990; 27: 280–288.
Jauss M, Krack P, Franz M, Klett R, Bauer R, Gallhofer B et al. Imaging of dopamine receptors with [123I]iodobenzamide single-photon emission-computed tomography in neuroleptic malignant syndrome. Movement Disord 1996; 11: 726–728.
Ishiguro H, Arinami T, Saito T, Akazawa S, Enomoto M, Mitushio H et al. Association study between the –141C Ins/Del and TaqI A polymorphisms of the dopamine D2 receptor gene and alcoholism. Alc Clin Exp Res 1998; 22: 845–848.
Katsuragi S, Kiyota A, Tsutsumi T, Isogawa K, Nagayama H, Arinami T et al. Lack of association between a polymorphism in the promoter region of the dopamine D2 receptor and personality traits. Psychiatry Res 2001; 105: 123–127.
Ohara K, Nagai M, Tani K, Nakamura Y, Ino A, Ohara K . Functional polymorphism of –141C Ins/Del in the dopamine D2 receptor gene promoter and schizophrenia. Psychiatry Res 1998; 81: 117–123.
Pohjalainen T, Någren K, Syvälahti EKG, Hietala J . The dopamine D2 receptor 5′-flanking variant, −141C Ins/Del, is not associated with reduced dopamine D2 receptor density in vivo. Pharmacogenetics 1999; 9: 505–509.
Mihara K, Kondo T, Suzuki A, Yasui-Furukori N, Ono S, Sano A et al. Relationship between functional dopamine D2 and D3 receptors gene polymorphisms and neuroleptic malignant syndrome. Am J Med Genet 2003; 117B: 57–60.
Kawanishi C, Hanihara T, Shimoda Y, Suzuki K, Sugiyama N, Onishi H et al. Lack of association between neuroleptic malignant syndrome and polymorphisms in the 5HT1A and 5HT2A receptor genes. Am J Psychiatry 1998; 155: 1275–1277.
Kawanishi C, Furuno T, Onishi H, Sugiyama N, Suzuki K, Matsumura T et al. Lack of association in Japanese patients between neuroleptic malignant syndrome and debrisoquine 4-hydroxylase genotype with low enzyme activity. Psychiatric Genet 2000; 10: 145–147.
We thank Dr Shinsuke Washizuka for his helpful advice.
About this article
Cite this article
Kishida, I., Kawanishi, C., Furuno, T. et al. Association in Japanese patients between neuroleptic malignant syndrome and functional polymorphisms of the dopamine D2 receptor gene. Mol Psychiatry 9, 293–298 (2004) doi:10.1038/sj.mp.4001422
- TaqI A
- −141C Ins/Del
- dopamine D2 receptor (DRD2)
- genetic polymorphism
- neuroleptic malignant syndrome
Evaluation of association of DRD2 TaqIA and -141C InsDel polymorphisms with food intake and anthropometric data in children at the first stages of development
Genetics and Molecular Biology (2018)
Molecular Neurobiology (2015)
European Journal of Pediatrics (2014)
Journal of korean Academy of Child and Adolescent Psychiatry (2013)
The Journal of Nervous and Mental Disease (2013)