human psychopharmacology

Hum. Psychopharmacol Clin Exp 2011; 26: 120124. Published online 16 March 2011 in Wiley Online Library (wileyonlinelibrary.com) DOI: 10.1002/hup.1176

Correlation of adenosinergic activity with superior efcacy of clozapine for treatment of chronic schizophrenia: a double blind randomised trial
Ali Ghaleiha1, Navid Honarbakhsh2, Mohammad-Ali Boroumand3, Morteza Jafarinia2, Mina Tabrizi4, Farzin Rezaei5, Maedeh Raznahan6 and Shahin Akhondzadeh2*
1 2

Research Center for Behavioral Disorders and Substance Abuse, Hamadan University of Medical Sciences, Hamadan, Iran Psychiatric Research Center, Roozbeh Hospital, Tehran University of Medical Sciences, Tehran, Iran 3 Department of Clinical Research, Tehran Heart Center, Tehran University of Medical Sciences, Tehran, Iran 4 Department of Medical Genetics, Faculty of Medicine, Tehran University of Medical Sciences, Tehran, Iran 5 Qods Hospital, Kurdistan University of Medical Sciences, Sanandaj, Iran 6 Faculty of Medicine, Tehran University of Medical Sciences, Tehran, Iran

Objective It has been proposed that a decit of adenosinergic activity could contribute to the pathophysiology of schizophrenia. The authors undertook this study to further evaluate the level of adenosine deaminase (ADA) in patients with chronic schizophrenia treated with monotherapy of haloperidol, risperidone or clozapine and correlation between the ADA level with response to treatment. Methods The trial was a prospective, 8-week, double blind study of parallel groups of patients with chronic schizophrenia. Eligible participants in the study were 51 patients with chronic schizophrenia with ages ranging from 20 to 45 years. All participants were inpatients, in the active phase of illness, and met DSM-IV-TR criteria for schizophrenia. Patients were randomly allocated (17 patients in each group) to risperidone (6 mg/day) or haloperidol 15 mg/day or clozapine (300 mg/day). Serum ADA activity was measured at baseline and week 8. Results The plasma levels of ADA in patients with chronic schizophrenia who received clozapine were signicantly higher than patients who received haloperidol. In addition, response to treatment was positively correlated with plasma levels of ADA only in the clozapine group (r 0.46 and p 0.04). Conclusion The results indicate an increased activity of the enzyme ADA in the serum of schizophrenic patients being treated with clozapine and this increase may be correlated with clozapines superior antipsychotic efcacy. Copyright # 2011 John Wiley & Sons, Ltd. key words adenosine; adenosine deaminase; schizophrenia

INTRODUCTION Schizophrenia is a debilitating illness, rating as one of the leading causes of lost years of life quality. The illness imposes a disproportionate burden on patients, their families, the healthcare system and the society (Akhondzadeh, 2006; Mohammadi and Akhondzadeh, 2001). Although the dopamine hypothesis of schizophrenia remains the leading neurochemical hypothesis, other neurotransmitter receptors may also be involved in the pathogenesis of schizophrenia (Akhondzadeh, 2006; Dixon et al., 1999). Adenosine, a purine nucleoside, has been known to possess both inhibitory and excitatory neuromodulatory function, and to be
* Correspondence to: S. Akhondzadeh, Psychiatric Research Center, Roozbeh Psychiatric Hospital, Tehran University of Medical Sciences, South Kargar Street, Tehran 13337, Iran. Tel: 98-21-88281866; Fax: 98-2155419113. E-mail: s.akhond@neda.net

involved in the ne-tuning of other neurotransmitters (Boison, 2008). There is a large amount of data showing that adenosine plays an opposing role to dopamine in the brain (Fuxe et al., 2010; Shen and Chen, 2009). Adenosine agonists and antagonists produce behavioural effects similar to dopamine antagonists and dopamine agonists, respectively (Akhondzadeh et al., 2000; Lara et al., 2006). There are increasing putative therapeutic applications for adenosine receptor agonists and antagonists that act at the four adenosine receptor subtypes A1, A2A, A2B and A3 (Boison, 2008). Preclinically, adenosine and its analogs exert antipsychotic, anxiolytic, sedative, anticonvulsant and antiaggressive effects (Boison, 2008; Shen and Chen, 2009). A strong case has been made in the literature for the involvement of adenosine receptors, particularly the A1 and A2A types in the aetiology of schizophrenia (Lara and Souza, 2000;
Received 15 November 2010 Accepted 12 January 2011

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Lara et al., 2006). Diogo Lara has championed the idea that adenosinergic activity might be decient in schizophrenia (Lara and Souza, 2000; Lara et al., 2006). Altogether these observations support a putative role for decient levels of adenosine in the brain of schizophrenic patients and are supportive of the adenosine hypofunction hypothesis of schizophrenia (Lara and Souza, 2000; Lara et al., 2006). Dipyridamole, a reuptake inhibitor of adenosine was benecial for schizophrenia patients on 20 mg haloperidol daily for positive symptoms, which was suggested to be due to adenosinedopamine interactions (Akhondzadeh et al., 2000). A number of studies and clinical observations indicate that allopurinol as a xanthine oxidase inhibitor may reduce the symptoms of schizophrenia in patients who are also receiving antipsychotic medication (Akhondzadeh et al., 2005; Brunstein et al., 2005; Dickerson et al., 2009). Extracellular adenosine can be taken up by the nucleoside transporter and inactivated by deamination to inosine by adenosine deaminase (ADA) or phosphorylated to AMP by adenosine kinase (Boison, 2008). These processes are mostly intracellular, but ADA is also found associated with cell membranes (Boison, 2008). Brunstein et al. (2007) reported increased serum ADA activity in 26 male schizophrenic patients under antipsychotic monotherapy of either typical antipsychotics or clozapine and 26 healthy volunteers balanced for age and race. The increased serum ADA activity was more pronounced in the clozapine-treated group (Brunstein et al., 2007). The authors undertook this study to further evaluate the level of ADA in patients with chronic schizophrenia treated with monotherapy of haloperidol, risperidone or clozapine and correlation between the ADA level with response to treatment. METHODS Trial setting The trial was a prospective, 8-week, double blind study of parallel groups of patients with chronic schizophrenia and was undertaken in Roozbeh Psychiatric hospital in Iran, from June 2008 to May 2010. Participants Eligible participants in the study were 51 patients with chronic schizophrenia (18 women and 33 men) with ages ranging from 20 to 45 years. All participants were inpatients, in the active phase of illness, and met DSMIV-TR criteria for schizophrenia (American Psychiatric Association, 2000). Minimum score of 60 on the
Copyright # 2011 John Wiley & Sons, Ltd.

Positive and Negative Syndrome Scale (PANSS) was required for entry into the study (Kay et al., 1987). The patients did not receive neuroleptics for a week prior to entering the trial or long-acting antipsychotics for at least 2 months before the study. Patients were excluded from the study if they had a clinically signicant organic and neurologic disorder, serious psychotic disorders other than schizophrenia. Pregnant or lactating women and women of reproductive age without adequate contraception were also excluded. The trial was performed in accordance with the Declaration of Helsinki and subsequent revisions were approved by the ethics committee at Tehran University of Medical Sciences (grant No.: 7844). Written informed consents were obtained before entry into the study. Intervention Fifty-one patients were randomly allocated (17 patients in each group) to risperidone (6 mg/day) or haloperidol 15 mg/day or clozapine (300 mg/day) for a 8-week and double blind study. The patients had a 1-week antipsychotic washout period before entering the study. During the washout period, the patients received benzodiazepine if necessary and lorazepam was the drug of choice. Patients were randomised to receive haloperidol or clozapine or risperidone in a 1:1:1 ratio using a computer-generated code. Three patients dropped out of the study (one patient from each group). Patients also received biperiden if they faced extrapyramidal symptoms. Patients were assessed by a psychiatrist at baseline and after 2, 4, 6 and 8 weeks after the start of medication. Outcome The principal measure of outcome was the PANSS that has been used in several studies in Iran (Akhondzadeh et al., 2006, 2007). PANSS total score was considered as primary efcacy and secondary efcacy parameters included the PANSS subscales. The raters used standardised instructions in the use of PANSS. The mean decrease in the PANSS score from baseline was used as the main outcome measure of patient response to treatment. Treatment response was dened as at least 50% improvement in the PANSS total score. Five millilitres of peripheral blood at baseline and week 8 was obtained by venous puncture with vacuum tubes without anticoagulants. Serum was separated by centrifugation and frozen to 708C until assay. Serum ADA activity was measured by a commercial assay kit (Diazyme Laboratories, USA) performed by Hitachi 902 auto-analyzer with precisions of Intra assay %CV < 4.5% and Inter assay %CV < 6.0%.
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Statistical analysis Statistical differences among three groups were analysed by one-way analysis of variance (ANOVA) followed by the Tukeys test for multiple comparison. Correlations of response to treatment with plasma level of ADA were analysed by the Kendall t-test. To compare the demographic data, the Student t-test and Chi-square were used. Results are presented as mean SD. Differences were considered signicant with p 0.05. RESULTS Sixty-eight patients were screened for the study and 51 patients were randomised to trial medication (17 patients in each group). Seventeen patients were excluded from the study. One patient from each group dropped out due to issue of substance abuse leaving 16 patients in each group who completed the trial (Figure 1). Signicant differences were not identied between patients randomly assigned to the groups in regards to basic demographic data including age, gender, marital status, smoking, level of education, mean duration of illness and liver function tests (Table 1). PANSS total score and its subscales at baseline and endpoint among three groups The mean SD scores of the three groups of patients on PANSS total score and its three subscales at week 0 are shown in Table 2. A one-way ANOVA revealed that there were no signicant differences among the three groups at week 0 in the PANSS total score and its subscales.

A signicant difference was observed on the PANSS total score and general psychopathology score among the three groups at week 8 ( p 0.02). A Tukey posthoc test revealed a signicant difference between clozapine group and the haloperidol group ( p < 0.01) in the general psychopathology scale at week 8 ( p 0.004). In addition, a Tukey post-hoc test revealed a signicant difference between the clozapine or the risperidone groups and the haloperidol group ( p < 0.01) in the total PANSS score at week 8 ( p < 0.05). The changes of total PANSS score at the endpoint compared with baseline were: 37.87 8.04 (mean SD), 48.43 6.31 and 43.62 7.58 in the haloperidol, clozapine and risperidone group, respectively. A signicant difference was observed on the changes of PANSS total score among the three groups ( p 0.0009). A Tukey post-hoc test revealed a signicant difference between clozapine group and the haloperidol group in the changes of PANSS total score ( p < 0.001). ADA level at baseline and endpoint among three groups ADA level of the three groups of patients at week 0 are shown in Figure 2. A one-way ANOVA revealed that there were no signicant differences among the three groups at week 0. ADA level increased signicantly in all three groups after 8 weeks of treatment compared to baseline ( p < 0.001). In addition, a signicant differences were observed at week 8 among the three group ( p < 0.001). A Tukey post-hoc test revealed a signicant ADA level difference between the clozapine group and the haloperidol group at week 8 ( p < 0.001).

Figure 1. Trial prole.

Copyright # 2011 John Wiley & Sons, Ltd.

Hum. Psychopharmacol Clin Exp 2011; 26: 120124. DOI: 10.1002/hup

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Table 1. Baseline data Haloperidol group Gender Age (mean SD) (year) Marital status Level of education Duration of illness (year) Type of schizophrenia Smoking AST ALT Alkaline phosphatase Male: 9, female: 7 33.00 10.11 Single: 13, married: 2, divorced: 1 Under high school diploma: 14, diploma: 2 7.26 4.29 Paranoid: 11; residual: 2; undifferentiated: 2 7 22.7 6.5 33.5 8.9 71.2 10.8 Clozapine group Male: 11, female: 5 37.12 9.39 Single: 12, married: 3, divorced: 1 Under high school diploma: 15, diploma: 1 7.77 5.85 Paranoid: 12; residual: 3; undifferentiated: 1 5 22.2 5.85 34.6 9.2 75.65 11.0 Risperidone group Male: 10, female: 6 32.06 5.80 Single: 11, married: 3, divorced: 2 Under high school diploma: 14, diploma: 2 7.95 6.01 Paranoid: 12; residual: 2; undifferentiated: 2 7 22.9 6.2 35.8 7.8 79.25 10.6 p ns ns ns ns ns ns ns ns ns ns

Table 2. Mean SD of the three protocols on the total Week 0 Positive subscale Haloperidol 28.50 3.14 Clozapine 30.37 3.93 Risperidone 28.12 4.19 Negative subscale Haloperidol 20.80 2.56 Clozapine 22.37 3.44 Risperidone 20.81 2.83 General psychopathology subscale Haloperidol 48.93 4.44 Clozapine 47.93 3.94 Risperidone 45.87 6.30 Total score Haloperidol 97.62 5.76 Clozapine 100.68 7.04 Risperidone 95.43 8.04 PANSS scores and its three subscales. Week 8 16.03 3.28 16.00 2.94 13.62 3.87 13.37 2.66 12.87 3.09 12.81 2.42 30.42 5.62 23.87 4.30 26.37 5.90 59.81 8.50 52.93 6.03 52.81 8.84

Correlation between response to treatment and ADA level In clozapine, risperidone and haloperidol groups 7 (43.75%), 4 (25%) and 1 (6.25%) patient responded to treatment. A positive correlation was observed between ADA level at week 8 and response to treatment in the clozapine group (r 0.46 and p 0.04), but such a correlation was not observed in the risperidone and haloperidol groups (r 0.20 and p 0.37; r 0.33 and p 0.14, respectively).

DISCUSSION Adenosine is a modulator of brain function uniquely positioned to integrate excitatory and inhibitory neurotransmission (Boison, 2008). The past few years have brought a wealth of new data fostering our understanding of how the adenosine system is involved in the pathogenesis of neurological diseases (Lara et al., 2006; Salimi et al., 2008). It has been proposed that a decit of adenosinergic activity could contribute to the pathophysiology of schizophrenia (Dutra et al., in press; Lara and Souza, 2000). The present study indicates that plasma levels of ADA in patients with chronic schizophrenia that received clozapine were signicantly higher than patients who received haloperidol. In addition, response to treatment was positively correlated with plasma levels of ADA only in the clozapine group. The increase in ADA level would theoretically be associated with a reduction of adenosine, which is against the hypothesis of a hypoadenosinergic state in schizophrenia. However, this could be a compensatory change that may somehow reect a readjustment of the system and this increase could mean an increased adenosine
Hum. Psychopharmacol Clin Exp 2011; 26: 120124. DOI: 10.1002/hup

Figure 2. Adenosine deaminase level among the three groups of patients at baseline and endpoint. p < 0.001.

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turnover depending on how much is produced and released in the brain. Adenosine deaminase activity is a new target of investigation of pharmacotherapy in patients with schizophrenia (Salimi et al., 2008). Brunstein et al. (2007) have recently reported that either altered adenosine metabolism is present in schizophrenic patients or adenosinergic activity is inuenced by treatment with antipsychotics, particularly clozapine. The results of the present study are in line with Brunstein et al.s (2007) report. It has also been reported that adenosine may be involved in the mechanism(s) of action responsible for clozapines superior antipsychotic efcacy. Lara et al. (2001) reported that chronic treatment with clozapine, but not with haloperidol, enhances ecto-50 -nucleotidase activity in the striatum, suggesting that purinergic activity may be particularly affected by clozapine treatment. Caffeine intake was not evaluated in this study although there is no evidence that caffeine can affect serum adenosine levels (Brunstein et al., 2007). An ecto-50 -nucleotidase hydrolyses AMP to adenosine. Extracellular adenosine can be taken up by the nucleoside transporter and inactivated by deamination to inosine by ADA or phosphorylated to AMP by adenosine kinase. These processes are mostly intracellular, but ADA is also found associated with cell membranes (Boison, 2008). In this study, we focused on peripheral adenosine metabolism by determining the activity of serum ADA in schizophrenic patients under antipsychotic monotherapy. We did not measure serum 50 -nucleotidase in this study and this is considered a limitation of the present trial. Although clozapine in the present slightly increased liver function tests but the differences were not signicant. In addition, Brunstein et al. (2007), reported in their study that liver function test is not involved in the effect of clozapine in the ADA level. CONCLUSION The results indicate an increased activity of the enzyme ADA in the serum of schizophrenic patients being treated with clozapine. This increase may be correlated with clozapines superior antipsychotic efcacy. CONFLICT OF INTEREST The authors have declared no conict of interest.
ACKNOWLEDGEMENTS This study was the post graduate thesis of Dr. Navid Honarbakhsh toward the Iranian board of psychiatry. This
Copyright # 2011 John Wiley & Sons, Ltd.

study was supported by a grant from Tehran University of Medical Sciences (Grant No: 7844) to Prof. Shahin Akhondzadeh. We declare no conict of interest.

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Hum. Psychopharmacol Clin Exp 2011; 26: 120124. DOI: 10.1002/hup

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