Lack of association between the T→C 267 serotonin 5-HT6 receptor gene (HTR6) polymorphism and prediction of response to clozapine in schizophrenia
Introduction
Response to the atypical antipsychotic, clozapine, is highly variable among individuals who have treatment-refractory and/or intolerant schizophrenia (Bleehen, 1993). The ability to predict response to clozapine would be important from a clinical perspective: predicted responders to this drug could be treated preferentially, whereas those less likely to respond could be offered other possible treatments. However, finding specific and consistent predictors of response to clozapine has proven to be a difficult task [reviewed by Meltzer (1996)]. Clozapine's wide spectrum of unique clinical effects for the treatment of schizophrenia [reviewed by Meltzer (1995)] suggests that many factors, both biological and environmental, are involved in the phenotype of response.
Pharmacogenetics seeks to identify genetic variation (polymorphism) in or near the coding region of genes that encode protein structures involved in the action of a drug, and correlate these with variability in the clinical profile, e.g. its pattern of response and/or side effects. This paradigm can be applied to predict the variable responsiveness to clozapine. Recent advances in molecular genetics allow for the identification of genes involved in complex traits (Bennett et al., 1995). These techniques are well suited for examining the trait of clozapine response, as it is likely complex and multifactorial in nature.
Clozapine has a high affinity for many receptors from multiple neurotransmitter systems. All of these pharmacological interactions have been implicated in its mechanism of action [reviewed by Ashby and Wang (1996)]. In particular, clozapine has moderate to high affinity for at least five subtypes of serotonin receptors (Meltzer, 1994, Meltzer, 1999). The current study examined polymorphisms in the following candidate receptor genes from the serotonin system: 5-HT1A (HTR1A), 5-HT6 (HTR6), and 5-HT7 (HTR7). There are several lines of evidence implicating a role for these serotonin receptor proteins in the mechanism of action of clozapine.
Newman-Tancredi et al. (1996) have demonstrated that clozapine acts as a partial agonist at 5-HT1A receptors. The effect of clozapine to increase extracellular dopamine levels in rat prefrontal cortex is believed to be important in its ability to improve positive and negative symptoms (Kuroki et al. 1998). This effect is blocked by pretreatment with the 5-HT1A antagonist, WAY 100635 (Rollema et al., 1997; Ichikawa and Meltzer, unpublished data), suggesting it is mediated by the partial agonist effect of clozapine. The combination of a 5-HT1A agonist, 8-OH-DPAT, with haloperidol produces a clozapine-like pattern of activation of the early intermediate gene, c-fos, in rat prefrontal cortex, nucleus accumbens, and striatum (Tremblay et al., 1998). The ability of clozapine to decrease serotonin efflux in the rat ventral hippocampus also is partially dependent upon its 5-HT1A agonist properties (Bengtsson et al., 1998). Thus, there is significant preclinical evidence to suggest 5-HT1A agonism is a meaningful component of the action of clozapine.
5-HT1A agonism has also been shown to prevent and ameliorate neuroleptic-induced catalepsy in rodents (Broekkamp et al., 1988, Hicks, 1990, Invernizzi et al., 1988, Neal-Beliveau et al., 1993, Wadenberg, 1992, Wadenberg and Ahlenius, 1991). This effect has also been observed in primate models of extrapyramidal symptoms (EPS) (Casey, 1992, Casey, 1994, Liebman et al., 1989). For a review, refer to Kapur and Remington (1996).
From a neuroanatomical perspective, 5-HT1A receptor sites are localized in many areas of the brain, including the hippocampus, the septum, the amygdaloid, the neocortex, the hypothalamus, and the raphe nuclei, particularly the dorsal raphe [reviewed by Hoyer et al. (1994)]. Many of these areas have been implicated as neuroanatomical substrates of schizophrenia. Based on this information, the 5-HT1A receptor gene (HTR1A) is an important candidate to test with respect to clozapine response.
HTR1A is located on human chromosome 5q and a cytosine to thymidine (C→T) polymorphism in this gene, which causes an amino acid substitution of proline to leucine at position 16 of the 5-HT1A receptor protein (pro16leu), has been identified in a Japanese sample (Inayama et al., 1995). This polymorphism was genotyped in our group of clozapine-treated patients.
The human 5-HT6 receptor gene (HTR6) has been cloned and localized to chromosome 1p (Kohen et al., 1996). A silent thymidine to cytosine polymorphism at position 267 (T→C 267), within the first exon of HTR6, has been identified (Kohen et al., 1996). Given that a weak positive association has been reported between this candidate receptor gene variant and response to clozapine in a Chinese sample (Yu et al., 1999), we have evaluated this polymorphism in our sample of clozapine-treated patients.
mRNA for the 5-HT6 receptor has been found in several human brain regions, including the caudate nucleus, the hippocampus, and the amygdala; lower levels were observed in the thalamus, subthalamic nuclei, and the substantia nigra (Kohen et al., 1996). Several of these brain areas have been implicated in the pathophysiology of schizophrenia. Clozapine and several other atypical antipsychotic agents are antagonists at, and have demonstrated high affinity for, 5-HT6 receptors (Glatt et al., 1995, Kohen et al., 1996, Roth et al., 1994). Approximately, 40% of clozapine binding sites in the rat brain pharmacologically resemble the 5-HT6 receptor (Glatt et al., 1995), and based on relative affinities of clozapine for D2 and 5-HT2A receptors, 5-HT6 receptors should be highly occupied at clinically relevant doses of clozapine (Kohen et al., 1996). This suggests that 5-HT6 receptors may be important in the mechanism of action of clozapine and other atypical antipsychotic agents.
Clozapine is an antagonist at, and possesses high affinity for, 5-HT7 receptors (Roth et al., 1994). mRNA and in situ hybridization studies have demonstrated that 5-HT7 receptors may be expressed in the brain in the hypothalamus, the anteroventral and paraventricular thalamic nuclei, and the hippocampus [reviewed by Eglen et al. (1997)]. Expression of mRNA in these midline, thalamic, and limbic structures suggests a role of 5-HT7 in the regulation of emotion (Eglen et al., 1997) and some of these structures may be involved in the pathophysiology of schizophrenia. There is some evidence which suggests that the 5-HT7 receptor subtype may be involved in the regulation of circadian rhythm phase shifts (Lovenberg et al., 1993, Ying and Rusak, 1997). This role of the 5-HT7 receptor may be interesting, considering that some people afflicted with schizophrenia experience a reversal of the sleep–wake cycle accompanied by severe insomnia (Benca, 1996).
The gene encoding the 5-HT7 receptor has been localized to human chromosome 10q (Gelernter et al., 1995). A cytosine to thymidine (C→T) polymorphism, leading to a proline to leucine amino acid substitution at position 279 (pro279leu) in the third intracellular loop of the receptor protein, was genotyped in our group of clozapine-treated patients (Erdmann et al., 1996, Pesonen et al., 1998).
Section snippets
Clinical sample
Clinical data from patients with DSM-III-R diagnoses of schizophrenia (treatment-refractory or intolerant; Kane et al., 1988) were obtained from: Case Western Reserve University in Cleveland (HY Meltzer, n=105); Hillside Hospital in Long Island (JA Lieberman, n=65); and the Bronx Veteran's Administrative Medical Center in New York (S Sevy, n=16). Treatment response was evaluated at 6 months or more using the following criteria: (1) ≥20% reduction in Brief Psychiatric Rating Scale (BPRS) total
Results
There were no statistically significant differences in ethnicity and response rate between the patients from the three clinical sites. There was a significant difference observed between the mean age of the patients from the three clinical sites [F(2, 181)=3.43, P=0.007]; using Tukey's HSD test for post hoc comparisons, the mean ages of the Meltzer and Lieberman samples were significantly lower than that of the Sevy sample (SS). There was also a significant difference in the proportion of males
Discussion
We have found no evidence of an association between the T→C 267 polymorphism in the 5-HT6 receptor gene and clinical response to clozapine in our sample of schizophrenia patients. This is contrary to the findings of Yu et al. (1999) who observed that patients with the homozygous genotype T/T 267 were more likely to be responders to clozapine than those with either the C/T 267 or C/C 267 genotypes (total n=99). This discrepancy may be due to methodological differences between the studies. In
Acknowledgements
MM was supported by a Department of Psychiatry, Faculty of Medicine, University of Toronto Summer Scholarship. This work was supported by a Medical Research Council of Canada (MT15007) grant and a NARSAD Independent Investigator Award to JLK.
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2016, European NeuropsychopharmacologyCitation Excerpt :However, no significant heterogeneity (p<0.05) among studies was reported in our meta-analyses as well as no publication bias, according to Egger’s test results. Other SNPs out of the seven subtypes of serotonin receptors have been also analyzed in less than 3 studies considering CLZ response providing inconsistent results (HTR2A polymorphisms rs6305, rs1805055 (Nothen et al., 1995; Arranz, 2000b), HTR2C rs521018, rs1292371, rs5988072, rs12833104, rs498177, rs498207 (Arranz et al., 2000b; Bai et al., 2011; Schumacher et al., 2000; Sicard et al., 2010), HTR3A rs1176713, rs14396, rs1596, rs2276302, rs1150226 (Arranz et al., 2000b; Souza et al., 2010a; Gutierrez, 2002), on HTR3B gene (rs1176744, rs3782025, rs2276307, rs3758987, CA repeat) (Souza et al., 2010a, Gutierrez, 2002), 5-HT1A rs1800041 (Masellis et al., 2001), 5-HT5A 12 A/T (Arranz et al., 2000b; Birkett et al., 2000), 5-HT5A 19 G/C (Arranz et al., 2000b; Birkett et al., 2000 ), 5-HT6 267CT(Masellis et al., 2001; Yu et al., 1999). Studies involving other genetic variants found interesting associations but have to be replicated: on COMT, Met carriers of the Val158Met (472GA) rs4680 showed better improvement (Woodward et al., 2007), on NMDA Receptor subunit 2B GRIN2B C2664T, higher CLZ mean dose was reported with CC genotype in two studies (Chiu et al., 2003; Hong et al., 2001), on GDNF family receptor alpha 2 (Souza et al., 2010b); on Neurexin-1 NRXN1 (Lett et al., 2011; Souza et al., 2010a).
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