Tian-Dao Wang, Ming Pan, Xiang-Lai Liu, Qin-Wei Xu, Yu Zhou

Keywords:Schizophrenia Olanzapine Melanocortin 4 receptor Glucose and Lipid metabolism

ABSTRACT Objective: To investigate the effect of Melanocortin four receptor, MC4R rs489693 polymorphism on glucose and Lipid metabolism in schizophrenia patients treated with Olanzapine for 12 weeks. Methods: 171 patients with schizophrenia were divided into AA Group (N = 12) , AC group (N = 59) and CC Group (N = 100) according to the polymorphism of MC4R gene at 489693 locus detected by DNA sequencing. Blood Glucose and lipid levels were measured before and 12 weeks after treatment, the differences of variables among the 3 groups were compared, and the incidence of glucose and lipid abnormalities after treatment was statistically analyzed. Results: After 12 weeks of treatment, the net increase of blood glucose in AA group was greater than that in CC group(P<0.05), and the net increase of cholesterol and triglyceride in AA group was greater than that in AC group and CC group(all P<0.05), and the incidence of Blood Glucose and at least one dyslipidemia in AA Group was higher than that in AC and CC group (all P< 0.01) . Conclusion: The rs489693 gene polymorphism of MC4R gene is related to the disorder of glucose and lipid metabolism in schizophrenia treated with olanzapine.

1. Introduction

Schizophrenia (SCZ) is an important part of current mental health, and its harmfulness has been widely recognized. It has been reported that the life expectancy of SCZ patients is 10-20 years lower than that of the general population, about two thirds of deaths were due to cardio-cerebrovascular disease[1], suggesting that how to avoid or reduce the level of glucose and Lipid Disorders and other adverse reactions have long-term significance for the quality of life while the mental patients are receiving symptomatic treatment .In the prophase of this study, the Abnormal Status of Glucose and Lipid in long-term psychiatric inpatients in our hospital was analyzed. The incidence of hyperglycemia and hyperlipemia in our hospital were 12.94% and 53.74% , respectively, which were higher than the residents in this area[2]. It not only affect the quality of life of patients, but also affect the long-term prognosis. In order to further explore the genetic factors bisides environmental factors (lifestyle) that lead to abnormal glycolipid metabolism in mental patients,we selected the most extensive type of schizophrenia in our hospital, who were treated with second-generation antipsychotic Olanzapine which is the more common treatment for the disease. Then we analyze the polymorphism RS489693 of the drug-toxic gene melanocortin 4 receptor (MC4R) , which has great influence on the metabolic index of patients and explore the relationship between MC4R and abnormal metabolism of glucose and lipid in mental patients from a pharmacogenetic point of view. Finally our study can provide the basis which can implement individual medication plan and evaluate medication safety for clinicians .

2. Objects and methods

2.1 Objects

171 psychiatric patients who were hospitalized in Anning Hospital of Hainan Province from October 2018 to December 2019 were selected. Criteria for admission: ①Han patients who met the diagnostic criteria of the 4th edition of the Chinese classification and diagnostic criteria for mental disorders; ②patients who did not take or stopped taking any antipsychotic before admission and underwent drug washout period for 2 weeks; ③Olanzapine was given alone for one week and its minimum concentration was measured to meet the effective treatment concentration and repeated every two weeks to ensure compliance. ④cultural, social and educational background were sufficient to understand informed consent and the content of the study. Criteria for rejection: ①diagnosis other than schizophrenia; ②significant cardio-cerebrovascular disease, diabetes, and other serious physical disorders; ③history of chronic alcohol or other drug dependence and abuse;④pregnancy;⑤Plasma trough concentration levels of Olanzapine were not maintained effectively during the study.

2.2 Method

2.2.1 General Information Collection

The questionnaire was compiled to collect the patients'name, sex, age, nationality, course of disease, family history and smoking history. The polymorphism of MC4R gene RS489693 were detected by DNA sequencing and divided into CC group (100 cases) , AC group (59 cases) and AA group (12 cases) , including 139 males and 32 females, with an average age of 36.19±14.59 years.

2.2.2 Laboratory examination

Blood samples were collected on an empty stomach at the beginning of drug therapy and 12 weeks later. Serum total cholesterol(TC), triglyceride(TG), high density lipoprotein cholesterol (HDL-C), low density lipoprotein cholesterol(LDL-C), Fasting Glucose(GLU) were measured by HITACHI-7600 automatic biochemical analyzer. The plasma trough concentration of Olanzapine was determined by High-performance Liquid Chromatography 2701 to ensure effective treatment after one week of steady-state administration, and the treatment concentration was continuously monitored every 2 weeks.

2.2.3 Genotype Detection

Oral Mucosal epithelial cells were taken from patients with sterile cotton swabs and were examined by Shanghai Kangli Medical Laboratory. The samples were analyzed by Matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOFMS) using polymerase chain reaction and restriction fragment length polymorphism (RFLP) technique and Mas-sArray DNA time of flight mass spectrometry (MS) system, the genotype of RS489693 of MC4R was detected.

Table 1 General data analysis of different genomes

2.2.4 Statistical method

Excel was used for data entry and spss22.0 was used for statistical analysis. Hardy-weinberg equilibrium test was used to analyze the population representativeness of the selected samples, chi-square test or Fisher's exact test was used for the general count data, and One-Way ANOVA was used for the measurement data, expressed as x±s; used covariance analysis to analyze the differences of glucose and lipid levels between different groups, and used Bonferroni correction method to make multiple comparisons after the event for variables with significant differences, so as to determine the specific differences between groups; after treatment, the incidence of abnormal blood glucose and the incidence of at least one abnormal blood lipid in different groups were analyzed by Chi-square test or Fisher's exact test. P<0.05 was statistically significant.

3. Result

3.1 General data analysis of different genomes

There were no significant differences in sex, age, course of disease, family history and smoking history among the 3 groups (all P>0.05) , see Table 1. The distribution frequency of RS489693 gene was in accordance with Hardy-Weinberg equilibrium test (χ2=0.643，P =0.72) .

3.2 Analysis of Glucose and Lipid levels in patients after treatment

After 12 weeks of treatment, the course of disease, sex, age and Olanzapine concentration were used as covariates to analyze the net increase of glucose and lipid levels before and after treatment.There were significant differences in the net increment of Blood Glucose, cholesterol and triglyceride among the three groups (all P<0.05), the rest had no statistical difference; the Bonferroni correction method was used for multiple comparison, the net increase of Blood Glucose in AA group was higher than that in CC group (P=0.022) , the net increase of cholesterol in AA group was higher than that in AC group and CC group (P=0.031, P=0.011) , and the net increase of triglyceride in AA group was higher than that in AC group and CC group (P=0.008, P=0.016) , the differences were statistically significant, as shown in Table 2.

3.3 Analysis of the incidence of abnormal glucose and lipid in patients after treatment

After 12 weeks of treatment, the incidence of abnormal blood glucose and blood lipid was shown in table 3. Results in the abnormal rate of blood glucose and at least one abnormal rate of blood lipid, the AA group was higher than the other two groups (P<0.01) , the difference was statistically significant.

4. Discussion

Antipsychotic is currently the most effective treatment for schizophrenia. However, with the widespread use of second generation Atypical antipsychotic (SGA) in the treatment of schizophrenia, compared with the traditional antipsychotics, patients are more susceptible to adverse drug reactions[3], such as glycolipid metabolism disorders, and studies have shown that there are individual differences in adverse drug reactions between patients and that these adverse drug reactions are closely related to genetic factors, therefore, we can help clinicians better implement individualized medication by exploring these genetic factors[4].MC4R is an obesity related gene[5], and the association between RS489693 polymorphism and Metabolic Markers is thought to be common , and the AA genotype of MC4R is particularly associated with increased body mass index, waistline and other obesity markers in drug treated patients[6].Obesity causes the disorder of Glucose and Lipid metabolism in patients, then induces cardio-cerebrovascular disease, leads to the occurrence of Metabolic Syndrome (MS)[7].Malhotra and other researchers analyzed the whole genome of psychiatric children exposed to SGA for the first time, which also suggested that MC4R was involved in SGA induced weight gain and related Metabolic Disorders[8].

In this study, we selected the patients who were treated with olanzapine, the second generation antipsychotic drug commonly in our hospital as the study object to analyze the occurrence of glycolipid metabolism disorder with different alleles of MC4R rs489693. The results showed that after treatment, the net increase of blood glucose in homozygous AA carriers was higher than that in CC carriers, while the increase of cholesterol and triglyceride in homozygous A allele was more significant than that in C allele carriers, and the probability of occurrence of hyperglycemia and hyperlipidemia (at least one increase) in AA carriers was higher than that in the other two groups,which was basically consistent with the expected results of this study.The reason may be the rs489693 polymorphism which affects the regulatory function of MC4R by the change of MC4R expression,also the use of blood glucose and insulin sensitivity[9].In addition, olanzapine itself has the effect ofinfluencing the insulin secretion of islet β cells and binding to the glucose transporter protein during use procwess [10], thus causing the increase of blood glucose, cholesterol and triglyceride levels. It is predicted that with the development of the disease and the increase of drug treatment time, the risk of metabolic syndrome in patients may be greater, which is also proved by some domestic related studies[11,12].

Table 2 Analysis of Glucose and Lipid levels in patients with different genomes before and after treatment

Table 3 Analysis of the incidence of abnormal blood glucose and lipid after treatment

The disadvantage of this study is that the CYP1A2 of drug metabolism gene is either extensive or super-fast, and the sample of slow metabolism type can not be collected for multi-angle analysis. In addition, it may be more convincing to enlarge the sample size of AA type subjects and carry out MC4R Gene multi-locus analysis.However, from the domestic studyies about the MC4R polymorphism , Olanzapine drug treatment caused by schizophrenia patients with glycolipid metabolic disorders [13,14],which could explain that the probability of MS was not to be underestimated during the long-term drug treatment in patients with schizophrenia.It was extremely disadvantageous to the health and quality of life of patients. In order to reduce the occurrence of such problems, clinicians were advised to treat patients with the use of genotyping to predict more accurately the adverse drug reactions that may occur in patients ,targeting drug selection, monitoring individual drug treatment concentrations during treatment, avoiding side effects from high concentrations of the drug.In addition,Olanzapine may also be used in combination with Metformin or Aripiprazole to optimize treatment regimens in patients with schizophrenia[15,16], especially those with the AA genotype, which can reduce the incidence of adverse drug reactions, improve the safety of drug use.