1) To investigate the relationship between NrCAM polymorphisms and methamphetamine abuse in an ethnically homogenous Korean population. 2) To further support our findings by investigating the association among NrCAM gene variants, certain personality traits, and addictive symptoms of methamphetamine abusers.
Thirty-seven male methamphetamine abusers (age=43.3±7.8) and30 non-users (16 men, 14 women; age=59.8±10.4) were recruited. Ten single nucleotide polymorphisms (SNPs) in the NrCAM gene were assayed to compare genotype distributions between the 2 groups. Personality characteristics were measured using the Temperament and Character Inventory (TCI) and the NEO Personality Inventory, Revised (NEO PI-R). Addictive symptoms were assessed using the Diagnostic Interview for Genetic Studies (DIGS) and reviews of the subject's medical records.
Among the 10 SNPs in the NrCAM gene, the frequency of the TA genotype at rs1990162 was significantly lower in methamphetamine abusers compared to non-users (p=0.042). In the 3 NrCAM gene SNPs (rs381318, rs2072546, and rs6954366), the distribution of genotypes and alleles were significantly associated with some traits in the TCI and NEO PI-R. Genotypes and alleles at 5 gene SNPs (rs2142325, rs381318, rs1269621, rs1269634, and rs1990162) were associated with certain addictive symptom dimensions in the patients.
These findings support the idea that NrCAM is associated with genetic susceptibility of methamphetamine abuse and is also associated with certain personality characteristics that may increase disturbed addictive behavior.
Drug abuse is a persistent global phenomenon, affecting almost every country in the world. High levels of drug dependence
cause detrimental societal effects such as low productivity, transmission of infectious diseases, family and social
disorders, increased crime, and excessive utilization of healthcare systems.
In South Korea, statistics show that methamphetamine is the most widely abused non-prescription drug, followed by cannabis and opiates.
Diverse and complex genetic influences have been shown to contribute to an individual's vulnerability toward substance abuse.
NrCAM is a gene that encodes neuronal cell adhesion molecule, which produces a receptor involved in nervous system development expressed in the central nervous system and located in the 7q region.
Personality traits are also predisposing factors toward addiction or an affinity toward drug usage. Drug addiction may be considered a convergence between the physiological effects of a drug and environmental factors such as one's individual personality.
In this study, genetic polymorphisms in the NrCAM gene in South Korean methamphetamine abusers were compared with those of non-user controls. We then analyzed the associations between NrCAM gene variants and personality traits along with the addictive characteristics of methamphetamine abusers.
The subjects in this study included inpatients at the Inpatient Addiction Unit of the Bugok National Hospital in Korea, which has an established protocol for the evaluation, treatment, and follow-up of patients with a diagnosis of drug abuse. We identified 37 patients that fulfilled the DSM-IV diagnostic criteria for methamphetamine abuse or dependence.
Semi-structured interviews were performed by a psychiatrist to gather basic information such as age, marital status, and employment. Parameters for addictive behavior included the total duration of drug usage, typical dosage sizes, and the number of times arrested by police for methamphetamine-related offenses.
Our control group consisted of 30 normal healthy volunteers with no known history of psychiatric or substance use disorders. They were also checked for an absence of major relevant medical disorders such as thyroid disorder, diabetes mellitus, etc.
Samples sizes for treatment and control were determined by power computations; for the required power 0.80 (i.e., 1-β=0.80), we allowed the probability of type I error be 0.05 and Δ/σ=1, where Δ=p1-p2 under the alternative hypothesis, and σ was the standard deviation of the measurement in each patient.
There are 10 single nucleotide polymorphisms (SNPs) present in NrCAM genes, which are functionally relevant to neuropsychiatric disorders, on chromosome 7. These include 4 exonic SNPs (rs1269621, rs2072546, rs381318, and rs6958498), 4 intronic SNPs (rs1990162, rs2142325, rs6954366, and rs722519), and 2 putative regulatory SNPs (rs1269634 and rs17155059). Genotyping was carried out by laboratory personnel who were blinded to phenotype data, as described previously by Lee et al.
We assessed personality characteristics using both Temperament and Character Inventory (TCI) and NEO Personality Inventory, Revised (NEO PI-R).
The TCI is an inventory for evaluating distinct personality traits. It is closely related to the Tridimensional Personality Questionnaire (TPQ) and also derives elements from Zuckerman's and Eysenck's Dimensions of Personality. The TCI addresses 7 dimensions of personality traits with 4 so-called "temperaments" (Novelty Seeking, Harm Avoidance, Reward Dependence, and Persistence) and 3 "characters" (Self-Directedness, Cooperativeness, and Self-Transcendence). This test compliments the NEO PI-R, which is a psychological personality inventory assessment developed by Paul T. Costa Jr. and Robert R. McCrae for use with adult men and women. The NEO PI-R includes 5 major domains of personality (Neuroticism, Extraversion, Openness to Experience, Agreeableness, and Conscientiousness), as well as 6 sub-scales that further define each domain.
The genotype distribution was calculated for each individual polymorphism evaluated using Hardy-Weinberg equilibrium to confirm data quality.
The association between the case-control status and each individual SNP was measured using the odds ratio and its corresponding 95% confidence interval using logistic regression models (adjusting for sex and age as covariates). Case-control analysis was conducted using 4 models. All analyses were performed assuming dominant, recessive, additive, or allelic effects for each polymorphism. In the dominant model, both the heterozygous variant and the rare homozygous variant were combined; in the recessive model, the variant was defined as the rare homozygous genotype only; in the additive model, each genotype variant had the same effect; and in the allele model, the rare allele variant had an effect. The likelihood ratio test was used to test the effect of each SNP at a 5% significance level. All data were processed and analyzed using R software, version 2.6.2. Associations between personality dimensions or addictive symptoms and individual SNPs in abusers were then evaluated using Analysis Of Variance (ANOVA) with the SPSS system, version 11.5.
The demographics and clinical characteristics of methamphetamine abuse subjects are indicated in
The mean duration of methamphetamine abuse was 14.35±1.10 years, while the mean total dose of methamphetamine that had been consumed during the period of drug abuse was calculated to be 326.46±40.94 g, estimated solely on the basis of the patient's statements and uncorrected for drug purity. The largest total dose of methamphetamine intake was estimated to be 1000 g.
The mean number of arrests for methamphetamine-related offenses was 3.8±0.4, with the highest number being 10.
Genotype distributions at 10 NrCAM SNPs in methamphetamine abusers and non-user controls are compared in
In 3 NrCAM gene SNPs (rs381318, rs2072546, and rs 6954366), the distributions of particular genotypes or alleles were significantly associated with traits of the TCI and the NEO PI-R. Of these, rs6954366 was significantly associated with persistence (p=0.033) and self-transcendence (p=0.035) in the TCI, as well as extraversion (p=0.012) and friendliness (p=0.039) in the NEO PI-R. Significant associations were also found between rs381318 and novelty seeking (p=0.020), as well as between rs2072546 and reward dependence (p=0.420) (
In addition, genotypes or alleles at 5 of the SNPs (rs2142325, rs381318, rs1269621, rs1990162, and rs1269634) were associated with certain addictive characteristics. Of these, rs2142325 was associated with 3 addictive characteristics, including duration of abuse (p=0.025), total dose (p=0.046), and the number of arrests (p=0.038); rs1269634 was significantly associated with total dose (p=0.049) and the number of arrests (p= 0.023); and rs381318 (p=0.053), rs1269621 (p=0.013), and rs 1990162 (p=0.056) were significantly associated with the number of arrests. One SNP (rs381318) was also associated with both personality and addictive symptom dimensions (
For multiple comparisons, we used the false discovery rate (FDR) instead of the family-wise error rate (FWER), such as Bonferroni correction. FWER controls the probability of at least 1 false positive, and FDR controls the expected value of false positives in m simultaneous hypotheses. To be specific, we let
The NrCAM gene encodes a neuronal cell adhesion molecule with multiple immunoglobulin-like C2-type domains and fibronectin type III domains.
We assayed 10 NrCAM gene SNPs to determine the association between these changes and methamphetamine abuse in 37 male methamphetamine abusers. The frequency of the TA genotype in 1 SNP (rs1990162) in methamphetamine abusers was significantly lower as compared to that in the controls (p=0.042), while no significant differences were found at any of the other 9 SNPs present in the NrCAM gene. These observations lend some support to our hypothesis that NrCAM is a methamphetamine abuse-associated gene with variants that affect substance abuse predisposition in human subjects. However, our results could also be considered insignificant, if more stringently interpreted. However, several issues need to be addressed in the future. Validation studies on the 10 SNPs of NrCAM need to be performed to compliment our limited information on the association between the NrCAM gene and methamphetamine abuse. Ethnic differences also need to be taken into consideration. The phases of addiction vulnerability association of the 30 NrCAM 520Pro/Ala710c/þ1343a 3'UTR haplotype was similar in 2 previous European-American substance abuser/control comparisons but this was found to be opposite in the study on African-American subjects.
Molecular heterosis also deserves a mention; this occurs when heterozygous genotypes at a locus exhibit significantly larger (positive heterosis) or smaller (negative heterosis) effects on a quantitative or dichotomous trait than homozygous genotypes. There is accumulating evidence that molecular heterosis is common in humans and may occur in up to 50% of all genetic associations. Three explanations for molecular heterosis are proposed. The first is based on an inverted U-shaped response curve in which either too little or too much gene expression is deleterious; in this case, optimal gene expression would be associated with heterozygous genotypes. The second proposes an independent third factor causing a hidden stratification of the sample such that in 1 set of subjects homozygosity is associated with the highest phenotype score, while in the other set, homozygosity is associated with the highest phenotype score. The third explanation suggests greater fitness in heterozygotes because they show a broader range of gene expression than homozygotes. Allele-based linkage techniques usually neglect heterotic associations. Because up to 50% of association studies show a heterosis effect, this can significantly diminish the power of family-based linkage and association studies. This point could be complemented by 4 different genetic model analyses (dominant, recessive, additive, and allelic), such as those shown in
Several previous studies have reported personality differences between addicts and normal subjects.
Our finding of an association between the NrCAM gene and addictive symptoms in methamphetamine abusers supports the model that addiction vulnerability may be strongly influenced by a genetic predisposition.
After investigation of the relationship between the NrCAM gene and addictive symptoms, 5 SNPs (rs2142325, rs381318, rs1269621, rs1269634, and rs1990162) were found to be associated with addictive symptoms. In particular, rs2142325 was significantly associated with all defined addiction parameters (duration of drug use, total dose, and number of arrests). rs1269634 was also significantly associated with total dose and number of arrests, while rs381318, rs1269621, and rs1990162 were only significantly associated with the number of arrests. Although we found some associations between the NrCAM gene and addictive symptoms, the clinical significance of these findings needs to be confirmed in future studies.
This study has several limitations. First, given that we used a small sample size, the statistical power of our analyses was reduced, and thus, further studies, using larger study cohorts than those used in our study, should be undertaken. The second limitation was that demographic characteristics such as gender and age were not matched between methamphetamine abusers and controls. The third is that the 10 SNPs of NrCAM gene were not validated by assaying. In addition, the scale for measures of personality dimension and the criteria of addictive symptoms were broad, and could be more clearly refined. Finally, and most importantly, unlike other psychiatric disorders that are primarily caused by underlying etiologic factors, addiction is a disease that occurs only in people who have been exposed to a substance. Therefore, it is very difficult, but important to control for both environmental factors that are crucial in the onset of addiction, as well as genetic factors (i.e., an individual who is not addicted despite having been exposed to methamphetamine). Thus, the difference between patients and controls could simply be the result of different levels of exposure to methamphetamine; however, it is important to note that genetic factors can also dictate patterns of behavior that result in greater exposure to risk stimuli.
Future research should place emphasis on denser genotyping panels, sequencing, and functional studies. These may help to confirm whether NrCAM or other genes in linkage disequilibrium with this candidate, contain causal variants that influence the pathogenesis of addiction in Korean and other ethnic populations.
This study was supported by Biomedical Research Institute Grant (2009-30), Pusan National University Hospital.
Demographic and clinical characteristics in methamphetamine abusers and controls
NrCAM genotype frequencies in Korean methamphetamine abusers and controls
Freq: frequency, MA: methamphetamine abuser, OR: odds ratio, CI: confidence intervals, LCL: lower confidence limit, UCL: upper confidence limit, INF: infinite, NrCAM: neuronal cell adhesion molecule
Association analysis between NrCAM gene SNP genotypes and alleles and personality traits in methamphetamine abusers
TCI: Temperament & Character Inventory, NEO PI-R: Revised NEO Personality Inventory, NS: novelty seeking, HA: harm avoidance, RD: reward dependence, Per: persistence, SD: self-directiveness, Coo: cooperativeness, ST: self-transcendence, Ex: extraversion, A: agreeableness, C: conscientiousness, Ne: neuroticism, O: openness to experience, ns: non-significant, NrCAM: neuronal cell adhesion molecule, SNP: single nucleotide polymorphism
Association analysis between NrCAM gene SNP genotypes and alleles and addictive symptoms in methamphetamine abusers
DU: duration of use, TD: total dose, NA: number for arrests by police, ns: non-significant, NrCAM: neuronal cell adhesion molecule, SNP: single nucleotide polymorphism