Tamara L. Wall

Genomic screen for loci associated with alcohol dependence in Mission Indians

Alcohol dependence is a leading cause of morbidity and mortality in Native Americans, yet biological factors underlying the disorder in this ethnic group remain illusive. This studys aims were to map susceptibility loci for DSM‐III‐R alcohol dependence and two narrower alcohol‐related phenotypes in Mission Indian families. Each participant gave a blood sample and completed an interview using the Semi‐Structured Assessment for the Genetics of Alcoholism (SSAGA) that was used to make alcohol dependence diagnoses and the narrower phenotypes of withdrawal, and drinking severity. Genotypes were determined for a panel 791 microsatellite polymorphisms. Analyses of multipoint variance component LOD scores for the dichotomous DSM‐III‐R phenotype revealed no peak LOD scores that exceeded 2.0 at any chromosome location. Two chromosomes, 4 and 12, had peak LOD scores that exceeded 2 for the alcohol use severity phenotype and three chromosomes 6, 15, 16 were found to have peaks with LOD scores that exceeded 2 for the withdrawal phenotype. Evidence for linkage to chromosomes 4 and 15, and 16 have been reported previously for alcohol related phenotypes whereas no evidence has as yet been reported for chromosomes 6 and 12. Combined linkage and association analysis suggest that alcohol dehydrogenase 1B gene polymorphisms are partially responsible for the linkage result on chromosome 4 in this population. These results corroborate the importance of several chromosomal regions highlighted in prior segregation studies in alcoholism and further identify new regions of the genome that may be unique to either the restricted phenotypes evaluated or this population of Mission Indians.

Alcohol Metabolism in Asian-American Men with Genetic Polymorphisms of Aldehyde Dehydrogenase

Epidemiologic studies have found that rates of alcohol use and alcoholism in persons of Asian descent are lower than rates in other ethnic groups. One possible reason is that about half of certain Asians, including Chinese, Japanese, and Korean persons, have a deficiency of the low-Km mitochondrial aldehyde dehydrogenase (ALDH2) isoenzyme, which is responsible for metabolizing acetaldehyde. A deficiency of ALDH2 results from inheritance of the mutant ALDH2*2 allele, a dominant mutation that exerts its effect both by reducing enzyme activity and increasing the turnover of this activity [1, 2]. After ingestion of alcohol, the faces of Asians with one or both alleles of ALDH2*2 become visibly flushed. Asians who are homozygous for ALDH2*1 generally lack visible alcohol-induced flushing or experience only a mild flush response. The dominance of the ALDH2 mutation, however, does not seem to be complete; phenotypic differences are associated with the three ALDH2 genotypes. Asians who are homozygous for ALDH2*2 drink very little alcohol [3], and no studies have found alcoholic persons with this genotype [4-8]. Asians who are heterozygous for ALDH2*2 drink less alcohol and are also less likely to be alcoholic compared with Asians with ALDH2*1 alleles, but they are not fully protected from alcoholism. Approximately 12% of alcoholic Asians have the ALDH2*1/2*2 genotype [5]. In the context of alcoholism or lower alcohol intake, Asian persons who are heterozygous for ALDH2*2 may be more vulnerable to alcohol-associated conditions, including liver disease [6-9], asthma [9], and esophageal cancer [10]. The three ALDH2 genotypes are also associated with variability in response to alcohol [11]. Asians who are homozygous for ALDH2*2 are very sensitive to alcohol and have tachycardia, hypotension, and vomiting after ingesting a moderate amount of alcohol. Asians who are heterozygous for ALDH2*2 are more sensitive to alcohol than Asians with ALDH2*1 alleles, although the response of the former is not necessarily aversive. Among Asians with an ALDH2 deficiency, differences in sensitivity to alcohol may be mediated by differences in alcohol metabolism, slower elimination of alcohol, or accumulation of acetaldehyde in the blood [12]. Some studies [13-17] have measured blood levels of alcohol or acetaldehyde after ingestion of alcohol in Asians who were known to have ALDH2 genotypes, but these studies had an inadequate sample size, did not include a placebo control, or did not control for use of alcohol and cigarettes (which can alter alcohol metabolism). We measured blood levels of alcohol and acetaldehyde after ingestion of alcoholic or placebo beverages in Asian-American men who underwent genotyping at the ALDH2 locus. Particular attention was given to matching the groups for age, height, weight, history of alcohol use, and history of smoking. Methods Asian-American men 21 to 25 years of age were recruited from advertisements in university newspapers for our randomized, double-blind, crossover study. They completed a questionnaire that solicited information on demographic characteristics; patterns of and problems with alcohol and drug use; and family history of alcohol, drug, and psychiatric problems. We excluded persons who completely abstained from alcohol, persons who had consumed more than 60 standard alcoholic drinks per month during the previous 6 months, and persons who reported that either biological parent was not of Chinese, Japanese, or Korean descent. Thirty-five men who did not have a personal or family history of alcohol dependence and who had no evidence of other substance dependence, major psychiatric disorders, or medical disorders gave informed consent to participate in two test sessions. The study was approved by the institutional review board at the Scripps Clinic and Research Foundation. Participants were asked to refrain from using alcohol, cigarettes, and other drugs (including aspirin, nonsteroidal anti-inflammatory agents, and anti-histamines) that might alter alcohol metabolism for 3 days before testing. On test days, each participant arrived at the clinic at 7:30 a.m. after an overnight fast. He then ate a low-fat breakfast (two slices of dry toast and juice), and an indwelling heparin lock was inserted for drawing blood. At the first session, blood was drawn and genotyping at the ALDH2 locus was done by using polymerase chain reaction of DNA and allele-specific oligonucleotide probes [1]. At 9:00 a.m., each participant was given a placebo beverage (3 mL of 95% alcohol in a reservoir on top of noncaffeinated, sugar-free soda) or 0.75 mL of 95% alcohol (0.56 g/kg of body weight) as a 20%-by-volume solution in the same mixer. The alcohol and placebo were ingested over 7 minutes through a placebo alcohol apparatus [18]. Blood was drawn to determine levels of alcohol and acetaldehyde before beverage ingestion and 15, 30, 45, 60, 90, 120, and 150 minutes after beverage ingestion. Blood alcohol concentrations were determined by using a modified alcohol dehydrogenase assay [19]. The rate of alcohol elimination (mg/kg per hour) was calculated from the slope of the pseudolinear decline of the blood alcohol concentration-time curve (usually from the 90-, 120-, and 150- minute samples) by using linear least-squares regression. Blood acetaldehyde levels were determined by using a modified fluorigenic high-performance liquid chromatographic assay [20] that had a detection sensitivity in the picomole range and intra-assay and interassay precisions of 2.4% and 3.7%, respectively. Statistical analyses, done by using SYSTAT software (SYSTAT, Inc., Evanston, Illinois), focused on differences between participants with ALDH2*1/2*1 and those with ALDH2*1/2*2. Demographic information, data on recent alcohol and cigarette use, peak blood alcohol concentration, time to peak blood alcohol concentration, volume of distribution, and rate of alcohol elimination were analyzed by using one-way analysis of variance; ALDH2 genotype was a between-participant variable. Data on blood alcohol concentration and acetaldehyde level were analyzed by using separate 2 8 analysis of variance for alcohol and placebo sessions; ALDH2 genotype was a between-participant variable, and time was a repeated measurement. Significant interactions were then analyzed by using post hoc comparisons with contrast matrices. The Bonferroni correction was used to limit the familywise error rate to 0.05 for comparisons among the placebo session time points and among the alcohol session time points. Results Genotyping for ALDH2 revealed 20 participants who had ALDH2*1/2*1 genotype, 13 who had ALDH2*1/2*2 genotype, and 2 who had ALDH2*2/2*2 genotype. Three participants (1 with ALDH2*1/2*2 genotype and the 2 with ALDH2*2/2*2 genotype) became ill after ingesting alcohol and were excluded from analyses because of missing data. One participant with ALDH2*1/2*2 genotype whose acetaldehyde levels exceeded 4 SDs from the mean (most likely as a result of instrumentation error) was also excluded from data analyses. The Table 1 shows demographic information and patterns of recent alcohol and cigarette use for the remaining 31 men. The ALDH2 genotype groups did not differ significantly for any of these variables; this reflects participant selection. Table 1. Demographic Information and Recent Patterns of Alcohol and Cigarette Use in 20 Asian-American Men with ALDH2*1/2*1 Genotype and 11 Asian-American Men with ALDH2*1/2*2 Genotype* Mean peak blood alcohol concentration SD was 81.3 12.48 mg/dL; the peak occurred 43.1 15.85 minutes after ingestion of alcohol. Mean volume of distribution was 0.718 0.1124 L/kg of body weight, and the mean rate of alcohol elimination was 97.8 33.97 mg/kg per hour. The ALDH2 genotype groups did not differ significantly for any of these variables. Mean blood alcohol concentrations for the alcohol session and mean acetaldehyde levels for the placebo and alcohol sessions, measured over time according to ALDH2 genotype, are shown in the (Figure 1). Figure 1. Mean (SD) blood levels of alcohol and acetaldehyde before and after ingestion of a placebo beverage containing 3 mL of 95% alcohol and an alcoholic beverage (0. n n P Analysis of variance revealed that the main effects for ALDH2 genotype and the interaction between ALDH2 genotype and time were not significant for the data on blood alcohol concentration from the alcohol session. Analysis of variance also revealed that the main effects for ALDH2 genotype and the interaction between ALDH2 genotype and time were significant for the data on acetaldehyde levels from the placebo sessions (ALDH2 genotype, P < 0.005; interaction between ALDH2 genotype and time, P < 0.013) and the alcohol sessions (ALDH2 genotype, P < 0.002; interaction between ALDH2 genotype and time, P < 0.001). Post hoc analyses with Bonferroni corrections revealed significant group differences 30, 45, and 60 minutes after placebo ingestion and 60, 90, 120, and 150 minutes after alcohol ingestion. Discussion We found that Asian-American men who were heterozygous for the ALDH2*2 allele did not differ from carefully matched men who were homozygous for the ALDH2*1 allele in measures of blood alcohol concentration overall or at any time after alcohol ingestion. These findings are consistent with the results of one study [14] but differ from those of other studies [15-17] in which persons with ALDH2*2 alleles had significantly slower rates of alcohol elimination than did persons with ALDH2*1 alleles. These discrepancies may result from group differences on important variables, especially history of alcohol use, that we controlled for. We also found that, despite equivalent blood alcohol concentrations, participants with ALDH2*1/2*2 genotype had significantly higher blood acetaldehyde levels after ingesting the alcohol beverage than did participants with ALDH2*1/2*1 genotype. These findings suggest that blood acetaldehyde levels rather than bloo

Psychiatric Morbidity, Illicit Drug Use and Adherence to Zidovudine (AZT) Among Injection Drug Users with HIV Disease

This study describes the relationship between the need for psychiatric consultation, illicit drug use, and zidovudine (AZT) adherence in HIV-infected injection drug users (IDUs) in methadone maintenance treatment (MMT). The treatment records of 57 IDUs in MMT who had been prescribed AZT between May and August of 1991 were reviewed. Those who required psychiatric consultation (P+, N = 46, 81%) were compared with those who did not require psychiatric consultation (P-, N = 11, 19%) on adherence to AZT treatment (using the mean corpuscular volume [MCV] as a biological marker), on recent illicit drug use, and on CD4 lymphocyte (T cell) count changes from the beginning to the end of AZT treatment. The P+ subjects were less likely than P- subjects to adhere to AZT treatment: fewer in the P+ group had an MCV outside of the normal range, and P+ subjects had a lower average monthly increase in MCV since the beginning of AZT treatment. Recent illicit drug use and CD4 lymphocyte count changes from the beginning to the end of AZT treatment did not show group differences. Psychiatric morbidity among HIV-infected IDUs in MMT is common, and may contribute to poor adherence to AZT treatment. Psychiatric screening and adherence-enhancing interventions should be targeted to IDUs entering drug treatment programs.

Comorbidity of Select Anxiety and Affective Disorders With Alcohol Dependence in Southwest California Indians

BACKGROUND Native Americans, overall, have the highest prevalence of alcohol dependence of any US ethnic group. In several large national surveys, alcohol dependence has been significantly associated with higher rates of anxiety and affective disorders (comorbidity). However, the frequencies of these disorders and their comorbidity with alcohol dependence in Native American populations are relatively unknown. METHODS Demographic information and DSM-III-R diagnoses were obtained by using the Semi-Structured Assessment for the Genetics of Alcoholism developed for the Collaborative Study on the Genetics of Alcoholism from 483 Southwest California Indian adults residing on contiguous reservations. The Semi-Structured Assessment for the Genetics of Alcoholism allowed differentiation of each anxiety and affective disorder into one of three types: independent of substance use, concurrent with alcohol use, and concurrent with drug use. RESULTS Sixty-six percent of the men and 53% of the women sampled had a lifetime diagnosis of alcohol dependence. Fourteen percent of the sample had a lifetime independent anxiety disorder, and 14% of the sample had a lifetime independent affective disorder. Alcohol- and/or drug-concurrent major depression occurred in 8%, and other alcohol- and/or drug-concurrent anxiety and affective disorders each occurred in less than 1.1% of the sample. No significant comorbidity was found between alcohol dependence and independent agoraphobia, social phobia, or major depressive disorder. CONCLUSIONS In this Southwest California Indian sample, rates of anxiety and affective disorders were substantially similar to those reported in the National Comorbidity Survey; however, comorbidity of independent disorders with alcohol dependence was not as pervasive as in the National Comorbidity Survey. Rates of concurrent anxiety and affective disorders were low. These data support the hypothesis that despite high rates of alcohol dependence, Southwest California Indians do not have higher rates of anxiety and affective disorders or comorbidity of these disorders with alcohol dependence than those reported in large surveys of non-American Indian populations.

Genetic associations of alcohol dehydrogenase with alcohol use disorders and endophenotypes in white college students.

Associations of alcohol dehydrogenase (ADH) gene polymorphisms (ADH1B*2 and ADH1C*1) with a lifetime alcohol use disorder (AUD) were examined in White college students. Alcohol-related endophenotypes likely to be influenced by elevations in acetaldehyde were also assessed. Individuals with an ADH1B*2 allele had lower rates of AUDs, consumed a lower maximum number of drinks in a 24-hr period, reported a greater level of response to alcohol, were more likely to have experienced alcohol-induced headaches following 1 or 2 drinks, and reported more severe hangovers than those lacking this allele. These findings are consistent with the hypothesis that enhanced sensitivity to alcohol and lower levels of alcohol use reflect the mechanism by which ADH1B*2 protects against developing an AUD.

Association of ALDH1 promoter polymorphisms with alcohol-related phenotypes in southwest California Indians.

BACKGROUND Cytosolic aldehyde dehydrogenase (ALDH1A1) is an important enzyme in the metabolism of acetaldehyde and the synthesis of retinoic acid. Two polymorphisms in the promoter region of ALDH1A1-ALDH1A1*2 and ALDH1A1*3-have recently been identified and described in small samples of Asian, Caucasian, and African individuals. The aim of this study was to determine the prevalence of these polymorphisms in a sample of Southwest California Indians and to test for associations with alcohol dependence and other substance-related behaviors. METHODS The participants in this study were 463 adult men and women recruited from 8 contiguous Indian reservations. A structured interview was used to gather information on demographics, psychiatric diagnoses, and personal drinking and drug use history. A blood sample was obtained from each participant, and leukocyte DNA was extracted and used to genotype for the presence of the ALDH1A1 promoter polymorphisms. RESULTS Twenty-seven participants (6%) possessed ALDH1A1*2 (frequency, 0.03), two participants possessed ALDH1A1*3, and one participant displayed both of these alleles. Individuals with an ALDH1A1*2 allele had lower rates of alcohol dependence and regular tobacco use than those without this allele. Individuals with ALDH1A1*2 also reported a significantly lower maximum number of drinks ever consumed in a 24-hr period, reported drinking fewer drinks per occasion when they first started drinking regularly, and reported lower expectations of alcohols effects compared with individuals without this allele. CONCLUSIONS Results from this study suggest that ALDH1A1*2 may be associated with protection from the development of alcohol and other substance use disorders.

Effects of alcohol on the EEG in Asian men with genetic variations of ALDH2

Approximately 50% of Asians experience a facial flush following alcohol ingestion. These individuals have an inactive form of mitochondrial aldehyde dehydrogenase (ALDH) encoded by the ALDH2*2 allele. This study matched 15 flushing and 15 nonflushing Asian men on demographics and drinking histories. The 30 subjects were genotyped for ALDH2 and were evaluated both before and following placebo and 0.75 ml/kg alcohol. The two groups did not differ significantly on blood alcohol concentrations after drinking, but did differ in electroencephalographic (EEG) response on the falling phase of the blood alcohol curve. Nonflushing subjects displayed significant increases in slow-alpha EEG activity (7.5-9.0 Hz) at 90 and 150 min post-alcohol consumption, compared to flushing subjects who did not show characteristic increases in this frequency band at these timepoints. These data suggest flushers, those with at least one ALDH2*2 allele, have less of slow-alpha wave EEG response to alcohol than nonflushers with ALDH2*1/2*1 genotype.

ADH2 and alcohol-related phenotypes in Ashkenazic Jewish American college students.

A variety of genetically influenced alcohol-related phenotypes relate to risk for alcohol dependence. In Asians, variation in the alcohol dehydrogenase (ADH2) gene relates to alcohol dependence, alcohol consumption, and reported alcohol-related symptoms, even after controlling for variation in the aldehyde dehydrogenase (ALDH2) gene. The association of ADH2 polymorphisms with alcohol-related behavior, however, has not been well characterized in non-Asians. This study evaluated 84 Ashkenazic Jewish American college students to determine the prevalence of the ADH2*2 allele (0.31). Carriers of ADH2*2 reported significantly fewer drinking days per month. ADH2*2, however, was not related to alcohol use disorders, alcohol-induced flushing and associated symptoms, number of binge drinking episodes in the past 90 days, maximum number of drinks ever consumed, or self-reported levels of response to alcohol. Results suggest that Ashkenazic Jewish Americans with ADH2*2 alleles drink less frequently, which might contribute, in part, to the overall lower rates of alcoholism in this population.

Evaluation of Aldehyde Dehydrogenase 1 Promoter Polymorphisms Identified in Human Populations

BACKGROUND Cytosolic aldehyde dehydrogenase, or ALDH1A1, functions in ethanol detoxification, metabolism of neurotransmitters, and synthesis of retinoic acid. Because the promoter region of a gene can influence gene expression, the ALDH1A1 promoter regions were studied to identify polymorphism, to assess their functional significance, and to determine whether they were associated with a risk for developing alcoholism. METHODS Sequence analysis was performed in the promoter region by using Asian, Caucasian, and African American subjects. The resulting polymorphisms were assessed for frequency in Asian, Caucasian, Jewish, and African American populations and tested for associations with alcohol dependence in Asian and African American populations of alcoholics and controls. The functional significance of each polymorphism was determined through in vitro expression analysis by using HeLa and HepG2 cells. RESULTS Two polymorphisms, a 17 base pair (bp) deletion (-416/-432) and a 3 bp insertion (-524), were discovered in the ALDH1A1 promoter region: ALDH1A1*2 and ALDH1A1*3, respectively. ALDH1A1*2 was observed at frequencies of 0.035, 0.023, 0.023, and 0.012 in the Asian, Caucasian, Jewish, and African American populations, respectively. ALDH1A1*3 was observed only in the African American population, at a frequency of 0.029. By using HeLa and HepG2 cells for in vitro expression, the activity of the luciferase reporter gene was significantly decreased after transient transfection of ALDH1A1*3-luciferase compared with the wild-type construct ALDH1A1*1-luciferase. In an African American population, a trend for higher frequencies of the ALDH1A1*2 and ALDH1A1*3 alleles was observed in a population of alcoholics (p = 0.03 and f = 0.12, respectively) compared with the control population. CONCLUSIONS ALDH1A1*2 and ALDH1A1*3 may influence ALDH1A1 gene expression. Both ALDH1A1*2 and ALDH1A1*3 produce a trend in an African American population that may be indicative of an association with alcoholism; however, more samples are required to validate this observation. The underlying mechanisms contributing to these trends are still unknown.

Prevalence, Predictors, and Outcomes of Early Adherence after Starting or Changing Antiretroviral Therapy

The objectives of this research were to assess prevalence and predictors of early antiretroviral therapy adherence using multiple indicators and to estimate effects of early adherence on subsequent HIV viral load and CD4+ lymphocyte responses. Study subjects were adults with HIV infection referred to an antiretroviral therapy-monitoring clinic for initiation or change in therapy between March 1998 and June 1999. The design was a prospective observational cohort involving baseline interview followed by 30 days of electronic adherence monitoring (MEMS), 30-day interview, and follow-up viral load at 1, 3, and 6 months. Adherence indicators included MEMS therapeutic coverage, observed/expected cap openings, and self-reported adherence assessed at 30 days. Of 235 consenting patients, 60 (26%) failed to complete 30 days of electronic monitoring (noncompleters). At 6 months, mean change from baseline plasma viral load was inferior among noncompleters (0.5 log vs. 1.7 log). Predictors of adherence, varying by adherence metric, included: gender, race, prior antiretroviral therapy experience, substance abuse, prior adherence behavior, health beliefs, and pharmacist prediction of adherence. Self-reported adherence was more sensitive in predicting viral load responses than MEMS-based measures and identified poor adherence at earlier time points. Approximately a quarter of consenting patients were unable to complete 30 days of MEMS monitoring, and early drop out was a poor prognostic sign. Predictors of adherence varied depending upon how adherence was measured. Differences in virologic response between patients with optimal or poor adherence may not emerge until several months after regimen change or initiation. Structured assessment of self-reported adherence is an inexpensive and useful tool to assist clinicians in monitoring adherence.