Utility and relationships of biomarkers of smoking in African-American light smokers.

Although expired carbon monoxide (CO) and plasma cotinine (COT) have been validated as biomarkers of self-reported cigarettes per day (CPD) in heavy smoking Caucasians, their utility in light smokers is unknown. Further, variability in CYP2A6, the enzyme that mediates formation of COT from nicotine and its metabolism to trans-3'-hydroxycotinine (3HC), may limit the usefulness of COT. We assessed whether CO and COT are correlated with CPD in African-American light smokers (< or =10 CPD, n = 700), a population with known reduced CYP2A6 activity and slow COT metabolism. We also examined whether gender, age, body mass index, smoking mentholated cigarettes, or rate of CYP2A6 activity, by genotype and phenotype measures (3HC/COT), influence these relationships. At baseline, many participants (42%) exhaled CO of < or =10 ppm, the traditional cutoff for smoking, whereas few (3.1%) had COT below the cutoff of < or =14 ng/mL; thus, COT seems to be a better biomarker of smoking status in this population. CPD was weakly correlated with CO and COT (r = 0.32-0.39, P < 0.001), and those reporting fewer CPD had higher CO/cigarette and COT/cigarette, although the correlations coefficients between these variables were also weak (r = -0.33 and -0.08, P < 0.05). The correlation between CPD and CO was not greatly increased when analyzed by CYP2A6 activity, smoking mentholated cigarettes, or age, although it appeared stronger in females (r = 0.38 versus 0.21, P < 0.05) and obese individuals (r = 0.38 versus 0.24, P < 0.05). Together, these results suggest that CO and COT are weakly associated with self-reported cigarette consumption in African-American light smokers, and that these relationships are not substantially improved when variables previously reported to influence these biomarkers are considered.

Novel and established CYP2A6 alleles impair in vivo nicotine metabolism in a population of Black African descent.

Cytochrome P450 2A6 (CYP2A6) is a human enzyme best known for metabolizing tobacco-related compounds, such as nicotine, cotinine (COT), and nitrosamine procarcinogens. CYP2A6 genetic variants have been associated with smoking status, cigarette consumption, and tobacco-related cancers. Our objective was to functionally characterize four nonsynonymous CYP2A6 sequence variants with respect to their haplotype, allele frequency, and association with in vivo CYP2A6 activity. In vivo, nicotine was administered orally to 281 volunteers of Black African descent. Blood samples were collected for kinetic phenotyping and CYP2A6 genotyping. In vitro, nicotine C-oxidation catalytic efficiencies of heterologously expressed variant enzymes were assessed. The four uncharacterized sequence variants were found in seven novel alleles CYP2A6(*)24A&B ; (*)25, (*)26, (*)27, and *28A&B, most were associated with impaired in vivo CYP2A6 activity. Nicotine metabolism groupings, based on the in vivo data of variant alleles, were created. Mean trans-3'-hydroxycotinine/cotinine (3HC/COT) differed (P<0.001) between normal (100%), intermediate (64%), and slow (40%) groups. Systemic exposure to nicotine following oral administration also differed (P<0.001) between normal (100%), intermediate (139%), and slow (162%) metabolism groups. In addition, alleles of individuals with unusual phenotype-genotype relationships were sequenced, resulting in the discovery of five novel uncharacterized alleles and at least one novel duplication allele. A total of 7% of this population of Black African descent had at least one of the eight novel characterized alleles and 29% had at least one previously established allele. These findings are important for increasing the accuracy of association studies between CYP2A6 genotype and behavioral, disease, or pharmacological phenotypes.

A novel CYP2A6 allele, CYP2A6*23, impairs enzyme function in vitro and in vivo and decreases smoking in a population of Black-African descent.

OBJECTIVES: CYP2A6 is the main enzyme involved in nicotine metabolism in humans. We have identified a novel allele, CYP2A6*23 (2161C>T, R203C), in individuals of Black-African descent and investigated its impact on enzyme activity and association with smoking status. METHODS: Wild-type and variant enzymes containing amino acid changes R203C (CYP2A6*23), R203S (CYP2A6*16) and V365M (CYP2A6*17) were expressed in Escherichia coli. The effect of CYP2A6*23 in vivo was examined in individuals of Black-African descent given 4 mg oral nicotine. RESULTS: CYP2A6*23 occurred at an allele frequency of 2.0% in individuals of Black-African descent (N=560 alleles, 95% confidence interval, 0.8-3.1%) and was not detected in Caucasians (N=334 alleles), Chinese (N=288 alleles) or Japanese (N=104 alleles). In vitro, CYP2A6.23 had greatly reduced activity toward nicotine C-oxidation similar to CYP2A6.17, as well as reduced coumarin 7-hydroxylation. Conversely, CYP2A6.16 did not differ in activity compared with the wild-type enzyme. The trans-3'-hydroxycotinine to cotinine ratio, a phenotypic measure of CYP2A6 activity in vivo, was lower in CYP2A6*1/*23 and CYP2A6*23/*23 individuals (mean adjusted ratio of 0.60, n=5) compared with CYP2A6*1/*1 individuals (mean adjusted ratio of 1.21, n=150) (P<0.04). CYP2A6*23 trended toward a higher allele frequency in nonsmokers (3.1%, N=9/286 alleles) compared with smokers (0.7%, N=2/274 alleles) (P=0.06). CONCLUSION: These results suggest the novel CYP2A6*23 allele impairs enzyme function in vitro and in vivo and trends toward an association with lower risk of smoking.

Evidence for natural desensitization to mosquito salivary allergens: mosquito saliva specific IgE and IgG levels in children.

BACKGROUND: There are few epidemiologic data on the prevalence of mosquito allergy, although local reactions to mosquito bites are common. OBJECTIVE: To investigate the prevalence of mosquito allergy in children by measuring serum levels of mosquito saliva specific IgE and IgG antibodies that correlate well with the size of mosquito bite local reactions. METHODS: Using enzyme-linked immunosorbent assays to measure mosquito (Aedes vexans) saliva-specific antibodies, we investigated sensitization to mosquito bites in 402 children aged 1 month to 18 years and correlated mosquito saliva specific IgE and IgG levels with age and sex. Twenty-three serum samples from infants who had never been exposed to mosquitos were used as negative controls. RESULTS: Mean levels of mosquito saliva specific IgE and IgG were lowest in the 23 negative control serum samples. In the 402 samples from children who may have been exposed to mosquitos, mean saliva specific IgG levels were higher in boys than in girls (P < .008). Levels of IgE and IgG correlated with each other (P < .001). A significant inverse correlation was found between age and both IgE and IgG levels. IgE levels peaked at the age of 6 to 12 months of age, and IgG levels peaked at 1 to 6 months of age. Levels of IgE and IgG antibodies gradually declined after the age of 5 years. CONCLUSIONS: Based on the presence of mosquito saliva specific antibodies, exposed infants and young children are at increased risk of having allergic reactions to mosquito bites. Antibody levels decline throughout childhood and adolescence, suggesting that natural desensitization may occur.