Glucuronidation is an important pathway in the metabolism of nicotine, with previous studies suggesting that ∼22% of urinary nicotine metabolites are in the form of glucuronidated compounds. Recent in vitro studies have suggested that the UDP-glucuronosyltransferases (UGT) 2B10 and 2B17 play major roles in nicotine glucuronidation with polymorphisms in both enzymes shown to significantly alter the levels of nicotineglucuronide, cotinine-glucuronide, and trans-3′-hydroxycotinine (3HC)-glucuronide in human liver microsomes in vitro. In the present study, the relationship between the levels of urinary nicotine metabolites and functional polymorphisms in UGTs 2B10 and 2B17 was analyzed in urine specimens from 104 Caucasian smokers. Based on their percentage of total urinary nicotine metabolites, the levels of nicotine-glucuronide and cotinine-glucuronide were 42% (P < 0.0005) and 48% (P < 0.0001), respectively, lower in the urine from smokers exhibiting the UGT2B10 (*1/*2) genotype and 95% (P < 0.05) and 98% (P < 0.05), respectively, lower in the urine from smokers with the UGT2B10 (*2/*2) genotype compared with the urinary levels in smokers having the wild-type UGT2B10 (*1/*1) genotype. The level of 3HC-glucuronide was 42% (P < 0.001) lower in the urine from smokers exhibiting the homozygous UGT2B17 (*2/*2) deletion genotype compared with the levels in urine from wild-type UGT2B17 subjects. These data suggest that UGTs 2B10 and 2B17 play important roles in the glucuronidation of nicotine, cotinine, and 3HC and suggest that the UGT2B10 codon 67 SNP and the UGT2B17 gene deletion significantly reduce overall glucuronidation rates of nicotine and its major metabolites in smokers.

trans-3′-Hydroxycotinine (3HC) and its glucuronide (3HC-Gluc) are major nicotine metabolites excreted in the urine of smokers and other tobacco users. While several members of the UDP-glucuronosyltransferase (UGT) family of enzymes were previously shown to be active in catalyzing the formation of 3HC-Gluc, a comprehensive screening of all known human UGT1A and 2B enzymes for glucuronidation activity against 3HC was not previously performed. In the present study, 8 UGT1A and 6 UGT2B enzymes were screened for activity against 3HC. UGT2B17 exhibited the highest O-glucuronidation activity, exhibiting a 4-fold lower (p<0.005) KM (8.3 mM) than that observed for UGTs 1A9 (35 mM) or 2B7 (31 mM) and a KM smaller than that observed for human liver microsomes (HLM; 26 mM). The KM for 3HC-O-Gluc formation was 3.1-fold lower (p<0.0005) in HLM from male subjects exhibiting the wild-type genotype UGT2B17 (*1/*1) than that in HLM from subjects homozygous for the UGT2B17 deletion genotype [UGT2B17 (*2/*2)]. Both UGTs 2B10 and 1A4 exhibited 3HC-N-Gluc formation activity, with UGT2B10 exhibiting a 4-fold lower (p<0.05) KM (13 mM) than that observed for UGT1A4 (57 mM) and which was similar to the KM observed in HLM (14 mM). There was a 91% (p<0.0001) and 39% (p<0.001) decrease in 3HC-N-Gluc formation activities in HLM from subjects with the UGT2B10 (*2/*2) and UGT2B10 (*1/*2) genotypes, respectively, compared to that of HLM from subjects with the wild-type UGT2B10 (*1/*1) genotype. These results suggest that UGT2B17 and UGT2B10 play key roles in the glucuronidation of 3HC in the human liver and that functional polymorphisms in UGT2B17 and UGT2B10 are associated with significantly reduced glucuronidation activities against 3HC.

Glucuronidation is an important pathway in the metabolism of nicotine, with ∼28% of urinary nicotine metabolites in the form of glucuronidated compounds. Recent in vitro studies have suggested that the UGTs 2B10 and 2B17 play major roles in nicotine glucuronidation with polymorphisms in both enzymes shown to significantly alter the levels of nicotine-, cotinine-, and trans-3-hydroxy-cotinine (3HC)-glucuronides in human liver microsomes in vitro. In the present study, the relationship between the levels of urinary nicotine metabolites and functional polymorphisms in UGTs 2B10 and 2B17 were analyzed in urine specimens from 104 Caucasian smokers. The levels of nicotine-glucuronide and cotinine-glucuronide were 47 (p&lt;0.0005) and 47% (p&lt;0.0001) lower, respectively, in urine from smokers exhibiting the UGT2B10 (*1/*2) genotype, and 98 (p&lt;0.0001) and 99 (p&lt;0.0001), respectively, in urine from smokers with the UGT2B10 (*2/*2) genotype, as compared to the urinary levels in wild-type UGT2B10 smokers. The levels of 3HC-glucuronide was 64% (p&lt;0.0001) lower in the urine from smokers exhibiting the homozygous UGT2B17 (*2/*2) deletion genotype as compared to the levels in urine from wild-type UGT2B17 subjects. These data are consistent with previous in vitro studies and demonstrate that UGTs 2B10 and 2B17 play important roles in the glucuronidation of nicotine, cotinine and 3HC in vivo, and suggest the UGT2B10 codon 67 SNP and the UGT2B17 deletion significantly reduce overall glucuronidation rates of nicotine and its major metabolites in smokers. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 918.

3’-Hydroxycotinine (3HC) and its glucuronide (3HC-Gluc) conjugate are major nicotine metabolites excreted in the urine of smokers and other tobacco or nicotine users. Previous studies have demonstrated that human liver microsomes (HLM) were active in catalyzing the formation of both 3HC-O-Gluc and 3HC-N-Gluc. While not all human UDP-glucuronosyltransferases (UGTs) were previously screened, UGTs 2B7, 1A9, 2B4 and 2B15 were shown to be active in the formation of 3HC-O-Gluc and UGT1A4 active in 3HC-N-Gluc formation. In the present study, all known human UGTs except for UGT2B28 were screened for glucuronidation activity against 3HC. Homogenates from cell lines over-expressing UGTs 2B17 and 2B10 exhibited the highest O- and N-glucuronidation activities, respectively for the UGTs tested. Kinetic analysis for the formation of 3HC-O-Gluc demonstrated that the KM for UGT2B17 is 3.7- and 4.2-fold lower than that observed for UGTs 1A9 (p&lt;0.005) and 2B7 (p&lt;0.005); the KM of 3HC-N-Gluc formation was 4.4-fold lower (p&lt;0.05) for UGT2B10 than that observed for UGT1A4. Kinetic analysis demonstrated that HLM from subjects exhibiting the homozygous null UGT2B17 deletion [UGT2B17 (*2/*2)] exhibited a KM for 3HC-O-Gluc formation that was 3.1-fold higher (p&lt;0.001) than that observed in HLM from subjects exhibiting the wild-type UGT2B17(*1/*1) genotype. The KM of 15.6 μM for 3HC-O-Gluc formation in UGT2B17 (*2/*2) HLM was close to that observed for UGT2B17 (8.3 μM). 3HC-N-Gluc formation activity was decreased by 91% (p&lt;0.001) in HLM from subjects with the UGT2B10(*2/*2) knock-out genotype and 39% (p&lt;0.001) in HLM from subjects with the UGT2B10(*1/*2) genotype as compared to that observed for UGT2B10(*1/*1) HLMs. The KM of 13 μM for 3HC-N-Gluc formation in UGT2B10 (*1/*1) HLM (n=6) was close to that observed for UGT2B10 (13 μM). These results demonstrate that UGTs 2B17 and 2B10 play key roles in the hepatic glucuronidation of 3HC and that functional polymorphisms in UGT2B17 and UGT2B10 are associated with significantly reduced glucuronidation activities against 3HC in HLM. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2757. doi:10.1158/1538-7445.AM2011-2757