Prominent Stereoselectivity of NNAL Glucuronidation in Upper Aerodigestive Tract Tissues

Tobacco specific nitrosamines (TSNAs) are among the most potent carcinogens found in cigarettes and smokeless tobacco products. Decreases in TSNA detoxification, particularly 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), have been associated with tobacco-related cancer incidence. NNK is metabolized by carbonyl reduction to its major carcinogenic metabolite, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), which is detoxified by glucuronidation at the nitrogen within the pyridine ring or at the chiral alcohol to form four glucuronide products: (R)-NNAL-O-Gluc, (S)-NNAL-O-Gluc, (R)-NNAL-N-Gluc, (S)-NNAL-N-Gluc. Stereoselective NNAL-Gluc formation and the relative expression of NNAL-glucuronidating UGTs (1A4, 1A9, 1A10, 2B7, 2B10, 2B17) were analyzed in 39 tissue specimens from the upper aerodigestive tract (esophagus (n = 13), floor of mouth (n = 4), larynx (n = 9), tongue (n = 7), and tonsil (n = 6)). All pooled tissue types preferentially formed (R)-NNAL-O-Gluc in the presence of racemic-NNAL; only esophagus exhibited any detectable formation of (S)-NNAL-O-Gluc. For every tissue type examined, UGT1A10 exhibited the highest relative expression levels among the NNAL-O-glucuronidating UGTs, ranging from 36% (tonsil) to 49% (esophagus), followed by UGT1A9 > UGT2B7 > UGT2B17. UGT1A10 also exhibited similar or higher levels of expression as compared to both NNAL-N-glucuronidating UGTs, 1A4 and 2B10. In a screening of cells expressing individual UGT enzymes, all NNAL glucuronidating UGTs exhibited some level of stereospecific preference for individual NNAL enantiomers, with UGTs 1A10 and 2B17 forming primarily (R)-NNAL-O-Gluc. These data suggest that UGTs 1A10 and 2B17 may be important enzymes in the detoxification of TSNAs like NNK in tissues of the upper aerodigestive tract.