Clinical and pharmacogenetic predictors of circulating atorvastatin and rosuvastatin concentrations in routine clinical care.

BACKGROUND: A barrier to statin therapy is myopathy associated with elevated systemic drug exposure. Our objective was to examine the association between clinical and pharmacogenetic variables and statin concentrations in patients. METHODS AND RESULTS: In total, 299 patients taking atorvastatin or rosuvastatin were prospectively recruited at an outpatient referral center. The contribution of clinical variables and transporter gene polymorphisms to statin concentration was assessed using multiple linear regression. We observed 45-fold variation in statin concentration among patients taking the same dose. After adjustment for sex, age, body mass index, ethnicity, dose, and time from last dose, SLCO1B1 c.521T>C (P<0.001) and ABCG2 c.421C>A (P<0.01) were important to rosuvastatin concentration (adjusted R(2)=0.56 for the final model). Atorvastatin concentration was associated with SLCO1B1 c.388A>G (P<0.01) and c.521T>C (P<0.05) and 4ß-hydroxycholesterol, a CYP3A activity marker (adjusted R(2)=0.47). A second cohort of 579 patients from primary and specialty care databases were retrospectively genotyped. In this cohort, genotypes associated with statin concentration were not differently distributed among dosing groups, implying providers had not yet optimized each patient's risk-benefit ratio. Nearly 50% of patients in routine practice taking the highest doses were predicted to have statin concentrations greater than the 90th percentile. CONCLUSIONS: Interindividual variability in statin exposure in patients is associated with uptake and efflux transporter polymorphisms. An algorithm incorporating genomic and clinical variables to avoid high atorvastatin and rosuvastatin levels is described; further study will determine whether this approach reduces incidence of statin myopathy.

Resveratrol in combination with other dietary polyphenols concomitantly enhances antiproliferation and UGT1A1 induction in Caco-2 cells.

AIMS: The only FDA approved medication for colorectal cancer (CRC) prevention is celecoxib. Its adverse effects underline the need for safer drugs. Polyphenols like resveratrol are in clinical trials for this purpose. This study aimed at examining effects of resveratrol alone and in combination with curcumin or chrysin on UGT induction in Caco-2 cells. Phytochemical combinations were selected using drug combination analyses of various anti-proliferation ratios of resveratrol+curcumin and resveratrol+chrysin. MAIN METHODS: Cell proliferation and UGT1A1 induction assays were carried out with individual polyphenols and combinations. Cell viability was determined with AlamarBlue assays. UGT1A1 mRNA was quantified via real time RT-PCR. UGT activity was determined with 4-methylumbelliferone (4MU) glucuronidation. KEY FINDINGS: Cell proliferation IC(50) estimates (± SE) for resveratrol, curcumin and chrysin were 20.8 ± 1.2, 20.1 ± 1.1 and 16.3 ± 1.3µM respectively. Combination of anti-proliferative effects showed additivity for resveratrol+chrysin and resveratrol+curcumin. Resveratrol at its IC(50) mediated a four-fold induction of UGT1A1 mRNA in a concentration independent manner. Chrysin at its IC(50) induced UGT1A1 expression seven-fold while Curcumin at its IC(90) mediated a two-fold induction. The 20 µM:40µ M resveratrol+curcumin and 20 µM :32 µM resveratrol+chrysin combinations mediated the greatest increases in mRNA expression (12 and 22 folds respectively). Significant increase in 4-MU glucuronidation was observed with combinations exhibiting maximal mRNA induction. SIGNIFICANCE: Phytochemical combinations can offer greater chemoprevention than single agents. These chemicals might offer safer options than present synthetic therapeutics for CRC prevention.

Characterizing the effects of common UDP glucuronosyltransferase (UGT) 1A6 and UGT1A1 polymorphisms on cis- and trans-resveratrol glucuronidation.

The dietary polyphenol resveratrol (RES) exists as cis- and trans-isomers with known stereospecific and stereoselective glucuronidation at the 3 and 4' positions by distinct UGT1A isoforms. We examined cis-RES glucuronidation in various protein sources. UGT1A6 or UGT1A1 genotype-dependent cis-or trans-RES glucuronidation, respectively, was further determined. cis-RES exhibited partial substrate inhibition in UGT1A6 Supersomes and human embryonic kidney 293 cells overexpressing genetically variant UGT1A6 alleles. Cells expressing UGT1A6*4 had the highest activity with a V(max) of 612 +/- 27.36 nmol/min/mg, followed by UGT1A6*3. The *2 allozyme had a higher V(max) (1.6-fold) and K(m) (1.9-fold) than *1. In 51 human liver samples genotyped for UGT1A6, four alleles (frequencies) were identified as *1 (0.58), *2 (0.36), *3 (0.01), and *4 (0.05), leading to assignment of the following genotypes (frequencies): *1/*1 (0.29), *1/*2 (0.45), *1/*3 (0.02), *1/*4 (0.10), and *2/*2 (0.14). Up to 5-fold variability in trans-RES glucuronidation was observed in individual liver samples. In livers stratified by UGT1A6 genotype, a significant difference in cis-RES glucuronidation activity (p < 0.05) was seen between the *2 variants compared with homozygous *1 livers. The trans-RES glucuronidation was quantitated in a human liver bank genotyped for the UGT1A1 TATA box repeat polymorphism. There was no significant difference for formation of trans-RES 3-O-glucuronide. We were surprised to find that trans-RES 4'-O-glucuronide formation was higher in livers with the 7/7 genotype compared with 6/6 and 6/7 (p < 0.05). In conclusion, cis-RES glucuronidation exhibited atypical partial substrate inhibition kinetics in vitro. Whereas cis-RES glucuronidation varied with UGT1A6 genotypes, the UGT1A1*28 polymorphism did not explain variability in trans-RES glucuronidation.

Resveratrol (trans-resveratrol, 3,5,4'-trihydroxy-trans-stilbene) glucuronidation exhibits atypical enzyme kinetics in various protein sources.

The dietary polyphenol trans-resveratrol (3,5,4'-trihydroxy-trans-stilbene) is glucuronidated at the 3 and 4' positions to yield two major glucuronide conjugates, resveratrol-3-O-glucuronide (R3G) and resveratrol-4'-O-glucuronide (R4'G). The major enzymes catalyzing this conjugation reaction are members of the UDP-glucuronosyl transferase (UGT) 1A family and include UGT1A1 and UGT1A9, with minor contributions by UGT1A10. The kinetics of resveratrol glucuronidation in these three UGT1A isoforms as well as in human liver and intestinal microsomes were characterized across a wide concentration range. Atypical kinetics were observed for resveratrol glucuronidation in all the protein sources studied. The V(max) estimate per total protein for both glucuronides was higher in human intestinal microsomes compared with human liver microsomes (12.2 +/- 0.34 versus 7.4 +/- 0.25 nmol/min/mg for R3G and 8.9 +/- 0.14 versus 0.45 +/- 0.01 nmol/min/mg for R4'G). The kinetic profile for formation of R3G in both human liver and intestinal microsomes fits a substrate inhibition model, whereas that for R4'G exhibited a biphasic kinetic profile in human liver microsomes and substrate inhibition in human intestinal microsomes. In recombinant human UGT supersomes, for both glucuronides, UGT1A9 exhibited higher activity than UGT1A1, whereas the lowest activity was observed with UGT1A10. The kinetic profile for R3G exhibited substrate inhibition for all three isoforms, whereas that for R4'G differed, exhibiting substrate inhibition for UGT1A1 and UGT1A10 and Hill kinetics for UGT1A9. These results suggest that in vitro kinetics of resveratrol glucuronidation at high concentrations cannot be ignored in predicting in vivo clearance upon high-dose consumption of resveratrol.