ABSTRACT
BACKGROUND: Tamoxifen is hydroxylated by cytochrome P450 (CYP) 2D6 to the potent metabolites 4-hydroxytamoxifen (4OHtam) and 4-hydroxy-N-demethyltamoxifen (4OHNDtam), which are both conjugated by sulphotransferase (SULT)1A1. Clinical studies indicate that CYP2D6 and SULT1A1 genotypes are predictors for treatment response to tamoxifen. Therefore, we examined the relationship between CYP2D6 genotype, SULT1A1 genotype, SULT1A1 copy number and the pharmacokinetics of tamoxifen. PATIENTS AND METHODS: The serum levels of tamoxifen and metabolites of 151 breast cancer patients were measured by high-pressure liquid chromatography-tandem mass spectrometry. The CYP2D6 and SULT1A1 polymorphisms and SULT1A1 copy number were determined by long PCR, PCR-based restriction fragment length polymorphism, DNA sequencing and fluorescence-based PCR. RESULTS: The levels of 4OHtam, 4OHNDtam and N-demethyltamoxifen were associated with CYP2D6 predicted enzymatic activity (P < 0.05). The SULT1A1 genotype or copy number did not influence the levels of tamoxifen and its metabolites. However, the ratios of N-demethyltamoxifen/tamoxifen and N-dedimethyltamoxifen/N-demethyltamoxifen were related to SULT1A1 genotype. CONCLUSION: CYP2D6 and SULT1A1 genotypes may partly explain the wide inter-individual variations in the serum levels of tamoxifen and its metabolites. We propose that therapeutic drug monitoring should be included in studies linking CYP2D6 and SULT1A1 genotypes to clinical outcome.
Subject(s)
Antineoplastic Agents, Hormonal/pharmacokinetics , Arylsulfotransferase/genetics , Breast Neoplasms/drug therapy , Cytochrome P-450 CYP2D6/genetics , Selective Estrogen Receptor Modulators/pharmacokinetics , Tamoxifen/pharmacokinetics , Adult , Aged , Aged, 80 and over , Arylsulfotransferase/metabolism , Biotransformation/genetics , Breast Neoplasms/blood , Breast Neoplasms/enzymology , Cytochrome P-450 CYP2D6/metabolism , Female , Gene Dosage , Gene Frequency , Genotype , Humans , Middle Aged , Norway , Polymorphism, Restriction Fragment Length , Tamoxifen/analogs & derivatives , Tamoxifen/bloodABSTRACT
We have developed a method for the determination of tamoxifen (tam) and its metabolites 4-hydroxytamoxifen (4OHtam), N-demethyltamoxifen (NDtam), N-dedimethyltamoxifen (NDDtam), tamoxifen-N-oxide (tamNox), and 4-hydroxy-N-demethyltamoxifen (4OHNDtam) in 50 microl human serum. Serum proteins were precipitated with acetonitrile. Deuterated-tamoxifen (D5 tam) was added as internal standard. Sample supernatant was injected into an on-line reversed-phase extraction column coupled with a C18 analytical column and analytes were detected by tandem mass spectrometry. The lower limits of quantification were 0.25 ng/mL for 4OHtam, NDtam and tam, 1.0 ng/mL for NDDtam and tamNox. Ranges of within- and between-day variation were 2.9-15.4% and 4.4-12.9%, respectively.
Subject(s)
Chromatography, Liquid/methods , Mass Spectrometry/methods , Tamoxifen/blood , Breast Neoplasms/drug therapy , Chemical Fractionation/methods , Humans , Sensitivity and Specificity , Tamoxifen/analogs & derivatives , Tamoxifen/metabolism , Tamoxifen/therapeutic useABSTRACT
The influence of environmental (extracellular) pH on the sporulation rhythm in Neurospora crassa was investigated for wild-type (frq+) and the mutants chr, frq1, frq7, and frq8. In all mutants, including wild type, the growth rate was found to be influenced strongly by extracellular pH in the range 4-9. On the other hand, for the same pH range, the period length of the sporulation rhythm is little influenced in wild type, chr, and frq1. A loss of pH homeostasis of the period, however, was observed in the mutants frq7 and frq8, which also are known to have lost temperature compensation. Concerning the influence of extracellular pH on growth rates, a clear correspondence between growth rates and the concentration of available H2PO4- ion has been found, indicating that the uptake of H2PO4- may be a limiting factor for growth under our experimental conditions. The loss of pH compensation in the frq7 and frq8 mutants may be related to less easily degradable FRQ7,8 proteins when compared with wild-type FRQ. Results from recent model considerations and experimental results predict that, with increasing extra-and intracellular pH, the FRQ7 protein degradation increases and should lead to shorter period lengths.
