High performance liquid chromatography of tamoxifen and metabolites in plasma and tissues

Lim, Chang Kee; Chow, Loretta; Yuan, Zhi-Xin; Smith, Lewis L.

doi:10.1002/bmc.1130070606

Cited by 22 publications

(8 citation statements)

References 7 publications

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“…Levels of TAM and metabolites in both plasma and tumour tissue compared well with those previously reported, although there was a tendency for levels of 4OH and DMT to be somewhat higher than those detected by other investigators (Daniel et al, 1979;Robinson et al, 1989;Langan-Fahey et al, 1990;Lim et al, 1993;MacCallum et al, 1997). There could be several explanations for this: from recovery of spiked plasmas, 4OH as the first metabolite off the HPLC column was seen to be the metabolite most likely to be contaminated with spurious peaks, thus overestimation of plasma levels of all three metabolites, most especially 4OH (as the metabolite present at the lowest concentrations) is a possibility.…”

Section: supporting

confidence: 84%

True

MacCallum¹,

Cummings²,

Dixon

et al. 2000

Br J Cancer

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Patients treated with tamoxifen (TAM) for primary breast cancer often manifest de novo or acquired resistance, possibly through changes in drug metabolism. Using solid-phase extraction methods and reversed-phase high-performance liquid chromatography separations, levels of TAM and metabolites 4-hydroxytamoxifen (4OH) and desmethyltamoxifen (DMT) have been measured in plasma and tumour tissue from breast cancer patients treated with TAM for at least 3 months. Patients were categorized into those with tumours responding to TAM and those showing de novo or acquired resistance. Levels of TAM, 4OH and DMT in both plasma and tissue samples were correlated with clinical response, length of treatment and patient weight. Interesting results included accumulation of 4OH in tumour tissues over time in all patients, with significance reached in the acquired resistance group. In addition, significantly lower levels of 4OH and DMT were found in plasma taken from responding patients after 3 months of treatment when compared to non-responding patients, and a small group of ER-poor patients showed significantly lower levels of all three species in plasma when compared to other patients. Whilst not explaining TAM resistance in all cases, these differences could account for the development of resistance to TAM treatment in certain subgroups of patients. © 2000 Cancer Research Campaign

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Section: supporting

confidence: 84%

True

MacCallum¹,

Cummings²,

Dixon

et al. 2000

Br J Cancer

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“…A solution of tamoxifen (4.0 g, 10.78 mmol) in absolute ethanol (100 mL) and concentrated HCl (60 mL) was refluxed for 4 h. The reaction mixture was cooled and partitioned between aqueous NaOH (3 M, 200 mL) and ether (200 mL). The ether layer was washed (2 × 100 mL of water), concentrated, and dissolved in methanol (5 mL) to give a 1:1 mixture of cis and trans isomers as determined by HPLC analysis (22). Reversephase preparative HPLC was then used to isolate the pure cis isomer.…”

Section: Cis-(e)-1-[4-[2-(dimethylamino)ethoxymentioning

confidence: 99%

Determination of DNA Damage in F344 Rats Induced by Geometric Isomers of Tamoxifen and Analogues

Brown

Martin

et al. 1998

Chem. Res. Toxicol.

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To investigate the activation mechanisms involved in tamoxifen carcinogenicity, analogues of tamoxifen isomers modified at the ethyl group were synthesized and assessed for their ability to induce hepatic DNA damage following their administration to female F344 rats. The cis isomer was prepared by acid-catalyzed isomerization of tamoxifen and isolated by preparative HPLC. The active metabolite alpha-hydroxytamoxifen and geometric isomers of bromotamoxifen and C-desmethylenetamoxifen, analogues in which the ethyl group has been replaced by a bromine atom and methyl group, respectively, were synthesized according to published procedures. The levels of hepatic DNA adducts induced were determined by 32P-postlabeling. Bromotamoxifen and tamoxifen 1,2-epoxide caused no detectable DNA damage relative to controls. Trans isomers of tamoxifen, C-desmethylenetamoxifen, and alpha-hydroxytamoxifen all produced DNA adducts at a 5-90-fold higher level than the corresponding cis isomers. In contrast, both the cis and trans isomers of alpha-hydroxytamoxifen showed similar reactivity toward calf thymus DNA in vitro. Molecular models of alpha-hydroxytamoxifen isomers suggest this difference in DNA adduct-forming ability is due to steric hindrance of the enzymes involved in the activation of this metabolite. There were high adduct levels in the liver, but no uterine DNA adducts were detected in rats treated with alpha-hydroxytamoxifen. This suggests that in contrast to the liver, alpha-hydroxytamoxifen is not further activated in rat uterus. This may help to explain the absence of uterine tumors in rats following long-term tamoxifen treatment.

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“…Since pure DMT was unavailable, the identity of the presumed DMT peak was confirmed indirectly by inhibiting its metabolic pathway with a P450 3A inhibitor, ketoconazole, and by its relative retention time compared to the parent compound and 4-OH-TAM, as reported in previous studies that used similar HPLC conditions. 23,25,26 DMT formation was decreased by more than 50% by the addition of ketoconazole (0.5 µM). Neither 4-OH-TAM or DMT was formed when the NADPH generating system was omitted, indicating that both are cytochrome-dependent oxidative metabolites.…”

Section: Resultsmentioning

confidence: 97%

Influence of Polyethylene Glycol and Acetone on the in Vitro Biotransformation of Tamoxifen and Alprazolam by Human Liver Microsomes

Cotreau-Bibbo

Moltke

Greenblatt

1996

Journal of Pharmaceutical Sciences

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The use of in vitro hepatic microsomal models as a method of studying the biotransformation of xenobiotics can be complicated by the insolubility of a lipophilic substrate. Addition of solvent to an in vitro system is an approach used to increase solubility of such substrates, but solvents may interact with the microsomal enzymes and cause changes in the kinetic parameters. This study focused on the solvents polyethylene glycol-400 (PEG) and acetone and their influence on the human microsomal in vitro biotransformation of the antineoplastic agent tamoxifen (TAM). The antianxiety agent alprazolam (ALP), a verified P450 3A substrate, was also studied. TAM is a nonpolar drug that is metabolized to two oxidative metabolites, N-desmethyltamoxifen (DMT) and 4-hydroxytamoxifen (4-OH-TAM), by cytochrome P450 isoforms. DMT is formed primarily by 3A isoforms, and the pathway(s) responsible for 4-OH-TAM formation are unknown. Biotransformation of ALP is mediated by P450 3A isoforms, which form alpha-hydroxyalprazolam (alpha-OH-ALP) and 4-hydroxyalprazolam (4-OH-ALP). Both PEG and acetone at 5% of the total microsomal incubation volume were found to solubilize TAM in the in vitro system. However, both solvents had an effect on the P450 mediated metabolism of ALP and TAM. For ALP, PEG was a noncompetitive inhibitor of alpha-OH-ALP (mean Ki = 2.06%) and 4-OH-ALP (mean Ki = 2.37%) formation. Acetone stimulated the production of alpha-OH-ALP, but had no apparent influence on 4-OH-ALP formation. The solvent's influence on TAM metabolism varied. PEG decreased the amount of DMT formed by the microsomes as compared to the system containing no solvent (control); however, 4-OH-TAM formation in the presence of PEG was 146-226% of controls. Samples containing acetone produced smaller quantities of both DMT (39-63%) and 4-OH-TAM (45-69%) as compared to controls. Since PEG and acetone increased the solubility of TAM in the incubation buffer but inhibited or accelerated enzymatic reactions compared to buffer alone, actual solubility in buffer was not a determinant of the rate of TAM metabolism. TAM appeared to be taken up into the microsomal fraction, making it available for biotransformation by the P450 isoforms. Although solvents may increase the solubility of nonpolar agents in aqueous systems, careful evaluation of the effects of solvent on metabolite formation, as compared to buffer controls, is needed in order to properly evaluate an in vitro metabolic model.

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High performance liquid chromatography of tamoxifen and metabolites in plasma and tissues

Cited by 22 publications

References 7 publications

True

True

Determination of DNA Damage in F344 Rats Induced by Geometric Isomers of Tamoxifen and Analogues

Influence of Polyethylene Glycol and Acetone on the in Vitro Biotransformation of Tamoxifen and Alprazolam by Human Liver Microsomes

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