Breast Cancer Research and Treatment

, Volume 131, Issue 2, pp 473–481 | Cite as

Tamoxifen metabolites as active inhibitors of aromatase in the treatment of breast cancer

  • Wenjie Jessie Lu
  • Zeruesenay Desta
  • David A. Flockhart
Preclinical Study

Abstract

The mechanism of tamoxifen action in the treatment of breast cancer is believed to be via active metabolites that act as potent estrogen receptor antagonists. Attempts to identify relationships between active metabolite concentrations and clinical outcomes have produced mixed results. Since anti-estrogenic effects may be brought about not only by estrogen antagonism, but also by reduced estrogen synthesis, we tested the ability of tamoxifen and its principal metabolites to inhibit aromatase in vitro. The activity of human aromatase in both recombinant and placental microsomal preparations was measured using the rate of generation of a fluorescent metabolite in the presence and absence of multiple concentrations of tamoxifen, endoxifen, N-desmethyl-tamoxifen, and Z-4-hydroxy-tamoxifen. Aromatase inhibition was further characterized by measuring the inhibition of testosterone metabolism to estradiol. The biochemical mechanisms of inhibition were documented and their inhibitory potency was compared. Using recombinant human aromatase, endoxifen, and N-desmethyl-tamoxifen were able to inhibit aromatase activity with Ki values of 4.0 and 15.9 μM, respectively. Detailed characterization of inhibition by endoxifen and N-desmethyl-tamoxifen indicated non-competitive kinetics for both inhibitors. Similarly, endoxifen-inhibited testosterone metabolism via a non-competitive mechanism. No appreciable inhibition by tamoxifen or Z-4-hydroxy-tamoxifen was observed at similar concentrations. The relative inhibitory potency was: endoxifen > N-desmethyl-tamoxifen >>> Z-4-hydroxy-tamoxifen > tamoxifen. Similar data were obtained in human placental microsomes. Endoxifen and N-desmethyl-tamoxifen were found to be potent inhibitors of aromatase. Inhibition by these tamoxifen metabolites may contribute to the variability in clinical effects of tamoxifen in patients with breast cancer. Relationships between tamoxifen metabolite concentrations and clinical outcomes may be complex, and the biologic mechanisms that underlie these relationships may include aromatase inhibition.

Keywords

Tamoxifen Endoxifen Breast cancer Aromatase inhibitor Estrogen 

List of symbols

Endoxifen

4-Hydroxy-N-desmethyl-tamoxifen

4HT

Z-4-hydroxy-tamoxifen

NDMT

N-desmethyl-tamoxifen

CYP

Cytochrome P450

CYP19

Aromatase

MFC

7-Methoxy-4-trifluoromethylcoumarin

HFC

7-Hydroxytrifluoromethylcoumarin

HPLC

High performance liquid chromatography

UV

Ultraviolet

IC50

The half maximal inhibitory concentration

Km

The Michaelis constant

Vmax

The maximum reaction rate

Ki

The equilibrium dissociation constant of the inhibitor

KSapp

The apparent Michaelis constant

Vmaxi

The apparent maximum reaction rate in the presence of the inhibitor

Notes

Acknowledgments

The authors are grateful to Evan Ogburn MSc for providing technical assistance. This study was supported in part by the National Institutes of Health National Center for Research Resources [Grant K24RR020815] to DF, by the National Institute for General Medical Sciences [Grants T32GM008425, U01GM061373] to DF, and by the Department of Defense Breast Cancer Research Program Predoctoral Fellowship [W81XWH-11-1-0016] to WL.

Conflict of interest

WJL and DAF are authors of a patent submitted to the U.S. patent office that describes new uses of the chemical structures described herein entitled “Materials for inhibiting aromatase and method of using the same to diagnose, treat and monitor breast cancer”.

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Copyright information

© Springer Science+Business Media, LLC. 2011

Authors and Affiliations

  • Wenjie Jessie Lu
    • 1
  • Zeruesenay Desta
    • 2
  • David A. Flockhart
    • 2
  1. 1.Division of Clinical Pharmacology, Department of Pharmacology and ToxicologyIndiana University Simon Cancer Center, Indiana University School of MedicineIndianapolisUSA
  2. 2.Departments of MedicineIndiana University Simon Cancer Center, Indiana University School of MedicineIndianaUSA

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