Ultrasensitive Serum Estradiol Measurement by Liquid Chromatography-Mass Spectrometry in Postmenopausal Women and Mice

Abstract

Accurate measurement of very low circulating estradiol (E₂) (<5 pg/ml) in postmenopausal women and in mice is essential to investigating sex steroid action in target tissues. However, direct immunoassays are too inaccurate and conventional mass spectrometry-based measurement too insensitive at these serum E₂ levels. We report application of an ultrasensitive method using a novel estrogen-selective derivatization in liquid chromatography-mass spectrometry to measure serum E₂, with a detection limit of 0.25 pg/ml in small (0.2 ml) serum volumes that can quantify serum E₂ in 98% and serum E₁ in 100% of healthy postmenopausal women. Aromatase inhibitor (AI) treatment of postmenopausal women with breast cancer further reduces serum E₂ by 85% and serum estrone (E₁) by 80%. The wide scatter of circulating E₂ in AI-treated women suggests that the degree of sustained E₂ depletion, now quantifiable, may be an efficacy or safety biomarker of adjuvant AI treatment. This ultrasensitive method can also measure serum E₂ in most (65%) female but not in any male mice. Further studies are warranted using this and comparable ultrasensitive liquid chromatography-mass spectrometry estrogen measurements to investigate the relationship of circulating E₂ (and E₁) in male, postmenopausal female, and childhood health where accurate quantification of serum estrogens was not previously feasible. This will focus on the direct impact of estrogens as well as the indirect effects of androgen aromatization on reproductive, bone, and brain tissues and, notably, the efficacy and safety of AIs in adjuvant breast cancer treatment.