QUANTIFYING ESTROGEN METABOLISM - AN EVALUATION OF THE REPRODUCIBILITY AND VALIDITY OF ENZYME IMMUNOASSAYS FOR 2-HYDROXYESTRONE AND 16-ALPHA-HYDROXYESTRONE IN URINE

Rapid and simple enzyme immunoassays (EIAs) were recently developed to measure 2-hydroxyestrone and 16 alpha-hydroxyestrone in unextracted u rine. The balance between these competing estrogen metabolism pathways may serve as a biomarker of breast cancer risk. Before testing these assays in epidemiologic studies, we evaluated their reproducibility, a nd validity relative to gas chromatography-mass spectroscopy (GC-MS). Overnight 12-hr urine collections from five midfollicular premenopausa l women, five midluteal premenopausal women, and five postmenopausal w omen were aliquoted and stored at -70 degrees C. Two aliquots from eac h woman were assayed with the EIAs in a random, blinded order, monthly over 4 months and 1 year later. Reproducibility over 4 months was goo d for both metabolites in premenopausal women (coefficient of variatio n = 8-14%) and satisfactory in postmenopausal women (similar to 19%). Reproducibility over 12 months remained good in premenopausal women, b ut was poor in postmenopausal women, with mean readings increasing 50 to 100%. Wide variation in estrogen metabolite levels enabled a single EIA measurement to characterize individual differences among premenop ausal women in midfollicular (intraclass correlation coefficient = 98- 99%) and midluteal phase (85-91%). A narrower range in metabolite leve ls among postmenopausal women reduced discrimination (78-82%). The cor relation between EIA and GC-MS measurement was excellent for both meta bolites (r > 0.9), except for 2-hydroxyestrone in postmenopausal women (r = 0.6). Analysis of absolute agreement suggested that both EIAs we re less sensitive than GC-MS, and each detected nonspecific background . The tow concentration of estrogen metabolites in urine from postmeno pausal women may explain the problems with reproducibility and validit y in this menstrual group. Accordingly, more sensitive EIAs have been developed and are now being evaluated.