Review of Studies on Estrogen Biosynthesis in the Human1

While the gonads and placenta are usually thought of as the principal sites of estrogen synthesis, many other tissues are now known to contain the aromatase enzyme. Despite a wealth of information, the functional significance, if any, of this wide spread distribution is not yet clear. Nonetheless, elevated estrogen production resulting from either an apparent increase in enzyme activity or increased substrate availability in blood can have important effects on target tissues such as the breast, uterus, and bone. Our early studies suggesting that aromatase inhibitors can effectively reduce the impact of peripheral estro gen synthesis have been amply confirmed by the elegant studies of Santen and others. Many previous studies have suggested that aromatase activity may be present in breast tumors. The profound implications for growth of estrogen-sen sitive tumors led us to reevaluate this question. Using a sensi tive modification of our original radiometrie aromatase assay, we have found detectable activity in only about 40% of human breast tumors (n = 100). Only rarely (1/100) does the aro matase activity exceed 1.0 pm/hr/g tissue or 0.001 % of that found in the human placenta. These quantitative results contra dict several published reports which suggest that most breast tumors contain biologically significant levels of aromatase ac tivity. The mammalian aromatase (estrogen synthetase) enzyme catalyzes the formation of aromatic C)8 estrogenic steroids from dg androgens containing the A", 3-ketone grouping in Ring A of the steroid nucleus. This enzyme has received considerable attention because of the complexity of the chem ical reaction (Chart 1) and also because of the central impor tance of estrogens ¡nmany reproductive and metabolic pro cesses. The uniquely large capacity of the human placenta to synthesize estrogens provided not only convenient starting material (urine) for the isolation of steroid hormones in the early dawn of endocrinology but also a readily available tissue for biochemical studies of the aromatase enzyme. As became evident during these conference proceedings, interest in aro matase now extends far beyond human pregnancy to diverse areas such as its role in sexual behavior and its pharmacolog ical inhibition ¡ntreatment of estrogen-dependent cancer. In view of the many comprehensive reviews that are available (7, 10, 40, 42), I will confine this article to a brief account of my own interests in aromatase and its products over the past 20 years. Following completion of my Ph.D. work with Seymour Lieberman at Columbia University, I joined Paul MacDonald in the Department of Obstetrics and Gynecology at Southwestern Medical School in Dallas in 1962. We set up a laboratory and immediately began to work on the problem of estrogen produc tion and metabolism in pregnant women. While still in New York, I had conceived a new approach to the difficult problem of isolating estrogens from the urine of pregnant women by gradient elution chromatography on Celite using ethylene glycol and mixtures of isooctane and ethyl acetate. This system, which was adapted to microcolumn purification of steroids prior to radioimmunoassay by Abraham er al. (1), made it possible for us to isolate estrone, estradici, and estriol in pure form from 5 to 10 liters of urine in about 5 days. However, when we hit upon the idea that placental estrogens may be derived from circulating DS,2 it took only about 48 hr to obtain the answer from the urine of the first subject who had received 3H-labeled DS and 14C-labeled estradiol. When both 3H and '"C channels of the scintillation counter lit up, our shouts and whoops of excitement went unheard, as it was about 4 a.m. We quickly confirmed this discovery and published both the methodology and preliminary findings in December 1963 (36, 38). The unique experimental design, using 2 isotope-labeled tracers, which allows quantitation of conversion and production rates of precursor and product, proved to be a powerful tool that has solved many problems during the ensuing 18 years. We continued our studies of pregnancy and established the following major points: (a) placental conversion of maternal DS increases from about 1 to 2% at 6 to 7 weeks to 30 to 40% at term of normal gestation and accounts for about 50% of total estradiol production (38, 39); (b) estriol is formed by 16 ahydroxylation of DS prior to aromatization, and only about 10% of the total is derived from maternal precursors (39); (c) estro gen production during anencephalic pregnancy is derived al most exclusively from maternal DS and is proportional to the level of adrenal secretory activity (24); (cOtrophoblastic tissues in women pregnant with hydatidiform moles or in patients with choriocarcinoma retain the capacity to aromatize DS (20, 25); and (e) the conversion of DS to estrogens is negligible in nonpregnant individuals (25). This work firmly established the important principle of hormone production from inactive pre cursor molecules in or near target tissues which is now rec ognized in many other endocrine systems. In about 1966, we began to study the human placental aromatase system in vitro. We were particularly interested in the kinetics of the overall aromatization process but were initially discouraged by the painfully tedious assays involving isolation of radiolabeled estrogenic products by chromatogra phy. After reading several papers in which 1,2-3H-labeled androgens were used in metabolic studies and yields of estro gen had to be corrected for losses, it occurred to me that this loss could be used to advantage in assaying aromatase activity. Our first description of a tritium release (radiometrie) assay was presented at the 1969 meeting of the Society for Gyne1Presented at the Conference "Aromatase: New Perspectives for Breast Cancer," December 6 to 9, 1981, Key Biscayne, Fla. 1 The abbreviations used are: DS, dehydroisoandrosterone sulfate; 19-OHA, 19-hydroxyandrostenedione; 19-OXOA. 19-oxoandrostenedione.