[CANCER RESEARCH 43, 3940-3943, August 1983] Plasma Levels of Estrone, Estrone Sulfate, and Estradici and the Percentage of Unbound Estradici in Postmenopausal Women with and without Breast Disease1 M. J. Reed,2 R. W. Cheng, C. T. Noel, H. A. F. Dudley, and V. H. T. James Department of Chemical Pathology [M. J. R., R. W. C., C. T. N., V. H. T. J.] and Academic Surgical Unit [H. A. F. D.¡,St. Mary's Hospital Medical School, London W2 1PG, England ABSTRACT A preliminary account of some of the results obtained in this study has been presented (11). To investigate the possibility of increased tissue exposure to estrogen in breast cancer patients, plasma levels of estrogens and the percentage of unbound estradiol were measured in SUBJECTS postmenopausal women with benign or malignant breast disease Patients for the present study were recruited from postmenopausal and compared with levels in normal postmenopausal women. women [66 ±10 (S.D.) years old] attending a breast clinic. Blood (20 ml) The percentage of unbound estradiol in breast cancer patients was taken between 2 p.m. and 4:30 p.m. Steroid analyses were carried [1.85 ±0.35% (S.D.)] was significantly higher (p < 0.001) than out before the subject's clinical status was known. It was subsequently in normal postmenopausal women [1.52 ±0.33%] and was still established that some of the women had breast cancer, some had benign significantly higher when patients were matched with control breast disease, and others were women who had previously had breast subjects for weight (p < 0.001) or ideal body weight (p < 0.001). cancer (4 months to 18 years previously) who were undergoing follow- The binding capacity of sex hormone binding globulin was similar up examination. No distinction has been made in the results between women with breast cancer and those who had previously had breast in both groups of women. No significant differences in the plasma cancer. levels of estrone, estradiol, or estrone sulfate were detected Blood samples were obtained from normal postmenopausal women between breast cancer and normal subjects. (57 ±11 years old) between 9 a.m. and 6 p.m. These women were in It is concluded that, given similar concentrations of estradiol good health and without any endocrinological disorder. None of the in plasma of normal and breast cancer subjects, the significant subjects had received any hormone replacement therapy in the 3 months increase found in the unbound estradiol fraction may result in a preceding the study. very small increment in tissue exposure to estrogens in breast Blood obtained from patients and control subjects was centrifuged, cancer subjects. However, even such a small increase in tissue and the plasma was removed and stored at -20° until assayed. exposure to estradiol may be significant, given the length of time required for breast tumor development. MATERIALS AND METHODS INTRODUCTION Plasma levels of estrone, estradiol, and estrone sulfate were measured by methods described previously (4, 15). The percentage of unbound The circumstantial evidence suggesting that estrogens may estradiol was measured in undiluted plasma using a Dianorm dialysis be involved in the development of tumors in hormone-dependent machine (23). Analysis of a plasma pool gave values of 8.3% (n = 10) and 8.7% (n = 15) for the intra- and interassay coefficients of variation tissues such as the breast has resulted in a search for evidence for the measurement of the unbound fraction by this method. The fraction of estrogen excess in women with breast cancer (10). Several of estradiol not bound to SHBG3 was measured using a precipitation studies have been made of urinary estrogen excretion by women technique (16), and intra- and interassay coefficients of variation were with and without breast cancer (24), but few investigations have 4.2% (n = 21) and 5.5% (n = 24), respectively. The binding capacity of been carried out to measure plasma levels of estrogens. In the SHBG was measured by the method of Rosner (20) as modified by present study, plasma levels of estrone and estradiol have been Anderson er al. (2). A detailed description of the techniques used to measured in postmenopausal women with benign or malignant measure the fractions of unbound and non-SHBG-bound estradiol has been published previously (11). A subject's ideal body weight was breast disease and have been compared with levels in normal calculated by comparison of a subject's weight with tables for the postmenopausal women. Plasma levels of estrone sulfate have average weight of women of the same age (6). also been measured in subjects with breast cancer, inasmuch Statistics. Data were analyzed using Student's t test and linear as this hormone is a potential source of unconjugated estrone regression, using the method of least squares. and estradiol. Because measurement of plasma estrogen con centrations may not indicate the level of biologically available estrogen (21), the percentage of unbound estradiol was meas RESULTS ured. In addition, for some patients, the fraction of estradiol Plasma Estrogen Concentration. Plasma levels of estrone, bound to albumin was also determined. estradiol, and estrone sulfate are shown in Table 1. Plasma levels of estrone and estradiol in postmenopausal women with 1This investigation was supported by grants from the Cancer Research Cam benign or malignant breat disease and plasma levels of estrone paign (CP2) and Wellcome Trust (7964/1.5). 2 To whom requests for reprints should be addressed. Received November 17, 1982; accepted May 6, 1983. 3The abbreviation used is: SHBG, sex hormone binding globulin. 3940 CANCER RESEARCH VOL. 43 Downloaded from cancerres.aacrjournals.org on October 1, 2021. © 1983 American Association for Cancer Research. Estrogen and Breast Cancer Table1 = -0.36; p < 0.01) and also between SHBG binding capacity Plasma concentrations of estrone, estrone sulfate, and estradici in and subjects' percentage of ideal body weight (r = -0.33; p < postmenopausal breast disease patients and normal postmenopausal women 0.05). sulfate(pg/mi)302 (pg/ml)28.2±11.2"(35)i> (pg/ml)14.0 35 Normal ±197(46) ±6.0(32) Benign 21 .8 ± 5.4 ( 5) 11.4 ±5.2(10) CancerEstrone 31 .4 ±13.4 (26)Estrone 307 ±177(17)Estradici13.5 ±6.1(43) ' Mean ±S.D. ' Numbers in parentheses, number of subjects. 50 60 70 80 90 100 Body weight (Kg) Chart 2. Correlation between body weight and plasma levels of estradiol (E2)in normal postmenopausal women (•)and in postmenopausal women with benign (A) or malignant (•)breast disease. 40 50 60 70 80 90 100 700 Body weight (Kg) Chart 1. Correlation between body weight and plasma levels of estrone (E,) in 600 normal postmenopausal women (•)and in postmenopausal women with benign (A) or malignant (•)breast disease. 500 sulfate in patients with breast cancer did not differ from the 400 levels in normal postmenopausal women. Significant correlations CD were found between plasma levels of estrone (Chart 1) and a subjects' body weight and also between plasma levels of estra 300 dici and body weight (Chart 2). No such correlation was found between plasma levels of estrone sulfate and body weight (Chart 200 3) for patients with breast disease (r = 0.07, not significant). 100 r=oi3 Percentage of Unbound Estradici, Percentage of Non- N.S. SHBG-bound Estradici and SHBG Binding Capacity. The frac tion of unbound estradici in plasma from normal women and 40 50 60 70 80 90 100 patients with breast disease is shown in Chart 4 and Table 2. Body weight (Kg) The fraction of unbound estradici was significantly higher (p < Chart 3. Lack of significant correlation between body weight and plasma levels 0.001 ) in the breast cancer group than in normal women and of estrone sulfate (£,S)in postmenopausal women with breast cancer. also in women with benign breast disease (p < 0.02). Because the binding capacity of SHBG is reduced in obese women, the unbound estradiol fraction was compared in a group (16) (33) (60) of weight-matched controls and breast cancer patients and also rP<0-02n rP<0-001, in a group matched for ideal body weight (Chart 5), and a - significant difference (p < 0.001) was still found. ui 2 The fraction of estradiol not bound to SHBG (I.e., unbound plus mainly albumin bound) was also measured in the women with breast disease (Table 2), but there was no significant difference between women with benign or malignant disease. There was no significant difference in the binding capacity of SHBG (expressed as ng 5 a-dihydrotestosterone per 100 ml plasma) in patients with breast cancer [2.1 ±1.2 (S.D.)], in those with benign breast disease [2.5 ±1.8], or in normal postmeno Benign Normal Breast Cancer pausal women [2.0 ±1.4] (Table 2). For patients with breast Chart 4. Percentage of unbound estradiol (E¡)in plasma from normal postmen disease, significant negative correlations were found between opausal women and those with benign or malignant breast disease. Numbers in the fraction of unbound estradiol and SHBG binding capacity (r parentheses, number of subjects. AUGUST 1983 3941 Downloaded from cancerres.aacrjournals.org on October 1, 2021. © 1983 American Association for Cancer Research. M. J. Reed et al. Table 2 Tabte3 Percentage ol unbound estradici, percentage oínon-SHBG-bound estradiol, and Concentrations of unbound, non-SHBG-bound, SHBG-bound, and albumin-bound SHBG binding capacity in postmenopausal breast disease patients and normal estradiol in postmenopausal women with benign or malignant breast disease postmenopausal women Albumin- binding ca Non-SHBG- SHBG-bound bound pacity (¿ig5a-dihy- Unbound estradiol bound estradiol estradiol estradiol of unbound of non-SHBG- drotestosterone/ (pg/100 ml) (pg/100 ml) (pg/100 ml) (pg/100 ml) estradiol1.52 boundestradiol31 100ml)2.0 19.5±10.0s(10)" Benign 411±331(8) 838±227(8)434±318(8) ±0.33" (33)" Normal ±1.4 (32) Cancer 24.1±12.4(42) 540±363(24)1056±408(24)530±346(24) 1.79±0.22C(16) Benign .2 ±12.7 (10) 2.5 ±1.8 (15) a Mean ±S.D.