Adrenal Dehydroepiandrosterone and Human Mammary Cancer1

[CANCER RESEARCH 38. 4036-4040. November 1978] 0008-5472/78/0038-OOOOS02.00 Adrenal Dehydroepiandrosterone and Human Mammary Cancer1

John B. Adams,2 Lesley Archibald, and Christine Clarke

School ot Biochemistry. University of New South Wales, Sydney. Australia

Abstract tion to weight (13, 15). Calculations suggested that over one-half of the differences in breast cancer incidence be A hypothesis implicating adrenal dehydroepiandroster- tween regions in Holland and Japan can be attributed to one (DHEA) (sulfate) in the etiology of human breast cancer of the "adrenal" or "Western" type has been differences in body weight and height (15). As pointed out by de Waard, such a combination can be interpreted as a presented (Adams, J. B. Steroid Hormones and Human measure of body surface area (15). A recent hypothesis has Breast Cancer. An Hypothesis. Cancer, 40: 325-333, implicated adrenal DHEA3-DHEAS in the etiology of human 1977). High concentrations of DHEA sulfate in the blood breast cancer, particularly of the "Western" or "adrenal" provide a potentially high flux of the free steroid to type (2). It was demonstrated that the metabolism of these mammary tumors, due to the presence therein of a sulfa steroids by human breast carcinoma tissue was very similar tase. The free steroid, in turn, is metabolized by human to that by skin; one possibility for this similarity is the mammary tumors in vitro to 5-androstene-3/i,17/J-diol reputed derivation of the mammary gland from primitive (ADIOL) and 7-hydroxydehydroepiandrosterone. It has sweat glands. It is possible that DHEA-DHEAS, the exact now been found that ADIOL when administered s.c. to physiological function of which remains unknown, may immature female rats will deplete the estrogen receptor in utilize skin as a major target organ and hence may also the uterine cytosol. Similarly, dimethylbenzanthracene- influence the mammary gland. induced rat mammary tumors, when incubated in vitro Recently, it has been shown that ADIOL, which is pro with 1 /IM ADIOL, show translocation of the estrogen duced from DHEA-DHEAS in both skin (20) and mammary receptor from cytosol to the nucleus, as measured by cancer tissue (7, 22), can compete very effectively for the exchange assays. The magnitude of depletion of cytosol estrogen receptor (28). Thus ADIOL could conceivably be estrogen receptor by ADIOL was less than that obtained one of the factors linking the observations of de Waard on with 17/i-estradiol, studied at 0.03 /.IM concentrations, but body weight and height (surface area) with the proposed greater than that with 1 fiM dihydrotestosterone. Among a involvement of adrenal DHEA-DHEAS in postmenopausal wide group of C steroids examined as possible inhibitors breast cancer (2). In this paper the ability of ADIOL to cause of estrogen sulfotransferase, both DHEA and ADIOL translocation of the estrogen receptor and to act as a potent showed marked inhibitory properties. By contrast, both inhibitor of estrogen sulfotransferase is demonstrated. 7a- and 7/i-hydroxydehydroepiandrosterone showed neg ligible inhibitory effects. This paralleled previous findings on the influence of a 7-hydroxyl group in modifying the Materials and Methods ability of ADIOL to compete effectively for the estrogen Chemicals. DHEA, ADIOL, 17/3-estradiol, and DHT used receptor. Thus the high levels of 7-hydroxylase found in in animal experiments were obtained from Sigma Chemical human mammary tumors, and acting on both DHEA and Co., St. Louis, Mo. Steroids used in the enzyme inhibition ADIOL, may function in controlling the intracellular con studies were obtained from the following sources: testos centrations of these steroids. terone, androsterone, and epiandrosterone from Koch- Light Laboratories, Colnbrook, England; 1-androstene- Introduction 3,17-dione, DHT, and 5a-androstane-3,17-dione from Ster- aloids, Wilton, N. H. 4-Androstene-3,17-dione, 50-andros- De Waard (14, 16) first suggested that 2 types of human tan-3a-ol-11,17-dione, DHEA, ADIOL, and Norit A from breast cancer may exist, one associated with ovarian hor Sigma; etiocholanolone, 5-androstene-3ÃŸ,16Â«-diol-17-one, mones in the premenopausal period and the other with a and 5-androstene-30,16/3-diol-17-one from Ikapharm, Ra- postmenopausal hormonal environment. He termed the mat-Gan, Israel; 7Â«-hydroxytestosterone, 5Â«-androstane- latter the "Western" environmental type, due to the low 3a,16/3,17/3-triol, 5a-androstane-3Â«,16o,17/3-triol, 7Â«- and incidence of this type of breast cancer in Asian countries, 7/3-hydroxydehydroepiandrosterone, and 5-androstene- particularly Japan. Variation in international breast cancer 3/3,17a-diol kindly supplied by the Medical Research Coun incidence was suggested to be related to body weight and cil, Steroid Reference Collection, London, England; 17/3- nutritional status (14, 16). More recently, the emphasis on [2,4,6,7-3H]estradiol (specific activity, 96 Ci/mmol) from the weight alone has been modified to include height in addi Radiochemical Centre, Amersham, England; Eagle's basal medium (dry powder) through the Commonwealth Serum ' Presented at the John E. Fogarty International Center Conference on Hormones and Cancer, March 29 to 31, 1978, Bethesda, Md. Supported by grants from the Australian National Health and Medical Research Council. 3The abbreviations used are: DHEA, dehydroepiandrosterone; DHEAS. 2To whom requests for reprints should be addressed, at School of dehydroepiandrosterone 3-sulfate; ADIOL. 5-androstene-30.170-diol; DHT, Biochemistry. University of N.S.W.. P. O. Box 1. Kensington, N. S. W. 2033 17ÃŸ-hydroxy-5u-androstan-3-one; DMBA, 1.12-dimethylbenz(a)anthracene; Australia. i g intragastric.

4036 CANCER RESEARCH VOL. 38

Laboratory, Melbourne, Australia; Bio-Gel HTP (DMAgrade) from stock solutions in propylene glycol. The concentration from Bio-Rad Laboratories, Richmond, Calif. of the latter was held constant at 6% (v/v). Estrogen Receptor in Rat Uteri. The experimental pro Liquid Scintillation Counting. This was carried out with a cedure for studying the influence of ADIOL on rat uteri Triton-toluene phosphor system (5) and computerized cytosol receptor was based on that described by Rochefort Packard 2650 instrument. Quench corrections were carried and Garcia (24) in studies of the effects of DHT on estrogen out by the external standard method. receptor translocation. Immature female Wistar rats (20- to 22-day-old) were used in groups of 5. ADIOL was adminis Results tered s.c. in doses of 8 to 10 mg in 1 ml of peanut oil containing 10% (v/v) ethanol. Controls received the vehicle Although ADIOL will compete effectively for the estrogen alone. Cytosol was prepared by centrifugation at 100,000 x receptor in cytosol preparations of human mammary tu g for 1 hr. Estrogen receptor was assayed, after brief mors and myometrium (28), to our current knowledge no treatment of cytosol with charcoal to remove any endoge experiments have been reported wherein the effects of nous free steroid, by a single saturating dose procedure ADIOL have been examined, either in intact animals or in with 20 nM 17/3-[3H]estradiol in the presence and absence tissue preparations. Rochefort and Garcia (24) have shown of 2 fj.M cold 17/3-estradiol. Incubations were continued for that high doses of DHT provoked the in vivo translocation 20 hr at 2Â°,and unbound steroid was removed by dextran- of estrogen receptor in the immature rat uterus. Under coated charcoal (24). similar conditions high doses of ADIOL caused a depletion In Vitro Studies on Estrogen Receptor Translocation in of accessible estrogen receptor sites in the rat uterine DMBA-induced Rat Mammary Tumors. Mammary tumors cytosol. This reached a maximum after some 4 hr when the were induced in 50-day-old female Sprague-Dawley rats by estrogen receptor sites were reduced to 30% of that in a single i.g. feeding of 20 mg of DMBA in 1 ml of peanut oil. control animals (Chart 1). A dose of 8 mg was administered Animals were sacrificed, and tumors (weighing about 2 g) since a dose of this order achieved the maximum effect in were removed, washed in 0.9% NaCI solution, and minced the case of DHT (24). finely with scissors. The tissue pieces were then mixed DHT at concentrations of 1 to 50 Â¡J.Mwill cause the thoroughly to provide a homogeneous sample. Weighed translocation of estrogen receptor in immature rat uteri in portions (usually 0.5 g) were placed in Eagle's medium (3 vitro (25, 26). In DMBA-induced rat mammary tumors in ml), buffered with bicarbonate to pH 7.2. Steroid was added vitro, 17/3-estradiol, ADIOL, and DHT all individually caused to the empty flasks from stock solutions, solvent was depletion of total (exchangeable) estrogen receptor sites in evaporated with N2, and propylene glycol (5 /nl) was added. the cytosol; their relative effectiveness is in the order listed After being gassed with carbogen, the flasks were stop (Charts 2 and 3). pered and shaken at 37Â°for1 hr. With the newly described exchange assay for mammary The method of isolation of cytoplasmic and nuclear tumor nuclear estrogen receptor, which overcomes the fractions, and the assay of estrogen receptor in these problem of nuclear receptor destruction by proteases (19), fractions, was that of Carola and McGuire (19), the only evidence has been obtained that depletion of cytosol estro exception being that 1 mM dithiothreitol was used in place gen receptor is accompanied by an increase in receptor in of monothioglycerol in the buffers. Essentially, the assay the nucleus (Chart 3). It would appear that estrogen recep involves an exchange at 25Â°(cytosol) and 30Â°(nuclear tor accumulation in the nucleus may be more prolonged extract), the receptor in the nuclear extract being absorbed with ADIOL. However, at the time of writing for this meeting, onto hydroxyapatite prior to incubation with [3H]estrogen these results must be regarded as preliminary only and will to prevent destruction by proteases present in such extracts need further confirmation. from mammary tumors (18, 19). Estrogen receptor ab sorbed onto hydroxyapatite (Bio-Gel HTP) was incubated for 3.5 hr at 30Â°with5.6 nM 17/3-[3H]estradiol, plus or minus 100 0.5 fjiM unlabeled 17/3-estradiol, contained in 0.5 ml of TK I buffer (10 mM Tris: MCI (pH 8.5), 0.6 M KCI, 1.0 mM dithio 0 threitol, containing 10% (v/v) glycerol). Frequent vortexing 2 was carried out during this period. Cytosol estrogen recep I tor was assayed, after prior treatment with charcoal at 2" J BO for 10 min (24), by incubation for 20 hr at 25Â°with 5.6 nM Ã® 170-[3H]estradiol, plus or minus 0.5 /J.M unlabeled 17ÃŸ- estradiol. Tubes were then placed in ice prior to addition of dextran-coated charcoal, and the procedure of Carola and McGuire was followed (19). Inhibition Studies with Estrogen Sulfotransferase. Es trogen sulfotransferase (3'-phosphoadenylylsulfate:estrone 1 2 3 4 5 TIME HR Sulfotransferase, EC 2.8.2.4) was isolated from bovine pla Chart 1. Effect of ADIOL on estrogen receptor in immature rat uterine centa (6) and assayed as described previously (4). Inhibition cytosol in vivo. Groups of rats received 8 mg ADIOL s.c , and available estrogen receptor sites in cytosol, prepared from pooled uteri, were assayed by various nonsubstrate steroids was carried out with 25 at time intervals shown. The percentage of depletion of estrogen receptor /Â¿Mconcentrations of the steroids, the latter being added with time is shown compared to controls receiving vehicle alone.

NOVEMBER 1978 4037

3OO Sulfotransferase activities in human mammary tumors, measured with DHEA and 17ÃŸ-estradiolas substrates, have been reported to correlate with the individual patient's CONTROL prognosis and response to hormone ablation. A lack of Å’ 200 measurable activity or activities in which the ratio of DHEAS DHT to estradici sulfate was <1 reflected the poorest prognoses (12). Sulfurylation of estrogen in the endometrium is under hormonal control, and estrogen sulfotransferase is

10O switched on after the proliferative phase of the cycle (9, 23). In this instance then, the sulfurylation represents an inacti-

DIOL vation process. From the data in Table 1, nonsubstrate C,,, steroids can inhibit purified estrogen sulfotransferase to n differing degrees; DHEA, and particularly ADIOL, exhibits Chart 2. Depletion of exchangeable estrogen receptor (flÂ£)in cytosol of high inhibition. This is also shown in Chart 4A. High DMBA-induced rat mammary tumors in vitro. Tumor tissue was incubated concentrations of a 7-hydroxylase acting on DHEA and for 1 hr in Eagle's medium containing either 0.03 Â¿iM17/3-estradiol(Â£),1 Â¿iM ADIOL and found in human mammary tumors (11, 22) 5-androstene-3/3.170-diol (DIOL), or 1 MMDHT. prompted an examination of the influence of 7-hydroxyl- ation of DHEA on its ability to inhibit estrogen sulfotransfer TUMOR 1 ase. It is seen that 7-hydroxydehydroepiandrosterone ex hibited negligible inhibition of the enzyme (Table 1; Chart DHT 200 D CYTOSOL 4B). â€¢¿NUCLEUS TUMOR , 10Oi B DHT

100 -I Chart 3. Distribution of estrogen receptor following in vitro incubation of DMBA-induced rat mammary tumors with various steroids. Conditions as in Chart 2. Exchangeable estrogen receptor (RE) sites assayed in both cytosol and nucleus. Tumor 1 was more fibrous than Tumor 2, which may have O.2 delayed access of steroid. E. 17/a-estradiol; DIOL, 5-androstene-3/3,17/3-diol; C, control.

Table 1 2 4 6 8 1O 2 4 6 8 1O Inhibition of estrogen sulfotransferase by nonsubstrate C,,, E2 juM E2 /Â¿M steroids Chart 4. Effect of nonsubstrate C,, steroids on rate of formation of All steroids were tested at a concentration of 25 /Â¿M.The estradici 3-sulfate by purified estrogen sulfotransferase. A: â€¢¿control;A, concentration of estrogen was 7 Â¿Â¿M. testosterone; D, DHEA; O, ADIOL. B: â€¢¿.control;D, DHEA; A, 7a-hydroxy- Sulfurylation dehydroepiandrosterone. E2, estradici. All steroids were tested at 25 //M relative to con- concentrations. In ÃŸ,3times the enzyme concentration of A was used. Steroid trol as 1.0 CELL 1.0 BLOOD 4-Androsten-17/3-ol-3-one (testosterone) 0.83 4-Androsten-7a, 17ÃŸ-diol-3-one 0.99 4-Androstene-3,17-d ione 0.99 7-OH DHEA 7-TRIOL 1-Androstene-3,17-d ione 0.89 \ / 7-hydroxylase 5a-Androstan-17/3-ol-3-one5a-Androstan-3a-ol-17-one (androsterone)5a-Androstane-3/3-ol-1 DHEA (epiandrosterone)5a-Androstane-3,17-dione5a-And7-one DHEAS â€”¿-DHEA â€”¿ DIOLâ€”â€¢ RE...__

7/3-triol5a-Androstane-3a ,16/3,1 7/3-triol5/3-Androstan-3a-ol-1rostane-3a ,16a ,1 DHEAS- (etiocholanolone)5ÃŸ-Androstan-3a-ol-17-one ,17-dione5-Androstene-3j3-ol-11 Eâ€”Â» ANDROGENS 7-one5-Androstene-30,7a-diol-1 7-one5-Androstene-30,7j8-diol-1 7-one5-Androstene-3/3,17a-diol5-And Chart 5. Summary of main reactions involving DHEA-DHEAS metabolism iol5-Androstene-3/3,16a-diol-1rostene-3/3,170-d in the mammary tumor cell. RA and RE, androgen and estrogen receptors, 7-one5-Androstene-3/3,16/3-diol-1 respectively. E, 17/3-estradiol; ES. estradiol 3-sulfate; 7-OH DHEA, 7a- 7-one0.750.740.770.870.900.820.891.060.700.951.040.970.610.850.94hydroxydehydropiandrosterone; 7-TRIOL, 5-androstene-3/3,7a,17/3-triol; broken arrows, inhibition of estradiol sulfotransferase.

4038 CANCER RESEARCH VOL. 38

Discussion (3). Such a situation would lead to higher intracellular levels of free DHEA which, in turn, would be expected to inhibit It is well documented that the major urinary metabolites the sulfurylation of estrogen (Chart 4) and thus maintain its of DHEA-DHEAS, namely, androsterone and etiocholano- concentration. lone, are lowered in patients with breast cancer. Evidence A scheme summarizing the possible involvement of has also been presented that these metabolites are subnor DHEA-DHEAS at the level of the tumor cell is shown in Chart mal at the preclinical stage of the disease (10). Since body 5. weight and height are positively correlated with increased risk of breast cancer (13, 15), it is feasible that altered References metabolism may be responsible. While this could occur via 1. Abul-Hajj, Y. J. Metabolism of Dehydroepiandrosterone by Hormone the liver (8), involvement of body weight and height would Dependent and Hormone Independent Human Breast Carcinoma. Ste perhaps rather implicate lean body mass or skin (surface). roids, 26. 488-500, 1975. An increase in conversion of DHEA-DHEAS to ADIOL would 2. Adams, J. B. Steroid Hormones and Human Breast Cancer. An Hypoth esis. Cancer, 40: 325-333, 1977. be one metabolic route compatible with these changes. 3. Adams, J. B., and Chandra, D. P. Dehydroepiandrosterone Sultotrans- Although ADIOL would seem to be able to cause translo ferase as a Possible Shunt for the Control of Steroid Metabolism in Human Mammary Carcinoma. Cancer Res., 37: 278-284, 1977. cation of estrogen receptor in rat uteri in vivo (Chart 1) and 4. Adams, J. B., Ellyard, R. K., and Low, J. Enzymic Synthesis of Steroid rat mammary tumor in vitro (Charts 2 and 3), it remains to Sulphates IX. Physical and Chemical Properties of Purified Estrogen Sulphotransferase from Bovine Adrenal Glands, the Nature of Its Isoen be seen whether such translocations are accompanied by a zymic Forms and a Proposed Model to Explain Its Wave-like Kinetics. biological response. The work of Garcia and Rochefort (17) Biochim. Biophys. Acta, 370: 160-188. 1974. has demonstrated that translocation of estrogen receptor 5. Adams, J. B., and Li, K. Biosynthesis of 17/3-Oestradiol in Human Breast Carcinoma Tissue and a Novel Method for Its Characterization. Brit. J. in rat uteri by DHT in vivo was accompanied by stimulated Cancer, 31: 429-433, 1975. protein synthesis, including the specific uterine protein 6. Adams, J. B., and Low, J. Enzymic Synthesis of Steroid Sulphates X. normally induced by estrogen. However, induction of this Isolation of Oestrogen Sulphotransferase from Bovine Placenta and Comparison of Its Properties with Adrenal Estrogen Sulphotransferase. uterine-induced protein by DHT in this system in vivo could Biochim. Biophys. Acta, 370.-189-196, 1974. not be demonstrated by other workers, although induction 7. Adams, J. B., and Wong, M. S. F. Paraendocrine Behaviour of Human Breast Carcinoma: In vitro Transformation of Steroids to Physiologically was established by in vitro experiments (27). Active Hormones. J. Endocrinol., 41: 41-52, 1968. It remains a possibility that the translocation of estrogen 8. Brooks, S. C. The Metabolism of Steroid Hormones in Breast Cancer: A receptor by ADIOL may occur by prior conversion to estro Reappraisal. In: K. M. J. Menon and J. R. Reel (eds.). Steroid Hormone Action and Cancer, pp. 36-50. New York: Plenum Press, 1976. gen in the tissue. However, although low conversions of 9. Buirchell, B. J., and HÃ¤hnel,R. Metabolism of Estradiol-17/3 in Human DHEA to 17j3-estradiol have been demonstrated in some Endometrium during the Menstrual Cycle. J. Steroid Biochem., 6. 1489- human mammary tumors in vitro (1), in our experience with 1494,1975. some 15 tumors incubated with labeled DHEA of high 10. Bulbrook, R. D., Hayward, J. L., and Spicer, C. C. Relation between Urinary Androgen and Corticoid Excretion and Subsequent Breast specific activity, no conversion to estrogen could be de Cancer. Lancet, 2. 395-398, 1971. tected (K. Li and J. B. Adams, unpublished observations). 11. Couch, R. A. F., Skinner, S. J. M., Tobler, C. J. P., and Douss, T. W. The in vitro Synthesis of 7-Hydroxy Dehydroepiandrosterone by Human In addition, the enhanced effect of ADIOL, compared to Mammary Tissues. Steroids, 26: 1-15, 1975. DHT, in translocating estrogen receptor (Charts 2 and 3) is 12. Dao, T. L., and Libby, P. R. Steroid Sulfate Formation in Human Breast Tumors and Hormone Dependency. In: T. L. Dao (ed.), Estrogen Target consistent with its higher affinity for the estrogen receptor Tissues and Neoplasia, pp. 181-200. Chicago: Chicago University Press, as compared to DHT (28). The major metabolite of labeled 1972. DHEA produced by both rat mammary tumors (21) and 13. De Waard, F. Breast Cancer Incidence and Nutritional Status with human mammary tumors (22) was 7Â«-hydroxydehydro- Particular Reference to Body Weight and Height. Cancer Res., 35. 3351- 3356,1975. epiandrosterone. In the latter case conversions to 7-oxy- 14. De Waard, F., Baanders-van Halewijn, E. A., and Huizinga, J. The genated DHEA derivatives ranged from 10 to 50%. ADIOL Bimodal Age Distribution of Patients with Mammary Cancer. Cancer, 17: 141-151, 1964. was formed in yields averaging 4% (22). Such high conver 15. De Waard, F., Cornelius, J. P., Aoki, K., and Yoshida, M. Breast Cancer sions to 7-oxygenated derivatives are of considerable inter Incidence According to Weight and Height in Two Cities of the Nether est. For example, it was found that the ability of ADIOL to lands and in Aichi Prefecture, Japan. Cancer, 40: 1269-1275, 1977. 16. De Waard, F., Laive, J. W. J., and Baanders-van Halewijn, E. A. On the compete for the estrogen receptor in uterine cytosol in vitro Bimodal Age Distribution of Mammary Carcinoma. Brit. J. Cancer, 14: was markedly reduced by the introduction of a 7a-hydroxyl 437-451,1960. group (22). A close parallel is seen when, once again, the 17. Garcia, M., and Rochefort, H. Androgens on the Estrogen Receptor IIâ€”¿ Correlation between Nuclear and Uterine Protein Synthesis. Steroids, introduction of a 7a-hydroxyl group into DHEA reduces its 29: 111-126, 1977. ability to inhibit estrogen sulfotransferase (Table 1; Chart 18. Carola, R. E., and McGuire, W. L. Estrogen Receptor and Proteolytic Activity in Human Breast Tumor Nuclei. Cancer Res., 37. 3329-3332, 4A). It is evident that a polar substituent in position 7 must 1977. diminish binding to a hydrophobic area in the steroid- 19. Carola, R. E., and McGuire, W. L. An Improved Assay for Nuclear binding sites of both receptor protein and estrogen sulfo Estrogen Receptor in Experimental and Human Breast Cancer. Cancer transferase. The 7-hydroxylase may then function to control Res., 37: 3333-3337, 1977. 20. Hsia, S. L. Potentials in Exploring the Biochemistry of Human Skin. intracellular levels of ADIOL and DHEA. Essays Biochem., 7: 1-38, 1971. As mentioned earlier, steroid sulfotransferase activities in 21. Li, K., Adams, J. B., and Chandra, D. P. In Vitro Metabolism of Dehydroepiandrosterone by Mammary Gland and Mammary Tumours in human breast tumors correlate with prognosis. A situation the Rat. J. Steroid Biochem., 7: 501-504, 1976. in which the ratio of DHEAS to estradiol sulfate is <1, 22. Li, K., Foo, T., and Adams, J. B. Products of Dehydroepiandrosterone Metabolism by Human Mammary Tumors and Their Influence on Estra reflecting a poor prognosis (12), may be due to the low diol Receptor Binding. Steroids, 37: 113-127, 1978. levels of DHEA sulfotransferase observed in many tumors 23. Pack, B. A., and Brooks, S. C. Cyclic Activity of Estrogen Sulfotransfer-

NOVEMBER 1978 4039

ase in the Gilt Uterus. Endocrinology, 95: 1680-1690, 1974. 27. Schmidt, W. N., Sadler, M. A., and Katzenellenbogen, B. S. Androgen 24. Rochefort, H., and Garcia, M. Androgen on the Estrogen Receptor Iâ€”¿ Uterine Interaction-Nuclear Translocation of the Estrogen Receptor Binding and in vivo Nuclear Translocation. Steroids, 28: 549-560, 1976. and Induction of the Synthesis of the Uterine-Induced Protein (IP) by 25. Rochefort, H., Lignon, F., and Capony, F. Formation of Estrogen High Concentrations of Androgens in Vitro but Not in Vivo. Endocrinol Nuclear Receptor in Uterus: Effect of Androgens, Estrone and Nafoxi- ogy, 98: 702-716, 1976. dine. Biochem. Biophys. Res. Commun., 47: 662-670, 1972. 28. Thijssen, J. H. H., Poortman, J., and Schwarz, F. Androgens in Postmen- 26. Ruh, T. S., Wassilak, S. G., and Ruh, M. F. Androgen-lnduced Nuclear opausal Breast Cancer: Excretion, Production and Interaction with Accumulation of the Estrogen Receptor. Steroids, 25: 257-273, 1975. Estrogens. J. Steroid Biochem., 6: 729-734, 1975.

4040 CANCER RESEARCH VOL. 38

John B. Adams, Lesley Archibald and Christine Clarke

Cancer Res 1978;38:4036-4040.

Updated version Access the most recent version of this article at: http://cancerres.aacrjournals.org/content/38/11_Part_2/4036

E-mail alerts Sign up to receive free email-alerts related to this article or journal.

Reprints and To order reprints of this article or to subscribe to the journal, contact the AACR Publications Subscriptions Department at [email protected].

Permissions To request permission to re-use all or part of this article, use this link http://cancerres.aacrjournals.org/content/38/11_Part_2/4036. Click on "Request Permissions" which will take you to the Copyright Clearance Center's (CCC) Rightslink site.