Soybean Phytoestrogen Intake and Cancer Risk12C. HERMAN, T. ADLERCREÜTZ,3* BARRY R. GOLDIN, SHERWOOD L. GORBACH,KRISTER A. V. HÖCKERSTEDT,** SHAW WATANABE, ESA K. HÄMÄLÄINEN,***M. HELENE MARKKANEN,* TARÜ H. MÄKELÄ,KRISTIINA T. WÄHÄLÄ,TAPIO A. HASE AND THEODORE FO7S/S****
*Department of Clinical Chemistry (Fliï-00290),**IVSurgical Department, Helsinki (Jniuersity CentralHospital (FIN-00130), and Department of Chemistry (FIN-00100), University of Helsinki, Helsinki,Finland; Department of Community Health, Nutrition/Infection Unit, Tufts University School ofMedicine, Boston, MA 02] 11; Epidemiology Division, National Cancer Center Research Institute,Tsukiji 5-chome, Chuo-ku, Tokyo, Japan; ***Department of Clinical Chemistry, University CentralHospital, FIN-70211 Kuopio, Finland; ****Department of Oncology and Immunology, Children'sUniversity Hospital, Ruprecht-Karls-Universitat, D-6900 Heidelberg, Germany
ABSTRACT Because many Western diseases are hormone-dependent cancers, we have postulated that theWestern diet, compared with a vegetarian or semi-vegetarian diet, may alter hormone production, metabolism or action at the cellular level. Recently, our interest has been focused on the cancer-protective roleof some hormone-like diphenolic phytoestrogens of dietary origin, the lignans and isoflavonoids. The precursors of the biologically active compounds originate insoybean products (mainly isoflavonoids but also lignans), as well as whole grain cereals, seeds, probablyberries and nuts (mainly lignans). The plant lignan andisoflavonoid glycosides are converted by intestinal bacteria to hormone-like compounds with weak estrogenicand antioxidative activity; they have now been shownto influence not only sex hormone metabolism and biological activity but also intracellular enzymes, proteinsynthesis, growth factor action, malignant cell proliferation, differentiation and angiogenesis, making themstrong candidates for a role as natural cancer protectivecompounds. Epidemiológica! investigations supportthis hypothesis, because the highest levels of thesecompounds are found in countries or regions with lowcancer incidence. This report is a review of results thatsuggest that the diphenolic isoflavonoids and lignansare natural cancer-protective compounds. J. Nutr. 125:757S-770S, 1995.
INDEXING KEY WORDS:
•soybean •phytoestrogens •cancer risk
Many epidemiological and migrant studies supportthe view that the Western diet is one of the main factors causing the high incidence of the so-called Western diseases (Rose et al. 1986, Trowell and Burkitt
1981), including the major hormone-dependent cancers, colon cancer and coronary heart disease. Becauseof the fact that these diseases are related to some extentto sex hormones or sex hormone metabolism (Adler-creutz et al. 1982, Adlercreutz 1984, 1990), we havepostulated that the Western diet, compared with thevegetarian or semivegetarian diet in developing andAsian countries, may alter hormone production, metabolism or action at the cellular level by some biochemical mechanisms. Recently, our interest has beenfocused on the biological role of two groups of hormone-like diphenolic phytoestrogens of dietary origin,the lignans and isoflavonoids. These compounds,which mainly occur in soybean and whole-grain products and various seeds, have now been shown to influence not only sex hormone metabolism and biological activity but also intracellular enzymes, proteinsynthesis, growth factor action, malignant cell prolif-
1Presented at the First International Symposium on the Role of
Soy in Preventing and Treating Chronic Disease, held in Mesa, AZ,February 20-23, 1994. The symposium was sponsored by ProteinTechnologies International, the soybean growers from Nebraska,Indiana and Iowa and the United Soybean Board. Guest editors forthis symposium were Mark Messina, 1543 Lincoln Street, PortTownsend, WA 98368, and John W. Erdman, Jr., Division of Nutritional Sciences, University of Illinois, Urbana, IL 61801-3852.
2 Supported mainly by the Medical Research Council of the
eration and angiogenesis, making them strong candidates for a role as cancer-protective compounds.
Phytoestrogens in Soy
It has been well known since 1931 that soybeanscontain high amounts (up to 100-300 mg/100 g) ofthe glycosides of the two isoflavones, daidzein andgenistein (Eldridge and Kwolek 1983; Walz 1931). Athird major compound, glycitein, was found later as aglycoside (Nairn et al. 1973). Small amounts of thesethree compounds occur in the free form. Fermentedsoy may contain a catecholic conversion product ofglycitein, 6,7,4-trihydroxyisoflavone (Györgyet al.1964, Klus et al. 1993). Furthermore, small amounts(~5 ^ig/100 g) of the isoflavone coumestrol have beenfound (Lookhart et al. 1978). Soy sauce does not contain any isoflavones, but it does contain the lignanprecursor, coniferyl alcohol (Yokotsuka 1986). Recently, we developed new methodologies for the assayof phytoestrogens in meal, flour and other plant products. Using this gas chromatographic-mass spectro-metric method, we found preliminary evidence suggesting the presence of secoisolariciresinol in soy meal.
Phytoestrogens Identified in Man
After consuming the plant isoflavonoids and lig-nans, many metabolic conversions occur in the gutresulting in the formation of hormone-like compoundswith estrogen activity and the ability to bind weaklyto estrogen receptors (Price and Fenwick 1985, Setchelland Adlercreutz 1988). In 1979, two cyclically occurring unknown compounds, now called enterolactoneand enterodiol, were detected in the urine of the female vervet monkey and in women, and subsequentlyidentified separately and independently by two groups(Setchell et al. 1980, Stitch et al. 1980). They wereshown to be diphenols with a lignan structure, butlacking the para-oxygen substitution, differing in thisway from plant lignans. Small amounts of four plantlignans, matairesinol, lariciresinol, isolariciresinol andsecoisolariciresinol, were also found and identified inhuman urine. Furthermore, we have tentatively identified 7a-hydroxymatairesinol and 7'-hydroxyentero-
lactone in human urine (Adlercreutz et al. 199Id,Setchell and Adlercreutz 1988).
The isoflavonoid phytoestrogens are heterocyclicphenols with a close similarity in structure to estrogens. Their diphenolic character makes them similarto lignans. They occur in numerous plants. Manystudies have shown that they have hormonal effectsin animals (Price and Fenwick 1985). The followingisoflavonoid phytoestrogens have been identified ordetected in human urine in this laboratory: formo-nonetin, methylequol, daidzein, dihydrodaidzein, O-desmethylangolensin, genistein, and 3',7-dihydroxy-
isoflavan. Equol was identified independently in twolaboratories (Adlercreutz et al. 199Id).
Metabolism of Phytoestrogens in Man
The literature has been reviewed on the origin, formation and metabolism of the phytoestrogens in animals (Price and Fenwick 1985) and in man (Adlercreutz 1988a, Adlercreutz et al. 199Id, Setchell andAdlercreutz 1988) and, therefore, only a few pointswill be dealt with in this connection.
Equol and O-desmethylangolensin are most likelyformed, in humans as in sheep, by intestinal bacterialaction on formononetin and daidzein present in foodstuffs like soy products. Some people are not able toproduce equol (Setchell and Adlercreutz 1988), this isalso the case in Japanese subjects accustomed to consuming large amounts of soy (Adlercreutz et al. 1991e).Among the isoflavonoids daidzein, O-desmethylangolensin, equol and genistein can be measured inplasma (Adlercreutz et al. 1993b), urine (Adlercreutzet al. 1991c) and feces (Adlercreutz et al. 1995), permitting studies on the absorption and metabolism ofthese compounds.
The mammalian lignans, enterolactone and enterodiol, are formed from plant precursors by the actionof intestinal bacteria. The lignan matairesinol is converted to enterolactone and secoisolariciresinol to enterodiol, and the latter is oxidized to enterolactone(Setchell and Adlercreutz 1988). After further development of our methodology (Adlercreutz et al. 1991c)we can now analyze the following lignans in urine,plasma and feces: matairesinol, secoisolariciresinol,enterodiol and enterolactone. Many of these compounds have been identified and measured in cow milk(Adlercreutz et al. 1986a) and cow urine (Fotsis, T.and Adlercreutz, H., unpublished results). Enterolactone concentration is high in human and bovine semen(Dehennin et al. 1982), some lignans and isoflavonoidshave been identified and measured in saliva, breastaspirate or cyst fluid and prostatic fluid by gas chro-matographt/mass spectroscopy (Finlay et al. 1991).
With regard to lignans, the macrobiotics living inBoston had the highest values, followed by the othervegetarian groups. The lowest lignan values amongthe subjects in Western societies were found in the
1Values are geometric means with SEM-rangein parentheses. For each subject the mean of two 72-h urinary collections with about 6 mointerval were used for calculating group means.
2The women have been described in Adlercreutz et al. (1986b).3The women have been described in Coldin et al. (1986).4 Data from Adlercreutz et al. (1991e).
breast cancer groups and the Boston omnivorouswomen. In an earlier study the differences were evenlarger (Adlercreutz et al. 1982, 1986b). However, evenlower lignan values were found in the Asian immigrantwomen living in Hawaii; low values were found in theJapanese men and women, although they had very highisoflavonoid values. The recent immigrants to Hawaiihad similar isoflavonoid values as those women livingin Boston, indicating that after immigration theyeliminate soy products from their diet; however, theyhave higher values than the Finnish women. Becausethe Hawaiian immigrants do not consume any whole-grain bread, the result is very low urinary lignan andisoflavonoid values. They consume a low-fat diet
(Goldin et al. 1986), which may explain why they arestill protected from breast cancer. This may also produce higher fecal excretion of phytoestrogens andlower values in urine, similar to the pattern found forendogenous estrogens. It must be remembered thatthese Asian women are shorter than Western womenand they weigh less. In relation to their body weight,the phytoestrogen levels may be higher than judgedfrom the urinary excretion values.
Main Biological Effects of DiphenolicPhytoestrogens
Estrogenic and antiestrogenic activity. We focus on the biological effects of only those compounds
that are derived from soy products and have beenmeasured in humans. Other compounds are discussedin a recent review (Whitten and Naftolin 1991).
The lignans, enterolactone and enterodiol, bindweakly to rat uterine cytosol (Clark, J. H. and Adler-creutz, H., unpublished observations) but have no detectable estrogenic activity in vivo in mice (Setchell etal. 1981). However, four sensitive assays in tissue culture, including breast cancer cell lines, showed thatthe lignans were stimulatory and the effect could beblocked by tamoxifen. No antiestrogenic propertiescould be observed (Jordan et al. 1985). In another studyenterolactone inhibited estrogen-stimulated RNAsynthesis in rat uterine tissue in vivo when administered 22 h before estradiol (Waters and Knowler 1982).These diverging results are difficult to explain, but ithas been suggested (Adlercreutz 1990, Whitten andNaftolin 1991) that the effect of weak exogenous estrogens may be either agonistic or antagonistic depending on the level of endogenous estrogens. Thishas been experimentally confirmed with regard tocoumestrol (Whitten and Naftolin 1991).
Numerous studies have shown that the isoflavonoidphytoestrogens bind to estrogen receptors and haveweak estrogenic activity (Jordan et al. 1985, Martinet al. 1978, Shutt and Cox 1972). All of the main is-oflavonoids (genistein, daidzein, equol, O-desmethyl-angolensin) and their precursors (biochanin A, for-mononetin) detected in human and animal urine bindto the estrogen receptor. Many studies suggest thatisoflavonoids or soy products containing high amountsof these compounds have significant estrogenic effectsin animals and man (Gavaler et al. 1991, Mäkeläet al.1991, Setchell et al. 1984, Van Thiel et al. 1991, Whitten and Naftolin 1991). Soy, containing isoflavonoids,has been found to have both estrogenic and antiestrogenic effects in mice (Mäkeläet al. 1991). The bestknown estrogenic effect of phytoestrogens is the"clover disease" in .Australian sheep (Bennets et al.
1946, Price and Fenwick 1985). On the other hand,definite antiestrogenic effects have also been observedin vivo. High levels of synthetic estrogens seem to becounteracted by isoflavonoids administered or by theirpresence in the diet (Folman and Pope 1966, 1969,Mäkeläet al. 1991).
Binding to the type n estrogen nuclear bindingsite. Several isoflavonoids and lignans compete withestradiol for the rat uterine nuclear type II estrogen-binding site (Adlercreutz et al. 1992a) (sometimescalled the bioflavonoid receptor). Because of theirmoderate affinity and relatively high capacity, thesesites have not been regarded as true receptors. Thesesites are, however, in some way involved in regulatingestrogen-stimulated uterine growth (Markaverich andClark 1979, Markaverich et al. 1981). It was foundthat some flavonoids like luteolin, quercetin and pel-argonin inhibit estradiol binding to these binding sitesand the growth of MCF-7 cells in culture, and in vivo
the estradiol stimulation of the immature rat uterus(Markaverich et al. 1988b). The structures of theseflavonoids are very similar to those of the isoflavones.The highest affinity of type II binding of the diphenoliccompounds in human urine is shown by the isoflavonedaidzein and the isoflavan equol, but also some lignanslike matairesinol, isolariciresinol and enterolactoneshow competition. The binding, however, is weakerthan that demonstrated for luteolin, but the concentrations in blood may be very high (Adlercreutz et al.1993c). For luteolin no quantitative data in biologicalfluids is available. In preliminary experiments wefound that this compound occurs in urine in very lowamounts.
Subsequently, an endogenous inhibitor of the nuclear type II binding site, methyl p-hydroxyphenyllac-
tate (Markaverich et al. 1988a), was identified. It waspostulated that proliferation of malignant cells is directly related not only to the permanent stimulationof nuclear type II binding sites by estrogens or othercompounds, but also to very low to nonmeasurablelevels of the competitive inhibitor methyl p-hydroxy-phenyllactate (Markaverich et al. 1988a). This compound seems to be derived from endogenous bioflavonoid metabolism. Many of these phenolic compounds may have a synergistic action since it isunlikely, because of close structural similarities, thatonly one of them inhibits cell growth. Recently, a ty-rosinase-like enzyme was found to be associated withthe type II binding catalyzing the conversion of estradiol to a catechol estrogens, which then is probablyconverted in whole or in part to the respective ortho-quinones (Garai and Clark 1992). It was also foundthat luteolin and 4,7-dihydroxyflavone bind to the typeII nuclear estrogen-binding sites with high affinity (Kd5-10 nM) (Markaverich and Gregory 1993) and thatthe binding of luteolin is probably covalent.
Aromatase inhibition. Plant lignans have beenshown to possess anticarcinogenic, antiviral, bacteri-cidic and fungistatic activities (Setchell and Adlercreutz 1988). Only those related to cancer will be discussed here.
Enterolactone, the most abundant mammalian lig-nan, is a moderate inhibitor of placental aromataseand competes with the natural substrate androstene-dione for binding to the enzyme (Adlercreutz et al.1993a). Experiments with a choriocarcinoma cell line(JEG-3) showed that enterolactone is very readilytransferred from cell culture media into the cells inhibiting the aromatase enzyme (Adlercreutz et al.1993a). Enterolactone together with other diphenolslike the flavonoids, which occur in high amounts inthe diet and are inhibitors of the aromatase enzyme(Kellis and Vickery 1984), may reach sufficient concentrations in fat cells to reduce conversion of andro-stenedione to estrone. In fact, isoflavonoids and lignans show weak aromatase inhibiting activity in hu-
man preadipocytes in vitro (Campbell and Kurzer1993, Wangetal. 1994).
Effect on tyrosine kinase, growth factors andother enzymes. Genistein is an isoflavone that wasidentified by us in human urine (Adlercreutz et al.1991 c). It is found in large amounts both in urine andplasma of Japanese subjects consuming a traditionalJapanese diet (Adlercreutz et al. 1991e, 1993e) and inmacrobiotics (Tables 1 and 2). Genistein is a specificinhibitor of tyrosine protein kinases (except the p40protein-tyrosine kinase), topoisomerase II (see Adlercreutz 1990) and protein histidine kinase (Huang etal. 1992). Protein-tyrosine kinase activity is associated
with cellular receptors for epidermal growth factor,insulin, insulin-like growth factor I, platelet-derived
growth factor and mononuclear phagocyte growthfactor. The tyrosine kinases seem to play an importantrole in cell proliferation and transformation. Theseenzymes have been associated with oncogene productsof the retroviral src gene family and are correlated withthe ability of retrovirus to transform cells (Akiyamaet al. 1987, Markovits et al. 1989, Ogawara et al. 1989,Teraoka et al. 1989). Tyrosine kinase activity is alsoassociated with breast cancer oncogene expression (LeCam 1991,Lehtolaetal. 1992). Furthermore, genisteinstimulates differentiation of leukemic and other malignant cells (Table 3). Lignans and isoflavonoids aswell as foods containing large amounts of these compounds or their precursors, seem to inhibit cancer cellgrowth (Table 3).
Stimulation of sex hormone binding globulinsynthesis. Lignans and isoflavonoids seem to stimulate sex hormone binding globulin synthesis in theliver and in this way they may reduce the biologicaleffects of sex hormones (Adlercreutz et al. 1987,1988b). An increase in sex hormone binding globulindecreases the relative amount of free testosterone andfree estradici and reduces both the albumin-bound andthe free fraction of the sex hormones. This reducesthe metabolic clearance rate of the steroids and therebylowers their biological activity. In Finnish women totalfiber intake, total fiber intake/kg body weight andgrain fiber intake/kg body weight correlate positivelywith urinary excretion of total lignans and isoflavonoids (Adlercreutz et al. 1987, 1988b). The urinaryexcretion of the two groups of compounds and alsoenterolactone alone in both pre- and postmenopausalFinnish women correlate positively with plasma sexhormone binding globulin and negatively with plasmapercentage free estradiol and percentage free testosterone (Adlercreutz et al. 1987, 1988b). In vitro studiesusing HepG2 liver cancer cells showed that enterolactone (Adlercreutz et al. 1992a), genistein (Mousavi andAdlercreutz 1993) and daidzein (Carson, M., Louko-vaara, M., Palotic, A., and Adlercreutz, H., unpublished results) stimulate sex hormone binding globulinsynthesis. This may explain the higher sex hormone
binding globulin values found in vegetarians (Adlercreutz et al. 1988b; Armstrong and Doll 1975).
Lignans, Isoflavonoids and Cancer
Breast cancer. The lignans and isoflavonoid phy-toestrogens are normal constituents of human urine,plasma and feces and occur in large amounts in plasma,urine and feces, particularly in vegetarians, in subjectsconsuming large amounts of whole-grain products,vegetables, berries, fruits, linseed (flaxseed) and sesameseeds and in Japanese consuming a traditional Japanesediet (Adlercreutz et al. 1982, 1986b, 1987, 1988b,c,199le, 1992b, 1993c, Axelson et al. 1982, Shultz etal. 1991). Plasma levels are higher in vegetarian womencompared with omnivorous women (Adlercreutz et al.1993b) and urinary excretion is higher in North Kareliacompared with southwest Finland (Adlercreutz et al.1986b), correlating negatively with rates of breast andprostate cancer risk.
Thus, in populations at low risk for hormone-dependent cancer the urinary and plasma levels of thediphenols are usually high (Tables 1 and 2). The recentAsian immigrants to Hawaii seem to be an exception(Tables 1 and 2) because they have a very low excretionbut also a low risk of breast cancer. This may be because the fecal excretion of the metabolites might behigh as was found for endogenous estrogens (Goldinet al. 1986) diverting some part of the metabolites fromurine to feces. Subjects with breast cancer (Adlercreutzet al. 1982) or those at high risk of breast cancer (omnivorous women living in Boston, MA) (Tables 1 and2) excrete low amounts of lignans and isoflavonoids.Finnish subjects with medium risk have relatively highlignan excretion but low isoflavonoid excretion, andJapanese subjects at low risk of breast cancer have highisoflavonoid excretion, but relatively low lignan excretion. However, in plasma the concentrations ofbiologically active free and sulfate-conjugated lignansare higher in Japanese compared with Finnish men despite lower urinary excretion in the Japanese men (unpublished results). This is most likely due to less conjugation of the lignans with glucuronic acid, becausethe plasma levels of glucuronic acid conjugates aremuch lower than in the Finnish men. Plasma assayshave not yet been carried out in Japanese women sowe do not know whether the same phenomenon occursin them. Because in Boston, MA, and in Finland thelignan excretion is mainly associated with the intakeof grain fiber or whole-grain products (Adlercreutz etal. 1986b, 1987, and unpublished data), it seems thatin these countries the risk of breast cancer may dependon the intake of such products. The significantly positive association between lignan excretion and intakeof whole-grain products and total fiber is altered bythe intake of various seeds with relatively low fiberbut high content of lignan precursors. This occurs particularly in vegetarians. Another factor affecting the
association between intake of whole-grain productsand lignan excretion is the fact that many subjectsconsume purified grain fiber directly or in the form ofwhole-meal bread which is a mixture of bran and whitemeal containing only small amounts of meal from thealeurone layer of the grain where the lignan precursorsoccur. Intake of fruits and berries in Finnish womenalso has a positive correlation with lignan excretion(Adlercreutz et al. 1987). Berries contain the seeds ofthe plant and these may be rich in lignan precursors(Adlercreutz et al. 1987). In Japanese subjects lignanexcretion shows the strongest correlation with the intake of whole soybeans (Adlercreutz et al. 1991c). Thissuggests that the lignan precursors are lost during processing of the soybean proteins.
In this connection we will not discuss in detail therole of fiber intake in the prevention of breast cancerand the possible mechanisms (Adlercreutz 1990,1991b). Intake of fiber-rich food definitely affects theplasma levels of estrogens, increases phytoestrogenintake, but also reduces energy intake (Arts et al.1992), which may be an important risk-reducing factor(Weindruch et al. 1991). All these factors may affectmenarcheal age (Adlercreutz 199la, Arts et al. 1992,de Ridder et al. 1991), which is a strong determinantof breast cancer risk (Henderson and Bernstein 1991).
Some recent prospective epidemiological studies donot show any protective effect of fiber (Willett et al.1992) with regard to breast cancer risk. However, inthis study only the amount of total dietary fiber wasdetermined. Total fiber intake tells us relatively littleabout the lignan content of the diet or the effect onthe enterohepatic circulation of estrogens, which affects plasma estrogen levels (Adlercreutz 1990). Results of studies in postmenopausal women in Boston(Adlercreutz et al. 1989a) and in premenopausalwomen in Helsinki (Adlercreutz et al. 1989b) showedthat the main and, in fact, the only really significantdifference between the diets of the breast cancer patients and the omnivorous and vegetarian controlwomen was a lower intake of grain products and fiberin the subjects with breast cancer. If the diets of theBoston and Finnish women studied by us are compared, the main difference is in the grain and grainfiber intake, being much higher in the Helsinki womenwith a lower risk for breast cancer than in the Bostonwomen. This disparity results in differences in lignanexcretion. In both groups the intake of soy productsseemed to have been low.
In 1986 we included the isoflavonoid phytoestro-gens in the theory concerning protective effect of lig-nans in breast cancer (Adlercreutz et al. 1986b, Adlercreutz 1988a), although we discussed the possibilitypreviously (Adlercreutz et al. 1982). The structural differences among these compounds are relatively smallbecause both groups are diphenols with molecularweights close to those of endogenous estrogens. Ourview is supported by the finding of high excretion of
isoflavonoids in urine and high plasma levels of Japanese subjects consuming a traditional diet (Adlercreutz et al. 1987, 1988c, 1991e, 1993e). Japanesewomen consuming this diet have very low incidenceof breast cancer.
Our hypothesis with regard to the protective roleof these compounds in breast cancer has received support from numerous studies (Table 3). Some show thatheated or unheated powdered soybean chips or heatedsoy decrease mammary tumor formation or inhibitprogression of such tumors in rat breast cancer models(Barnes et al. 1990, Hawrylewicz et al. 1991). Furthermore, linseed containing high amounts of lignansinhibits mammary carcinogenesis in rats (Serraino andThompson 1992a). Genistein, identified by us in human urine and plasma, is anticarcinogenic perhaps dueto its inhibitory effect on protein tyrosine kinase(Akiyama et al. 1987, Reddyetal. 1992), its inhibitionof angiogenesis (Fotsis et al. 1993) or its antioxidativeproperties (Wei et al. 1993) (see also above). Genisteinalso induces differentiation of malignant cells (Table3). The concentrations needed are relatively high; onthe other hand the total concentration of genistein inplasma in Japanese men may exceed 2 ¿tmol/1(Adlercreutz et al. 1993c). Genistein and daidzein have beenshown to be antiproliferative with regard to breastcancer cells (Table 3). Long ago it was shown that isoflavonoids are competitive with regard to estrogenbinding to its receptor in breast cancer cells (Martinet al. 1978). Because genistein also inhibits growth ofreceptor-negative breast cancer cells, its mechanismof action may not be directly related to the estrogenreceptor. Furthermore, epidemiological evidence obtained in Singapore indicates that soy intake is associated with lower breast cancer risk in women (Lee etal. 1991). There is an inverse relation between the urinary excretion of enterolactone and plasma luteinizinghormone in Finnish women (Adlercreutz 1990). Soyintake also prolongs the menstrual cycle length inwomen (Cassidy et al. 1993) and flaxseed intake inrats prolongs the cycle and luteal phase length inwomen (Phipps et al. 1993, Orcheson et al. 1993).These observations suggest that these compounds mayaffect the hormonal system via the hypothalamus-hy-pophyseal endocrine system as previously suggested(Adlercreutz 1990).
Recently, we found that enterolactone alone (0.5-10 mmol/1) stimulates the growth of MCF-7 cells, butin the presence of estradiol, added both in concentrations stimulating growth or in lower amounts, growthdid not differ from the control or tended to be slightlyless (Mousavi and Adlercreutz 1992). The mechanismof this phenomenon is unknown, but it may be relatedto the type II nuclear estrogen-binding site.
Prostatic cancer. Despite the same incidence oflatent and small or noninfiltrative prostatic carcinomasas in the Western countries, the mortality in Japanand some other Asian countries is low (Breslow et al.
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1977, Ota and Misu 1958, Yatani et al. 1982). We suggested that this may be due to the effect of phytoes-trogens, particularly isoflavonoids, which inhibit thegrowth of latent cancers (Adlercreutz 1990). In epi-demiological studies fat and meat show a positive association and cereals a negative association with pros-tatic cancer mortality (Rose et al. 1986). Decreasedprostatic cancer risk has been found in Adventists men(Mills et al. 1989) having high consumption of beans,lentils, peas and some dried fruits (all dietary sourcesof flavonoids) and in men of Japanese ancestry in Hawaii (Severson et al. 1989) consuming much rice andto fu, a soybean product containing isoflavonoids. Themean consumption of soy products (except soy sauce)was 39.2 ±36.4 g/d in Japanese men living in a smallvillage outside Kyoto where the people consume a traditional Japanese diet based on their own products(Adlercreutz et al. 1991e). The intake of various soyproducts in men and women in the same study showeda strong positive association with urinary excretion ofisoflavonoids. Lignan excretion in Japanese subjects isrelatively low (Table 4) and shows only a positive association with the intake of pulses and beans andboiled unprocessed soybeans (Adlercreutz et al. 1993c).Preparation of tofu products seems, therefore, toeliminate the lignan precursors from the beans.
Soy has been found to have a protective effectagainst prostatitis in rats (Sharma et al. 1992). To ourknowledge, prostatitis has not been associated withprostatic cancer. Recently, however, it was found thatsoy is protective against prostatic dysplasia in a mousemodel (Mäkeläet al. 1991). Genistein and biochaninA, the precursor of genistein, inhibit, in cell culturesthe growth of androgen-dependent and -independentprostatic cancer cells (Peterson and Barnes 1993). Thiseffect was not caused by inhibition of the tyrosine ki-nase associated with the epidermal growth factor receptor. The well-known therapeutic effect of estrogensin prostatic cancer suggests that phytoestrogens mayinhibit prostatic cancer cell growth during the promotional phase of the disease or they may influencedifferentiation as shown for genistein with leukemiccells and other cancer cells (Table 4). Recently, an ep-idemiological study showed that environmental factors later in life can substantially impact the likelihoodof developing clinically detectable prostatic cancer(Shimizu et al. 1991).
Despite high fat intake the prostatic cancer incidence in Finland, particularly in northeast Finland(Teppo et al. 1980), is much lower than in UnitedStates, but higher than in Japan. The higher productionof lignans in the gut, due to relatively high intake ofwhole-grain products, particularly rye bread, in thelow-incidence rural areas in Finland, may perhaps explain this phenomenon. The lignans are weaker estrogens than the isoflavonoids but a protective effect maywell be independent of the estrogenic effect. A recentstudy suggests that there is an increase in type II nu-
clear estrogen-binding sites in the dysplastic dorsolat-eral part of the prostate glands of Noble rats (Ho andYu 1993). Consequently bioflavonoids and lignans maybe involved in regulating the growth of prostatic cancer via these binding sites.
Thus, epidemiological studies as well as cell cultureand animal experiments provide evidence which suggests that isoflavonoids and perhaps other phytoestro-gens like lignans are protective and lower the risk ofprostatic cancer during the promotional phase of thedisease.
Colorectal and other cancers. In 1984 one of ussuggested that the lignans may be protective with regard to both breast and colon cancer (Adlercreutz1984). Recently, we observed a high lignan excretionin subjects with a low risk of colon cancer (Korpela etal. 1992). Lignan excretion is also higher in Finnishsubjects (Adlercreutz et al. 1986b) living in areas withlower colon cancer risk (Teppo et al. 1980). Epidemiological evidence obtained in Japan (Watanabe andKoessel 1993) points to lower colon cancer incidencein areas with high tofu consumption. This is now beingfurther investigated. Diets containing 5% linseed (w/w), rich in lignans, seem to protect against colon cancerin rats (Serraino and Thompson 1992b). Colon cancercell lines and primary colorectal tumors have type IInuclear estrogen-binding sites and quercetin, as shownalso for MCF-7 cells, inhibits the growth of colon cancer cells in culture (Ranelletti et al. 1992). Quercetinis a flavonoid occurring in high amounts in certainplants, but in our experience very little is excreted inhuman urine. Due to their phenolic structure, lignansand flavonoids have antioxidative properties (Adlercreutz 1984, Lu and Liu 1992, Nairn et al. 1976, Weiet al. 1993) and may prevent conversion of procarcin-ogens to carcinogens or eliminate free radicals inthe gut.
Synthetic genistein and extracts from human urinecontaining genistein have been shown to inhibit thegrowth of cells from solid pediatrie tumors like neu-roblastomas (with both normal and enhanced MYCNexpression), rhabdomyosarcomas and Ewings sarcomas(Schweigerer et al. 1992). Such extracts and synthesized genistein inhibited bFGF-stimulated endothelialcell (bovine brain-derived capillary endothelial cells)proliferation and in vitro angiogenesis (Fotsis et al.1993). Genistein reduces the production of plasmin-ogen activator and plasminogen activator inhibitor-1(Fotsis et al. 1993) in cloned bovine microvascular endothelial cells from the adrenal cortex. Genistein alsoinhibits the growth of gastric cancer cells (Matsukawaet al. 1993) and stimulates differentiation in manymalignant cells including melanoma cells (Kiguchi etal. 1990). Genistein seems to modulate decreased drugaccumulation in non-P-glycoprotein mediated mul-tidrug resistance (Versantvoort et al. 1993).
Soy products that contain isoflavonoids and lignansmay play a role in the prevention of several types of
cancer. The concentrations in plasma of these compounds may easily reach biologically active levelswithout toxic effects. By inhibiting the effect of growthfactors and angiogenesis, genistein may be a generalinhibitor of cancer growth. By modulating drug transport, genistein may prove to be a good addition toestablished cancer therapy. The described biologicaleffects may also be used as a preventive strategy forother Western diseases not discussed in this connection, such as cardiovascular diseases and osteoporosis,due to the estrogenic and antioxidative effects. Theevidence obtained is not yet sufficient for any specificdietary recommendations and further work is neededto establish the role of these natural compounds inhuman health and disease.
LITERATURE CITED
Adlercreutz, H., Fotsis, T., Heikkinen, R., Dwyer, J. T., Woods, M.,Goldin, B. R. & Gorbach, S. L. (1982) Excretion of the lignansenterolactone and enterodiol and of equol in omnivorous andvegetarian women and in women with breast cancer. Lancet 2:1295-1299.
Adlercreutz, H. (1984) Does fiber-rich food containing animallignan precursors protect against both colon and breast cancer?An extension of the fiber hypothesis. Gastroenterology 86: 761-
Brunow, G. & Hase, T. (1986a) Assay of lignans and phy-toestrogens in urine of women and in cow milk by GC/MS (SIM).In: Advances in Mass Spectrometry-85. Proceedings of the 10thInternational Mass Spectrometry Conference (Todd, J. F. J., ed.),pp. 661 -662. John Wiley, Chichester, Sussex, England.
Adlercreutz, H., Fotsis, T., Bannwart, C., Wähälä,K., Mäkelä,T.,Brunow, G. & Hase, T. (1986b) Determination of urinary lignans and phytoestrogen metabolites, potential antiestrogens andanticarcinogens, in urine of women on various habitual diets. J.Steroid Biochem. 25: 791-797.
Adlercreutz, H., Höckerstedt, K., Bannwart, C., Bloigu, S., Hämä-läinen,E., Fotsis, T. & Ollus, A. (1987) Effect of dietary components, including lignans and phytoestrogens, on enterohepaticcirculation and liver metabolism of estrogens, and on sex hormone binding globulin (SHBG). J. Steroid Biochem. 27: 1135-1144.
Adlercreutz, H. (1988a) Lignans and phytoestrogens. Possiblepreventive role in cancer. In: Progress in Diet and Nutrition(Rozen, P., Horwitz, C., eds.), vol. 14, pp. 165-176. S. Karger,Basel, Switzerland.
Adlercreutz, H., Höckerstedt, K., Bannwart, C., Hämäläinen,E.,Fotsis, T. &. Bloigu, S. (1988b) Association between dietaryfiber, urinary excretion of lignans and isoflavonic phytoestrogens,and plasma non-protein bound sex hormones in relation to breastcancer. In: Progress in Cancer Research and Therapy: Hormonesand Cancer 3. (Bresciani, F., King, R. J. B., Lippman, M. E., Ray-naud, J. P., eds.), vol. 35, pp. 409-412. Raven Press, New York.
Adlercreutz, H., Honjo, H., Higashi, A., Fotsis, T., Hämäläinen,E.,Hasegawa, T. & Okada, H. (1988c) Lignan and phytoestrogenexcretion in Japanese consuming traditional diet. Scand. J. Clin.Lab. Invest. 48(Suppl 190): 190(abstr.).
Adlercreutz, H., Hämäläinen,E., Gorbach, S. L., Goldin, B. R.,Woods, M. N. & Dwyer, J. T. (1989a) Diet and plasma an-drogens in postmenopausal vegetarian and omnivorous womenand postmenopausal women with breast cancer. Am. J. Clin.Nutr. 49: 433-442.
Adlercreutz, H., Fotsis, T., Höckerstedt, K., Hämäläinen,E., Bannwart, C., Bloigu, S., Valtonen, A. & Ollus, A. (1989b) Diet
and urinary estrogen profile in premenopausal omnivorous andvegetarian women and in premenopausal women with breastcancer. J. Steroid Biochem. 34: 527-530.
Adlercreutz, H. (1990) Western diet and Western diseases: somehormonal and biochemical mechanisms and associations. Scand.J. Clin. Lab. Invest. 50(Suppl 201): 3-23.
Adlercreutz, H. (1991 a) Effect of diet and exercise on hormones:implications for monitoring training in women. Clin. J. SportsMed. 1: 149-153.
& Hase, T. (1991 c) Isotope dilution gas chromatographic-massspectrometric method for the determination of lignans and iso-
flavonoids in human urine, including identification of genistein.Clin. Chim. Acta 199: 263-278.
Adlercreutz, H., Mousavi, Y., Loukovaara, M. &. Hämäläinen,E.(l99Id) Lignans, isoflavones, sex hormone metabolism andbreast cancer. In: The New Biology of Steroid Hormones (Hoch-berg, R. B., Naftolin, F., eds.), vol. 74, pp. 145-154. Raven Press,
New York.Adlercreutz, H., Honjo, H., Higashi, A., Fotsis, T., Hämäläinen,E.,
Hasegawa, T. & Okada, H. (1991e) Urinary excretion of lignans and isoflavonoid phytoestrogens in Japanese men andwomen consuming traditional Japanese diet. Am. J. Clin. Nutr.54: 1093-1100.
Adlercreutz, H., Mousavi, Y., Clark, J., Höckerstedt, K., Wähälä,K., Mäkelä,T. & Hase, T. (1992a) Dietary phytoestrogens andcancer: in vitro and in vivo studies. J. Steroid Biochem. Mol.Biol. 41:331-337.
Adlercreutz, H., Hämäläinen,E., Gorbach, S. L. &. Goldin, B. R.(1992b) Dietary phyto-oestrogens and the menopause in Japan.
Hase, T., Arosemena, P. J., Kellis, J. T., Jr. & Vickery, L. E.(1993a) Inhibition of human aromatase by mammalian lignansand isoflavonoid phytoestrogens. J. Steroid Biochem. Mol. Biol.44: 147-153.
Adlercreutz, H., Fotsis, T, Lampe, J., Wähälä,K., Mäkelä,T., Brunow, G. ¿XHase, T. (1993b) Quantitative determination oflignans and isoflavonoids in plasma of omnivorous and vegetarianwomen by isotope dilution gas-chromatography mass-spectrom-etry. Scand. J. Clin. Lab. Invest. 53(Suppl. 215): 5-18.
Adlercreutz, H., Markkanen, H. & Watanabe, S. (1993c) Plasmaconcentrations of phyto-oestrogens in Japanese men. Lancet 342:1209-1210.
Adlercreutz, C. H. T., Fotsis, T., Kurzer, M. S., Wähälä,K. T., Mäkelä,T. H. & Hase, T. A. (1995) Isotope dilution gas chromatographic-mass spectrometric method for the determinationof unconjugated lignans and isoflavonoids in human feces, withpreliminary results in omnivorous and vegetarian women. Anal.Biochem., in press.
Armstrong, B. &.Doll, R. (1975) Environmental factors and cancerincidence and mortality in different countries, with special reference to dietary practices. Int. J. Cancer 15: 617-631.
Arts, C. J. M., Covers, C. A. R. L., Vandenberg, H. & Thijssen,J. H. H. (1992) Effects of wheat bran and energy restrictionon onset of puberty, cell proliferation and development of mammary tissue in female rats. Acta Endocrinol. 126: 451-459.
Axelson, M., Sjövall,J., Gustafsson, B. E. & Setchell, K. D. R. (1982)Origin of lignans in mammals and identification of a precursorfrom plants. Nature 298: 659-660.
Barnes, S., Grubbs, C., Setchell, K. D. R. &.Carlson, J. (1990) Soybeans inhibit mammary tumors in models of breast cancer. In:Mutagens and Carcinogens in the Diet. Progress in Clinical andBiological Research. (Pariza, M. W., Aeschbacher, H. U., Eton,J. S. and Sato, S., eds.), vol. 347, pp. 239-253. Wiley-Liss, Inc.,New York.
Bennets, H. W., Underwood, E. J. & Shier, F. L. (1946) A specificbreeding problem of sheep on subterranean clover pastures inWestern Australia. Aust. Vet. J. 22: 2-12.
Breslow, N. E., Chan, C. W., Dhom, G., Drury, R. A. B., Franks,L. M., Gellei, B., Lee, Y. S., Lundberg, S., Sparke, B., Sternby,N. H. & Tulinius, M. (1977) Latent carcinoma of prostate atautopsy in seven areas. Int. J. Cancer 20: 680-688.
Campbell, D. R. & Kurzer, M. S. (1993) Flavonoid inhibition ofaromatase enzyme activity in human preadipocytes. J. SteroidBiochem. Mol. Biol. 46: 381-388.
Cassidy, A., Bingham, S., Carlson, J. & Setchell, K. D. R. (1993)Biological effects of plant estrogens in premenopausal women.FASEB J. 7(3 Ft II): 5000(abstr.).
Chae, Y. H., Coffing, S. L., Cook, V. M., Ho, D. K., Cassady, J. M.fin.Baird, W. M. (1991 ) Effects of biochanin A on metabolism,DNA binding and mutagenicity of benzo<a>pyrene in mammalian cell cultures. Carcinogenesis 12: 2001-2006.
Constantinou, A., Kiguchi, K. &. Huberman, E. (1990) Inductionof differentiation and DNA strand breakage in human HL-60and K-562 leukemia cells by genistein. Cancer Res. 50: 2618-
2624.de Ridder, C. M., Thijssen, J. H. H., Van't Veer, P., Van Duuren,
R., Bruning, P. F., Zonderland, M. L. & Erich, W. B. M. (1991)Dietary habits, sexual maturation, and plasma hormones in pu-bertal girls—a longitudinal study. Am. J. Clin. Nutr. 54: 805-
813.Dehennin, L., Reiffsteck, A., Joudet, M. & Thibier, M. (1982)
Eldridge, A. & Kwolek, W. F. (1983) Soybean isoflavones: effectof environment and variety on composition. J. Agrie. Food Chem.31:394-396.
Finlay, E. M. H., Wilson, D. W., Adlercreutz, H. &. Griffiths, K.(1991) The identification and measurement of phyto-oestrogensin human saliva, plasma, breast aspirate or cyst fluid, and prostaticfluid using gas chromatography-mass spectrometry. J. Endocrinol.
129(Suppl): 49(abstr.|.Folman, Y. &. Pope, G. S. (1966) The interaction in the immature
mouse of potent oestrogens with coumestrol, genistein and otherutero-vaginotrophic compounds of low potency. J. Endocrinol.34:215-225.
Folman, Y. & Pope, G. S. (1969) Effect of norethisterone acetate,dimethylstilboestrol, genistein and coumestrol on uptake of[3H]oestradiol by uterus, vagina and skeletal muscle of immaturemice. J. Endocrinol. 44: 213-218.
Fotsis, T., Pepper, M., Adlercreutz, H., Fleischmann, G., Hase, T.,Montesano, R. Su.Schweigerer L. (1993) Genistein, a dietary-derived inhibitor of in vitro angiogenesis. Proc. Nati. Acad. Sci.USA 90: 2690-2694.
Garai, J. & Clark, I. H. (1992) Tyrosinase-like activity and estra-diol binding in rat uterine nuclear extracts. Steroids 57: 248-
256.Gavaler, J. S., Galvao-Teles, A., Monteiro, E., Van Thiel, D. H. &.
Rosenblum, E. (1991) Clinical responses to the administrationof bourbon phytoestrogens to normal postmenopausal women.Hepatology 14: 87A (abs.).
Goldin, B. R., Adlercreutz, H., Gorbach, S. L., Woods, M. N., Dwyer,J. T., Cordon, T., Bohn, E. & Gershoff, S. N. (1986) The relationship between estrogen levels and diets of Caucasian American and Oriental immigrant women. Am. J. Clin. Nutr. 44: 945-
953.György,P., Murata, K. & Ikehata, H. (1964) Antioxidants isolated
from fermented soybeans (Tempeh). Nature 203: 870-872.Hawrylewicz, E. J., Huang, H. H. & Blair, W. H. (1991) Dietary
soybean isolate and methionine supplementation affect mammary tumor progression in rats. J. Nutr. 121: 1693-1698.
Henderson, B. E. & Bernstein, L. (1991) The international variation in breast cancer rates: an epidemiological assessment. BreastCancer Res. Treat. 18:S11-S17.
Hirano, T., Oka, K., Kawashima, E. &.Akiba, M. |1989a) Effectsof synthetic and naturally occurring flavonoids on mitogen-in-duced proliferation of human peripheral-blood lymphocytes. LifeSci. 45: 1407-1441.
Hirano, T., Oka, K. &.Akiba, M. (1989b) Antiproliferativeeffectsof synthetic and naturally occurring flavonoids on tumor cellsof the human breast carcinoma cell line, ZR-75-1. Res. Commun.Chem. Pathol. Pharmacol. 64: 69-78.
Hirano, T., Wakasugi, A., Oohara, M., Oka, K. & Sashida, Y. (1991 )Suppression of mitogen-induced proliferation of human peripheral blood lymphocytes by plant lignans. Planta Med. 57: 331-334.
Ho, S. M. &YU, M. (1993) Selective increase in Type-II Estrogen-Binding sites in the dysplastic dorsolateral prostates of noblerats. Cancer Res. 53: 528-532.
Honma, Y., Okabe-Kado, J., Kasukabe, T., Hozumi, M. &. Umes-zawa, K. (1990) Inhibition of abl oncogene tyrosine kinaseinduces erythroid differentiation of human myelogenous leukemia K.562 cells. Jpn. J. Cancer Res. 81: 1132-1136.
Huang, J. M., Nasr, M., Kim, Y. H. &. Matthews, H. R. (1992)Genistein inhibits protein histidine kinase. J. Biol. Chem. 267:15511-15515.
Jing, Y. K., Nakaya, K. a Han, R. (1993) Differentiation of pro-myelocytic leukemia cells HL-60 induced by daidzein in vitroand in vivo. Anticancer Res. 13: 1049-1054.
Kellis, J. T., Jr., & Vickery, L. E. (1984) Inhibition of human estrogen synthetase (aromatase) by flavones. Science 225: 1032-
1034.Kiguchi, K., Constantinou, A. I. &.Huberman, E. (1990) Genistein-
induced cell differentiation and protein-linked DNA strandbreakage in human melanoma cells. Cancer Commun. 2: 271-
278.Klus, K., Borgerpapendore, G. & Barz, W. (1993) Formation of
6,7,4'-trihydroxyisoflavone (factor-2) from soybean seed isoflavones by bacteria isolated from tempe. Phytochemistry 34: 979-
981.Kondo, K.,Tsuneizumi, K., Watanabe, T. & Oishi, M. (1991) In
duction of in vitro differentiation of mouse embryonal carcinoma(F9) cells by inhibitors of topoisomerases. Cancer Res. 51: 5398-
5404.Korpela, J. T., Korpela, R. & Adlercreutz, H. (1992) Fecal bile
acid metabolic pattern after administration of different types ofbread. Gastroenterology 103: 1246-1253.
Kuriu, A., Ikeda, H., Kanakura, Y., Griffin, J. D., Druker, B., Yagura,H., Kitayama, H., Ishikawa, J., Nishiura, T., Kanayama, Y.,Yonezawa, T. & Tarui, S. (1991) Proliferation of human my-eloid leukemia cell line associated with the tyrosine-phospho-rylation and activation of the proto-oncogene c-kit product. Blood78: 2834-2840.
Le Cam, A. (1991 ) Natural tyrosine-kinase inhibitors. Pathol. Biol.39: 796-800.
Lee, H. P., Gourley, L., Duffy, S. W., Esteve, J., Lee, J. &. Day,N. E. (1991) Dietary effects on breast-cancer risk in Singapore.Lancet 337: 1197-1200.
Lehtola, L., Lehväslaiho, H., Koskinen, P. & Alitalo, K. (1992) Achimeric EGFR/neu receptor in functional analysis of the neuoncoprotein. Acta Oncol. 31: 147-150.
Lookhart, G. L., Jones, B. L. & Finney, K. F. (1978) Determinationof coumestrol in soybeans by high-performance liquid and thin-layer chromatography. Cereal Chem. 55: 967-972.
Lu, H. &. Liu, GT. (1992) Anti-oxidant activity of dibenzocy-clooctene lignans isolated from schisandraceae. Planta Med. 58:311-313.
Makishima, M., Honma, Y.; Hozumi, M, Sampi, K., Hattori, M.,Umezawa, K. & Motoyoshi, K. (1991) Effects of inhibitors ofprotein tyrosine kinase activity and/or phosphatidylinositolturnover on differentiation of some human myelomonocyticleukemia cells. Leukemia Res. 15: 701.
Makishima, M., Honma, Y., Hozumi, M., Nagata, N. & Motoyoshi,K. (1993) Differentiation of human monoblastic leukemiau937 cells induced by inhibitors of myosin light chain kinaseand prevention of differentiation by granulocyte-macrophagecolony-stimulating factor. Biochim. Biophys. Acta 1176: 245-
249.Markaverich, B. M. & Clark, J. H. (1979) Two binding sites for
estradici in rat uterine nuclei: Relationship to uterotropic response. Endocrinology 105: 1458-1462.
Markaverich, B. M., Upchurch, S. & Clark, J. H. (1981) Progesterone and dexamethasone antagonism of uterine growth: a rolefor a second nuclear binding site for estradici in estrogen action.J. Steroid Biochem. 14: 125-132.
Markaverich, B. M., Gregory, R. R., Alejandro, M. A., Clark, J. H.,Johnson, G. A. & Middleditch, B. S. (1988a) Methyl p-hy-
droxyphenyllactate. An inhibitor of cell growth and proliferationand an endogenous ligand for nuclear type II binding sites. J.Biol. Chem. 263: 7203-7210.
Markaverich, B. M., Roberts, R. R., Alejandro, M., Johnson, G. A.,Middleditch, B. S. &. Clark, J. H. (1988b) Bioflavonoid interaction with rat uterine type II binding sites and cell growth inhibition. J. Steroid Biochem. 30: 71-78.
Markaverich, B. M. &.Gregory, R. R. (1993) Preliminary assessment of luteolin as an affinity ligand for type-II estrogen-bindingsites in rat uterine nuclear extracts. Steroids 58: 268-274.
stein on mammalian DNA topoisomerase II. Cancer Res. 49:5111-5117.
Martin, P. M., Horwitz, K. B., Ruyan, D. S. & McGuire, W. L.(1978) Phytoestrogen interaction with estrogen receptors inhuman breast cancer cells. Endocrinology 103: 1860-1867.
tsumoto, K., Nishino, H. &.Aoike, A. (1993) Genistein arrestscell cycle progression at G(2)-M. Cancer Res. 53: 1328-1331.
Mills, P. K., Beeson, W. L., Phillips, R. L. &. Fraser, G. E. (1989)Cohort study of diet, lifestyle and prostate cancer in adventistmen. Cancer 64: 598-604.
Mousavi, Y. & Adlercreutz, H. (1992) Enterolactone and estradiolinhibit each other's proliferative effect on MCF-7 breast cancercells in culture. J. Steroid Biochem. Mol. Biol. 41: 615-619.
Mousavi, Y. & Adlercreutz, H. |1993) Genistein is an effectivestimulator of SHBG production in Hep-G2 human liver cancercells and suppresses proliferation of these cells in culture. Steroids58:301-304.
Mäkelä,S., Pylkkänen,L., Saniti, R. & Adlercreutz, H. (1991) Roleof plant estrogens in normal and estrogen-related altered growth
of the mouse prostate. In: EURO FOOD TOX III. Proceedingsof the Interdisciplinary Conference on Effects of Food on theImmune and Hormonal Systems, pp. 135-139. Institute of Toxicology, Swiss Federal Institute of Technology &. University ofZürich,Schwerzenbach, Switzerland.
Nairn, M., Gestetner, B., Kirson, I., Birk, Y. & Bondi, A. (1973)A new isoflavone from soya beans. Phytochemistry 22: 237-239.
Nairn, M., Gestetner, B., Bondi, A. & Birk, Y. (1976) Antioxidativeand antihemolytic activities of soybean isoflavones. J. Agrie. FoodChem. 24: 1174-1177.
Ogawara, H., Akiyama, T., Watanabe, S. L, Ito, N., Kobori, M. &Seoda, Y. (1989) Inhibition of tyrosine protein kinase activityby synthetic isoflavones and flavones. J. Antibiot. 42: 340-343.
Okura, A., Arakawa, H., Oka, H., Yoshinari, T. & Monde, Y. (1988)Effect of genistein on topoisomerase activity and on the growth
Orcheson, L., Rickard, S., Seidl, M., Cheung, F., Luyengi, L., Fong,H. & Thompson, L. U. (1993) Estrus cycle and organ changesin rats fed flaxseed, its mammalian lignan precursor or tamoxifen.FASEB J. 7(3 Pt II): 1686(abstr.(.
Ota, K. & Misu, Y. A. (1958) A study on latent carcinoma of theprostate in Japanese. Gann 49(Suppl): 283-284.
Peterson, G. & Barnes, S. (1991) Genistein inhibition of thegrowth of human breast cancer cells—independence from estrogen receptors and the multi-drug resistance gene. Biochem. Biophys. Res. Commun. 179: 661-667.
Peterson, G. &.Barnes, S. (1993) Genistein and biochanin-A inhibitthe growth of human prostate cancer cells but not epidermalgrowth factor receptor tyrosine autophosphorylation. Prostate22: 335-345.
Phipps, W. R., Martini, M. C., Lampe, J. W., Slavin, J. L. &. Kurzer,M. S. (1993) Effect of flax seed ingestion on the menstrualcycle. J. Clin. Endocrinol. Metab. 77: 1215-1219.
Price, K. R. &.Fenwick, G. R. (1985) Naturally occurring oestro-gens in foods—a review. Food Addit. Contam. 2: 73-106.
Ranelletti, F. O., Ricci, R., Larocca, L. M., Maggiano, N., Capelli,A., Scambia, G., Benedettipanici, P., Mancuso, S., Rumi, S. &Piantelli, M. (1992) Growth-inhibitory effect of quercetin andpresence of type-II estrogen-binding sites in human colon-cancercell lines and primary colorectal tumors. Int. J. Cancer 50: 486-
492.Reddy, K. B., Mangold, G. L., Tandon, A. K., Yone'da, T., Mundy,
G. R., Zilberstein, A. & Osborne, C. K. (1992) Inhibition ofbreast cancer cell growth in vitro by a tyrosine kinase inhibitor.Cancer Res. 52: 3636-3641.
Rose, D. P., Boyar, A. P. & Wynder, E. L. (1986) Internationalcomparison of mortality rates for cancer of the breast, ovary,prostate, and colon, and per capita food consumption. Cancer58:2363-2371.
Schweigerer, L., Christeleit, K., Fleischmann, G., Adlercreutz, H.,Wähälä,K., Hase, T., Schwab, M., Ludwig, R. & Fotsis, T. (1992)Identification in human urine of a natural growth inhibitor forcells derived from solid paediatric tumours. Eur. J. Clin. Invest.22: 260-264.
Serraino, M. & Thompson, L. U. (1991) The effect of flaxseedsupplementation on early risk markers for mammary carcino-genesis. Cancer Lett. 60: 135-142.
Serraino, M. &. Thompson, L. U. (1992a) The effect of flaxseedsupplementation on the initiation and promotional stages ofmammary tumorigenesis. Nutr. Cancer 17: 153-159.
Serraino, M. &. Thompson, L. U. (1992b) Flaxseed supplementation and early markers of colon carcinogenesis. Cancer Lett.63: 159-165.
Setchell, K. D. R., Lawson, A. M., Mitchell, F. L., Adlercreutz, H.,Kirk, D. N. &.Axelson, M. (1980) Lignans in man and in animalspecies. Nature 287: 740-742.
Setchell, K. D. R., Lawson, A. M., Borriello, S. P., Harkness, R.,Gordon, H., Morgan, D. M. L., Kirk, D. N., Adlercreutz, H.,Anderson, L. C. & Axelson, M. (1981) Lignan formation inman—microbial involvement and possible roles in relation tocancer. Lancet 2: 4-7.
Setchell, K. D. R., Borriello, S. P., Hulme, P. & Axelson, M. (1984)Nonsteroidal estrogens of dietary origin: possible roles in hormone-dependent disease. Am. J. Clin. Nutr. 40: 569-578.
Severson, R. K., Nomura, A. Y. M., Grove, J. S. & Stemmerman,G. N. (1989) A prospective study of demographics and prostatecancer among men of Japanese ancestry in Hawaii. Cancer Res.49: 1857-1860.
Sharma, O. P., Adlercreutz, H., Strandberg, J. D., Zirkin, B. R., Cof-fey, D. S. &. Ewing, L. L. (1992) Soy of dietary source plays apreventive role against pathogenesis of prostatitis in rats. J. Steroid Biochem. Mol. Biol. 43: 557-564.
Shimizu, H., Ross, R. K., Bernstein, L., Yatani, R., Henderson, B. E.&.Mack, T. M. (1991) Cancers of the prostate and breast amongJapanese and white immigrants in Los Angeles County. Br. J.Cancer 63: 963-966.
Shultz, T. D., Bonorden, W. R. & Seaman, W. R. (1991 ) Effect ofshort-term flaxseed consumption on lignan and sex hormonemetabolism in men. Nutr. Res. 11: 1089-1100.
Shutt, D. A. &Cox, R. I. (1972) Steroid and phytoestrogen bindingto sheep uterine receptors in vitro. J. Endocrinol. 52: 299-310.
Stitch, S. R., Toumba, J. K., Groen, M. B., Funke, C. W., Leemhuis,J., Vink, J. & Woods, G. F. (1980) Excretion, isolation andstructure of a phenolic constituent of female urine. Nature 287:738-740.
Teraoka, H., Ohmura, Y. &Tsukada, K. (1989) The nuclear matrixfrom rat liver is capable of phosphorylating exogenous tyrosine-containing substrates. Biochem. Int. 18: 1203-1210.
Thompson, L. U. &. Serraino, M. (1990) Lignans in flaxseed andbreast and colon carcinogenesis. In: Proceedings of the Flax Institute of the U.S., January 25-26, 1990. pp. 30-35. Fargo, NorthDakota.
Tráganos, F., Ardelt, B., Halko, N., Bruno, S. & Darzynkiewicz, Z.(1992) Effects of genistein on the growth and cell cycle progression of normal human lymphocytes and human leukemicMOLT-4 and HL-60 cells. Cancer Res. 52: 6200-6208.
Trowell, H. C. & Burkitt, D. P. (1981) Western Diseases: TheirEmergence and Prevention. Edward Arnold Ltd, London, England.
Van Thiel, D. H., Galvaoteles, A., Monteiro, E., Rosenblum, E. &Gavaler, J. S. (1991) The phytoestrogens present in de-etha-nolized bourbon are biologically active—a preliminary study ina postmenopausal woman. Alcohol. Clin. Exp. Res. 15: 822-823.
Versantvoort, C. H. M., Schuurhuis, G. J., Pinedo, H. M., Eekman,C. A., Kuiper, C. M., Lankelma, J. &. Broxterman, H. J. (1993)Genistein modulates the decreased drug accumulation in non-P-Glycoprotein mediated multidrug resistant tumour cells. Br.J. Cancer 68: 939-946.
Walz, E. (1931) Isoflavon- und saponin-glucoside in Soja hispida.Justus Liebigs Ann. Chem. 498: 118-155 (in German).
Wang, C., Mäkelä,T. H., Hase, T. A., Adlercreutz, C. H. T. &.Kurzer,M.S. (1994) Lignans and isoflavonoids inhibit aromarase enzyme in human preadipocytes. J. Steroid Biochem. Molec. Biol.50: 205-212.
Watanabe, S. & Koessel, S. (1993) Colon cancer: an approach frommolecular epidemiology. J. Epidemici. 3: 47-61.
Waters, A. P. & Knowler, J. T. "(1982) Effect of a lignan (HPMF)
Wei, H. C., Wei, L. H., Frenkel, K., Bowen, R. & Barnes, S. (1993)Inhibition of tumor promoter-induced hydrogen peroxide formation in vitro and in vivo by genistein. Nutr. Cancer 20: 1-12.
Weindruch, R., Albanes, D. & Kritchevsky, D. (1991) The roleof calories and caloric restriction in carcinogenesis. Hematol.Oncol. Clin. North Am. 5: 79-89.
Whitten, P. L. & Naftolin, F. (1991) Dietary estrogens—a biologically active background for estrogen action. In: New Biologyof Steroid Hormones (Hochberg, R. B., Naftolin, P., eds.), pp.155-167. Raven Press, New York.
Wille«, W. C., Hunter, D. J., Stampfer, M. J., Colditz, G., Manson,J. E., Spiegelman, D., Rosner, B., Hennekens, C. H. & Speizer,F. E. (1992) Dietary fat and fiber in relation to risk of breastcancer. J. Am. Med. Assoc. 21: 2037-2044.
Yatani, R., Chigusa, I., Akazaki, K., Stemmerman, G. N., Welsh,R. A. a Correa, P. (1982) Geographic pathology of latentprostatic cancer. Int. J. Cancer 29: 611-616.
Yokotsuka, T. (1986) Soy sauce biochemistry. Adv. Food Res. 30:195-329.
