Clinical Abstracts on Soy

Clinical Abstracts 1

Role of the Main Components of Whole Soybean Products, Soy Protein and Soy Oil, in Reducing Hypercholesterolemia

E.M. Kurowska* J. Jordan*, J.D.Spencer*, S. Wetmore*, L.Piche**, M. Radzikowski**, and KK.Carroll*
*University of Western Ontario, Canada
**.Brescia College, Ontario, Canada
Presented at The Second International Symposium on the Role of Soy In Preventing and Treating Chronic Disease, Brussels, Belgium, September 15-18, 1996.

Dietary substitution of soy protein for animal protein is known to be associated with a reduction of plasma total and LDL cholesterol levels, especially in individuals with hypercholesterolemia. It was hypothesized that this reduction would be greater if whole soybean products (containing soy protein and soy oil) were substituted for animal products (containing animal protein and fat). To investigate this, effect of whole soybean products versus an animal product, milk, was studied in 34 hypercholesterolemic individuals (17 men, 17 women).

Using a three-treatment three period, cross-over design, the subjects received sequentially three types of beverages, each for a period of 4 weeks, with two week washout periods: I) 2% cow's milk products, II) a combination of skim milk products and soy oil, III) whole soybean products containing soy protein and 2% soy oil. Daily intake of calories, protein, carbohydrates and total fat was unaffected by treatment as assessed by three-day food records.

The whole soybean treatment significantly increased HDL cholesterol (by 7%, p<0.04) and reduced LDL/HDL cholesterol ratio (by 14%, p<0.007). In 24 participants, called "responders", treatment with whole soybean products significantly reduced LDL cholesterol (by 11%, p<0.05) and LDL/HDL cholesterol ratio (by 19%, p<0.008).

The magnitude of observed beneficial changes in plasma lipoprotein profile depended on baseline characteristics of the subjects. Changes in LDL/HDL cholesterol ratio induced by the intake of soybean products were inversely related to the initial LDL/HDL cholesterol ratio (r=-0.56, p<0.0007) and positively correlated with the initial HDL cholesterol concentrations (r=0.52, p<0.002).

In summary, our results demonstrated that 1) in individuals with moderate hypercholesterolemia, intake of whole soybean products can improve plasma lipid profile and the effect appears to be mostly but not entirely due to soy protein, 2) susceptibility to treatment with soybean products is greater in subjects with higher initial LDL and lower initial HDL cholesterol levels.

Clinical Abstracts 2

Soy and Hypocholesterolemic Effects: Clinical Experience and Molecular Mechanisms

Cesare R. Sirtori*, Christina Manzoni+, Elisabetta Giannazza+ and Marla R. Lovatti+
*Director, Centre E. Grossi Paoletti, +Institute of Pharmacological Sciences, University of Milano, Italy.
Presented at The Second International Symposium on the Role of Soy In Preventing and Treating Chronic Disease, Brussels, Belgium, September 15-18, 1996.

The first clinical study reporting significant cholesterol-lowering properties of a soy-protein-based diet was by Hodges et.al. in 1967 in moderately hypercholesterolemic patients after substitution of animal proteins with a textured soybean product (TVP), had a fall of cholesterolemia of 20% or more. Our first clinical trial involved 30 hypercholesterolemic (type IIA) patients, 20 underwent a crossover study of a TVP versus animal protein diet, resulting in a 20-24% reduction of total and LDL-cholesterol. Patients with cholesterolemias > 250-280mg/dl (>6.5- 7.0 mmol/L) are most responsive. In a meta-analysis, Anderson et.al.[NEJM 1995;333:276-282] confirmed that the percent LDL cholesterol reduction ranges from -3.3% in normolipidemic subjects, up to -26% or more in patients with marked elevations (>335mg/dl).

Elucidation of the mechanism/s of plasma cholesterol reduction has been the object of stimulating debate. The search for the mechanism has been more recently focused on the protein components. Changing from animal to soy protein is followed by significant increases in the LDL-receptor activity in the liver in animals and lympho-monocytes in man. Studies at the molecular level have suggested that soy globulins, specifically the 7S, may exert a cholesterol lowering effects in animals, and, in vitro, can significantly increase the maximal binding of LDL to liver high-affinity receptors.

Clinical Abstracts 3

Metanalysis Review of Soy Protein in Reducing the Risk of Coronary Heart Disease

James W. Anderson, M.D. Professor of Medicine and Clinical Nutrition University of Kentucky, Lexington, KY, United States.
Presented at the American Dietetic Association 80th Annual Meeting and Exhibition, held October 27-30, 1997, in Boston, Massachusetts.

Soy protein exerts several anti-atherogenic effects. First, it decreases LDL-cholesterol levels significantly 1}. Second, it tends to increase HDL-cholesterol levels this is rather unique since most dietary interventions such as oat bran intake or creased saturated fat intake significantly decrease HDL-cholesterol levels. Third, soy isoflavones, plant chemicals unique to soybeans, have antioxidant properties, which protect LDL from oxidation. Fourth, soy isoflavones have favorable effects on blood vessel function.

Our recent meta-analysis of soy protein studies clearly indicates the potency of soy protein in decreasing LDL-cholesterol levels. In this study we analyzed the results of 38 studies using over 730 research volunteers conducted as controlled clinical studies of which were published in peer-reviewed journals.

Thirty-four studies included only adults while four studies included only children. As a soy protein source, 20 used isolated soy protein, 15 used textured soy protein, and three used a combination of isolated and textured soy protein. Soy protein intake averaged 47 grams per day (range 17-124 grams/day) and 15 studies (40%) used <31 grams of soy protein daily. In 14 of the studies the test diets resembled a typical Western diet while in 21 studies the diets were low in fat (<30% of energy) and low in cholesterol (<200 mg/day). In 19 studies the soy protein and control diets were considered to be comparable with respect to total fat intake, saturated fat intake, cholesterol intake, and weight maintenance.

Soy protein intake was associated with a 9.3% reduction in serum cholesterol, a 12.9% reduction in serum LDL-cholesterol, and a 10.5% reduction in serum triglycerides. All of these decreases were statistically significant. Serum HDL-cholesterol levels increased by 2.4%, a non-significant increase. These findings had a strong consistency because 34 of 38 studies reported that soy protein intake decreased serum cholesterol levels (P < 0.001). Those volunteers with normal initial levels had LDL-cholesterol decreases of 7.7% while those with severe hypercholesterolemia had LDL-cholesterol decreases of 24%. This indicates that individuals with more severe hypercholesterolemia are likely to show a greater response to soy protein use.

Based on this scientific evidence we recently proposed these guidelines for soy protein intake. For persons in good general health a suggestions is to have 7 servings of soy protein per week. This would provide an average of approximately 8 to 10 grams of soy protein daily with 16-20 mg of soy isoflavones daily. For persons with diabetes or risk factors for coronary heart disease or persons with strong family histories of heart disease, osteoporosis or diabetes we suggest a goal of 14 servings of soy protein per week. This would provide an average of approximately 16 to 20 grams of soy protein/day with 32 to 40 mg of soy isoflavones/day.

Conclusion:

Since every one percent reduction in serum cholesterol decreases estimated risk of heart attack by two to three percent, this serum cholesterol reduction has the potential to reduce risk for CHD by 18-28%. Statistical analysis indicates that the intake of 25 grams of soy protein per day would reduce serum cholesterol by 8.9 mg/dl, 50 grams would reduce it 17.4 mg/dl, and 75 grams of soy protein would reduce serum cholesterol by 26.3 mg/dl. Our study, however, indicated that intake of 17-25 grams of soy protein per day could have a meaningful effect on serum cholesterol levels.

Clinical Abstracts 4

Cholesterol Lowering Effects by Soy: Potential Mechanisms

Shigeru Yamamoto, Takashi Yamamoto*, Hei-Mei Chung**, Ming-Fu Wang**, Sumie Shinjo, Tatsushi Komatsu***
*Fuji Oil Co., Izumisano, Osaka 598, Japan, **Department of Food and Nutrition, Province University, Taichung, Taiwan
*** Unit of Nutrition in Hospital, University of Occupational and Environmental Health, Kitakyushu, 807 Japan.
Presented at The Second International Symposium on the Role of Soy In Preventing and Treating Chronic Disease, Brussels, Belgium, September 15-18, 1996.

Many researchers in animals and humans have confirmed the hypocholesterolemic effect of soybean. The following experiments were conducted to study the relationships and the mechanisms involved.

Experiment 1

Sugano and his colleagues have shown that peptide fractions of soybean protein (SP-Peptide) produced in the lumen bind steroids and excrete them into the feces in rats. They tried to confirm it in humans and young female volunteers with slightly high concentration of serum cholesterol were given either diets containing casein or SP-Peptides for about a month. The results showed that, in the SP-Peptide group demonstrated a higher excretion of fecal steroids, higher concentration of HDL-cholesterol and lower concentration of LDL-cholesterol than in the casein group.

Experiment 2

Conducted to assess growth, nitrogen balance and other nutritional parameters in rats fed SP or various SP-Peptide fractions. No differences among all the groups. These results together with the results of Experiment 1 show that SP-Peptides transport steroids to the lower part of intestine to be excreted and then are themselves absorbed.

Experiment 3

Rats were fed with either SP or Casein for two months. The SP-diet group indicates a lower serum cholesterol concentration but also lower abdominal fat than in the casein group.

Experiment 4

To confirm the results of Experiment 3, obese ladies were given low energy diets or skimmed milk or SP for 3 weeks. It was found that in the SP group demonstrated more losses in body weight and fat were greater than in the skimmed milk group.

Experiment 5

The effect of SP on body fat cannot be explained by the increase of steroids in the feces, therefore, Shigeru et.al. hypothesize that very small quantities of SP-Peptides goes into the circulatory system and affect the lipid metabolism. To clarify this, they administered saline, SP-Peptides (10mg) or amino acid mixture patterned after SP-Peptides (10mg) to mice through the tail vein daily for 2 weeks. Serum cholesterol concentration was lower in the SP-Peptide group than in the other two groups.

Conclusion

The results suggest two mechanisms of the anticholesterolemic effect of soy protein: 1) Peptides produced from soybean protein in lumen bind steroids and excrete them into feces but the peptides themselves are absorbed into the circulatory system, 2) Small quantities of the peptides goes into the circulatory system and affects lipid metabolism.

Clinical Abstracts 5

Effect of Active Peptides from Soy Proteins on Cholesterol Homeostasis in Cell Cultures.

M. R. Lovati1, C. Manzoni1, E. Gianazza1, E. M. Kurowska2, and C. R. Sirtori1; 1Institute of Pharmacological Sciences, Faculty of Pharmacy, Milano, Italy; 2Centre for Human Nutrition, Department of Biochemistry, University of Western Ontario, London, ON, Canada. Presented at Third International Symposium on the Role of Soy in Preventing and Treating Chronic Disease October 31 November 3, 1999 Omni Shoreham Hotel ,Washington, DC,USA

An activation of LDL receptors was recently described in a human hepatoma cell line (Hep G2) exposed to purified (a+a') subunits from 7S soy globulin; ß chains were ineffective (Lovati et al. 1998). Further experiments in HepG2 cells demonstrated, by using a mutant soy cultivar devoid of the a' a subunits (Keburi), a marked LDL-receptor activation by the a' vs. subunit. A commercial isoflavone-free, heat-hydrolyzed soy concentrate, found effective in type II hypercholesterolemic patients, showed an LDL-receptor activation similar to that of the 7S globulin when incubated under the same conditions (Manzoni et al. 1998).

To assess the final identity of the putative peptide (or peptides) responsible for the biochemical effect, experiments were performed in Hep G2 cells exposed either to synthetic peptides corresponding to specific sequences differing among the a+ a' and ß subunits or peptides coming from the in vitro digestion (trypsin + pepsin) of soy concentrate Moreover, the ability of the whole 7S globulin, its a + a' and ß subunits, and whole soy concentrate interfere in the apoB secretion in the medium as well as in sterol biosynthesis was evaluated in the same model.

Hep G2 cells were preincubated for 24 h at 37°C in minimal essential medium with 5% lipoprotein-deficient serum in the presence or absence of synthetic and in vitro-produced peptides at different concentrations (10-4 or 10-6 mol/L for synthetic peptides, 0.5 mg/mL for digested Croksoy). After addition of 125I-LDL, cells were incubated at 37 %C for a further 4 h to determine the uptake and degradation of LDL. Increased 125I-LDL uptake (+41%) and degradation (+10%) vs. controls were shown after Hep G2 incubation with a synthetic peptide (10-4 mol/L, MW 2271) corresponding to the 127-150 positions of the consensus sequence present in the a' and not in the ß subunit of the 7S globulin.

Cells exposed to digested soy concentrate showed a marked up-regulation of LDL receptors vs. that in controls as well as vs. HepG2 cells incubated with undigested soy concentrate (125I-LDL uptake: +58% and +17%, respectively; 125I-LDL degradation: +113% and +26%, respectively). Among soy-derived products, only the 7S soy globulin showed an inhibitory activity on apolipoprotein B secretion (- 80% secretion) and 14C-acetate incorporation (-70%, -60%, and -48% into free and esterified cholesterol and triacylglycerol, respectively) when tested in Hep G2 cells at concentration of 1.0 mg/mL. These findings support the hypothesis (Sirtori et al. 1998) that one or more peptides escaping detection in vivo may reach the liver after intestinal digestion, thus eliciting a cholesterol-lowering effect. Moreover, these data indicate that the protein moiety, devoid of isoflavone components, is responsible for the biochemical effect.

Clinical Abstracts 6

Effects Independent of Cholesterol Reduction: Anti-atherogenic Effects of Soybean Protein. Viewpoints from Peroxidizability and Molecular Size of LDL and From Anti-Platelet Aggregation

Takemichi Kanazawa, The Second Department of Internal Medicine, Hirosaki Unviersity School of Medicine, Hirosaki, Japan. Presented at Third International Symposium on the Role of Soy in Preventing and Treating Chronic Disease Septermber 15-18, 1996, Brussels, Belgium.

To investigate the biological function of soy protein for prevention of vascular disease, the following studies were carried out. Low density lipoprotein (LDL) was separated from plasma of the patient with myocardial infarction (22 cases), cerebral infarction (18 cases), familial hypercholesterolemia (8 cases) and healthy (18 persons) by ultracentrifugation. Peroxidized LDL was evaluated by estimation of peroxidized cholesteryl linoleate on thin layer chromatography or of lipid peroxide by Determiner Kit. Lipids were measured by enzymatic method. Molecular structure of soybean peptide was determined by IH-NMR, HPLC and MS. Molecular size of LDL was calculated by planimeter from photograph of negative stain. Soy crème contained 10.6% protein was prepared by our previous method (J. Nutr. 125,639, 1995). Soybean peptide was separated by protease treatment, ODS-AQ column chromatography and HPLC.

The results:

Normal LDL obtained from healthy persons did not enhanced platelet aggregation induced by ADP, collagen and epinephrine, but LDL of patient with vascular disease enhanced clearly platelet aggregation.

Administration of soy crème suppressed an markedly peroxidizability of LDL due to Cu++ (in rabbit).
Soy crème administered orally suppressed an enlargement of LDL during cholesterol feeding (in rabbit)
Soy crème administration reduced LDL cholesterol level (in human and rabbit).
A peptide consisted of six kinds of amino acids, which was prepared from soybean suppressed markedly platelet aggregation, and composition and structure of amino acid was Asp-Glu-Gly-Leu-Phe-Arg.

Conclusion:

Various molecular constituents with useful function such as anti-peroxidizability to LDL, anti-platelet aggregability, suppression of enlargement of LDL molecular size during cholesterol administration and cholesterol reducing effect were contained in soybean. Thus, soybeans are very useful food for preventing vascular disease.

Clinical Abstract 7

Effect of Soy Protein on LDL Cholesterol in Hypercholesterolemic Costa Rican Children.

G. Arauz, G. Padilla, M. Roselló, S. Guzmán, S. Rodriguez, and L. Cunningham, J. M. Ordovás
Costa Rican Institute for Research and Education on Nutrition and Health (Inciensa), Tres Rios, Costa Rica.

A diet low in cholesterol, total fat, and saturated fat is the first step in the treatment of hypercholesterolemia. However, many children cannot achieve plasma cholesterol goals by using this dietary approach. It has been long recognized that there is a hypocholesterolemic effect of soy protein when it is partially substituted for animal protein. The objective of our study was to evaluate the effect of soy protein on LDL cholesterol reduction when soy products are used as part of a regular diet at home.

The product used were Archer Daniels Midland isolated protein and textured protein (soy meat and soymilk). We carried out a clinical random study including 26 hypercholesterolemic children; 13 were in the control group and 13 were in the intervention group (mean age 10.3 ±-15 y). During the 8-wk experimental period, both groups consumed a step 1 diet (< 30% total fat, < 10% saturated fat, < 250 mg cholesterol). The intervention group also consumed the soy products as part of their regular meals. Food intake and adherence to the protocol were assessed during weekly home visits. Daily soybean protein intake in the intervention group was 20.88 ±-33 g/d.

Preliminary data from 18 children (9 in each group) show that the mean LDL-cholesterol reduction was significantly (P < 0.005) higher in the intervention group (15.5%) than in the control group (5.5%). There were no significant differences in total cholesterol reduction. The addition of soy protein to a step 1 diet in hypercholesterolemic children was associated with a specific reduction in LDL cholesterol. We therefore propose that soy protein may offer a nonpharmacological alternative to reducing elevated LDL cholesterol in children.

Clinical Abstract 8

Molecular Effects of Soy-Isoflavone Genistein in Prostate Cancer Cells

Joanne N. Davis, Omer Kucuk, and Fazlul H. Sarkar
Departments of Cancer Biology1, Internal Medicine2, and Pathology3, Karmanos Cancer Institute, Wayne State University School of Medicine, Detroit, MI, USA. Third International Symposium on the Role of Soy in Preventing and Treating Chronic Disease. October 31 - November 3, 1999 Omni Shoreham Hotel Washington, DC USA

Prostate cancer is the second leading cause of cancer-related deaths in men in the United States, accounting for 36% of all male cancers and 13% of cancer-related deaths in men. Epidemiologic data provide convincing evidence that dietary factors play an important role in the etiology of cancer. We previously demonstrated that the dietary isoflavone genistein inhibits proliferation, induces apoptosis, and modulates important cell cycle regulatory molecules, particularly p21WAF1 and cyclin B, in prostate cancer cells, and therefore may be a potential chemopreventive or therapeutic agent. To further elucidate the molecular mechanism by which genistein elicits its effects, we first investigated the role of a transcription factor NF-B, and second, measured prostate-specific antigen (PSA) levels in prostate cancer cells. NF-B was shown to protect cells against apoptosis by initiating prosurvival mechanisms. We investigated whether genistein modulates NF-B, particularly the inactivation, which may lead to the apoptosis observed in genistein-treated cells. Here we show that genistein decreases NF-B activity in prosate cancer cells in a dose-dependent manner. Prostate cancer cells treated with genistein at 30 and 50 mol/L for 24 h resulted in reduced NF-B DNA binding. Using confocal microscopy, we showed that genistein blocks the translocation of NF-B p50 and p65 subunits from the cytoplasm to the nucleus, preventing NF-B activation and prohibiting DNA binding. Additionally, we demonstrated that genistein abrogates NF-B activation by two known inducers, H2O2 and tumor necrosis factor- (TNF-). Prostate cancer cells pretreated with 50 mol genistein/L for 48 h inhibited NF-B DNA binding and blocked translocation of NF-kB subunits to the nucleus when stimulated with either H2O2 or TNF-. These results suggest that the inactivation of NF-B by genistein may lead to the cell growth inhibition and apoptosis observed in genistein-treated cells.

The most valuable tumor marker used for the detection and monitoring of prostate cancer is PSA. PSA, a member of the kallidrein family, is a serine protease secreted by prostate epithelial cells. PSA is able to cleave the predominant seminal vesicle protein and was proposed as a candidate growth factor, cytokine, or growth factor regulator and was linked to tumor progression. Therefore, we investigated whether genistein has any effect on PSA expression and secretion in the androgen-sensitive prostate cancer cell line, LNCaP. LNCaP cells were treated with genistein at 0, 30, and 50 mol/L for 3 d. The medium was collected and assayed for the presence of PSA. We observed that treatment with genistein at 30 mol/L reduced PSA secretion by 50% and 50 mol/L reduced PSA by 80% compared with control. Using immunohistochemistry and Western blot analysis, we determined that genistein inhibits PSA protein expression levels but did not affect the protein expression levels of another tumor- associated antigen, prostate-specific membrane antigen. These results indicate that genistein lowers the PSA levels in prostate cancer cells in vitro.

In conclusion, the inactivation of NF-B and downregulation of PSA by genistein provide encouraging evidence to support genistein's role as a chemopreventive and therapeutic agent for prostate cancer. These results also indicate that NF-B may play a pivotal role in genistein-induced apoptosis, providing a mechanism by which genistein promotes cell death.

Clinical Abstract 9

The Specific Role of Genistein in Estrogen Metabolism

N. B. Kumar, K. Allen, A. Cantor, G. Shaw, and C. E. Cox; H. Lee Moffitt Cancer Center and Research Institute at the University of South Florida, Tampa, FL, USA. Third International Symposium on the Role of Soy in Preventing and Treating Chronic Disease. October 31 - November 3, 1999 Omni Shoreham Hotel Washington, DC USA

Aim. To evaluate the individual effectiveness of supplementing a group of premenopausal, breast cancer--free women with a dietary supplement of the isoflavone genistein (40 mg/d) in producing a change in sex hormones that are implicated in the initiation and promotion of breast cancer.

Methods. Sixty-eight consecutively recruited premenopausal omnivorous women, of all races and ethnicities, aged 25--55 y were admitted to the study and randomly assigned to an experimental group supplemented with soy (40 mg genistein/d) or to a control group consuming a placebo for a 12-wk period. Changes in their anthropometric, nutritional, and hormonal biomarkers from the early follicular phase were analyzed at baseline and after intervention,

Results. Preliminary analysis indicates that hormonal concentrations of free estradiol decreased by 78% in the group consuming genistein compared with 44% in the placebo group. Serum estrone and sex hormone-binding globulin levels were elevated in 47% and 46% of the subjects, respectively, in the experimental group compared with 26% and 40%, respectively, in the placebo group. In the experimental group the menstrual cycle of 58% of the subjects increased by more than 2 d compared with 36% in the placebo group.

Conclusions. This data suggest that increased genistein intake affects estrogen metabolism by altering the sex hormone concentrations that are implicated in breast cancer promotion or inhibition.

Clinical Abstract 10

Effect of Soybean Saponins on the Growth and Antioxidant Defense of Human Hepatocarcinoma Cells.

M.-K. Sung and M.-Y. Park; Department of Food and Nutrition, Sookmyung Women's University, Seoul, Korea. Third International Symposium on the Role of Soy in Preventing and Treating Chronic Disease. October 31 - November 3, 1999 Omni Shoreham Hotel Washington, DC USA

Carcinogenesis is a multistep process including initiation, promotion, and progression. Recent studies indicated that oxygen free radicals, byproducts of normal cellular respiratory processes as well as lipid peroxidation processes, induce cellular DNA damages, which is a most plausible mechanism for the initiation of carcinogenesis. Lipid peroxides also were shown to promote tumor cell growth. Saponins are amphiphilic compounds present in a variety of edible and nonedible plants.

Recent studies indicated that saponins extracted from soybeans inhibit the formation of lipid peroxides in corn oil samples; this may be due to their ability to scavenge radicals. In this study, effects of soybean saponins on the growth, cellular lipid peroxidation, and antioxidative enzyme activities of HepG2 cells were investigated. Effects of saponins were compared with -tocopherol and ascorbic acid. Cells (1-2 ( 107) were incubated for 24 h and then treated with tert-butylhydroperoxide (0.5 nmol/L for 45 min) to initiate lipid peroxidation followed by saponin treatment (300 g/plate for 48 h). Cellular superoxide dismutase (SOD), glutathione peroxidase (GPX), and glutathione S-transferase (GST) activities were measured.

Results showed that tert-butylhydroperoxide treatment significantly increased cellular malondialdehyde content. Cell growth was significantly decreased with saponin, -tocopherol, and ascorbic acid treatment. Malondialdehyde content was significantly reduced by saponin (72%) and -tocopherol (40%). Soybean saponins significantly increased cellular SOD, GPX, and GST activities. Ascorbic acid significantly decreased GPX activity. However, the activity of GST was not affected by either -tocopherol or ascorbic acid. These results indicate soybean saponins possess considerable antioxidative capacity, exerting antiproliferative effects on tumor cells.

Clinical Abstract 11

Soybean Saponins Inhibit the Formation of DNA Adducts in Human Colon and Liver Cells.

H.-S. Jeon and M.-K. Sung; Department of Food and Nutrition, Sookmyung Women's University, Seoul, Korea. . Third International Symposium on the Role of Soy in Preventing and Treating Chronic Disease. October 31 - November 3, 1999 Omni Shoreham Hotel Washington, DC USA

Numerous chemical carcinogens activated to form electrophilic agents react with DNA, which explains the induction of a heritable change in a cell leading to malignant transformation. This may be a main event in the initiation of carcinogenesis.

Soybeans contain up to 2% saponins. Soybean saponins were shown to inhibit the growth of human colon carcinoma cells with low toxicity. Also, they were shown to decrease the ornithine decarboxylase activity that is directly related to cancer cell proliferation. These results indicate that soybean saponins are important modulators in the promotion stage of carcinogenesis. This study was performed to examine the effects of soybean saponins on DNA adducts formation, which is the most important reaction of carcinogens with cellular macromolecules initiating carcinogenesis. CCD-18Co and HepG2 cells were used as models for colon and liver cells, respectively. Cells (4-5 (105) were seeded and allowed to attach. After 18 d (CCD-18Co) and 2 d (HepG2) in culture, soybean saponins at a concentration of 0-50 g/mL were added and incubated for 1 h. Preincubated tritiated aflatoxin B1(12 nmol/L; specific activity 25 Ci/mmol) was added to each plate and incubated for a further 48 h. DNA was purified and aliquots of DNA samples were used to measure radioactivity by liquid scintillation counting.

Results showed that soybean saponins significantly inhibited the formation of DNA-aflatoxin B1 adducts in CCD-18Co cells by 23.7%, 50.7%, and 49.4%, respectively, when 10, 30, and 50 g saponins/mL were used. Amounts of DNA adducts in HepG2 cells were also significantly decreased by 37.3%, 50.1%, and 49.8%, respectively.

These results indicate that soybean saponins may effectively reduce cellular DNA damages by carcinogens and can be regarded as potential chemopreventive agents.

Clinical Abstract 12

Antimitotic and Cancer Preventive Properties of a Soybean Peptide

Ben O. de Lumen and Alfredo F. Galvez; Nutritional Sciences and Toxicology, University of California at Berkeley, Berkeley, CA, USA. . Third International Symposium on the Role of Soy in Preventing and Treating Chronic Disease. October 31 - November 3, 1999 Omni Shoreham Hotel Washington, DC USA

Epidemiologic evidence suggesting the correlation between diets high in soybean and overall low cancer mortality rates, especially those of colon, breast, and prostate, has given impetus to identifying components in soybean responsible for its anticancer properties.

We isolated a soybean cDNA encoding the small subunit peptide of a cotyledon-specific 2S albumin (Gm2S-1). The peptide (named lunasin) has a unique, highly acidic carboxyl end. A chimeric gene encoding the lunasin peptide tagged with green fluorescent protein arrested cell division and caused abnormal spindle fiber elongation, chromosomal fragmentation, and cell lysis when transiently transfected into murine embryo fibroblast, murine hepatoma, and human breast cancer cells. Deletion of the acidic carboxyl end abolished the antimitotic effect. Immunolocalization of lunasin and an immuno-binding assay using synthetic peptides revealed the preferential adherence of lunasin to chromatin.

Lunasin is the first antimitotic peptide whose cDNA was cloned and the first from a common food source. We also showed that the exogenous application of lunasin peptide to as low as 125 nmol/L inhibits the in vitro transformation of mouse embryo fibroblast cells (C3H 10T 1/2) into tumorous foci by the carcinogens 3-methyl cholanthrene and 7,12-dimethylbez[a]anthracene.

The lunasin gene has potential application as an antimitotic cancer therapeutic agent, and the lunasin peptide may be an important cancer-preventive compound in soybeans. Furthermore, its antimitotic property suggests that lunasin could play a native role in arresting mitosis that initiates the cell expansion phase of seed development where DNA endoreduplication and synthesis of storage proteins, lipids, and carbohydrates occur.

Clinical Abstract 13

Dietary Intake of Phyto-Estrogens and Metabolic Cardiovascular Risk Profile: The Framingham Study

De Kleuijn Miriam, J., Van Der Schouw Yvonne, T., Wilson Peter,W.,Crobbee Dieederick, E, Jacques Paul, F , 30-03-2001

Women with hypertension, central obesity and elevated blood lipids are at high risk of cardiovascular disease. Menopause is associated with adverse changes in plasma lipid levels and body fat distribution. Hormone replacement therapy (HRT) can partly reverse these changes. Adverse effects such as vaginal bleeding and the associated risk of breast cancer in users of combined HRT prompted the search for alternative estrogen based preventive treatments.

Plant derived phyto-estrogens (isoflavones) are structurally and functionally comparable to estrogens made in the body. Animal studies showed that phyto-estrogens in soy imporve plasma lipoprotein and decrease cholesterol content of the aorta, and there are indications that phyto-estrogens protect against breast and uterine cancers.

The aim of the present study was to investigate the association between intake of the two main classes of dietary phtyo-estrogens, isoflavones and lignans, and metabolic cardiovascular risk factors, in particular blood pressure, waist hip ratio, and plasma lipoprotein levels.

We analyzed data obtained from 939 postmenopausal, Caucasion women participating n the Framingham Offspring Study. The waist hip ratio was lower in women with the highest intakes of lignans compared to women with the lowest intakes. Women with the highest intake of isoflavones or lignans had lower plasma triglyceride levels and higher plasma HDL cholesterol levels than women with the lowest intakes of isoflavones or lignans.

High intake of phyto-estrogens in postmenopausal women appears to be associated with a favorable metabolic cardiovascular risk profile.

Soy and Human Health