Suppressive effects of dietary genistin and daidzin on rat prostate carcinogenesis.

High intake of phytoestrogens through soybeans and their products is thought to be associated with low incidences of prostate and / or breast cancer in Asian countries. Possible chemopreventive effects of genistin or daidzin on rat prostate carcinogenesis were therefore investigated. Male F344 rats were given 10 biweekly subcutaneous injections of 3,2'-dimethyl-4-aminobiphenyl (DMAB) and then either genistin or daidzin in the diet at a concentration of 0.1% for 40 weeks. Other groups of rats given DMAB were treated with genistin or daidzin together with a high dose of testosterone propionate (TP). Both genistin and daidzin reduced the numbers of ventral prostate carcinomas (P < 0.05), with a tendency for decrease in incidence. Invasive carcinomas which developed in the anterior prostate and seminal vesicles with TP were, however, not influenced by the two isoflavones. Thus, the present data suggest that genistin and daidzin possess anti-cancer effects at relatively early stages of prostate cancer development, providing experimental support for epidemiological findings.     

Induction of invasive carcinomas in the accessory sex organs other than the ventral prostate of rats given 3,2'-dimethyl-4-aminobiphenyl and testosterone propionate

The promotion effects of testosterone propionate (TP) on prostate carcinogenesis were investigated in F344 rats given the prostatic carcinogen, 3,2'-dimethyl-4-aminobiphenyl (DMAB). One group of animals received s.c. DMAB injections at a dose of 50 mg/kg body weight at 2-week intervals for a total of 10 injections along with s.c. implantations of TP-containing Silastic tubes. A second experimental group of rats was given DMAB at the same dose and intervals but each injection of DMAB was combined with 3 prior consecutive daily 100-mg/kg body weight s.c. injections of TP. After cessation of carcinogen administration, animals in these two groups received TP implants from week 21 to the end of the experiment. All surviving animals were killed at week 56 and accessory sex gland tumor incidences were compared to those in DMAB alone and other appropriate control groups. The groups given TP plus DMAB and subsequent long term administration of TP developed lesions of the dorsolateral prostate, seminal vesicles, and coagulating glands which were all invasive adenocarcinomas. Incidences were 84.2% (16 of 19 rats) and 66.7% (12 of 18 rats), respectively. Macroscopic large tumors were induced in 13 animals among which 8 demonstrated metastasis to the abdominal cavity, liver, or lung. None of the control groups except for the group given TP injections plus DMAB had equivalent tumors. Development of carcinomas of the ventral prostate, which were all of in situ type, were not increased by subsequent treatment with TP. These data thus clearly showed that TP can exert strong enhancing effects on tumor development in the dorsolateral prostate, seminal vesicles, and coagulating glands but not in the ventral prostate.     

Beef Tallow, but Not Perilla or Corn Oil, Promotion of Rat Prostate and Intestinal Carcinogenesis by 3,2-Dimethyl-4-aminobiphenyl

The modifying effects of three kinds of fat (corn oil, beef tallow or perilla oil, each at 20% in the diet) on F344 rat prostate carcinogenesis induced by 3,2-dimethyl-4-aminobiphenyl (DMAB) were investigated. Non-invasive carcinomas of the ventral prostate were induced by DMAB alone and invasive carcinomas of the other prostate lobes and seminal vesicles by DMAB and testosterone propionate (TP). Eight groups of F344 rats were initiated with 50 mg/kg body weight of DMAB at 2-week intervals for the first 20 weeks, four also receiving TP, extended until week 60. The animals received basal chow powder diet or one of three high fat diets throughout the experiment (60 weeks). One further group served as a non-carcinogen-treated control maintained on basal chow powder diet. Beef tallow significantly increased the development of ventral prostate carcinomas with DMAB alone (from 15 to 45%, P<0.05), while perilla oil reduced the incidence of prostatic intraepithelial neoplasia (PIN) in the ventral lobe of rats given DMAB + TP (from 70 to 10%, P<0.01), but not in those given DMAB alone. No other effects of high fats were observed regarding PIN or invasive cancers of the dorsolateral and anterior prostate or seminal vesicles. A satellite experiment demonstrated that all high fat diets for 4 weeks increased the 5-bromo-2-deoxyuridine (BrdU) labeling index of prostate epithelial cells, suggesting that a high fat intake, irrespective of the fatty acid composition, may accelerate cell kinetics in the prostate. Of the three high fat diets, beef tallow was also found to increase intestinal carcinogenesis. Thus, the present data revealed carcinogenesis in the prostate and intestine to be promoted by beef tallow.     

Beef tallow, but not perilla or corn oil, promotion of rat prostate and intestinal carcinogenesis by 3,2'-dimethyl-4-aminobiphenyl.

The modifying effects of three kinds of fat (corn oil, beef tallow or perilla oil, each at 20% in the diet) on F344 rat prostate carcinogenesis induced by 3,2'-dimethyl-4-aminobiphenyl (DMAB) were investigated. Non-invasive carcinomas of the ventral prostate were induced by DMAB alone and invasive carcinomas of the other prostate lobes and seminal vesicles by DMAB and testosterone propionate (TP). Eight groups of F344 rats were initiated with 50 mg / kg body weight of DMAB at 2-week intervals for the first 20 weeks, four also receiving TP, extended until week 60. The animals received basal chow powder diet or one of three high fat diets throughout the experiment (60 weeks). One further group served as a non-carcinogen-treated control maintained on basal chow powder diet. Beef tallow significantly increased the development of ventral prostate carcinomas with DMAB alone (from 15 to 45%, P < 0.05), while perilla oil reduced the incidence of prostatic intraepithelial neoplasia (PIN) in the ventral lobe of rats given DMA + TP (from 70 to 10%, P < 0.01), but not in those given DMAB alone. No other effects of high fats were observed regarding PIN or invasive cancers of the dorsolateral and anterior prostate or seminal vesicles. A satellite experiment demonstrated that all high fat diets for 4 weeks increased the 5-bromo-2-deoxyuridine (BrdU) labeling index of prostate epithelial cells, suggesting that a high fat intake, irrespective of the fatty acid composition, may accelerate cell kinetics in the prostate. Of the three high fat diets, beef tallow was also found to increase intestinal carcinogenesis. Thus, the present data revealed carcinogenesis in the prostate and intestine to be promoted by beef tallow.     

Decrease of Prostaglandin E2 and 5–Bromo–2'–deoxyuridine Labeling but Not Prostate Tumor Development by Indomethacin Treatment of Rats Given 3,2'–Dimethyl-4-aminobiphenyl and Testosterone Propionate

The modifying effects of indomethacin (IM) on rat prostate carcinogenesis induced by 3,2'–dimethyl–4–aminobiphenyl (DMAB) were investigated. F344 rats were given 50 mg/kg body weight of DMAB at 2–week intervals for 20 weeks and then received IM at a dose of 20 ppm in the drinking water for 37 weeks. Separate groups additionally received testosterone propionate (TP) in Silastic tubes throughout the experiment. DMAB alone induced carcinomas in situ in the ventral lobe and in combination with TP caused invasive carcinomas of the dorso–lateral and anterior lobes and seminal vesicles. No clear suppression by IM of development of in situ carcinomas or in vasive carcinomas was observed. In a short–term satellite experiment, it was revealed that prostaglandin E₂ (PGE₂) levels in the dorso–lateral prostate and seminal vesicles, but not the ventral prostate, were significantly reduced by IM and that TP itself also suppressed PGE₂ levels. The 5–bromo–2'–deoxyuridine labeling index in the ventral prostate was significantly decreased by IM administration. These results indicate that while IM can efficiently suppress tissue PGE₂ levels, it does not inhibit tumor development in the prostate or seminal vesicles of rats in the present model.     

Direct Effects of Testosterone, Dihydrotestosterone and Estrogen on 3,2'-Dimethyl-4-aminobiphenyl-induced Prostate Carcinogenesis in Castrated F344 Rats

The present experiment was carried out to explore the effect of endogenous androgen on rat prostate carcinogenesis induced by 3,2'-dimethyl-4-aminobiphenyl (DMAB) and testosterone propionate (TP) or 5α-dihydrotestosterone (DHT) with or without ethinyl estradiol (EE). In order to eliminate the influence of endogenous androgen, F344 rats were orchiectomized just after initiation with the prostate carcinogen, DMAB, and then given TP, DHT, TP plus EE or DHT plus EE for 40 weeks. The results demonstrated that while administration of TP following DMAB treatment causes invasive carcinomas in the lateral and anterior prostate and seminal vesicles, DHT does not exhibit equivalent effects. Synergistic enhancement was also evident with TP plus EE, but not with DHT plus EE. The incidences of prostatic and seminal vesicle lesions in all groups of the present experiment, except for the group given castration without hormonal supplement, were equivalent to those previously found in non-castrated animals. Therefore, the present findings indicate that endogenous testosterone may not be required for promotion hy TP/EE of DMAB-initiated prostate carcinogenesis and that it may not contribute to the actions of DHT.     

Lobe specific effects of testosterone and estrogen on 3,2'-dimethyl-4-aminobiphenyl-induced rat prostate carcinogenesis

We have previously shown that chronic administration of a pharmacological dose of testosterone propionate (TP) after treatment with the carcinogen, 3,2'-dimethyl-4-aminobiphenyl (DMAB), results in development of invasive and metastatic adenocarcinomas arising from the dorso-lateral and anterior prostate, as well as the seminal vesicles. Co-administration of ethinyl estradiol (EE) with TP increased the yield of carcinomas in the lateral and anterior lobes. In the present experiment, male F344 rats were treated with DMAB for 20 weeks and then co-administered a pharmacological dose of TP together with various doses of EE for 40 weeks. Without hormone(s) administration, carcinomas were confined to the ventral prostate and all were of intra-acinar type. TP administration suppressed development of the ventral prostate carcinomas but caused invasive carcinomas of the lateral and anterior lobes and of seminal vesicles and intra-acinar carcinomas in the dorsal prostate. The appearance of carcinomas in the lateral and anterior prostate was increased by co-administration of EE in a dose-related fashion but carcinomas of the seminal vesicles were inversely reduced. The suppressive influence of TP on ventral carcinoma development was overcome by only the highest dose of EE. It is concluded that estrogen can modify the enhancing effects of TP on induction of rat prostate and seminal vesicle carcinomas in a dose-related fashion with lobe specificity.     

Enhancing Effect of Cadmium on Rat Ventral Prostate Carcinogenesis Induced by 3,2'-Dimethyl-4-aminobiphenyl

The effects of cadmium given at different stages during 3,2'-dimethyl-4-aminobiphenyl (DMAB)-induced rat prostate carcinogenesis were investigated using male F344 rats. Animals were given 10 subcutaneous injections of 50 mg/kg body weight of DMAB or the corn oil vehicle at two-week intervals. In addition, cadmium was administered at doses of 0, 10, or 30 /μmol/kg body weight as single intramuscular injection on the 1st day of the experiment or one day after the last injection of DMAB at week 20. Two further groups were subjected to administration of cadmium at 10 μmol/kg at week 20 and then 5 μmol/kg at week 40, or 10 μmol/kg at week 20 and then 5 μmol/kg at weeks 30, 40 and 50. At the termination, 60 weeks after the beginning of the experiment, the incidences and multiplicity of ventral prostate carcinomas in the groups given cadmium plus DMAB demonstrated a consistent tendency for increase over control values (groups receiving DMAB or cadmium alone). The numbers of carcinomas per rat and per unit area of prostate section were significantly elevated in the two groups given low doses of cadmium after cessation of DMAB administration. Cadmium alone also induced a few prostate carcinomas. The influence on development of prostate tumors did not appear to be a result of the induced severe testicular atrophy because serum testosterone levels were not affected. The results indicate that cadmium and DMAB can act synergistically to cause rat prostate carcinogenesis.     

Lack of tumorigenic response in the prostate gland of castrated F344 rats by 3,2'-dimethyl-4-aminobiphenyl given with methyltestosterone

In an attempt to induce a high incidence of prostate carcinoma, 3,2'-dimethyl-4-aminobiphenyl (DMAB), a prostatic carcinogen, was given during the period of cell proliferation of the prostate gland induced by the administration of methyltestosterone (MT) to castrated F344 rats. Three weeks after the surgical castration, rats were given diet containing 300 ppm of MT for 2 weeks and basal for 2 weeks alternately 12 times. During each treatment with MT, one (group 1) or two (group 2) subcutaneous injections of 50 mg/kg body wt. of DMAB was given. After the last treatment of MT, a pellet of testosterone propionate (TP) was implanted in the subcutis of all animals until the end of the experiment (week 60). No carcinomas developed in the prostate gland of any of the rats. Atypical hyperplasia of the ventral lobe of prostate was found in 4 of 22 rats in group 1 and 2 of 20 rats in group 2. The incidences of atypical hyperplasia of the seminal vesicles in groups 1 and 2 were 64% and 75%, respectively. No pathological lesions in the prostate were observed in 32 rats given DMAB without MT treatment.     

Lack of chemopreventive effects of lycopene and curcumin on experimental rat prostate carcinogenesis

The chemopreventive efficacy of lycopene and curcumin with regard to prostate carcinogenesis was investigated using 3,2'-dimethyl-4-aminobiphenol (DMAB)- and 2-amino-1-methylimidazo[4,5-b]pyridine (PhIP)-induced rat ventral prostate cancer models. Three 60 week experiments with male F344 rats were carried out. In the first DMAB was given for the first 20 weeks and lycopene or curcumin were administered concomitantly or subsequently at dietary doses of 15 and 500 p.p.m., respectively. In the second experiment lycopene and curcumin were given to rats pretreated with DMAB at doses of 5, 15 or 45 p.p.m. or 100 or 500 p.p.m. In the third PhIP was selected as an initiator for prostate carcinogenesis and administered for 20 weeks. Rats were then fed a diet containing lycopene at a dose of 45 p.p.m. or curcumin at a dose of 500 p.p.m. or both together. Chemopreventive effects of lycopene and curcumin on development of DMAB-induced ventral prostate carcinomas were observed only in the first experiment and no confirmation of inhibition potential was obtained in the following studies. Neither summational nor synergistic chemoprevention was evident. It is concluded from the present data that, overall, neither lycopene nor curcumin can consistently prevent rat prostate carcinogenesis.     

Sequential Observation of Rat Prostate Lesion Development Induced by 3,2'-Dimethyl-4-aminobiphenyl and Testosterone

3,2'-Dimethyl-4-aminobiphenyl (DMAB), when combined with high doses of testosterone propionate (TP) induces invasive adenocarcinomas with metastatic potential in the rat prostate. The processes underlying this tumor development, including the involvement of atypical hyperplasias, were sequentially investigated in F344 rats. DMAB was given subcutaneously at a dose of 50 mg/kg body weight 10 times at 2-week intervals. TP was administered chronically (in Silastic tubes) from the beginning of the experiment or after the DMAB administration until termination (week 60). Invasive adenocarcinomas were induced in the lateral and anterior prostate as well as the seminal vesicles. Atypical hyperplasias appeared from an early stage, with the later appearance of cancers being closely associated with such foci of morphological alteration. The findings confirm that combined administration of DMAB and pharmacological doses of TP yields invasive adenocarcinomas in the rat prostate and provide further support for the conclusion that atypical hyperplasias are premalignant lesions.     

Rat prostate as one of the target organs for 3,2'-dimethyl-4-aminobiphenyl-induced carcinogenesis: effects of dietary ethinyl estradiol and methyltestosterone

Twenty consecutive weekly sc injections of 50 mg/kg body weight of 3,2'-dimethyl-4-aminobiphenyl (DMAB), a multipotential carcinogen, were given to male F344 rats and subsequently groups of animals were treated with dietary ethinyl estradiol (EE, 2.5 ppm) or methyltestosterone (MT, 300 ppm) for up to 40 weeks. Prostate carcinomas were found in 4 out of 32 rats given DMAB followed by MT and in 2 out of 29 rats given DMAB alone. Atypical hyperplasia of the prostate epithelium in these two groups was found in 22% and 14%, respectively. Neither carcinoma nor atypical hyperplasia was seen in the prostate of animals given DMAB followed by EE. In other organs, tumors were frequently found in the ear duct, skin, and large intestine and less frequently in the lung, preputial glands, small intestine and liver. EE significantly suppressed tumor incidence of the ear duct and sebaceous glands while increasing the incidence of liver tumor and mesothelioma. The present data indicates DMAB to be a useful carcinogen for the induction of prostate carcinomas in rats.     

Low tumorigenic response to 3,2'-dimethyl-4-aminobiphenyl administration in the prostate of rats castrated at birth

Six-week-old rats which had been orchiectomized at birth were given 3,2'-dimethyl-4-aminobiphenyl (DMAB) at various doses combined with a stimulus to prostate epithelial cell proliferation in the form of oral administration of methyltestosterone (MT) for 4 weeks. Thereafter MT treatment was continued or the animals received subcutaneous implants of testosterone propionate (TP) and were maintained until sacrifice at week 60. Although prostatitis and prostatic enlargement were frequently observed, especially in the TP group, numbers of atypical hyperplastic lesions were low and only one prostatic carcinoma in situ developed. Thus, despite the presence of proliferation, castration brought about a significant reduction in susceptibility to DMAB.     

Low Tumorigenic Response to 3,2'-Dimethyl-4-aminobiphenyl Administration in the Prostate of Rats Castrated at Birth

Six-week-old rats which had been orchiectomized at birth were given 3,2'-dimethyl-4-aminobiphenyl (DMAB) at various doses combined with a stimulus to prostate epithelial cell proliferation in the form of oral administration of methyltestosterone (MT) for 4 weeks. Thereafter MT treatment was continued or the animals received subcutaneous implants of testosterone propionate (TP) and were maintained until sacrifice at week 60. Although prostatitis and prostatic enlargement were frequently observed, especially in the TP group, numbers of atypical hyperplastic lesions were low and only one prostatic carcinoma in situ developed. Thus, despite the presence of proliferation, castration brought about a significant reduction in susceptibility to DMAB.     

Development of Androgen-independent Carcinomas from Androgen-dependent Preneoplastic Lesions in the Male Accessory Sex Organs of Rats Treated with 3,2'-Dimethyl-4-aminobiphenyl and Testosterone Propionate

Two kinds of cancer can be induced in rat male accessory sex organs, one a non-invasive carcinoma arising in the ventral lobe and the other an invasive lesion which develops in the dorsolateral and anterior lobe as well as the seminal vesicles. In the present study, one group of male rats were given biweekly s.c. injections of 3,2'-dimethyl-4-aminobiphenyl (DMAB) for 20 weeks for induction of non-invasive carcinomas and the other group received DMAB with 40-week testosterone propionate for induction of invasive carcinomas. Half of the animals in each group were then subjected to bilateral orchiectomy at week 41 to remove testicular androgen, in order to examine the androgen dependence of both types of carcinomas as well as precancerous lesions. Animals were killed at weeks 41, 46 and 60. All parts of the prostate complex showed involution and significant weight reduction after castration, with a complete disappearance of atypical hyperplasias and carcinomas of the ventral prostate. However, in spite of suppression of development of atypical hyperplasias in the anterior prostate and seminal vesicles, the incidence of invasive carcinomas was not changed. Normal epithelial cells and atypical hyperplasias of all parts of the prostate and seminal vesicles and carcinomas of the ventral prostate were immunohistochemically positive for nuclear androgen receptor, while invasive carcinomas that developed in either castrated or non-castrated animals were negative. These findings suggest that in the ventral prostate, both precancerous and cancerous lesions are androgen-dependent, but in the anterior and seminal vesicles, cancerous lesions (invasive carcinomas) are androgen-independent while precancerous lesions are hormone-dependent.     

Development of androgen-independent carcinomas from androgen-dependent preneoplastic lesions in the male accessory sex organs of rats treated with 3,2'-dimethyl-4-aminobiphenyl and testosterone propionate.

Two kinds of cancer can be induced in rat male accessory sex organs, one a non-invasive carcinoma arising in the ventral lobe and the other an invasive lesion which develops in the dorsolateral and anterior lobe as well as the seminal vesicles. In the present study, one group of male rats were given biweekly s.c. injections of 3,2'-dimethyl-4-aminobiphenyl (DMAB) for 20 weeks for induction of non-invasive carcinomas and the other group received DMAB with 40-week testosterone propionate for induction of invasive carcinomas. Half of the animals in each group were then subjected to bilateral orchiectomy at week 41 to remove testicular androgen, in order to examine the androgen dependence of both types of carcinomas as well as precancerous lesions. Animals were killed at weeks 41, 46 and 60. All parts of the prostate complex showed involution and significant weight reduction after castration, with a complete disappearance of atypical hyperplasias and carcinomas of the ventral prostate. However, in spite of suppression of development of atypical hyperplasias in the anterior prostate and seminal vesicles, the incidence of invasive carcinomas was not changed. Normal epithelial cells and atypical hyperplasias of all parts of the prostate and seminal vesicles and carcinomas of the ventral prostate were immunohistochemically positive for nuclear androgen receptor, while invasive carcinomas that developed in either castrated or non-castrated animals were negative. These findings suggest that in the ventral prostate, both precancerous and cancerous lesions are androgen-dependent, but in the anterior and seminal vesicles, cancerous lesions (invasive carcinomas) are androgen-independent while precancerous lesions are hormone-dependent.     

Lack of modification by naturally occurring antioxidants of 3,2'-dimethyl-4-aminobiphenyl-initiated rat prostate carcinogenesis

The modifying effects of 6 naturally occurring antioxidants on 3,2'-dimethyl-4-aminobiphenyl (DMAB)-initiated rat prostate carcinogenesis were investigated in male F344 rats. Animals were pretreated with DMAB in a 20-week initiation protocol and then administered basal diet containing 0.8% catechol, 0.8% resorcinol, 0.8% hydroquinone, 2 ppm selenium, 2% gamma-orysanol or 1% alpha-tocopherol for 40 weeks. The experiment was terminated at week 60 for histopathological assessment of lesion development. Atypical hyperplasias and carcinomas of the prostate were observed in the ventral lobe in all groups treated with DMAB. However, the incidences of these lesions were not significantly different between carcinogen control and antioxidant-treated groups. There were also no significant increases or decreases in the incidences of tumors in any other organs.     

Promoting action of prolactin released from a grafted transplantable pituitary tumor (MtT/F84) on rat prostate carcinogenesis

The potential modifying effects of high prolactinemia on rat prostate carcinogenesis was investigated. Male F344 rats were treated at 5 times of 5-week intervals with s.c. injections of 3,2'-dimethyl-4-aminobiphenyl (DMAB), each injection following 3 weeks pretreatment with dietary ethinyl estradiol. After completion of the carcinogen administration stage, rats received multiple s.c. transplantations of a prolactin producing transplantable pituitary tumor, MtT/F84 until sacrifice at week 51. The effects of additional or single treatment with bromocriptine, a prolactin suppressing agent, were also investigated. The body, liver and kidney but prostate weights were significantly increased in the groups given MtT/F84. Although the development of prostate carcinomas was not affected by the observed hyperprolactinemia, the incidences of atypical hyperplasia of both ventral and lateral prostate were significantly enhanced. The findings thus indicate that prolactin may have promoting potential for prostate carcinogenesis.     

The effect of disulfiram on the carcinogenicity of 3,2' dimethyl-4-aminobiphenyl in Syrian golden hamsters and rats

3,2' -Dimethyl-4-aminobiphenyl (DMAB) was administered s.c.to Syrian golden hamsters and CDF rats with and without disulfiramin the diet. In a group of male hamsters given only weekly injectionsof DMAB over a period of 18–22 months, 10 of 25 hamstersdeveloped adenocarcinomas of the small or large intestine. Threehad carcinomas in both the large and small intestine, 3 hadonly colon carcinomas and 4 only small intestinal carcinomas.In the group fed disulfiram and carcinogen, 10 of 25 hamstersdeveloped adenocarcinomas of the colon, but only one carcinomaof the small intestine occurred. Furthermore, the appearanceof intestinal tumors was delayed. Urinary bladder carcinomasoccurred in both groups exposed to DMAB, with the appearancebeing slightly delayed in the group also receiving disulfiram.In male rats given DMAB s.c. once a week for 10 months, theaddition of disulfiram in the diet reduced the incidence ofcancers of the small intestine, similar to the effects in hamsters,but resulted in the occurrence of a small number of bladdercancers. An unusual occurrence was the development of prostaticcarcinoma in rats given DMAB.     

Variation in tumor yield in the prostate and other target organs of the rat in response to varied dosage and duration of administration of 3,2'-dimethyl-4-aminobiphenyl

The effects of varying dosages of 3,2'-dimethyl-4-aminobiphenyl (DMAB) in combination with cyclic dietary administration of ethinyl estradiol (EE) on induction of prostate carcinoma were investigated in male F344 rats. Animals received repeated treatment with 0.75 ppm of EE for 3 wk with intervals of 2 wk on basal diet. The cycle was repeated 10, 5, and 3 times in Groups 1, 2, and 3, respectively, a single s.c. injection of DMAB being given 2 days after each change to basal diet at a dose of 50 mg/kg of body weight in Group 1, 100 mg/kg of body weight in Group 2, and 167 mg/kg of body weight in Group 3. With this dosing schedule, the total dose of DMAB (500 mg/kg of body weight) per rat was the same in each group. Subsequent to the last treatment with EE, all rats were given basal diet until the end of the experiment (Wk 60) when all surviving animals were sacrificed for histological examination. Carcinoma of the prostate was found in 58.6, 45.0, and 25.9% of rats surviving for 60 wk in Groups 1 to 3, respectively, the incidences of atypical hyperplasia being 86.2, 85.0, and 74.1%. However, tumors of the small and large intestines, preputial gland, and pancreas developed in a dosage-dependent manner, the largest incidences being found in the group given 167 mg of carcinogen 3 times. Thus the present experiment confirmed that administration of DMAB combined with cyclic treatment with EE induces a high incidence of prostate carcinoma in rats and demonstrated that a low dosage of DMAB given over a long period is superior to a high dosage over a short period for specific induction of prostate lesions.