Serotherapy of primary rat mammary carcinoma: inhibition by ethylenedinitrilotetraacetic acid but not by [ethylenebis(oxyethylenenitrilo)]tetraacetic acid

We reported inhibition of growth of primary rat mammary carcinomas after infusions of tumor-bearer plasma absorbed with Protein A-Sepharose or inactivated CNBr Sepharose. Absorbed plasmas were depleted of the third component of complement (C3) (other complement components defined similarly) and C5 but not C1, C4, or C2. These results suggested that activation of the alternative pathway of complement might be involved in the observed antitumor effects. To test this concept sera were treated with ethylenedinitrilotetraacetic acid or [ethylenebis(oxyethylenenitrilo)]tetraacetic acid before absorption with Protein A-Sepharose. Ethylenedinitrilotetraacetic acid, by chelating calcium and magnesium, prevents activation of both the alternative and classical complement pathways. [Ethylenebis(oxyethylenenitrilo)]tetraacetic acid, by chelating calcium but not magnesium, permits activation of the alternative pathway but inhibits activation of the classical complement pathway. Sera in the presence or absence of chelating agent were absorbed with Protein A-Sepharose twice at room temperature. After absorption calcium was added to the sera. Rats were treated by i.v. injection of sera twice a week for 2 weeks. Measurements of tumor size were made weekly for 5-7 weeks and then tumor weight was determined. Groups were compared both for size of index and total tumors. The results can be summarized as follows: tumor-bearer sera before absorption did not inhibit the growth of rat primary mammary carcinomas; tumor-bearer sera after absorption with Protein A-Sepharose showed significant consumption of C3 and did inhibit tumor growth; tumor-bearer sera absorbed in the presence of ethylenedinitrilotetraacetic acid did not show a decrease in C3 functional activity and did not inhibit tumor growth; tumor-bearer sera absorbed in the presence of [ethylenebis(oxyethylenenitrilo)]tetraacetic acid did show a decrease in C3 functional activity and did inhibit tumor growth; sera from normal adult female rats after absorption with Protein A-Sepharose did inhibit tumor growth. The results are consistent with a role for the alternative pathway of complement in the inhibition of growth of rat primary mammary carcinomas observed after treatment with absorbed sera.     

Mammary tumor induction by N-methyl-N-nitrosourea in genetically resistant Copenhagen rats.

The Copenhagen rat is completely resistant to mammary cancer induction by N-methyl-N-nitrosourea (MNU) when the carcinogen is administered during sexual development, a period when other strains of rats are normally susceptible to mammary gland carcinogenesis. Here we administered 30 mg/kg MNU i.p. to two groups of neonatal (2-3-day-old) Copenhagen rats. One group (group B, 18 animals) received no further treatment, while the other group (group C, 17 animals) received a second dose of 30 mg/kg MNU via the tail vein at 50 days of age. About 30% of the rats in group B and about 70% of those in group C developed mammary carcinomas before they were 1 year of age. About one-half of the tumors in both groups were cribriform adenocarcinomas and one-half were adenosquamous carcinomas. The latter tumor type has not been observed previously in susceptible rat strains. The ability to induce these mammary tumors in the Copenhagen rat suggests that the putative mammary carcinoma suppressor gene is functionally inactive in neonatal animals or is inactivated when these animals are treated with MNU.     

Synergism between neutron radiation and diethylstilbestrol in the production of mammary adenocarcinomas in the rat.

When young female A X C rats were given 9.6 rads of 0.43-MeV neutrons, 32 of 33 survived a 50-week follow-up period, 2 rats developed a total of 3 mammary adenocarcinomas, and 3 rats developed a total of 4 mammary fibroadenomas. For 25 rats implanted with a 20-mg pellet containing 5 mg diethylstilbestrol and 15 mg cholesterol, average survival was 284 days; 22 rats developed a total of 182 mammary adenocarcinomas, and 21 rats developed a pituitary tumor. When diethylstilbestrol was given 2 days before neutron radiation to 35 rats, the average survival was 239 days; 32 rats developed a total of 842 mammary adenocarcinomas, 1 rat developed a single mammary fibroadenoma, and 34 rats developed a pituitary tumor. All of the 31 control rats survived the 50-week study period, and none developed tumors. Twenty-one of the rats that received both diethylstilbestrol and neutron radiation and 1 rat that received only diethylstilbestrol exhibited a multiple mammary adenocarcinoma response with a range of 18 to 72 mammary adenocarcinomas per rat. These results were interpreted to mean that a synergistic interaction between diethylstilbestrol and neutron radiation on mammary adenocarcinoma formation occurs in terms of an earlier onset and a larger number of mammary adenocarcinomas. These results confirm and complement a previously reported synergistic interaction between diethylstilbestrol and X-radiation on mammary adenocarcinoma formation in A X C female rats.     

Influence of caffeine on development of benign and carcinomatous mammary gland tumors in female rats treated with the carcinogens 7,12-dimethylbenz(a)anthracene and N-methyl-N-nitrosourea

The effect of chronic caffeine consumption (500 mg/liter of drinking water) on the initiation and promotion stages of 7,12-dimethylbenz(a)anthracene (DMBA) (a low dose, 0.5 mg/100 g body weight, i.v.) and N-methyl-N-nitrosourea (MNU) (a standard dose, 2.5 mg/100 g body weight, i.v.) induced mammary gland tumorigenesis in female Sprague-Dawley rats was determined. In the initiation studies, caffeine was administered for 30 days prior to and for 3-4 days after carcinogen treatment (carcinogens administered at 55-57 days of age); in the promotion studies, caffeine was administered beginning 3-4 days after carcinogen treatment and until experiment termination (DMBA study and MNU study, 48 and 26 weeks after carcinogen treatment, respectively). In the DMBA study, there were 62-73 rats/group, in the MNU study, 40 rats/group. Eighty-nine % of the mammary tumors induced by DMBA were benign (adenomas, fibroadenomas, often with cystic secretory activity), 11% were carcinomas (intraductal and invasive); virtually all of the MNU-induced mammary tumors were carcinomas (approximately 99%). Caffeine consumption during the initiation stage in the DMBA-treated rats resulted in a significant decrease in the mean number of mammary carcinomas per rat (50% reduction, P less than 0.01) and mean number of benign mammary tumors per rat (28% reduction, P less than 0.05); caffeine consumption during the promotion stage significantly decreased the mean number of benign mammary tumors per rat (57% reduction, P less than 0.001) while not significantly influencing mammary carcinoma number. In contrast, caffeine consumption during either the initiation or promotion stages of MNU-treated rats did not significantly influence this tumorigenic process. The influence of caffeine on urinary and fecal excretion of tritiated DMBA and on rat mammary gland development at the time of carcinogen treatment also was determined. Slightly reduced levels of tritium in 24-h urinary samples were observed in caffeine-treated animals (P = 0.06). No significant effect of caffeine on 24- to 96-h fecal or 48- to 96-h urinary excretion of the isotope was observed. No apparent effect of caffeine on rat mammary gland development (number of ducts, degree of lobuloalveolar development) was observed. That caffeine significantly suppresses the initiation stage of DMBA-induced rat mammary gland tumorigenesis, while not influencing this stage when MNU is used as a carcinogen, suggests that caffeine acts via an alteration in carcinogen (DMBA) activation. The lack of a pronounced effect of caffeine on tritiated DMBA excretion, however, does cast some doubt on this mechanism.(ABSTRACT TRUNCATED AT 400 WORDS)     

Influence of caffeine consumption on 7,12-dimethylbenz(a)anthracene-induced mammary gland tumorigenesis in female rats fed a chemically defined diet containing standard and high levels of unsaturated fat

The effect of caffeine (430-500 mg/liter of drinking water) on the initiation and promotion phases of 7,12-dimethylbenz(a)anthracene (DMBA)-induced mammary gland tumorigenesis in female Sprague-Dawley rats fed a chemically defined diet containing standard (5%) or high (20%) levels of fat (corn oil) was examined. In the initiation studies, caffeine and the standard or high fat diet treatments were provided for 34 days, from 24-29 days of age to 58-63 days of age. Three days prior to termination of caffeine-fat diet treatments, each rat received a single dose of DMBA. In the promotion studies, caffeine and the standard or high fat diets were provided commencing 3 days after a single dose of DMBA (at 56-61 days of age) and until termination of the study. Caffeine consumption, during the initiation phase significantly (P less than 0.05) reduced mammary carcinoma multiplicity (number of tumors/rat), in rats fed either a standard or high fat diet. In the promotion studies, prolonged consumption of caffeine in rats fed either a standard or high fat diet did not significantly effect mammary carcinoma multiplicity. In the early stages of promotion, an apparent increase in mammary carcinoma multiplicity was observed; this increase in mammary carcinoma multiplicity did not, however, reach the 5% level of statistical probability. When caffeine was administered during both the initiation and promotion phases, no significant effect on mammary carcinoma multiplicity was observed. Treatment of rats during the initiation or promotion phases with caffeinated coffee (via drinking water) mimicked the mammary tumor modulating activities of caffeine. Decaffeinated coffee consumption did not effect either the initiation or promotion phases of this tumorigenic process. In both the initiation and promotion studies, caffeine and/or coffee consumption did not significantly affect the incidence of mammary carcinomas (percentage of rats bearing mammary carcinomas) or the mean latency period of mammary tumor appearance. Thus, in female rats fed a chemically defined standard or high fat diet, caffeine consumption can significantly influence chemical carcinogenesis of the mammary gland; an effect that is dependent upon the duration and time-span of caffeine administration.     

Serotherapy of cancer: cellular changes in primary rat mammary carcinomas after infusion of syngeneic sera absorbed with protein A-Sepharose

Serotherapy and plasma therapy have proved to be effective in the treatment of diverse neoplasms. The mechanisms of the tumoricidal or growth-inhibitory effects are unknown. We previously reported that activation of the alternative pathway of complement in absorbed sera correlated with the presence of anti-tumor activity. Complement components generated during absorption may serve as the initial mediators of cytotoxicity; for example, C5a may function in its role as a chemo-attractant. To further investigate the anti-tumor mechanisms, we undertook a series of sequential histological studies of in vivo changes in tumors following i.v. serotherapy. We found diffuse inflammatory cellular infiltrates in the interstitial compartments of primary mammary carcinomas of rats within 3-4 hr of administration of protein A-Sepharose absorbed syngeneic serum. The number of inflammatory cells was significantly higher in tumors from treated rats: total infiltrating cells (p = 0.002), eosinophils (p = 0.001), neutrophils (p = 0.001), macrophages (p = 0.001), lymphocytes (p = 0.004) and plasma cells (p = 0.001). Also, the mitotic index of tumor cells was significantly lower 4 hr after serotherapy when compared with that of untreated rat tumor cells. C3 in tumor tissue was decreased at 4 hr following serotherapy. Fibrosis was present in tumor nodules with retarded growth 5 weeks after the start of serotherapy. Localization of the infiltrating cells to tumor interstitial compartments prevents direct contact between inflammatory cells and neoplastic cells, making it unlikely that direct cell-cell killing occurs. Indirect cell killing within the tumor bed apparently occurs through several mechanisms involving interactions between serotherapy-initiated humoral mediators and inflammatory cells. The resulting anti-tumor effects include microvascular injury leading to localized ischemia, tumor infarction, and fibroblastic reactions obstructing tumor invasion and growth.     

Post-initiation treatment of rats with indole-3-carbinol or beta-naphthoflavone does not suppress 7, 12-dimethylbenz[a]anthracene-induced mammary gland carcinogenesis

Indole-3-carbinol (I3C) and β-naphthoflavone (β-NF), blocking agents of 7,12-dimethylbenz[a]anthracene (DMBA)-initiated mammary gland carcinogenesis, were examined as potential post-initiation suppressing agents. Treatment of female Sprague–Dawley rats with I3C (250 mg/kg body weight (b.w.)), β-NF (20 mg/kg b.w.) or the vehicle ethanol:corn oil (2:3) (2.5 ml/kg b.w.), three times weekly by gavage, started 3 weeks after the initiation with one oral dose of DMBA (20 mg/rat at 7 weeks of age) and continued for up to 12 weeks. I3C- or β-NF- or vehicle-treated groups did not differ significantly in the overall outcome of mammary tumorigenesis including cumulative mammary tumor incidences and multiplicities, latent periods and number and weight of mammary tumors per tumor-bearing rat for malignant, benign and/or malignant + benign tumors. A tendency of the I3C-treated rats to develop fewer mammary adenocarcinomas with a greater average weight per tumor per rat (2.32±1.50 g) than in the β-NF- (1.52±1.58 g) or vehicle- (1.55±1.53 g) treated groups suggests an effect, yet to be confirmed, of I3C on tumor development and growth. A 12-week treatment with I3C or β-NF significantly increased the P450-dependent activities of ethoxy-, methoxy-, benzyloxy- and pentoxy-(with I3C only) resorufin O-dealkylase in hepatic microsomes indicating induction of several P450s. The alterations in the P450 complement may affect endogenous estrogen metabolism and mammary gland and tumor characteristics at the molecular level, e.g. estrogen receptor status and/or proliferative activity, which require further studies.     

Specific inhibitory effect of dietary eicosapentaenoic acid on N-nitroso-N-methylurea-induced mammary carcinogenesis in female Sprague-Dawley rats

We have investigated the effects of two qualitatively different types of unsaturated fatty acids on N-nitroso-N-methylurea (NMU)-induced mammary carcinogenesis in female Sprague-Dawley rats. Semipurified diets containing 4.7% eicosapentaenoic acid (EPA) plus 0.3% linoleic acid or 5% linoleic acid were prepared. Animals maintained on these diets were given an i.v. injection of NMU (50 mg/kg body wt) at 50 days of age and killed 20 weeks later. Both tumor incidence and tumor number per rat were significantly lower in the EPA diet group (60.0% and 2.3 +/- 2.5 versus 93.3% and 5.1 +/- 4.5 respectively) for the 5% linoleic acid diet. Furthermore, the average weight of tumor material (total) per rat was significantly lower in the EPA as compared to linoleic acid diet group (2.9 +/- 4.2 g and 11.4 +/- 12.2 g respectively). Analysis of phospholipid fatty acids in the mammary tumors in the EPA diet group showed a higher proportion of C16:0, C18:2, omega-3 fatty acids C20:5 and C22:6 and a lower proportion of C20:4. Furthermore, mammary tumors in rats fed the EPA diet demonstrated significant reduction in prostaglandins. The results thus suggest that inhibition by EPA of NMU-induced mammary carcinogenesis may be mediated via the modulation of lipid metabolism and associated reduction in prostaglandin synthesis.     

Failure of ascorbic acid to inhibit growth of transplantable and dimethylbenzanthracene induced rat mammary tumors

The effect of ascorbic acid on the growth of 7,12-dimethylbenzanthracene (DMBA)-induced mammary tumor and on growth of R3230AC and MT/W9-B transplantable rat mammary tumors was investigated. High doses of ascorbic acid averaging about 540 mg/day/rat administered orally in drinking water had no effect on the growth of both R3230A and MT/W9a-B transplantable mammary tumors. Furthermore, vitamin C was unable to postpone tumor induction, reduce tumor incidence or prolong survival time of rats treated with DMBA.     

Post-initiation treatment of rats with indole-3-carbinol or β-naphthoflavone does not suppress 7,12-dimethylbenz[a]anthracene-induced mammary gland carcinogenesis

Indole-3-carbinol (I3C) and β-naphthoflavone (β-NF), blocking agents of 7,12-dimethylbenz[a]anthracene (DMBA)-initiated mammary gland carcinogenesis, were examined as potential post-initiation suppressing agents. Treatment of female Sprague–Dawley rats with I3C (250 mg/kg body weight (b.w.)), β-NF (20 mg/kg b.w.) or the vehicle ethanol:corn oil (2:3) (2.5 ml/kg b.w.), three times weekly by gavage, started 3 weeks after the initiation with one oral dose of DMBA (20 mg/rat at 7 weeks of age) and continued for up to 12 weeks. I3C- or β-NF- or vehicle-treated groups did not differ significantly in the overall outcome of mammary tumorigenesis including cumulative mammary tumor incidences and multiplicities, latent periods and number and weight of mammary tumors per tumor-bearing rat for malignant, benign and/or malignant + benign tumors. A tendency of the I3C-treated rats to develop fewer mammary adenocarcinomas with a greater average weight per tumor per rat (2.32±1.50 g) than in the β-NF- (1.52±1.58 g) or vehicle- (1.55±1.53 g) treated groups suggests an effect, yet to be confirmed, of I3C on tumor development and growth. A 12-week treatment with I3C or β-NF significantly increased the P450-dependent activities of ethoxy-, methoxy-, benzyloxy- and pentoxy-(with I3C only) resorufin O-dealkylase in hepatic microsomes indicating induction of several P450s. The alterations in the P450 complement may affect endogenous estrogen metabolism and mammary gland and tumor characteristics at the molecular level, e.g. estrogen receptor status and/or proliferative activity, which require further studies.     

Effect of moderate vitamin A supplementation and lack of dietary vitamin A on the development of mammary tumors in female rats treated with low carcinogenic dose levels of 7,12-dimethylbenz(a)anthracene

We examined the effect of moderately increased and of marginal continued dietary supplementation of vitamin A (retinyl acetate) and the effect of lack of dietary vitamin A on the initiation and promotion stages of mammary tumorigenesis in female Sprague-Dawley rats treated with a single low (0.5 mg/100 g body weight) or very low (0.1 mg/100 g body weight) dose of i.v.-administered 7,12-dimethylbenz(a)anthracene. The number of mammary tumors was significantly (P less than 0.05) reduced if prior to and during initiation with 7,12-dimethylbenz(a)anthracene the rats were fed a moderately increased (30 micrograms/day) or marginal (3 micrograms/day) amount of vitamin A, compared to rats fed an adequate (10 micrograms/day) amount of vitamin A. The number of mammary tumors was also significantly (P less than 0.05) reduced when a moderately increased or marginal amount of vitamin A was provided during the tumor promotion phase. In addition, the number of mammary tumors was significantly (P less than 0.05) reduced by the lack of dietary vitamin A during both the initiation and promotion stages of this tumorigenic process, when compared to vitamin A adequate, ad libitum-fed rats, but not when compared to vitamin A adequate, food-restricted controls. The reduction in numbers of mammary tumors observed in these studies was reflected primarily in significant (P less than 0.05) decreases in mammary fibroadenomas; the number of mammary carcinomas was often reduced, but due to a low frequency of the carcinomatous lesions, this reduction did not reach the 5% level of statistical probability. Plasma and liver vitamin A levels were determined during both the initiation and promotion stages. As the dietary supplementation of vitamin A increased from 0 to 30 micrograms/day, there was an increase in mean liver and plasma vitamin A levels. No consistent correlation between plasma and liver vitamin A levels and the occurrence of mammary tumors was observed, except with the moderately increased (30 micrograms/day) intake of vitamin A, that resulted in a small, but statistically significant (P less than 0.05) increase of serum retinol at initiation; this may account for the observed reduction in mammary tumors. These results provide evidence that moderate alterations in vitamin A consumption can modulate low-dose chemically induced mammary gland tumorigenesis. Most importantly, suppression of mammary gland tumorigenesis can be achieved by moderately increased, frequent, and regular consumption of vitamin A; prolonged consumption of vitamin A-deficient diets or diets marginal in vitamin A does not enhance the risk of mammary tumor development.     

A mammary cancer suppressor gene and its site of action in the rat

Fifty-day-old female rats of the inbred Osborne-Mendel (OM) and Copenhagen (COP) strains were exposed to a single dose of either of 2 highly effective mammary chemical carcinogens, 7,12-dimethylbenz[a] anthracene (DMBA) or 1-methyl-1-nitrosourea (MNU). Female OM rats are highly susceptible to both of these carcinogens developing greater than 5 mammary adenocarcinomas per rat following a single exposure to either chemical. In contrast, female COP rats are completely resistant to both DMBA and MNU mammary cancer induction. Genetic breeding analysis of the F1 and F2 hybrids produced by crossing COP to OM rats demonstrated that the resistance of the female COP rat to DMBA and MNU is due to the presence of a single dominant autosomal allele in the germ line of the COP rat. Transplantation experiments demonstrated that the site of action of this COP gene is within the mammary epithelial cells themselves, not systemically or at the local mammary gland level within nonepithelial mammary cells. The resistance to DMBA-induced mammary carcinogenesis affected by the COP gene does not involve prevention of the initial interaction of DMBA with the mammary epithelial cells, but suppression of the progression of these initiated mammary cells to full cancer. This suppression does not involve paracrine release of diffusible factor(s). This gene does not suppress the development of MNU-induced renal or bladder cancers in the COP female rats. Thus, this newly identified autosomal dominant gene is specifically a mammary cancer suppressor gene. Analysis of the response of female feral rats to DMBA or MNU exposure demonstrates that this mammary cancer suppressor gene is also functional in feral rats. This suggests that this mammary cancer gene is functionally inactivated either by mutation or deletion in the germ line of highly susceptible strains of rats like the OM and inbred Sprague-Dawley rats, but functionally retained in resistant strains like the COP.     

Suppression of growth of 7,12-dimethylbenzanthracene-induced mammary carcinomas in female rats by repeated whole-body hyperthermia (41-42 degrees C)

The purpose of this study was to examine effects of repeated whole-body hyperthermia on the growth of carcinogen-induced rat mammary carcinomas. Female Sprague-Dawley rats were treated intragastrically with a single dose (100 mg/kg) of the carcinogen 7,12-dimethylbenzanthracene (DMBA) at 2 months of age. Subsequently, when each rat had a minimum of one palpable mammary carcinoma, the animals were divided into two groups, one to receive hyperthermia treatments and the other to serve as controls. The hyperthermia-treated rats received 14-20 1-hour episodes of whole-body hyperthermia (averaging 41.4 degrees C, colonic) over a period of 3-4 weeks. The animals were anesthetized with sodium pentabarbital during these treatments to suppress their thermoregulation and to minimize stress mediated by higher brain functions. Their temperature was raised by exposure to high air temperatures. Control rats were anesthetized on the same schedule, and with the same doses of barbital, as hyperthermia-treated rats, but during anesthetization were placed at lower air temperatures such that their body temperatures remained normal. Tumor volumes were measured once per week. By the end of the first week, hyperthermia had suppressed carcinoma growth: whereas 78% of control carcinomas had increased in volume, 73% of hyperthermia-treated ones had decreased in volume. The average volume of mammary carcinomas in hyperthermia-treated rats after 1 week was significantly smaller than the average volume of mammary carcinomas in control rats: 0.34 vs. 0.66 cm3 (effects of differences in initial volume statistically removed). This difference had enlarged by the end of four weeks to 0.11 cm3 (hyperthermia-treated) vs. 0.80 cm3 (control). Overall, this study provides compelling evidence that whole-body hyperthermia in the range of 41-42 degrees C suppresses growth of DM-BA-induced rat mammary carcinomas, and it describes a technique for hyperthermia induction in which stresses specific to hyperthermia are minimized or avoided.     

Suppression of mammary gland carcinogenesis by post-initiation treatment of rats with tamoxifen or indole-3-carbinol or their combination.

This study examined whether suppression of mammary gland carcinogenesis elicited by low doses of tamoxifen (TAM) can be enhanced by concomitant treatment of rats with indole-3-carbinol (I3C), a component of cruciferous vegetables and a dietary supplement used for its putative antiestrogenicity. Two weeks after one oral dose of 7,12-dimethylbenz[a]anthracene (DMBA) at 65 mg/kg body weight, female Sprague-Dawley rats started treatment with TAM (10 microg/rat) by subcutaneous injection, I3C (250 mg/kg body weight) by oral gavage, TAM+I3C or their respective vehicles three times per week, for up to 20 weeks. Significant increases in the median latency of malignant mammary tumors and decreases in the mean tumor mass per rat were due to TAM. Significant decreases in the mean tumor number per rat in TAM, I3C and TAM+I3C-treated rats indicated a cooperative effect of the two compounds. In both DMBA-initiated and uninitiated rats, significant increases in the ratios of liver to body weight in I3C and TAM+I3C-treated groups coincided with I3C-dependent increases of hepatic cytochrome P450 levels and activities (1A1, 1A2 and 2B1/2). The ratios of uterus to body weight decreased with the number of treatments and the decreases effected by TAM were greater than those by I3C. The levels of circulating estrone were increased in response to I3C treatment and were greater in DMBA-initiated rats than in uninitiated rats, which may contribute to the preventive effect of I3C. Chemoprevention may be accomplished through up-regulation of apoptotic enzyme (caspase) activities in the mammary gland or mammary tumors. Treatment with TAM, I3C or TAM+I3C had no effect on caspase-3&7, caspase-6, caspase-8 and caspase-9 activities in the mammary tumors or mammary gland of tumor-bearing rats or that of uninitiated rats. In the mammary gland of DMBA-initiated tumor-free rats, however, I3C treatment increased the levels of caspase-3&7 and caspase-9 activities, suggesting an I3C-mediated protective effect. Even though I3C alone is a much less effective suppressing agent of mammary carcinogenesis than TAM, I3C in combination with TAM does not weaken but may foster the benefits of chemoprevention with TAM.     

Caffeine inhibits development of benign mammary gland tumors in carcinogen-treated female Sprague-Dawley rats

Summary The purpose of this study was to assess the influence of caffeine on the incidence of benign mammary tumors in carcinogen (DMBA) treated female Sprague-Dawley rats. Four different animal models were used in these studies, i.e., the administration of DMBA to: [1] 55 day old virgin rats; [2] 53 day old ovariectomized, estrogen treated virgin rats; [3] 135 day old virgin rats and [4] 135 day old parous rats. A high incidence of benign mammary fibroadenomas was observed in each of the four animal models. In addition, in the estrogen treated ovariectomized animals, a high incidence of secretory mammary gland cysts was observed. Caffeine (500 mg/L drinking water) was administered daily throughout the study commencing 3–31 days after carcinogen treatment. Caffeine treatment significantly (P<0.05 to P<0.001) reduced the incidence of benign mammary fibroadenomas in the 55 day old virgin rat model (P<0.01), in the 53 day old estrogen treated ovariectomized virgin rat model (P<0.05 to P<0.001) and in the 135 day old virgin rat model (P<0.05). The number of benign mammary fibroadenomas was reduced by caffeine in the 135 day old parous rat model but this reduction was not significant (P<0.10). In addition, in the estrogen treated ovariectomized virgin rat model, caffeine significantly (P<0.05 to P<0.001) reduced the incidence of mammary gland cysts. Caffeine treatment either increased or had no significant effect on body weight gains, depending upon the animal model. Thus, caffeine consumption can influence the development of benign mammary tumors (fibroadenomas and cysts) in carcinogen treated female Sprague-Dawley rats, an influence that was shown to be consistently inhibitory.     

Dietary effects of conjugated docosahexaenoic acid on N-methyl-N-nitrosourea-induced mammary carcinogenesis in female Sprague-Dawley rats

The dietary effects of conjugated docosahexaenoic acid (CDHA) were examined in an N-methyl-N-nitrosourea (MNU)-induced rat mammary carcinogenesis model. Female Sprague-Dawley rats were administered 50 mg/kg MNU intraperitoneally at 49 days of age. A powdered AIN-76A diet containing 0, 0.2 or 1.0% CDHA was fed to the rats from 21 to 49 days of age (before MNU; pre-initiation phase) or from 49 days to 40 weeks of age (after MNU; post-initiation phase). Rats were sacrificed when their largest mammary tumor was > or =1 cm in size or when they reached 40 weeks of age. All histologically detected mammary carcinomas were evaluated. In rats that received CDHA after MNU, development of mammary carcinoma > or =1 cm was inhibited, and there was a significant decrease in the final mammary cancer incidence and multiplicity, compared with rats that did not receive CDHA. Consumption of the 0.2% CDHA diet after MNU significantly prolonged latency. Suppression of mammary cancer yield by consumption of a CDHA diet after MNU administration was not dose-dependent. In rats that received CDHA before MNU, suppression of mammary cancer was not observed. These results indicate that CDHA administration in the post-initiation period suppressed mammary carcinogenesis, whereas CDHA administration in the pre-initiation period was ineffective.     

Immune responses of adenovirus type-9 infected (tumor-free) male rats against adenovirus type-9 induced mammary fibroadenomas and a chemically induced mammary carcinoma

Lymphocytes from five non-tumor-bearing male W/Fu rats infected with human adenovirus type-9 as newborns or as adults share a common reactivity against rat mammary fibroadenoma target cells and rat mammary carcinoma target cells, as demonstrated by microcytotoxicity tests. Mammary fibroadenomas were induced by adenovirus type-9 infection of newborn female W/Fu rats, littermates of three of the male lymphocyte donors. The mammary carcinoma target cells were derived from a single rat mammary carcinoma induced by 3,2′-dimethyl-4-amino-biphenyl (DMABP). Lymphocytes from the male adenovirus type-9 infected rats were not cytotoxic to normal rat breast epithelial cells, nor to target cells explanted from a polyoma-virus-induced sarcoma. Sera from each of five mammary fibroadenoma-bearing female rats inhibited the cytotoxic effect of lymphocytes from the adenovirus type-9 infected males against both mammary fibroadenoma and carcinoma target cells. Sera from the female W/Fu rat bearing the DMABP induced mammary carcinoma also inhibited the male lymphocyte cytotoxicity against both fibroadenoma and carcinoma target cells. Serum from rats bearing a polyoma-virus-induced sarcoma, though actively blocking in its own system, had no effect on lymphocytes from the adenovirus type-9 infected males. Anti-viral antibody response was demonstrated 8 days following adenovirus type-9 infection of adult males. No anti-tumor humoral response was demonstrated in either newborn-infected or adult adenovirus type-9 infected W/Fu male rats. Sera were checked for cytotoxicity against fibroadenoma and mammary carcinoma target cells in the presence of active homologous complement. No complement-dependent cytotoxicity was evident. Sera from infected males were admixed to known blocking sera from mammary fibroadenoma- and carcinoma-bearing rats, with no significant abrogation of the blocking effect. These results indicate that, despite the lack of tumor development, infection of male W/Fu rats by adenovirus type-9 has produced an antigen which is at least partially common to the antigen (s) shared by adenovirus type-9 induced mammary fibroadenomas and a DMABP induced mammary carcinoma.     

Suppressive effect of sesamin against 7,12-dimethylbenz[a]-anthracene induced rat mammary carcinogenesis.

The effects of dietary supplementation of sesamin on 7,12-dimethylbenz[a]anthracene (DMBA)-induced mammary carcinogenesis in female Sprague-Dawley rats were studied. Experimental diets containing 0.2% sesamin (an equiweight mixture of sesamin and episesamin) or 0.2% alpha-tocopheryl acetate were given to rats starting 1 week before intragastric administration of DMBA (10 mg/rat). Sesamin significantly (p less than 0.05) reduced the cumulative number of palpable mammary cancers by 36% at 12 weeks post-DMBA administration compared with animals on a control diet. Alpha-tocopheryl acetate inhibited both the incidence and the cumulative number of mammary tumors by 20% and 45%, respectively. Concentrations of lipid peroxides in plasma, liver and tumors were all decreased in both sesamin and alpha-tocopheryl acetate groups. The activity of peripheral blood mononuclear cells (PBMC) increased in rats fed sesamin (140 to 150% of the control and alpha-tocopheryl acetate groups). Fatty acid compositions of plasma, liver and tumor phosphatidylcholine showed a decreased tendency of the metabolism of linoleic acid to arachidonic acid and hence of the plasma concentration of prostaglandin E2 in the sesamin group. The inhibitory effect of sesamin on DMBA-induced mammary carcinogenesis may be ascribed, at least in part, to immunopotentiation and increased antioxidative activity.     

Preventive effects of antioestrogen on mammary and pituitary tumorigenesis in rats

Six groups of inbred male Wistar/Furth (WF) rats were castrated at 40 days of age and group I received no further treatment. Groups 3 and 5 received 5.0 mg diethylstilboestrol (DES) pellets. Groups 4 and 6 were given both DES and 5.0 mg anti-oestrogen (antiE) clomiphene citrate pellets. At 50-55 days of age groups 2, 5, and 6 were exposed daily to drinking water containing 5.0 mg N-nitrosobutylurea (NBU), for 30 days. None of the castrated rats given NBU alone developed mammary or pituitary tumours (MT, PT). When antiE was administered, both MT and PT incidences were reduced in rats given DES alone or in combination with NBU. Furthermore, in antiE-treated rats receiving DES and NBU the mean number of MT per rat was also significantly decreased. Similarly a marked reduction in the mean pituitary weight was observed in antiE-treated groups. These results indicate that antiE treatment was effective in the prevention of both mammary and pituitary tumorigenesis in rats receiving DES alone or receiving a combination of DES and NBU, and its inhibitory effect on mammary tumorigenesis may be mainly due to competitive antagonism for DES-induced pituitary tumorigenesis by antiE.     

Effect of wheat bran fiber on the development of mammary tumors in female intact and ovariectomized rats treated with 7,12-dimethylbenz(a)anthracene and in mice with spontaneously developing mammary tumors

We examined the effect of consumption of graded increases of dietary fiber (soft white wheat bran) on the development of mammary gland carcinomas in intact female Sprague-Dawley rats during the promotion stage of carcinogenesis, induced with 7,12-dimethylbenz(a)anthracene (DMBA). The percent of rats with mammary carcinomas, the total number of mammary carcinomas and the mean number of mammary carcinomas per rat were reduced significantly at all fiber levels examined compared to rats fed a control diet. Inclusion of 9.6% fiber in the diets of ovariectomized rats that had been treated with a single i.v. dose of 2.5 mg DMBA/100 g body weight 2 weeks prior to removal of the ovaries resulted in a significant decrease of carcinomatous and benign mammary tumors compared to ovariectomized rats fed a control diet. Development of spontaneous mammary carcinomas in virgin C₃H/HeOuJ female mice and growth of a transplantable mammary gland tumor in such mice were reduced by inclusion of 9.6% fiber in the diet, a reduction that was significant or just barely missed significance, depending on the source of the fiber. Our observations provide evidence that inclusion of soft white wheat bran in the diet is effective in the suppression of mammary gland tumorigenesis in an array of experimental animal models. Int. J. Cancer 75:439–443, 1998. © 1998 Wiley-Liss, Inc.