Effect of D,L-2-difluoromethylornithine and endocrine manipulation on the induction of mammary carcinogenesis by 1-methyl-1-nitrosourea

The purpose of this investigation was to establish an efficientroute and dose regime for the long-term administration of tamoxifenin the study of mammary tumorigenesis in the rat. The secondobjective of this work was to determine whether treatment withD, L-2-difluoromethylornithine (DFMO), a synthetic inhibitorof the enzyme ornithine decarboxylase, would reduce the occurrenceof mammary cancers in tamoxi-fen-treated or ovariectomized rats.A total of 265 female Sprague-Dawley rats were assigned to oneof two experimental protocols. All animals were injected with50 mg 1-methyl-1-nitrosourea (MNU) per kg body wt at 50 daysof age. In experiment 1, beginning 7 days after the injectionof the carcinogen, animals were assigned to one of six groupswhich received either 0, 1 or 5 mg tamoxifen citrate per kgAIN-76A purified diet in addition to either no DFMO or a 0.125%w/v solution of DFMO as the drinking water. The experiment wasterminated 180 days following carcinogen treatment. Treatmentwith tamoxifen resulted in a dose-dependent reduction in cancerincidence, and the number of cancers induced and significantlyprolonged the median cancer-free time. This effect was alsoaccompanied by a decrease in the rate of body weight gain. Treatmentwith DFMO delayed latency and reduced tumor number. DFMO inaddition to tamoxifen (1 mg/kg diet) further prolonged latency.In experiment 2 each animal was assigned to one or four treatmentgroups when its first palpable mammary tumor was detected. Atthat time each was either ovariectomized or sham-operated. Inaddition, the rats were either provided no DFMO or a 0.5% w/vsolution of DFMO as the drinking water. The study was terminated35 weeks following carcinogen injection. Ovariec-tomy significantlyinhibited the occurrence of additional mammary tumors. Ovariectomyplus DFMO was more effective than ovariectomy alone in reducingtumor number. Collectively, these observations indicate thatsuppression of polyamine biosynthesis via the systemic administrationof DFMO inhibits the development of ovarian hormone insensitivemammary tumors.     

Effect of D,L-alpha-difluoromethylornithine on murine mammary carcinogenesis

The development of chemically-induced mammary gland carcinomasin rats was dramatically suppressed by provision of a 1% solutionof D,L--difluoromethylornithine (DFMO) in drinking water. Treatmentwith DFMO significantly reduced cancer incidence and the averagesize and number of cancers per rat and prolonged the cancer-freetime. DFMO appears to be effective in blocking some aspect ofthe promotion stage of chemically induced mammary carcinogenesisin the rat.     

Effect of tamoxifen and D,L-2-difluoromethylornithine on the growth, ornithine decarboxylase activity and polyamine content of mammary carcinomas induced by 1-methyl-1-nitrosourea

The effect of combined treatment with D,L-2-difluoromethylornithine(DFMO) and tamoxifen on the growth status, ornithine decarboxylase(ODC) activity and polyamine content of established 1-methyl-l-nitrosourea(MNU)-induced mammary tumors was investigated. DFMO treatment,a 0.125% solution provided as drinking water, inhibited therate of tumor occurrence and reduced the number of mammary tumorsinduced by a high dose of MNU (50 mg/kg body weight) duringthe first 120 days post-carcinogen treatment. Tamoxifen wasadministered daily via s.c. injection (25 µ/100 g bodyweight) to tumor-bearing rats in both treatment groups, i.e.control and DFMO-treated, for a 30-day period beginning 120days after carcinogen. Tamoxifen treatment induced tumor regressionbut the percentage of regressing, static or growing tumors wasno different in the presence or absence of DFMO. Whereas themammary tumors of DFMO-treated rats had reduced ODC activityand lower polyamine concentrations in comparison to the tumorsof untreated animals, tamoxifen had no effect on these parametersindependent of its effect on tumor growth status. DFMO did notincrease the efficacy of tamoxifen in inducing tumor regression.     

Effects of D,L-2-difluoromethylornithine and indomethacin on mammary tumor promotion in rats fed high n-3 and/or n-6 fat diets

Virgin female Sprague-Dawley rats (50 days of age) were administered a single intragastric 10-mg dose of 7,12-dimethylbenz(a)anthracene (DMBA). Twenty-one days later they were placed on diets containing either 20% corn oil (CO), 15% menhaden oil plus 5% corn oil (MO + CO), 20% CO plus 0.5% w/w of the irreversible ornithine decarboxylase inhibitor, D,L-2-difluoromethylornithine (CO + DFMO), 20% CO plus 0.004% w/w of the cyclooxygenase inhibitor indomethacin (CO + INDO), 20% CO + 0.004% INDO + 0.5% DFMO (CO + INDO + DFMO), or 15% MO + 5% CO + 0.5% DFMO (MO + CO + DFMO). The incidence of DMBA-induced mammary tumors was significantly reduced in rats fed diets containing DFMO but not in rats fed the diet containing indomethacin. Incidences of mammary tumors at 16 weeks post-DMBA were 86% in rats fed the CO diet, 83% in rats ingesting the diet containing CO + INDO, 28% in rats fed CO + DFMO, 32% in rats fed diet containing CO + INDO + DFMO, 59% in rats fed the MO + CO diet, and 24% in rats fed the MO + CO + DFMO diet. The average number of tumors and tumor burden per tumor-bearing rat were reduced and tumor latency was increased in all rats fed diets containing DFMO. Body weight gain, but not food intake, of rats fed the 20% fat + 0.5% DFMO diets was significantly less than in rats fed the 20% fat diets. Prostaglandin E and leukotriene (LTB4) syntheses, ODC activity and mammary tumorigenesis were significantly inhibited by feeding the diet containing menhaden oil or by adding 0.5% DFMO to any of the high fat diets. Feeding a 20% CO diet containing 0.004% INDO significantly reduced prostaglandin synthesis and ODC activity and increased LTB4 synthesis of mammary tumors but did not inhibit mammary tumorigenesis. This study suggests that the 5-lipoxygenase product LTB4 may be involved in mammary tumor production. Whereas a decrease in LTB4 appears to be associated with a decrease in tumorigenesis, an increase (as seen in the indomethacin group) was not associated with any change in the tumorigenic response.     

Effect of canthaxanthin on chemically induced mammary carcinogenesis

Canthaxanthin, a carotenoid with no vitamin A activity, was evaluated for its efficacy in the prevention of chemically induced mammary cancers. Canthaxanthin was administered in the diet at two dose levels (3,390 or 1,130 mg/kg diet). In the dimethylbenzanthracene-induced mammary cancer model, diet supplementation with canthaxanthin for 3 weeks prior to the carcinogen resulted in a 65% reduction in the number of mammary cancers. The feeding of canthaxanthin after the administration of methylnitrosourea had no significant effect on mammary carcinogenesis. These data demonstrate that canthaxanthin, at least in these models of mammary cancer, is active in preventing cancer initiation and not promotion. Analysis of tissues by high-pressure liquid chromatography revealed that canthaxanthin levels in the liver are very high when compared to those in the mammary gland. The observation that canthaxanthin is highly effective in preventing cancer initiation without toxicity suggests that carotenoids not possessing vitamin A activity should be further evaluated as chemopreventive agents.     

Effect of exercise on the induction of mammary carcinogenesis

Although data reported in several epidemiological investigations indicate that reduced consumption of dietary fat and increased levels of physical activity are associated with reduced risk for breast cancer, the results of some studies do not support these observations. Underlying this situation is the unanswered question about whether degree of body fatness, which is affected by dietary composition, total caloric intake, and energy expenditure, is the critical determinant affecting breast cancer risk. The objective of this work was to establish whether increasing energy expenditure by exercise would reduce the occurrence of mammary carcinomas induced by 7,12-dimethylbenz[a]anthracene (DMBA) in animals consuming a high fat diet to the level of occurrence observed in sedentary animals consuming a low fat diet. Female Sprague-Dawley rats were obtained at 21 days of age and maintained on a 5% (w/w) corn oil diet (AIN-76A) until they were 64 days of age. At 50 days of age, rats received either 5 mg DMBA or the solvent in which the carcinogen was dissolved. Fourteen days after DMBA intubation they were randomized into one of three groups: 5% fat (w/w), sedentary; 24.6% fat (w/w), sedentary; or 24.6% fat (w/w), exercised. Animals were exercised on a motor-driven treadmill at a belt speed of 20 m/min and a 1-degree incline for 15 min/day, 5 days/week for 18 weeks. Feeding a high fat versus a low fat diet increased the number of breast cancers induced and the rate at which they appeared in agreement with previous investigations. However, rather than retarding the development of tumors as was hypothesized, moderate treadmill exercise increased the incidence and number of cancers induced and shortened cancer latency in comparison to animals that received either the high fat or low fat diet and were sedentary. Body composition was not altered by the exercise regime imposed, although these animals weighed more than either sedentary group. These data document a heretofore unreported effect of a moderate level of aerobic work on breast cancer induction.     

Identification of murine mammary stem cells: implications for studies of mammary development and carcinogenesis

The epithelial components of the mammary gland are thought to arise from a stem cell capable of both self-renewal and multi-lineage differentiation. Furthermore, there is increasing evidence that mammary carcinomas originate in these cells or their immediate progeny. The recent identification of murine mammary stem cells should facilitate their molecular characterization and help to elucidate their role in mammary carcinogenesis. In addition, an understanding of the biology of these cells including the pathways that regulate their self-renewal and differentiation may suggest new approaches for the prevention and treatment of breast cancer.     

Stage-specific inhibition of mammary carcinogenesis by 1alpha-hydroxyvitamin D5

Active metabolites of vitamin D are well recognised as cancer chemopreventive and chemotherapeutic agents. However, they are toxic at effective concentrations. Earlier, we reported that a non-toxic analogue of vitamin D, 1alpha-hydroxyvitamin D5(1alpha(OH)D5), inhibited carcinogen-induced mammary lesion formation in mouse mammary organ cultures (MMOC) and in N-methyl-N-nitrosourea (MNU)-induced rat mammary carcinogenesis. In the present study, we determined if 1alpha (OH)D5 action is selective during the initiation or promotion phases in MMOC and in vivo. In MMOC, 1 microM 1alpha (OH)D5 suppressed both ovarian hormone-dependent and -independent mammary lesions by more than 60%. Inhibition of alveolar lesions was observed only during the promotion stage (p=0.0016). In a 7,12-dimethylbenz(a)anthracene (DMBA)-induced mammary carcinogenesis experiment, 1alpha (OH)D5 (40 microg/kg diet) inhibited cancer incidence by 37.5% (p<0.05) if 1alpha (OH)D5 was present in food during the promotion phase (+1 to end). However, a D5-supplemented diet during the initiation phase (-2 to +1 week) did not provide any protection. These results clearly show, for the first time, that the effects of vitamin D may be mediated selectively during the promotion or progression phases of carcinogenesis.     

Effect of human chorionic gonadotropin on mammary gland differentiation and carcinogenesis

The observation that mammary carcinogenesis is inhibited in rats which completed a pregnancy prior to exposure to the chemical carcinogen 7,12-dimethylbenz[a]anthracene (DMBA) led us to determine whether the protective effect of pregnancy could be mimicked by treatment with the placental hormone chorionic gonadotropin (hCG). We also studied the effect of this treatment on mammary gland structure and differentiation, and determined whether hCG exerts toxic or collateral effects on body weight and endocrine organs. The systemic effect of hCG on body wt and endocrine organs and mammary gland was studied in outbred virgin Sprague-Dawley rats which at the age of 50 days started receiving 100 IU hCG i.p. daily for 21 days. The animals were subdivided into nine groups of five animals each; one group was killed on the first day of and the others at 5, 10, 15 and 21 days of injection and 5, 10, 15 and 21 days post injection. The effect of the hormonal treatment on the estrous cycle was determined by studying the vaginal smears taken during and after the injection period. The following parameters were determined: body wt, weight and morphology of pituitary gland, adrenals, ovaries and uterine horns. Mammary glands were processed for histology, autoradiography for determination of DNA labeling index (DNA-LI) and whole mount preparation for morphometric studies. The effect of hCG on mammary carcinogenesis was studied in two groups of virgin rats; group I, which at the age of 50 days started receiving a daily i.p. injection of 100 IU hCG for 21 days; 21 days after the last injection they were given 8 mg DMBA/100 g body wt. Group II animals received DMBA only. hCG treated animals gained weight as a function of age at the same rate as controls. Treatment did not modify the weight of adrenal glands. The weight of ovaries, uterus and pituitary gland were transitorily increased by the 15th day of treatment, but had returned to the same values of controls by the time of DMBA administration. Treatment stimulated mammary gland differentiation, measured as a progressive reduction in number of terminal end buds and increase in the number of alveolar buds and lobules. The DNA-LI was significantly depressed in all terminal structures in the glands of treated animals. In group I animals hCG treatment decreased incidence of adenocarcinomas to 6.15 from 43.8% in group II animals.(ABSTRACT TRUNCATED AT 400 WORDS)     

Effect of dietary energy restriction on vascular density during mammary carcinogenesis

Inhibition of mammary carcinogenesis by dietary energy restriction is associated with a decrease in cell proliferation and the induction of apoptosis. Although changes in the metabolism of insulin-like growth factor I and glucocorticoids have been proposed to modulate these cellular processes, limitations in blood supply could induce similar effects. To investigate this possibility, female Sprague Dawley rats were given an injection of 1-methyl-1-nitrosourea and fed purified diets ad libitum or at 60% ad libitum intake, i.e., 40% dietary energy restriction. Premalignant mammary pathologies and mammary adenocarcinomas obtained from these rats were processed for vascular density analysis via CD-31 immunostaining. Vascular density, measured as vessels/unit area, of premalignant mammary pathologies and adenocarcinomas from dietary energy restriction rats was reduced 31 and 39%, respectively (P < 0.01). This effect, which was observed in a 50-microm wide band of tissue surrounding each pathology, was exerted on blood vessels > 25 microm2. Conversely, intratumoral vascular density was unaffected by dietary energy restriction. cDNA microarray and Western blot analyses of adenocarcinomas for evidence of dietary energy restriction-mediated effects on vascularization revealed that only the level of vascular endothelial growth factor receptor protein Flk-1 was significantly reduced (P < 0.001). It appears that dietary energy restriction imposes limitations in the supply of blood to developing pathologies, an effect that could directly inhibit the carcinogenic process. The vascular density data imply that dietary energy restriction inhibited the growth of endothelial cells but leave unresolved the question of whether dietary energy restriction had a specific effect on angiogenesis. The factors that account for the failure of dietary energy restriction to limit intratumoral vascularization are not obvious and merit additional investigation.     

Effect of vitamin D3 on carcinogen-modified liver enzymes and tumour incidence in experimental rat mammary carcinogenesis.

The anticarcinogenic effect of vitamin D3 in relation to biochemical and morphological markers in 7,12-dimethylbenz(alpha)anthracene (DMBA)-induced mammary carcinogenesis was investigated in two different sets of experiments. For each set, female Sprague-Dawley rats were divided into four groups, to allow comparison among treated and non-treated groups. At 50 days of age, animals of group B and C were given DMBA injection (0.5 mg/100 g body weight) through the tail vein, and normal control (group A) animals received the oil emulsion vehicle alone. Vitamin D3 at the dose of 0.3 microgram/0.1 ml propylene glycol was given orally twice a week, in carcinogen as well as non-carcinogen treated animals (group C and c), until the termination of the experiments (22-24 weeks for biochemical markers, and 35 weeks for morphology). At approximately 22-24 weeks, when marked lobular hyperplasia in DMBA control groups were confirmed through histology, the biochemical markers were modulated towards normal value for vitamin D3 in the treatment group, in comparison to the disturbed values caused by carcinogen administration in group B animals. Again, vitamin D3 supplementation was effective in reducing the tumour incidence (70% in comparison to 90% in group B). The results thus clearly concluded the antineoplastic potential of vitamin D3, and the existing correlation between biological and biochemical markers.     

Effect of Epitalon and Vilon treatment on mammary carcinogenesis in transgenic erbB-2/NEU mice

Female transgenic FVB mice transfected with the mammary erbB-2/neu oncogene were injected 0.1 ml 0.9% solution of sodium chloride (control), 1 meg Vilon peptide (Lys-Glu) or Epitalon peptide (Ala-Glu-Asp-Glu), s.c., 5 days in succession once a month, beginning from the age of 2 months. The characteristics of mammary tumor induction in the control and experimental groups did not differ until the age of 9 months. Later on, Epitalon-treated mice revealed distinct inhibition of carcinogenesis. One tumor per animal was detected in 7% (control), 4% (Vilon) and 16% (Epitalon) (p < 0.05). Two or more tumors per animal were in 75%, 95% and 56%, respectively (p < 0.05). Largest diameter of mammary adenocarcinoma in the Epitalon group was smaller than in controls by 33% (p < 0.05). Although the number of mice with metastases to the lung in all three groups was practically identical, their incidence in the Vilon group was 2.6 times higher than in Epitalon-treated animals (p < 0.05). Largest diameter of metastasis in the Epitalon group was the smallest, too. Our data point to inhibition of mammary carcinogenesis by Epitalon in transgenic erbB-2/neu mice.     

Dose-response effects of the COX-2 inhibitor, celecoxib, on the chemoprevention of mammary carcinogenesis

Recent chemopreventive studies in our laboratories showed that the COX-2 inhibitor, celecoxib, inhibited the induction of mammary cancer by 7,12-dimethylbenz(a)anthracene (DMBA). In this study, we examined the relative chemopreventive effect of varying doses of celecoxib on the development and growth of DMBA-induced rat mammary tumors. At 10 days prior to receiving a single intragastric dose of 15 mg DMBA/rat, female Sprague-Dawley rats were fed a control chow diet or diets containing 250, 500, 1000 or 1500 ppm celecoxib until termination of the experiment. Administration of increasing doses of celecoxib inhibited mammary tumor incidence and multiplicity as well as tumor volume in a dose-dependent manner. At 122 days post DMBA-intubation, mammary tumor incidence was 100% in the control rats compared to 80%, 50%, 45% and 25% in rats receiving 250, 500, 1000 or 1500 ppm celecoxib, respectively (p<0.001). Similarly, tumor multiplicity and tumor volume were significantly reduced by increasing the dose of celecoxib from 250 to 1500 ppm in the diet. The control rats had an average of 3.46 tumors/rat compared to 1.80, 1.00, 0.75 and 0.50 tumors/rat in animals receiving 250, 500, 1000 or 1500 ppm celecoxib, respectively (p<0.001). Average tumor volumes in rats fed 250, 500, 1000 or 1500 ppm celecoxib were 0.42, 0.34, 0.31 and 0.16 cm3 compared to 1.29 cm3 in the control rats (p<0.001). There was a concomitant increase in the steady-state serum concentration of celecoxib with the dose. These results indicate that, in this rat model, the chemopreventive effect of celecoxib against breast cancer is dose-dependent and that celecoxib is effective even at lower dose levels.     

Effect of high-dose, fractionated local irradiation on MNU-induced carcinogenesis in the rat mammary gland

The effect on chemical carcinogenesis in the mammary gland ofhigh-dose fractionated, local irradiation, as is used in thetreatment of human breast cancer, was examined in a rat modelof this disease process. For this purpose, a highly reproduciblemethod was employed for administering a therapeutic dose andfractionation schedule via an anterior portal to a single mammarygland chain of rats in a manner that minimized whole body irradiation.This approach offers significant advantages over whole bodyirradiation techniques previously used for investigations ofradiation mediated effects on the carcinogenic process. Amongthe advantages are that higher doses of radiation can be administeredto the target tissue with minimal side effects and that thecontralateral mammary gland chain can serve as a ‘withinanimal control’. When this approach was used to studythe effect of high-dose fractionated radiation on the risk ofdevelopment of mammary cancer in rats given 1-methyl-1-nitrosoureaprior to radiation, an enhanced tumorigenic response was observedthat greatly exceeded the response resulting from either radiationor carcinogen administered alone. This result was unanticipated,based on data from animal studies of the effects of whole bodyirradiation on mammary tumor development and the outcome ofclinical series. Possible reasons for the discrepancy are presented.     

Effect of tamoxifen on preneoplastic cell proliferation in N-nitroso-N-methylurea-induced mammary carcinogenesis.

N-Nitroso-N-methylurea (NMU) is an effective carcinogen for the induction of mammary carcinoma in the rat. Tamoxifen (TAM), used as a chemopreventive agent to reduce tumor incidence, has been well studied using this model. We have utilized the rat mammary carcinoma model to assess the effect of TAM on preneoplastic changes. Fifty-day-old virgin female Sprague-Dawley rats were randomized by weight and divided into the following five groups: Group 1, normal controls (n = 24); Group 2, TAM (n = 20); Group 3, NMU-short term (n = 24); Group 4, NMU-short term + TAM (n = 26); and Group 5, NMU-long term (n = 23). Seven weeks after the exposure to NMU, rats in Groups 1, 2, 3, and 4 were given injections of [3H]thymidine and sacrificed 4 h later for autoradiographic determination of thymidine labeling index (TLI). The rats from Group 5 were observed for 30 weeks after NMU exposure to confirm mammary tumor development. TLI in both terminal ducts and terminal end buds was modulated by treatment with TAM. Carcinogen administration induced higher TLI relative to the normal controls [18.3 +/- 1.8% (SD) versus 15.5 +/- 2.1%, P less than 0.001] in terminal end buds. The effect of carcinogen on TLI was also apparent in the terminal ducts (15.8 +/- 1.1% versus 9.5 +/- 1.1%, P less than 0.001). TAM administration was able to suppress both constitutive and NMU-induced TLI increases in terminal end buds (15.5 +/- 2.1% versus 2.8 +/- 1.1% and 18.3 +/- 1.8% versus 6.8 +/- 1.4%, respectively, P less than 0.001). Similar effects were observed in terminal ducts. In addition to its antiproliferative effect on nontransformed mammary tissue, TAM was effective in suppressing NMU-induced mammary tumor incidence and frequency. NMU-induced hyperproliferation is an intermediate stage in NMU carcinogenesis in the rat and is suppressed by TAM. Mammary epithelial hyperproliferation may provide a useful quantitative intermediate end point to evaluate chemopreventive efficacy.     

Combination therapy with 2-difluoromethylornithine and a polyamine transport inhibitor against murine squamous cell carcinoma

Using a recently developed autochthonous mouse model of squamous cell carcinoma (SCC), a combination therapy targeting polyamine metabolism was evaluated. The therapy combined 2-difluoromethylornithine (DFMO), an inhibitor of ornithine decarboxylase (ODC), and MQT 1426, a polyamine transport inhibitor. In 1 trial lasting 4 weeks, combination therapy with 0.5% DFMO (orally, in the drinking water) and MQT 1426 (50 mg/kg i.p., bid) was significantly more effective than with either single agent alone when complete tumor response was the endpoint. In the combination group, 72% of SCCs responded completely vs. 21 and 0% for DFMO and MQT 1426, respectively. A second trial involved a 4-week treatment period followed by 6 weeks off-treatment. With apparent cures as an endpoint, combination therapy was again more effective than either agent alone: a 50% apparent cure rate was observed in the combination group vs. 7.7% in the DFMO group. MQT 1426 had no inhibitory effect on SCC ODC activity nor did it enhance the inhibition by DFMO, but SCC polyamine levels declined more rapidly when treated with combination therapy vs. DFMO alone. The apoptotic index in SCCs was transiently increased by combination therapy but not by DFMO alone. Thus, targeting both polyamine biosynthesis and polyamine transport from the tumor microenvironment enhances the efficacy of polyamine-based therapy in this mouse model of SCC.     

Effect of PSC 833, an inhibitor of P-glycoprotein on N-methyl-N-nitrosourea induced mammary carcinogenesis in rats

Studies in our laboratory on the role of P-glycoprotein (Pgp, coded by mdr1 gene) have led to the hypothesis that over-expression of Pgp is closely associated with the development of cancer. It was conceived therefore that inhibitors of Pgp should inhibit the development of cancer. We have reported that PSC833 (PSC), a potent inhibitor of Pgp, inhibits the development of liver cancer in rats. Similarly, based on the intrinsic over-expression of Pgp in experimental mammary carcinogenesis, we studied the effect of PSC on N-methyl-N-nitrosourea induced mammary cancer in female Sprague-Dawley rats. The study indicates that PSC at daily dietary doses of 15 (PSC15) and 30 mg/kg (PSC30) body wt resulted in dose-dependent inhibition of the incidence as well as the growth of mammary tumors. Compared with controls, PSC15 and PSC30 inhibited: (i) mean tumor multiplicity by 32 and 67%, (ii) median tumor burden by 46 and 93% and (iii) incidence of ulcerated tumors by 40 and 82%, respectively. Most remarkably, PSC delayed median tumor incidence by 8 weeks, and exerted a 100% inhibitory effect on the incidence of large tumors, 4 cm(3) and greater. In all the cases, although the inhibitory effect of PSC was evident at both doses, only PSC30 exhibited statistical significance. A possible compounding effect that was also observed in PSC30-treated rats was a decrease in body weight gain not attributed to diminished food consumption. All in all, consistent with recent reports, which have demonstrated inhibition of cancer development by compromising Pgp function, this study introduces a novel role for Pgp in breast cancer and potentially an unexplored therapeutic approach in treating the disease.