Inheritance of a genetic factor from the Copenhagen rat and the suppression of chemically induced mammary adenocarcinogenesis

Female rats of ten different inbred strains were tested for their susceptibility to 7, 12-dimethylbenz[a]anthracene (DMBA)-induced mammary adenocarcinogenesis. Five of these strains (OM, NSD, WF, LEWIS, BUFF) were demonstrated to be highly susceptible to DMBA exposure developing greater than 2 continuously growing, macroscopically detectable mammary adenocarcinomas/rat following gastric intubation with DMBA. In contrast, the COP female rat is completely resistant. Cross-breeding COP to any of the highly susceptible strains produced F1 hybrids which are resistant to DMBA-induced mammary adenocarcinogenesis. Cross-breeding the same highly susceptible strains to the Fischer strain (i.e., a strain only intermediately susceptible to DMBA) produced F1 hybrids which were just as highly inducible as their highly susceptible non-Fischer parental strain. The resistance of the F1 hybrids produced by cross-breeding these highly susceptible strains to the COP strain therefore is not due to a recessive lack of susceptibility but to an active process of suppression of the high susceptibility of these F1 hybrids to DMBA-induced mammary adenocarcinogenesis. To determine if the site of action of the COP gene responsible for this suppression is in the mammary gland itself or at the host systemic level, a series of factors (i.e., host growth rate, mammary gland growth rate, systemic hormone level during the estrus cycle, serum and tissue levels of DMBA and its metabolites) were compared between female rats of the highly susceptible NSD versus the resistant COP strain. These results suggested that host systemic factors are not involved. To test this directly, donor mammary glands from the highly susceptible NSD and the resistant COP strains were transplanted into F1 hybrids produced by cross-breeding these two strains. The resultant donor glands were then directly exposed to DMBA, the animals were followed, and the incidence of mammary adenocarcinomas was determined. Seventy % of the NSD donor glands developed continuously growing cancers while only 10% of COP donor glands did the same. These results are clearly incompatible with host systemic factors being the major determinant in the resistance of NSD X COP F1 hybrids to DMBA-induced mammary adenocarcinogenesis. Instead, these results directly demonstrate that it is the genetic makeup of the donor mammary gland itself which determines its response to DMBA exposure.     

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.     

Spontaneously metastasizing variants derived from MNU-induced rat mammary tumour.

N-methyl-N-nitrosourea (MNU) given i.v. to female rats of inbred strains induces mammary adenocarcinomas which are hormone-sensitive but do not spontaneously metastasize (Williams et al., 1981). Tissue culture and selection techniques have been used to derive metastasizing tumours from a mammary tumour induced with MNU in F344/N rats. Histologically, primary tumours and metastases in the lung and lymph nodes were similar. These systems may constitute useful models for the study of breast-cancer metastasis.     

Species and rat strain variation in pancreatic nodule induction by azaserine.

Subtoxic doses of azaserine induced atypical acinar cell nodules (AACN) in the pancreases of outbred Wistar rats, inbred W/LEW and F344 rats, and outbred Charles River CD-1 albino mice 4-6 months after initiation of treatment in the growing animal. These AACN apparently represented preneoplastic lesions, some of which have the potential to develop into adenomas or adenocarcinomas. Wistar and W/LEW rats were highly responsive to nodule induction; AACN developed in about 90% of the outbred Wistar rats and in all of the W/LEW rats tested. F344 rats were less susceptible and developed about 10% as many AACN as the Wistar rats. Female rats developed approximately half as many AACN as males. The mouse was intermediate in response between the F344 and the two Wistar rats. Syrian golden hamsters and strain 13 guinea pigs were relatively unresponsive. These studies of azaserine-induced AACN provided a basis for selection of carcinogenic azaserine regimens and suggested that the young male W/LEW rat was the most sensitive of the animals studied.     

Comparison of 7,12-dimethylbenz[a]anthracene metabolism and DNA binding in mammary epithelial cells from three rat strains with differing susceptibilities to mammary carcinogenesis

The capacity of polycyclic aromatic hydrocarbons such as 7,12-dimethylbenz[a]anthracene(DMBA) to induce mammary carcinomas has been studied in threerat strains. Wistar/Furth (WF) rats are highly susceptible toDMBA-induced mammary carcinogenesis, Copenhagen (Cop) rats arecompletely resistant, and Fischer 344 (F344) rats have an intermediatesusceptibility. We have previously shown that WF rats possess‘enhancer genes’, which enhance susceptibility toinduced mammary cancer. Cop rats, however, possess a single‘suppressor’ gene which confers complete resistanceto mammary cancer. Both gene types are apparently absent inF344 rats. In order to determine possible mechanisms of actionof these enhancer and suppressor genes, we have examined DMBAmetabolism and DNA binding in mammary epithelial cells isolatedfrom each rat strain. Quantitative analyses of both metabolismand DNA binding indicate no significant differences among thestrains. In addition, HPLC analyses of DMBA metabolites andDMBA-DNA adducts were essentially identical. These data suggestthat the genes controlling susceptibility and resistance tomammary carcinogenesis in these rat strains are likely to beactive at later stages of the carcinogenic process.     

Power frequency magnetic fields increase cell proliferation in the mammary gland of female Fischer 344 rats but not various other rat strains or substrains

Epidemiological data have raised concerns about the relationship between exposure to power frequency magnetic fields (MFs) and breast cancer. We have shown previously that 50-Hz MFs at microtesla flux densities enhance mammary gland tumor development and growth in the 7,12-dimethylbenz[a]anthracene (DMBA) model of breast cancer in female Sprague-Dawley (SD) rats. We also demonstrated that MF exposure results in an enhanced proliferative activity of the mammary epithelium of SD rats, which is a likely explanation for the cocarcinogenic or tumor-promoting effects of MF exposure in the DMBA model. Comparison of different SD substrains indicated that the genetic background plays a pivotal role in these effects of MF exposure. This prompted us to compare the effects of MF exposure (100 microT, 2 weeks) on cell proliferation in the mammary gland in eight different strains and substrains of outbred and inbred rats. Proliferation of epithelial cells in the mammary tissue and adjacent skin was examined by labeling proliferating cells with bromodeoxyuridine (BrdU). In addition to the MF-sensitive SD substrain (SD1) previously used in our experiments, Fischer 344 rats were the only strain in which MF exposure significantly enhanced BrdU labeling in the mammary epithelium, indicating a marked increase in cell proliferation. The MF-induced increase in BrdU labeling in Fischer 344 rats was similar to that seen after DMBA application. Furthermore, whole mount analysis of mammary tissue from Fischer 344 rats demonstrated that MF exposure increased the number of terminal end buds, i.e. the site of origin of mammary carcinomas. By comparison with MF-insensitive inbred rat strains, Fischer 344 rats may serve to evaluate the genetic factors underlying sensitivity to cocarcinogenic or tumor-promoting effects of MF exposure.     

Exposure of Fischer 344 rats to a weak power frequency magnetic field facilitates mammary tumorigenesis in the DMBA model of breast cancer

The possibility that long-term exposure to relatively weak power frequency magnetic fields (MFs) emanating from the generation, transmission and use of electricity could increase the risk of breast cancer is a matter of ongoing debate. Laboratory studies using well-defined exposure conditions are useful to examine whether exposure to MF affects mammary tumorigenesis. Previous studies from different laboratories using the 7,12-dimethylbenz[a]anthracene (DMBA) model of breast cancer in female Sprague-Dawley (SD) rats have been inconclusive, which has been related to differences in MF sensitivity between SD substrains used in these studies. When we compared the effects of MF exposure on cell proliferation in the mammary gland of various outbred and inbred rat strains, Fischer 344 was the only inbred strain that exhibited a marked increase in cell proliferation. Based on these data, we suggested that MF exposure should significantly facilitate development and growth of mammary tumors in Fischer 344 rats, which was tested in the present study. Groups of 108 DMBA-treated rats were either MF exposed (100 muT, 50 Hz) or sham exposed for 26 weeks. MF exposure significantly facilitated mammary tumorigenesis. The incidence of rats with grossly recorded, histologically verified adenocarcinomas was increased by 45% (P = 0.0095). The most pronounced MF effect on tumor incidence was seen in the cranial inguinal complexes (L/R5). These data indicate that Fischer 344 rats are a suitable inbred strain to study the mechanisms underlying the effects of MF exposure on mammary tumorigenesis.     

Ability of partial hepatectomy to induce gamma-glutamyltranspeptidase in regenerated and transplanted hepatocytes of Fischer 344 and Wistar-Furth rats

The purpose of this study was to investigate the effect of a two-thirds partial hepatectomy on the expression of gamma-glutamyltranspeptidase (GGT) in parenchymal hepatocytes. Two to 3-month-old Fischer 344 (F344), Wistar-Furth (WF), and WF X F344 F1 rats of both sexes were used in this investigation. Partial hepatectomy in the F344 female rat significantly increased the percent of liver area positive for GGT from 1.3 to 20.4%, a 15-fold increase. In contrast, GGT was not induced by partial hepatectomy in either the female WF or WF X F344 F1 hybrid rat or in the male animals of these three strains of rats. This phenomenon, which was only observed in the F344 female rat, was blocked by oophorectomy 2 weeks prior to partial hepatectomy, indicating that the induction of GGT is in part dependent upon female sex hormones. By transplanting both F344 and WF hepatocytes into the axillary fat pads of isogeneic and WF X F344 F1 rats, we showed that the observed strain difference did not result from a variation in the animal hormonal environment but rather was due to the hepatocyte from the F344 rat being intrinsically more susceptible to GGT induction than that from the WF rat. Further, the extent of the enzyme induction was significantly greater when the F344 liver cells were transplanted into female recipient animals. Since hepatectomy is often used in hepatocarcinogenesis studies, it is important to note that in the F344 female animal, the surgical procedure itself increases hepatic GGT levels in both transplanted hepatocytes and the liver in situ. Thus, the F344 female rat should be used with caution in investigations where the carcinogenic potential of chemicals is based upon the formation of GGT-positive hyperplastic nodules in the liver.     

Inheritance and site of expression of genes controlling susceptibility to mammary cancer in an inbred rat model

An inbred rat model for genetically controlled susceptibility to chemically induced mammary cancer has been established. Wistar-Furth (W/Fu) rats were found to be more susceptible to 7,12-dimethylbenz(a)anthracene-induced mammary tumors than were Fischer (F344) rats. The susceptibilities of various F1, F2, and backcross generations of these strains were examined for susceptibility to 7,12-dimethylbenz(a)anthracene-induced mammary tumors. The data suggest that susceptibility is inherited as a dominant trait. Both a single locus autosomal model and an X-linked model have been ruled out. However, the data support the hypothesis that complete susceptibility is controlled by any one of a group of independently segregating genes; i.e., any one gene of this group is both necessary and sufficient to induce maximal susceptibility. It is not known if these genes are identical or different. In order to identify the role of these genes we asked if they were expressed in the mammary epithelial cells themselves or elsewhere in the rat. Chimeric animals were produced by transplanting mammary cells from either W/Fu or F344 rats into the white interscapular fat pad of female W/Fu X F344 F1 rats. One month after transplantation the animals were treated with 7,12-dimethylbenz(a)anthracene and then palpated weekly for tumor development at the graft site. Tumors developed more rapidly and in greater total frequency at sites grafted with W/Fu mammary cells. This result suggests that the genes controlling inherited susceptibility are expressed in the mammary cells. The role of these genes is now under investigation. We have thus far shown that they do not control carcinogen metabolism or activation.     

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.     

N-methyl-N-nitrosourea-induced rat mammary tumors. Hormone responsiveness but lack of spontaneous metastasis

N-Methyl-N-nitrosourea (MNU) given iv to rats 50-55 days old induced mammary tumors in 70% of F344/N and 91% W/ICRF inbred females with mean latency periods of 149 and 93 days, respectively. Reduction of the MNU dose did not affect tumor incidence in W/ICRF rats. Of the mammary tumors, 98% were classified histologically as adenocarcinomas, which grew progressively. Primary tumors of nonmammary origin were detected at low incidence. Upon histologic examination, no evidence was found for metastases of either the mammary or other primary tumors. No evidence for tumor-induced hypercalcemia was found. Oophorectomy at the time of MNU administration prevented tumor development; oophorectomy when at least 1 tumor/animal was palpable caused growth delay or regression. All MNU-induced and 7,12-dimethylbenz[a]anthracene-induced mammary tumors tested contained cytoplasmic estrogen receptor (ER) at similar concentrations and were indistinguishable histologically. MNU-induced tumors in F344 rats were transplantable and retained ER through three transplantations.     

Genetic bases of estrogen-induced tumorigenesis in the rat: mapping of loci controlling susceptibility to mammary cancer in a Brown Norway x ACI intercross

Exposure to estrogens is associated with an increased risk of breast cancer. Our laboratory has shown that the ACI rat is uniquely susceptible to 17beta-estradiol (E2)-induced mammary cancer. We previously mapped two loci, Emca1 and Emca2 (estrogen-induced mammary cancer), that act independently to determine susceptibility to E2-induced mammary cancer in crosses between the susceptible ACI rat strain and the genetically related, but resistant, Copenhagen (COP) rat strain. In this study, we evaluate susceptibility to E2-induced mammary cancer in a cross between the ACI strain and the unrelated Brown Norway (BN) rat strain. Whereas nearly 100% of the ACI rats developed mammary cancer when treated continuously with E2, BN rats did not develop palpable mammary cancer during the 196-day course of E2 treatment. Susceptibility to E2-induced mammary cancer segregated as a dominant or incompletely dominant trait in a cross between BN females and ACI males. In a population of 251 female (BN x ACI)F(2) rats, we observed evidence for a total of five genetic determinants of susceptibility. Two loci, Emca4 and Emca5, were identified when mammary cancer status at sacrifice was evaluated as the phenotype, and three additional loci, Emca6, Emca7, and Emca8, were identified when mammary cancer number was evaluated as the phenotype. A total of three genetic interactions were identified. These data indicate that susceptibility to E2-induced mammary cancer in the BN x ACI cross behaves as a complex trait controlled by at least five loci and multiple gene-gene interactions.     

Congenic rats reveal three independent Copenhagen alleles within the Mcs1 quantitative trait locus that confer resistance to mammary cancer

It has previously been shown that the Copenhagen (COP) rat contains several genetic loci that contribute to its mammary tumor-resistant phenotype after 7,12-dimethylbenz(a)anthracene (DMBA) administration. One of these loci, mammary carcinoma susceptibility 1 (Mcs1), is located on the centromeric end of chromosome 2 and appears to act in a semidominant fashion. To confirm the existence and independent action of this locus and also aid in the identification of the physical location of the Mcs1 gene, congenic lines were generated by transferring the Mcs1 COP allele onto a Wistar Furth (WF) genetic background. Male carriers were genotyped using microsatellite markers spanning 20-30 cM of the Mcs1 locus. One of the congenic lines minimally retained the COP allele at D2Mit29 on the centromeric end of chromosome 2 and extended distally to D2Rat201. Heterozygous Mcs1 carrier rats were interbred, and the female offspring were treated with DMBA. The female rats from the Mcs1 congenic line that carried one or two COP alleles of the Mcs1 region had a significantly reduced (65 and 85%, respectively) tumor development (P < 0.001) compared with rats carrying zero COP alleles at this locus. A WF.COP-D2Mit29/D2Rat201 homozygous congenic strain derived at the N10 generation was treated with DMBA, and the COP homozygous rats developed 1.5 +/- 0.3 carcinomas/rat versus 6.3 +/- 0.5 in WF control rats (P < 0.0001). Fine mapping of this congenic interval using several recombinant lines identified three genetic loci within the Mcs1 congenic region that independently supported a tumor resistance phenotype. These genetic loci have been termed Mcs1a, Mcs1b, and Mcs1c. In rats for which each locus was homozygous for the COP allele, tumor development was reduced by approximately 60% compared with littermate controls. The identification of these independent loci within the Mcs1 COP allele provide a model of the genetic complexity of cancer.     

Resistance of Copenhagen rats to chemical induction of glutathione S- transferase 7-7-positive liver foci

Copenhagen (Cop) rats are completely resistant to the chemical induction of mammary adenocarcinomas, but their susceptibility to hepatocarcinogenesis is virtually unknown. Rat liver is a well- characterized and easily manipulated tissue in which to study carcinogenesis. Therefore, if Cop rats are resistant to hepatocarcinogenesis, studies into resistance mechanisms may be feasible. Male Cop and F344 rats, 7-8 weeks old, were initiated using either N-nitrosodiethylamine (DEN) (200 mg/kg, i.p.) or a two-thirds partial hepatectomy (PH) followed by N-methyl-N-nitrosourea (MNU) (60 mg/kg, i.p.). The rats were then promoted using a modified resistant hepatocyte (RH) protocol (a combination of four doses of 2- acetylaminofluorene (2-AAF) and a single dose of CCl4 that provides a selective mitotic stimulus for initiated cells). Six weeks after initiation the rats were killed and liver sections were stained for glutathione S-transferase 7-7 (GST 7-7), a marker for putative preneoplastic hepatocytes. Cop rats were found to be highly resistant, having a approximately 9- and approximately 27-fold smaller percentage of liver area occupied by GST 7-7-positive foci than susceptible F344 rats following initiation by DEN and MNU respectively. Furthermore, gross liver nodules did not form in any of the Cop rats, whereas all F344 rat livers contained nodules. Hepatic necrosis caused by DEN during initiation, and CCl4 during promotion is necessary to stimulate compensatory hepatocyte division. We demonstrated that these agents do indeed increase serum transaminase levels and produce histologic evidence of necrosis in Cop rats. In order for liver foci to grow rapidly in the RH protocol, the surrounding normal hepatocytes must be mito-inhibited by 2-AAF. We found that the degree of mito-inhibition of normal hepatocytes by 2-AAF is the same in Cop and F344 rats. These results show that the Cop rat is highly resistant to the chemical induction of putative preneoplastic liver foci and nodules.      

Differential susceptibility to chemically induced thymic lymphomas in SENCARB and SSIN inbred mice

In the past 20 yr, several inbred strains have been derived from SENCAR outbred mice. These strains display different susceptibility to the induction of papillomas and progression to squamous cell carcinomas (SCC) in the skin after chemical carcinogenesis. In the present study, we showed that one of these strains SENCARB/Pt was highly susceptible to the development of N-methyl-N-nitrosourea (MNU)- and 7,12-dimethylbenz[a]anthracene (DMBA)-induced lymphomas. In contrast, the SSIN/Sprd inbred strain is completely resistant to T-cell lymphomagenesis by both carcinogens. Within 175 d after a single injection of 75 mg/kilogram body weight (kbw) of MNU, SENCARB/Pt mice exhibited a 91.6% incidence of lymphoma. In addition, during an independent tumorigenesis study with repeated doses of intragastric DMBA, SENCARB/Pt mice showed an incidence of 75% lymphoma development 300 d after the last treatment. Histopathological and flow cytometric parameters indicated that the lymphomas were of the T-cell lineage. In order to study the genetics of MNU-induced tumorigenesis, we generated F1 hybrid mice between SSIN/Sprd and SENCARB/Pt mice. Tumor incidence in MNU-injected F1 mice suggested that the high tumor incidence is a dominant trait. Loss of heterozygosity (LOH) analysis in these tumor samples revealed allelic imbalances on chromosomes 15 and 19. Given that these inbred strains are closely related, it is likely that a relatively small number of loci are responsible for the observed differences in susceptibility. Therefore, these SENCAR inbred strains constitute important new tools to study the genetic basis of resistance and susceptibility to chemically induced thymic lymphoma formation.     

Effect of retinyl acetate and 4-hydroxyphenylretinamide on initiation of chemically-induced mammary tumors

The anti-promotional effect of retinoids on chemically-induced mammary carcinogenesis in the rat is well established. The present studies were performed to determine the effect of long-term feeding of retinyl acetate and 4-hydroxyphenylretinamide (4-HPR) on initiation of mammary tumors induced by MNU or DMBA. Retinyl acetate (328 mg/kg of diet) or 4-HPR (782 mg/kg of diet) was added to the diet of female Sprague-Dawley rats for two months prior to the administration of the carcinogens. In the MNU model, a 50% increase in the number of mammary adenocarcinomas was observed in rats pretreated with retinyl acetate, while pretreatment with 4-HPR resulted in a 93% increase in the number of cancers. Continued treatment with 4-HPR throughout the study, however, caused a reduction in cancer number. In the DMBA mode, pretreatment with these retinoids significantly increased the number of benign mammary tumors, but not mammary cancers. These data suggest that newly synthesized retinoids should be evaluated for chemopreventive activity against mammary cancer initiation as well as for their anti promotional activity.     

ACCELERATED PAPER: Cloning, genetic mapping and expression studies of the rat Brca1 gene

The breast cancer gene BRCA1 has previously been cloned fromboth human and mouse. We cloned a fragment of the rat Brca1homologue in order to map it and explore its biological function.Partial cDNA fragments of the rat Brca1 homologue were isolatedby RT-PCR. Sequence analysis revealed that the RING-finger domainis well conserved among rat, mouse and human. Rat Brcal mRNAwas expressed in most tissues studied with the highest levelin testis, consistent with studies in human and mouse. Next,intron 6-containing DNA fragments were amplified by PCR fromWKY and WF strains. The splicing sites between exon 6 and exon7 are conserved between rat and human. Partial sequencing ofthe rat Brca1 intron 6 revealed a polymorphism of a pentanucleotideTTTTG repeat between the WKY and WF strains. With this intragenicmicrosatellite marker, we were able to map precisely the ratBrcal gene to chromosome 10 using a genetic linkage study of(WKYxWF)F1xWF baccross rats. Brca 1 cosegregates with markerBAND3A, and is flanked by R5123 and R5842. Using this polymorphicmarker, we also investigated the loss of heterozygosity (LOH)of the Brcal microsatellite marker in carcinogen- or radiation-inducedmammary carcinomas in (WFxF344)F1 female rats. No LOH or somaticmicrosatellite instability was detected in 18 DMBA-induced tumorsstudied. Only one LOH of the F344 allele was observed in 26radiation-induced tumors tested. Ribon-uclease protection assaysdemonstrated that Brca1 mRNA levels are similar in normal ratmammary glands and mammary carcinomas of various etiologies,including those induced by DMBA, NMU, activated-neu and activated-rasoncogenes.     

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)