Tumor promotion by fecapentaene-12 in a rat colon carcinogenesis model

Fecapentaenes are a group of fecal mutagens produced by anaerobicmicroflora of the colon. The potential of fecapentaene-12 (FP-12)to promote tumor development was tested in a rat colon carcinogenesismodel using N-methyl-N-nitrosourea (MNU) as the initiating agent.Two groups of female F-344 rats were initiated by intrarectalinstillations of MNU (2 mg in 0.5 ml H₂O, 3 times a week, for3 weeks; MNU and MNU + FP-12 groups). Two additional groups(FP-12 and Control) were given H₂O without carcinogen. In thepost-initiation phase, rats of the MNU + FP-12 and FP-12 groupswere intrarectally administered 400 ng of FP-12 in 0.5 ml T-Ebuffer, twice a week, for 24 weeks, whereas the MNU and Controlgroups received the vehicle only. Tumors were found only inthe MNU and MNU + FP-12 groups, their number being higher inthe latter. The number of carcinoma bearing rats as well asthe average number of carcinomas per rat were significantlyhigher (P< 0.05) in the MNU + FP-12 group as compared tothe MNU-alone values. Aberrant crypt foci (ACF) were found inall carcinogen-treated rats, including those that did not containtumors, whereas none were observed in the FP-12 and Controlgroups. The average number of ACF/cm² was also significantlyhigher in the MNU + FP-12 group, as was the case for the averagenumber of ACF containing >10 aberrant crypts per focus. Thesefindings suggest that FP-12 can express promoting activity inchemically induced colon carcinogenesis.     

Supplemental administration of 1{alpha}-hydroxyvitamin D3 inhibits promotion by intrarectal instillation of lithocholic acid in N-methyl-N-nitrosourea-induced colonic tumorigenesis in rats

The effect of 1-hydroxyvitamin D₃ [1(OH)D₃) on promotion byintrarectal instillation of lithocholic acid (LC) in N-methyl-N-nitrosourea(MNU)-induced colonic tumorigenesis was studied in a rodentmodel. Ninety-two female F344 rats received intrarectal injectionof 2.5 mg of MNU twice in one week followed by 1 mg of LC orits vehicle alone three times weekly for 48 weeks. Those whichreceived LC were given a concomitant intragastric administrationof 0.04 µg of 1(OH)D₃ or its vehicle alone three timesweekly. In the group receiving MNU alone (n=30) five rats borecolomc tumors; in the MNU + LC group (n=32) 15 and in the MNU+ LC + 1(OH)D₃ group (n=30) six rats bore colonic tumors (MNU+ LC versus MNU + LC + 1(OH)D₃ group, P<0.05). These resultsindicated that promotion of MNU-induced colonic tumorigenesisby LC was suppressed by supplemental administration of 1(OH)D₃.     

MOLECULAR EPIDEMIOLOGY AND CANCER PREVENTION: Differential activity of aspirin, ketoprofen and sulindac as cancer chemopreventive agents in the mouse urinary bladder

In vivo studies were conducted to compare the activity of threenon-steroidal anti-inflammatory drugs as inhibitors of urinarybladder carcinogenesis induced in B6D2F₁ (BDF) mice by N-butyl-N-(4-hydroxybutyl)nitrosamine(OH-BBN). Mice received continuous dietary exposure to non-toxicdoses of aspirin, sulindac or ketoprofen beginning 1 week priorto the first of eight weekly doses of 7.5 mg OH-BBN; studieswere terminated at 24 weeks after the first carcinogen dose.Both dose levels of sulindac (200 and 400 mg/kg diet) and bothdose levels of ketoprofen (40 and 80 mg/kg diet) reduced theincidence of transitional cell carcinoma of the urinary bladderby >70% from that seen in dietary controls. The high doseof sulindac conferred the greatest protection against bladdercancer induction. In contrast, when administered at 400 and800 mg/kg diet aspirin was inactive as a chemopreventive agentin the OH-BBN/BDF bladder cancer model. The significant potencyof sulindac and ketoprofen as inhibitors of urinary bladdercarcinogenesis, when considered with their history of safe humanuse, suggests that these agents merit further study as drugsfor cancer chemoprevention in this target tissue.     

Inhibition of aflatoxin B1 binding to hepatic DNA by dehydroepiandrosterone in vivo

Dehydroepiandrosterone (DHEA) a naturally occurring steroid,has been reported to inhibit the binding of N-dimethylnitrosamineand 7,12-dimethylbenz[a]anthracene to DNA in vivo and to increaseglutathione transferase activity. In this study, we have investigatedif DHEA could protect hepatic DNA from damage by the potenthepatocarcinogen aflatoxin B₁ (AFB₁). Young male Fischer 344(2-month-old) rats were fed a diet containing 0.8% DHEA for14 days. Control rats were pair-fed the same diet without DHEA.The rats were then administered a single i.p. dose of [³H]AFB₁in dimethylsulfoxide (0.6 mg/kg body weight; 200 mCi/mmol) andkilled after 3 h. Liver weight, mitochondrial, microsomal andcytosolic protein, cytochrome P450 content and glutathione transferaseactivity increased significantly (P < 0.001) in DHEA-fedrats; however, the hepatic DNA content was not altered. DHEAfeeding increased the total amount of AFB₁ bound to hepaticprotein but decreased the extent of DNA binding. In in vitroexperiments, there was less total binding to DNA and proteinby AFB₁ when using microsomes from DHEA-fed rats. These resultssuggest that DHEA inhibits the binding of AFB₁ to DNA by modifyingthe biotransformation of the carcinogen.     

The effects of methylnitrosourea (MNU) on natural killer (NK) cell cytotoxicity and cytokine production in rats

Eight-week-old male Sprague–Dawley rats were exposed tothe carcinogen methylnitrosourea (MNU) via gastric intubationat doses of either 10 or 20 mg/kg body wt. Rats were treatedonce a week for 4 weeks, then once every 2 weeks for 1 month,for a total of 6 treatments. MNU was found to exert no consistentsignificant immunosuppressive effects in vivo as measured byspleen natural killer (NK) cell cytotoxicity, interleukin-2(IL-2) production by splenic lymphocytes and prostaglandin E₂(PGE₂) production by adherent peritoiieal macrophages. In contrast,splenic NK cell cytotoxicity and IL-2 production of MNU-treatedrats were actually elevated at several of the later samplingperiods. PGE₂ production was also elevated in MNU-treated ratsin the later sampling periods. Body weights of MNU-treated ratswere markedly decreased as early as 4 weeks following the initialMNU treatment. This suppression persisted throughout the study.The most dramatic change in organ weights was seen in the thymus.Thymus weights of all MNU-treated rats were significantly decreased1 day after treatment and persisted for 4 weeks. By the 60 daysampling period, thymus weights were not significantly differentfrom controls. However, by 120 and 180 days, thymus weightsagain were significantly lowered In those rats receiving MNU.These changes in thymus weights were accompanied histologicallyby initial cortical thinning and progressive loss of corticalthymocytes followed by the appearance of hyperplastic and neoplasticcells. It thus appears that the carcinogenic effect of MNU isnot related to a depression of the Immune surveillance system,at least as measured by NK cell activity.     

Carcinogenic doses of methylnitrosourea induce dose response related delay in transit through S and G2phases in mouse epidermis: a cell kinetic study

The cell kinetic, tumorigemc and carcinogenic effects of theshort acting, alkylating carcinogen N-methyl-N-nitrosourea (MNU)on hairless mouse epidermis were investigated. The epidermalmitotic rate, the mitotic index, and the number of basal andsuprabasal cells were scored in histological sections. Incorporationof [³H] thymidine and flow cytometric analysis of cellular DNAand protein content were performed on isolated basal cells atintervals for up to 10 days after a single application of either1 or 10 mg MNU. The ensuing tumor rates and yields were observedfor up to 48 weeks after 1 mg MNU and 30 weeks after 10 mg MNU.Generally, MNU induced an initial delay in epidermal cell cycleprogression with an accumulation of cells in the S and G₂ phases.Some days after treatment the delayed cells were released andentered mitosis. One milligram MNU caused a moderate delay ofcells in S and G₂ lasting for 2–3 days, and this was followedby a release leading to an increased number of suprabasal cellson day 7. The highest dose of MNU caused a more pronounced delayin transit through S and G and seemed to be followed by rapidregenerative proliferation. The subsequent tumor crop after10 mg was significantly higher than that seen after the lowestdose. The present cell kinetic results are consistent with previousdata from the study of other carcinogens, all showing a carcinogen-inducedin itial reduction in DNA synthesis after appropriate doses.A delay in transit through G₂ phase was found as well, in dicatingthat a general delay in cell cycle progression may follow theapplication of most (or all) carcinogens.     

Effect of selective and non-selective muscarinic blockade on baclofen inhibition of gastric carcinogenesis induced by N-methyl-N'-nitro-N-nitrosoguanidine in Wistar rats

The effects of baciofen, a -amino-n-butyic acid receptor B agonist,on gastric carcinogenesis induced by N-methyl-N'-nitro-N-nitrosoguanidineand how its effects areinfluenced by selective (M₁) and non-selective(M₁ and M₂) pharmacological blockade of muscarinic receptorswere investigated in inbred Wistar rats. Rats were given s.c.injections of 8 mg/kg body wt baclofen with and without 0.5mg/kg body wt atropine (non-selective M¹ and M² muscarinic receptorantagonist)or 1.0 mg/kg body wt pirenzepine (selective M¹ muscarinic receptorantagonist every other day after a 25 week carcinogen treatment.At week 52 baclofen significantly decreased the incidence ofgastric cancers. Concomitant treatment with atropine significantlyattenuated the inhibition by baclofen of gastric carcinogenesis,but combined use with pirenzepine had no significant effecton the inhibition by baclofen of gastric carcinogenesis. Baclofenalso significantly decreased the labeling index of the antralomucosa. Baclofen plus atropine attenuated the decrease in thelabeling index of the antral mucosa due to baclofen, but baclofenplus pirenzepine had no significant effect on the labeling index.These results suggest that the inhibition of gastric carcinogenesisby baclofen is medicated through muscarinic receptors and M₂receptors, but not M₁ receptors, are involved in this response.     

DNA methylation adduct formation and H-ras gene mutations in progression of N-butyl-N-(4-hydroxybutyl)nitrosamine-induced bladder tumors caused by a single exposure to N-methyl-N-nitrosourea

After receiving 500 p.p.m. N-butyl-N-(4-hydroxybutyl) nitrosamine(BBN) in their drinking water for an initial 10 weeks, ratswere given a single i.p. injection of N-methyl-N-nitrosourea(MNU) at a dose of 50 mg/kg body wt at week 20 (at a stage whenbladder tumor development had already occurred), and then maintaineduntil they were killed at week 40. Three and six hours afterthe MNU injection, the DNA methylation adducts, O⁶-methyldeoxyguanine(O⁶-medG) and 7-methyldeoxyguanine (7-medG), were immunohistochemicallyrevealed to be markedly more frequent in urothelial preneoplasiasor neoplasias than in normal cells. These adducts were rapidlyrepaired, and although 7-dmeG in tumor cells still persistedafter 72 h, they appeared essentially to have returned to normallevels. At the termination, conversion of transitional cellcarcinomas (TCC) to squamous cell carcinomas (SCC) of the urinarybladder was significantly increased in the BBN + MNU group.The extent of invasion was also significantly greater with theadditional MNU treatment Expression of p21 protein, detectedby immunohistochemistry, was comparable between the groups.Mutations in the H-ras gene were observed in one case each ofthe BBN and BBN + MNU groups, and both cases showed a G:C toA:T transition at codon 12. The present study thus suggestedthat while an additional single treatment with MNU of rats bearingBBN-induced bladder neoplasias is associated with significant,possibly mutation-dependent tumor progression, H-ras mutationsare not necessary events.     

Possible involvement of arachidonic acid metabolism in phenobarbital promotion of hepatocarcinogenesis

The effects of inhibitors of arachidonic acid metabolism andantioxidants on the rat liver tumor promotion activity of phenobarbital(PB) were assessed using the enzyme-altered focus as the end-pointlesion. Fischer 344 male rats were initiated with N-nitrosodiethylamine(200 mg/kg) and then divided into five groups placed on basaldiet, diet containing 0.05% PB, diet containing 0.05% PB plus0.75%, 1% or 1.5% levels of various inhibitors of arachidonicacid metabolism or antioxidants, or diet containing 1% or 1.5%inhibitors or antioxidants alone for 10 weeks, and then killed.-Bromo phenacyl bromide, an inhibitor of phospholipase A₂ significantly inhibited the promotion activity of PB at dose levelsof 0.75% and 1.5%, reaching plateau at 0.75%. Both quercetin,an inhibitor of lipoxygenase, and morin, a dual inhibitor oflipoxygenase-cyclooxygenase, significantly reduced the promotionactivity of PB at the 1.5% but not 0.75% dose levels. Moreover,acetylsalicylic acid, an inhibitor of cyclooxygenase dose-dependentlyinhibited the promotion activity of PB. Among the antioxidantsinvestigated, vitamin E did not affect, but n-propyl gallateand ethoxyquin exerted a dose-dependent inhibition of PB promotion.These results are strongly suggestive of an involvement of phospholipaseA₂ lipoxygenase and cyclooxygenase arachidonic acid metabolicpathways in the mechanisms underlying PB enhancement of hepatocarcinogenesis.     

Carcinogenicity of N-nitroso(2-hydroxypropyl)(2-oxopropyl)amine, N-nitrosobis(2-hydroxypropyl)amine and cis-N-nitroso-2,6-dimethylmorpholine administered continuously in the Syrian hamster, and the effect of dietary protein on N-nitroso(2-hydroxypropyl)(2-oxopropyl)amine carcinogenesis

The effect of continuous week-long administration of the threepancreatic carcinogens N-nitroso(2-hydroxypropyl)(2-oxo-propyl)amine(HPOP), N-mtrosobis(2-hydroxypropyl)amine (BHP), and cis-N-nitroso-2,6-dimethylmorpholine(cis-NNDM), by a s.c. implanted osmotic pump, was examined inSyrian hamsters. HPOP at total doses of 220–250 mg/kgbody weight induced ductal adenocarcinomas in the pancreas (41%),and cholangiomas (18%) and cholangiocarcinomas (18%) in theliver, 25 weeks following the initiation of treatment. Higherdoses of HPOP resulted in severe hepatic injury and increasedmortality (LD₅₀=280 mg/kg). Cis-NNDM and BHP were less toxicthan HPOP and induced pancreatic lesions at doses of 950 mg/kg.These data document that a week-long schedule of continuousadministration of HPOP for the induction of pancreatic cancercompares favorably with those involving weekly injections. Applicationof this model to study the effect of dietary protein in HPOP-inducedcarcinogenicity showed that the number of cystic, intermediateand tubular complexes in the pancreas was significantly higherin animals fed a 20% as compared to an 8% protein diet 2 weeksprior to HPOP administration. Furthermore, the incidence ofpancreatic adenocarcinomas and in situ carcinomas was only 13%in the hamsters fed the low-protein diet as compared to 46%in those fed the high-protein diet.     

Modulation of DNA adduct formation by successive exposures of DNA to small and bulky chemical carcinogens

The effect of a carcinogen—DNA adduct on the formationof a second adduct upon subsequent exposure to a second carcinogenwas studied using (i) a modified Maxam-Gilbert chemical sequencingreaction and (ii) a DNA synthesis termination analysis. A DNAfragment of known sequence was reacted with micromolar concentrationsof N-methyl-N-nitrosourea (MNU), (+)-r-7,t-8-dihydroxy-t-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene(BPDE), N-acetoxy-2-acetylaminofluorene (N-acetoxy-AAF), oraflatoxin B₁-8,9-epoxide (AFB₁ epoxide) singly or in successivedouble reactions. N-Acetoxy-AAF adducts were sensitive to theMaxam-Gilbert sequencing reaction for guanine; these adductssignificantly blocked the formation of BPDE-guanine adducts.Treatment of DNA with BPDE, however, did not inhibit subsequentformation of N-acetoxy-AAF adducts. DNA synthesis terminationanalysis suggested that methylation of guanine altered subsequentarylation of guanine by N-acetoxy-AAF and AFB₁ epoxide, andthat combined treatment inhibited replication, an effect notseen after MNU treatment alone. Possible mechanisms for themodulation of carcinogen binding are discussed. It is concludedthat not only is DNA damage by genotoxic carcinogens dependentupon both the chemical nature of the carcinogens and the nearestneighbors to target guanine bases, but that the effect of subsequentexposures to carcinogens is not always additive.     

Failure of mitogen-induced cell proliferation to achieve initiation of rat liver carcinogenesis

Experiments were designed to determine whether mitogen inducedcell proliferation is as effective as regenerative cell proliferationin achieving initiation of liver carcinogenesis. To test thishypothesis male Wistar rats were injected with a single doseof diethylnitrosantine (DENA) or N-methyl-N-nitrosourea (MNU)during the peak of DNA synthesis following the administrationof the liver mitogen, lead nitrate, after partial hepatectomy(PH) or a necrogenic dose of CCl₄. The initiated hepatocyteswere monitored as -glutamyltransferase (GGT)-positlve foci usinga 2-week selection regimen consisting of 0.03% 2-acetylaminofluorene(2-AAF) coupled with a necrogenic dose of CCl₄. The resultsindicate that unlike compensatory cell proliferation such asthat induced by PH or CCl₄, mitogen-induced cell proliferationdid not result in any initiated hepatocytes despite the factthat in both types of models the extent of liver cell proliferationat the time of the administration of the carcinogen is similar.     

Chemoprevention of N-Nitroso-N-methylurea-induced Rat Mammary Carcinogenesis by Soy Foods or Biochanin A

We examined the effects of soybeans, a soy product (miso) and biochanin A, an isoflavone derivative, on N -nitroso- N -methylurea (MNU)-induced rat mammary carcinogenesis. Seven-week-old female CD/Crj rats received a single i.v. dose (40 mg/kg body weight) of MNU. After administration of MNU, rats were fed diet containing 0% (control), 2% or 10% soybeans, or 10% miso as a soy-supplemented diet, or 10 or 50 mg/kg biochanin A. All rats were observed for 18 weeks after MNU administration. At 18 weeks, the multiplicity (mean tumors/rat) of palpable mammary tumors was significantly decreased in the 10% soybean (1.1) and 10% miso (1.2) diet groups compared to the control (2.2) ( P <0.05, respectively). In the biochanin A-supplemented diet groups, the incidence (percentage of rats with tumors) was significantly decreased in the 50 mg/kg (32%) diet group compared to the control (80%) ( P <0.01), and the multiplicity was significantly decreased in both the 10 mg/kg (0.7) and 50 mg/kg (0.5) diet groups compared to the control (2.2) ( P <0.01 and P <0.001, respectively). The proliferative cell nuclear antigen labeling index of mammary tumors was significantly decreased in both biochanin A-supplemented diet groups compared to the control. The present results indicate that soybeans, miso, and biochanin A are useful for the prevention of mammary cancer.     

DNA and protein adducts as indicators of in vivo methylation by nitrosatable drugs

Exposure to methylating carcinogens may be monitored by measuringboth the formation of S-methylcysteine in haemoglobin and theurinary excretion of N-7-methylguanine (7-MeG), which is derivedin part from methylated nucleic acids. Female rats were exposedto methylmethanesulphonate, methylnitrosourea and to three drugs,aminopyrine, cimetidine and pyrilamine, which are potentialmethylating agents if nitrosation occurs in vivo. Because S-methylcysteinein haemoglobin and urinary 7-MeG occur naturally, the experimentswere carried out with stable isotope-labelled analogues whichcontained trideutero (d₃)-methyl groups. Gas chromatography-massspectrometry was used for the quantitative determination ofd₃-labelled adducts after their separation from the biologicalmatrix and chemical derivatization. Transfer of the intact d₃-methylgroup to cysteine and guanine was detected after intragastricadministration of d₃-methyl-methanesulphonate (50 mg/kg), d₃-N-methyl-N-nitrosourea(50 mg/kg), and d6-aminopyrine (AP) and sodium nitrite (both100 mg/kg). AP alone gave no detectable d₃-methyl adducts. Co-administrationof nitrite and d₆-pyrilamine or d₃-cimetidine yielded no d₃-7-MeG,although N-nitroso-d₃-cimetidine alkylated DNA in vitro in adose-dependent fashion. For AP and nitrite combinations urinaryexcretion of d₃-7-MeG was linearly related to the dose of nitriteand was essentially complete within 5 days. For d₃-methylmethane-sulphonate(50 mg/kg) the ratio of haemoglobin d₃-S-methyl-cysteine tourinary d₃-7-MeG was considerably (>9-fold) higher than foreither d₃-N-methyl-N-nitrosourea or AP/ nitrite (100 mg/kg)mixture. This is in accord with the S_N2 nature of the weak carcinogenmethylmethanesulphonate compared with the S_N1 nature of thereactive methylating agent derived from either one of the N-methyl-N-nitrosocompounds.     

Effects of sex difference, gonadectomy and estradiol on N-ethyl-N-nitrosourea-induced trigeminal nerve tumors in rats

The occurrence of trigeminal nerve tumors (TNTs) induced byneonatal administration of N-ethyl-N-nitrosourea (ENU) in WF? LE F₁ (F₁ rats was studied with special reference to sex difference,effect of gonadectomy and estradiol (E₂) administration. Experimentalgroups 1–6 were treated with 40 mg ENU/kg of body weightneonatally. They consisted of male, female, castrated male,ovariectomized female, E₂ pellet (0.1 mg, s.c.) supplementedand gonadectomized male and female rats respectively. Rats ofgroups 7–12 served as the respective controls withoutENU. All the rats were killed at 8 months of age. Levels ofserum E₂ and E₂ receptor (ER) of the TNTs were also examined.It was noted that the incidence of TNT was higher in males (79%)than in females (48%, P < 0.05) and did not change by castrationin males (91%) but increased in ovariectomized female rats (74%,P < 0.05). Administration of E₂ followed by gonadectomy inhibitedthe occurrence of TNTs in male rats (59%) but not in femalerats (60%). No TNT was observed in any control groups. Kidneytumors were the second most frequent tumors next to nervoussystem tumors in the present experiment. The incidence of kidneytumors was much higher in females (38%) than in males (4%, P< 0.05) and decreased by ovariectomy, whereas it increasedin male rats by E₂ administration. ER levels of TNTs and trigeminalnerve tissue were < 1 fmol/mg protein. These results suggestthat in rats treated with ENU neonatally, E₂ has an inhibitoryeffect on the induction of TNTs but may not be regulated throughER. E₂ also shows a promoting effect on kidney tumorigenesis.     

Positive influence of dietary deoxycholic acid on development of pre-neoplastic lesions initated by N-methyl-Nnitrosourea in rat liver

The effect of deoxycholic acid (DCA) treatment subsequent toinitiation of F344 male rats with N-methyl-N-nitrosourea (MNU),a wide spectrum carcinogen inducing tumors in many organs, wasinvestigated. Rats were initially given four doses of MNU (50mg/kg) i.p. within a 2-week period combined with a two-thirdspartial hepatectomy performed at day 7 and then placed on basaldiet containing DCA at concentrations of 0.313, 0.125, 0.050and 0.020% for 21 weeks prior to final sacrifice. All organsstudied were carefully examined histologkally and histochemicallyfor development of neoplastic and pre-neoplastic lesions. DCAenhanced the induction of glutathione S-transferase positive(GST-P+) liver cell foci in a dose-related manner. Furthermoregroups of rats given DCA without prior MNU administration alsodeveloped dosedependent numbers of pre-neoplastic liver lesions.In addition, increased numbers of small intestine tumors wereapparent in DCA-treated animals although the difference wasnot significant. Induction of tumors in the thyroids, Zymbalglands, skin and peripheral nerves was not affected. The resultsindicate that DCA is a strong promoter of hepatocarcinogenesiswith possible complete carcinogenkity in the liver and promotionpotential for tumor development in the small intestine.     

SHORT COMMUNICATION: Inhibition of rat mammary tumorigenesis by dietary cholesterol

The effects of dietary cholesterol and oxidized cholesterolon mammary tumor development were examined in female Sprague-Dawleyrats exposed to the carcinogen N-methyl-N-nitrosourea (MNU).Animals were administered 50 mg/kg MNU at 50 days of age andfed either a control (AIN-76) diet or the control diet supplementedwith 0.3% cholesterol or 0.3% oxidized cholesterol for up to26 weeks. The oxidized cholesterol was prepared by heating cholesterolat 110°C for 48 h. Gas chromatographic analysis of the oxidizedcholesterol revealed a 2% yield of oxidation products in additionto a large amount of unchanged cholesterol (>96%). Tumorincidence in the cholesterol group (67%) was significantly lowerthan in the control group (96%, P <0.05), but the oxidizedcholesterol group (79%) was not significantly different fromthe control or cholesterol groups. Average number of tumorsper animal was lower in the cholesterol group (1.5) than inthe control (2.8) or oxidized cholesterol groups (2.3, P <0.005).Serum low density lipoprotein (LDL) cholesterol was greaterin the cholesterol (185± 38 mg/dl) and the oxidized cholesterolgroups (160± 34 mg/dl) than in the control (55±4 mg/ dl, P <0.05), although there was no difference betweenthe cholesterol and the oxidized cholesterol groups. These resultsshow that dietary cholesterol inhibits mammary tumor developmentin this model. Elevated serum LDL cholesterol may inhibit denovo cholesterol synthesis in preneoplastic and/or tumor cells,thereby inhibiting their proliferation.     

Inositol and inositol hexaphosphate suppress cell proliferation and tumor formation in CD-1 mice

In previous studies, we have shown that inositol hexaphosphate(InsP₆), a constituent of cereal diet, inhibited azoxymethane-inducedexperimental large intestinal cancer (LIC) in Fischer 344 rats.We now report a similar antineoplastic action of InsP₆ in CD-1mice injected with 1,2-dimethylhydrazine (DMH). We had hypothesizedthat InsP₆ may bring about this effect by undergoing dephosphorylationto lower phosphorylated forms; the ready availability of Ins,to react with phosphates, may increase the total amount of thelower phosphorylated Ins and potentiate the action of InsP₆.LIC induced by DMH (15 mg/kg/week ? 13) in mice given a mixtureof 1% InsP₆ + 1% Ins show a significant reduction (P <0.005)in LIC prevalence over InsP₆ treatment. Surprisingly, Ins, anin vitro growth promoting agent also caused a significant (P< 0.001) suppression of LIC. InsP₆ ? Ins also showed a concomitantreduction in the mitotic rate in the non-neoplastic epithelium.Body weight data did not suggest any overt toxic effect of long-termadministration of InsP₆, Ins or InsP₆ ? Ins. Since InsP₆ isantineoplastic in two species of experimental animals, it should,in combination with Ins, be considered in our strategies forprevention of large intestinal cancer.     

The excretion of 7-methyladenine in the urine of rats exposed to carcinogenic methylating agents

Earlier studies showed that urine of rats which had been injectedwith the methylating agent N-[³H-methyl]-N-nitrosourea containeda previously undetected metabolic product, 7-[³H-methyl]adenine.This methylpurine, undoubtedly derived from alkylation of nucleicacids followed by depurination, was not labeled when¹⁴C-methyl-labeledmethionine was administered concurrently. To establish whetherurinary 7-methyladenine (7-MA) might serve as a marker of exposureto exogenous and carcinogenic methylating agents, the excretionof 7-MA following injection of methylating agents was measured.A GC-MS method, using pentafluorobenzyl derivatives and an internalstandard of tri-deutero-7-MA, was developed to assay levelsof 7-MA. Increasing the i.p. dose of N-methylnitrosourea (MNU)from 2 to 80 mg/kg/rat resulted in a linear increase in urinary7-MA, which at the highest dose was 1.6 µg during thefirst day and another 0.4 µg during day 2. Doses of 5mg/kg MNU led to elevated urinary levels of 7-MA (144 ng) comparedto controls (26 ng). Other methylating agents, such as dimethyl-nitrosamine,N-methyl-N-nitro-N-nitrosoguanidine and dimethyl sulfate, alsoprovided urinary 7-MA. To determine the fate of injected 7-MA,the administration of 2 µg 7-[³H-methyl]adenine led toan 80% recovery of radioactivity in the urine, almost all ofit during the first 24 h. No other labeled metabolites weredetected. At least for the rat, urinary 7-MA serves as an indicatorof exposure to methylating agents.     

Strong promoting activity by uracil on urinary bladder carcinogenesis and a possible inhibitory effect on thyroid tumorigenesis in rats initiated with N-methyl-N-nitrosourea

Uracil has been shown to cause a strong proliferative responsein the urinary bladder epithelium of rats and mice through calculusformation and, consequently, acts as a strong promoter in bladdercarcinogenesis. In this study, we examined the effect of uracilon two-stage carcinogenesis in various organs using N-methyl-N-nitrosourea(MNU) as the initiator. F344 rats were injected i.p. with MNUtwice weekly for 4 weeks and then given diet containing 3.0%uracil for 20 weeks. Uracil induced urinary bladder carcinomasin all rats pretreated with MNU, and it decreased the combinedincidence of adenomas and carcinomas in the thyroid. Althoughnot significant, uracil decreased the incidence of adenomasin the lung and increased that of lymphomas of the thymus. Apossible influence of a significantly decreased body weightgain caused by uracil treatment on the reduced tumor incidencein the thyroid and lung is discussed. The present data demonstratethe strong promoting activity of uracil for urinary bladdercarcinogenesis, and suggest a possible inhibitory effect onthyroid carcinogenesis.