Pathological abnormalities after long-term administration of medroxyprogesterone acetate in a queen.

In this report, multiple abnormalities (bilateral ovarian cysts, cystic endometrial hyperplasia and pyometra (CEH-P), mammary adenoma, fibrosarcoma and cystic-papillary adenocarcinoma) identified in a queen continually administered medroxyprogesterone acetate (MPA) for 9 years are described. An 11-year-old domestic shorthair intact queen was presented for reduced appetite, polyuria, polydipsia, abdominal distension, and mammary mass. Pyometra was diagnosed based upon clinical, ultrasonographic and radiographic examinations. Mastectomy and ovariohysterectomy (OHE) were performed for treatment. Bilateral ovarian cysts were detected during OHE. Histopathologically, follicular ovarian cysts, CEH-P, and benign and neoplastic mammary lesions were identified. We suggest that, the pathological abnormalities may have been attributed to adverse effects of prolonged administration of MPA.

No involvement of beta-catenin gene mutation in gastric carcinomas induced by N-methyl-N-nitrosourea in male F344 rats.

The agent N-methyl-N-nitrosourea (MNU) is a direct acting carcinogen and induces well-differentiated adenocarcinoma on the rat gastric mucosa. In this study, 27 histopathologically verified gastric carcinomas induced in male F344 rats were analyzed for mutations in the N-terminal phosphorylation sites (codons 1-51) of the beta-catenin gene by using polymerase chain reaction (PCR)-single strand conformation polymorphism (SSCP) assays. In parallel studies, the specific localization of the beta-catenin protein was also examined by immunohistochemical analysis. No mutations in the beta-catenin gene were found in any of 27 gastric carcinomas induced by MNU. Immunohistochemical analysis resulted in the beta-catenin protein to be localized in the plasma membrane but cytoplasmic and/or nuclear accumulation of beta-catenin was not identified in any of these carcinomas. These results suggest that mutations in the beta-catenin gene are less contributory to the development of rat gastric carcinomas induced by MNU. This animal model may provide a system for evaluating the mechanism of human gastric carcinogenesis that is not associated with beta-catenin gene mutations.

Inhibitory effect of loxiglumide (CR 1505), a cholecystokinin receptor antagonist, on N-nitrosobis(2-oxopropyl) amine-induced biliary carcinogenesis in Syrian hamsters.

We evaluated the cholecystokinin (CCK) receptor antagonist loxiglumide (CR1505) for a possible inhibitory effect on biliary carcinogenesis in a hamster model. Experimental group I underwent cholecystoduodenostomy and ligation of the distal end of the common bile duct, after which the animals were injected with N-nitrosobis(2-oxopropyl)amine (BOP) alone. Group II, after the same surgical procedure as in group I, were given injections of BOP and then given loxiglumide in their diet. The sham-operated group underwent simple laparotomy and then were given injections of BOP. Loxiglumide significantly inhibited BOP carcinogenicity in the gallbladder and extrahepatic bile duct but not in the intrahepatic bile ducts or pancreas. Autoradiography showed that loxiglumide significantly suppressed (125)I-Bolton-Hanter (BH)-CCK-8 binding to CCK receptors in the gallbladder and extrahepatic bile duct but not in the liver or pancreas, and CCK binding to its receptors was observed in an area identified as cancer tissue. CCK receptor antagonists have an inhibitory effect on BOP carcinogenesis in the extrahepatic biliary tract, including the gallbladder and extrahepatic bile duct, of Syrian hamsters. The difference in the inhibitory effect of loxiglumide on biliary carcinogenesis in hamsters according to site may be due to differences in CCK receptors or the affinity of loxiglumide for such biliary tract organs. A difference between carcinogenesis in the intrahepatic bile ducts and extrahepatic biliary tract may be another reason.

Selective mutation of K-ras by N-ethylnitrosourea shifts from codon 12 to codon 61 during fetal mouse lung maturation.

Fetal mouse lung before gestation day 17 shows unique sensitivity to causation of rapidly growing tumors by N-ethylnitrosourea (ENU). Since mouse lung tumors present a mutated K-ras oncogene, we hypothesized that this special susceptibility might reflect an unusual vulnerability of the K-ras gene. Of the lung tumors caused by ENU exposure of BALB/c mice on gestation day 14, 8/25 had a codon 12 mutation in K-ras, vs 4/25 in codon 61. Of 15 tumors after day 16 exposure, three had codon 12 and four codon 61 changes. Tumors from day 18 exposure had only codon 61 mutations (11/16), all A:T to G:C changes (CGA). By contrast, codon 12 (GGT) changes included G:C to T:A, to A:T, and to C:G. These results show significant (P<0.01) shift in the sensitivity of particular K-ras codons to ENU mutation, during fetal mouse lung maturation. In a test of a possible relationship to expression of K-ras, K-ras p21 was measured in lungs of fetal mice, and found to increase markedly on day 18 in comparison to days 14 and 16. Both alkylation of DNA and base damage due to reactive oxygen species are postulated as mechanisms for mutation by ENU, whose efficacies vary with state of fetal lung maturation and K-ras expression.

K-ras gene mutations in intrahepatic bile duct tumors of Syrian golden hamsters.

BACKGROUND AND OBJECTIVES: In our laboratory, we have developed a new model of carcinoma of the bile duct in Syrian golden hamsters, using N-nitrosobis(2-oxopropyl)amine (BOP). Morphologic and biologic characteristics of the carcinoma induced in this model are similar to those seen in humans. In order to examine the gene-related carcinogenesis of intrahepatic bile duct carcinoma, we investigated mutations in the K-ras gene in various early hyperplastic and neoplastic lesions of these hamsters, according to the original sites of the lesions. METHODS: Inbred female hamsters were given a subcutaneous injection of N-nitrosobis(2-oxopropyl)amine (BOP) following dissection of the extrahepatic bile duct on the distal end of the common duct and preparation of a cholecystoduodenostomy (CDDB) or simple laparotomy (SL). Neoplastic lesions arising from the intrahepatic bile duct were histologically examined, and K-ras mutations were investigated. RESULTS: Mutations of K-ras codon 12 were evident in 12% of tubular hyperplasias, 19% of tubular adenocarcinomas, 15% of papillary hyperplasias and 36% of papillary adenocarcinomas. In papillary adenocarcinoma arising from a large bile duct, K-ras mutations occurred more frequently than in tubular adenocarcinoma arising from ductule or ductular proliferation. K-ras mutations were present even in a hyperplasia; the positive rates of the mutations increased in the presence of a carcinoma. Genetic changes in carcinoma of the intrahepatic bile duct varied based on sites of the duct and the histological type. CONCLUSIONS: A part of the hyperplastic lesions of the intrahepatic bile duct presented K-ras gene mutation. This suggests that K-ras gene mutation is an early event in the carcinogenic process. In carcinoma of the intrahepatic bile duct, the lesion arising from a large bile duct of the hepatic hilum tended to exhibit a higher frequency of K-ras gene mutation than did a tubular lesion arising from ductule or ductal proliferation. This hamster model is useful to examine the carcinogenesis of human intrahepatic bile duct carcinoma.

Role of nuclear magnetic resonance spectroscopy (MRS) in cancer diagnosis and treatment: 31P, 23Na, and 1H MRS studies of three models of pancreatic cancer.

The role of nuclear magnetic resonance spectroscopy (MRS) in pancreatic cancer diagnosis and its treatment were assessed in three models of pancreatic neoplasms. Perfused MIA PaCa-2 human pancreatic cancer cells, s.c. implanted pancreatic tumors in hamsters, and pancreatic tumors induced in situ in rats by direct application of the carcinogen 7,12-dimethyl benzanthracene, were studied by phosphorous ((31)P), sodium ((23)Na), and proton ((1)H) MRS. (31)P spectra of pancreatic cancer were qualitatively similar to those of intact organs. There were, however, variations in peak intensities and ratios. Phosphomonoester signals were prominent in both normal pancreases and tumors, but their levels depended on the proliferation rate and on environmental conditions. Thus, the phosphomonoester:beta-nucleoside triphosphate ratio was 1.15 +/- 0.32 in 90% confluency and 1.31 +/- 0.43 in 70% confluency, and this ratio increased upon lowering the perfusion rate. Total (intra- and extracellular) sodium concentrations, measured in the solid tumors, were 39-40 micromol/g wet weight in normal pancreases. Contrary to a previous hypothesis that malignant transformation is associated with increased sodium content, our (23)Na MRS data showed that there were no significant differences between pancreatic tumors and intact organs. Proton spectra of perchloric acid extracts revealed several differences between tumors and control pancreases. The principal findings were elevated levels of the amino acid taurine, from 1.17 +/- 0.39 micromol/g wet weight in healthy pancreases, to 2.79 +/- 0.71 micromol/g wet weight in pancreatic carcinoma in rats, and lactate levels that increased from 0.92 +/- 0.2 to 6.19 +/- 1.93 micromol/g wet weight, respectively. On the other hand, creatine and glutamate were higher in the normal pancreases. Pancreatic cancer is usually resistant to chemotherapy, and we evaluated the effects of the metabolic inhibitors 2-deoxyglucose and lonidamine on the human pancreatic cancer cells by MRS and cytotoxicity studies. The IC50 of Adriamycin and 2-deoxyglucose were 1.49 +/- 0.18 x 10(6) and 136 +/- 17 microg/ml, respectively. These results were similar to data obtained previously in multidrug-resistant human breast cancer cells, which were highly resistant (33-fold) to Adriamycin but were more susceptible (9-fold) to 2-deoxyglucose than their parental cells.

Effect of synthetic protease inhibitor on histologic changes and free radical activity in hamsters with pancreatic cancer.

To investigate the effects of synthetic trypsin inhibitors on pancreatic cancer, camostat (FOY-305) was administered orally to hamsters with experimental pancreatic cancer induced by diisopropanol nitrosamine (DIPN). The effect of free radicals on carcinogenesis was examined by measuring the tissue levels of the scavengers superoxide dismutase (SOD) and glutathione peroxidase (GSX-Px), and pancreatic tissues were examined histologically. Cancers developed in all hamsters that survived 24 weeks in the DIPN group and the FOY group, but 80% of the cancers in the DIPN group were tubular adenocarcinomas, and 91% of those in the FOY group papillary adenocarcinomas. The SOD activity in the DIPN group was significantly lower in the cancerous area and the borderline region than in the non-cancerous region and normal tissue. SOD activity in the cancerous and borderline regions was higher in the FOY groups than in the DIPN group. GSH-Px levels in the borderline and non-cancerous regions were significantly higher in the FOY group than in the DIPN group. These results suggest that the synthetic protease inhibitor slows the progress of pancreatic cancer by its free radical scavenging activity.

Glucocorticoid hormone effect on transplacental carcinogenesis and lung differentiation: influence of histocompatibility-2 complex.

In the mouse, the histocompatibility-2 (H-2) haplotype influences induction of lung and intestinal tumors by N-ethyl-N-nitrosourea (ENU) treatment of fetuses or infant mice. The differentiation of lung and intestinal epithelium is known to be regulated by glucocorticoids. We show that glucocorticoid-induced development of alveolar lung volume is H-2 influenced and that glucocorticoid treatment of fetuses also influences prenatal ENU induction of lung and intestinal tumors. These glucocorticoid effects on tumorigenesis are also H-2 influenced. The number of papillary lung tumors increased in B10 (H-2b) and decreased in B10.A (H-2a) mice. In the intestine, the number of tumors increased in H-2b females and decreased in H-2b males. In H-2a mice, the number of intestinal tumors was unchanged but their location was altered. We propose that the H-2 complex influences tumorigenesis in lung and small intestine by affecting the hormonal regulation of differentiation of target epithelial cells.

Morphological and biological characteristics of mammary tumours induced by the direct application of DMBA powder to rat mammary glands.

Mammary tumours were induced by the direct dusting of 1 mg, 7,12-dimethylbenz(a)anthracene (DMBA) powder onto the mammary gland of both 30-day-old female and male Sprague-Dawley rats, and the tumours were examined histologically. Mammary tumours developed in 43/43 (100%) of the females 11 to 20 weeks after DMBA dusting and 16/23 (70%) of the males 18 to 28 weeks after dusting, while non-mammary spindle cell sarcomas occurred in 5/23 (22%) of the males 15 to 24 weeks after dusting. A variety of benign and malignant mammary tumours of epithelial and/or mesenchymal origin were induced, which are comparable to human mammary tumours. Different histological patterns were observed in different areas of the same tumours. Ovariectomy revealed hormone (ovary)-dependency in 10/17 (59%) of the tumours, revealing regressing epithelial and proliferating mesenchymal tumour elements on histological examination.

Experimental models for the sequential analysis of chemically-induced renal carcinogenesis.

In order to discriminate non-specific toxicity from the early precursor lesions of neoplasia, emphasis in these studies has been on the use of models requiring only a single administration of chemical. Our interests have focussed on three neoplastic entities in the kidney, renal mesenchymal neoplasia, renal cell carcinoma, and nephroblastoma. Dimethylnitrosamine administered as a single intraperitoneal injection to immature female Wistar rats, pre-conditioned for several days with a no-protein/sugar-only diet, has been used for investigating the complex morphological nature of renal mesenchymal tumors, their pathogenesis and the development of cell culture correlates. The near 100% tumor incidence and its facility for cell culture manipulation makes this a particularly potent model for studying chemical carcinogenesis and the evolution of cell transformation. Discovery that the rat kidney response to DMN was biphasic with respect to the time of treatment led to the subsequent development of a high incidence system for inducing renal adenocarcinoma, using older rats. Renal cell carcinomas could also be induced in mice by a single intravenous injection of streptozotocin. The tumor frequency in female CBA/H/T6J mice was almost 100%, providing a new model for the investigation of renal carcinogenesis in this species. Nephroblastoma has been a poorly comprehended neoplasm in both lower animals and man because of the lack of a high incidence model in conventional laboratory mammals. Recently, we have exploited an increased spontaneous predisposition of the Nb rat to nephroblastoma using a single intraperitoneal dose of N-ethylnitrosourea in pregnant females on day 18 of gestation, producing a frequency of 50% for this tumor type. More potent however, was a system which utilized the partially inbred IIIVO/J strain of rabbit using the same carcinogen and transplacental route of administration. The resultant incidence of nephroblastomas in the progeny was in excess of 90%, and like their counterparts in man, the neoplasms developed rapidly and had a potential for distant metastasis. Each one of these animal models is suitable for the sequential tracing of tumor pathogenesis, and in depth analysis of the biochemical and molecular mechanisms involved in the initiation and formation of different types of renal cancer.

Bilateral renal papillary adenocarcinoma in a young man on long-term captopril therapy.

A 31-year-old black man presented with bilateral multiple renal papillary adenocarcinomas. The patient had been maintained on captopril therapy for severe hypertension for three years. The possibility of drug-induced malignancy is discussed.

Mutagenicity and carcinogenicity of mono- and diacetyl hydrazine.

Two hydrazine derivatives, monoacetyl hydrazine (MAH) and diacetyl hydrazine (DAH), have been tested for mutagenic response in the Salmonella/mammalian microsome assay and micronucleus test. MAH but not DAH, increased the revertant mutants in TA100 and TA1535 and also increased the frequency of micronuclei in polychromatic erythrocytes. Gavage administration of MAH but not of DAH, resulted in increased incidence of lung tumors. These observations record for the first time the mutagenicity/carcinogenicity of MAH which is one of the metabolites of isoniazid in animals and humans.

[Experimental minute gastric cancer induced by N-ethyl-N'-nitro-N-nitrosoguanidine in dogs].

150 micrograms/day of N-ethyl-N'-nitro-N-nitrosoguanidine (ENNG) was administered to a total of 8 dogs, (4 mongrels at age of 4 months and 4 beagles at age of 6 months) over a period of 8 months by Kurihara 's method. As a result of the administration, we found development of minute cancer as follows: In 3 animals, male beagle killed at 575th day, male mongrel at 1, 105th days and male mongrel at 1, 245th days, a total of 20 neoplasms of the stomach was found (18 early cancers and 2 advanced cancers). 13 of which being the minute cancer measuring less than 0.5 cm. There were 11 mucosal cancers and 2 submucosal cancers. When classified by the macroscopic pathological type, none was classified as the elevated type (I, IIa types), 5 lesions as the flat type (IIb type), and 8 lesions as the depression type (IIc type). Two lesions of submucosal cancer belonged to IIc type. When classified by the histological type, 2 were classified as papillary adenocarcinoma, 2 as well differentiated tubular adenocarcinoma, 1 as moderately differentiated tubular adenocarcinoma, 5 as poorly differentiated adenocarcinoma and 3 as signet-ring cell carcinoma.

Organ-specific promoting effect of phenobarbital and saccharin in induction of thyroid, liver, and urinary bladder tumors in rats after initiation with N-nitrosomethylurea.

Tumor-promoting effects of phenobarbital (PB) and sodium saccharin (SS) were tested in rats pretreated with N-nitrosomethylurea (NMU) with special reference to the site of their action. Male F344 rats were initially given injections of NMU (20 mg/kg i.p.) twice a week for 4 weeks, then given basal diet containing 0.05% PB or 5% SS for the next 32 weeks, and then killed. Appropriate control studies were also done. Histological examination of whole organs of the rats showed that PB promoted thyroid carcinogenesis whereas SS did not. A significant increase in the incidences of total tumors as well as papillary adenocarcinoma of the thyroid was observed in the group given PB after NMU (p less than 0.001). The incidence of papillary adenoma with or without papillary adenocarcinomas was also high in the NMU-PB-treated group. The organ-specific promoting effect of PB in the induction of preneoplastic lesions, as demonstrated by development of gamma-glutamyl transpeptidase-positive foci in the liver and of SS in papillary or nodular hyperplasia in the urinary bladder, as reported previously, was also confirmed. The incidences of papillomas in the forestomach were similar in groups treated with NMU-PB, NMU-SS, or NMU alone. The results indicate that PB is a tumor promoter in the liver and thyroid and that SS is a tumor promoter in the urinary bladder of rats.

Induction of mammary tumors, estrous cycle abnormalities and endometrial hyperplasia in rats exposed to different doses of N-nitrosomethylurea.

Groups of female Sprague--Dawley strain rats were given 3 i.v. injections of N-nitrosomethylurea in doses of 0.5, 1, 2, 3, 4 or 5 mg/100 g body weight at 4-week intervals. The first dose was given when they were 50 days old. By 23 weeks after the first injection, mammary tumors had developed in 0, 0, 33, 54, 72 and 100% of animals respectively. There was a direct relationship between the total dose of carcinogen administered and the degree of tumor anaplasia observed on histological examination. All of the tumors contained assayable amounts of estrogen and progesterone receptors, and the receptor concentrations were not related to the dose of carcinogen. Twenty-one rats, all exposed to the 4 highest doses of N-nitrosomethylurea, had arrest of the estrous cycle at the stage of estrus. In 15 of the 21 the walls of the uterine horns were thickened and grossly distended by fluid. Histological examination demonstrated the presence of endometrial hyperplasia. These uterine abnormalities were usually accompanied by polycystic disease of the ovaries. Both endometrial hyperplasia and abnormal estrous cycles without uterine changes were associated with elevated progesterone receptor to estrogen receptor ratios in the corresponding mammary carcinomas.

The induction of immunologic tolerance in guinea pigs infused with dimethylbenz(a)anthracene.

The infusion of 7,12 dimethylbenz(a)anthracene (DMBA) dissolved in DMSO:glycofurol into adult guinea pigs rendered most recipients unresponsive to the induction of contact sensitization with DMBA, either by injection in adjuvant or topical application. The results were similar when guinea pigs infused with DMBA dissolved in Upjohn fat emulsion, were challenged with DMBA in adjuvant, but studies attempting topical sensitization to demonstrate unresponsiveness were inconclusive. Mammary tumors appeared after a latent period of 8 to 11 mo in 4 guinea pigs infused with DMBA in fat emulsion in which topical sensitization with DMBA was attempted.