Robert L. Morrissey

Difluoromethylornithine in combination with tamoxifen in female rats: 13-week oral toxicity study.

Purpose: Cancer chemoprevention is the use of pharmacologic or natural agents to inhibit the development of cancer. Difluoromethylornithine (DFMO) is an irreversible inhibitor of ornithine decarboxylase, the rate-limiting enzyme in the biosynthesis of polyamines. DFMO has demonstrated chemopreventive efficacy in animal models of tumorigenesis. Tamoxifen (TAM) is currently used for treatment of estrogen receptor-positive breast carcinoma and has demonstrated efficacy in chemoprevention of breast cancer in women at high risk for the disease. The administration of tamoxifen with DFMO is being considered for development by the National Cancer Institute as a potential drug regimen for the chemoprevention of breast carcinoma. Methods: The toxicity of DFMO in combination with TAM was evaluated in female rats following 13 weeks of daily administration by gavage. Dose groups were vehicle control, DFMO (1000 mg/kg per day), low TAM (0.25 mg/kg per day), high TAM (2.5 mg/kg per day), low combination (1000 + 0.25) and high combination (1000 + 2.5). Results: No mortalities occurred in the study. Clinical signs of toxicity were limited to dermal lesions consisting of scab formation and abrasions produced by DFMO. Administration of either DFMO or TAM resulted in decreased body weight gains, with coadministration having an additive effect. Serum albumin, total protein, cholesterol and triglyceride levels were decreased in all drug-treated dose groups, although histologic evidence of liver lesions were not seen. TAM resulted in increased numbers of red blood cells, whereas DFMO produced a slightly anemic response. DFMO produced lesions in the small intestine consisting of necrosis of crypt epithelium and crypt microabscess, which were enhanced by TAM coadministration. Administration of TAM resulted in histologic changes in the ovaries, fallopian tube, vagina, cervix and uterus, indicating that inhibition of ovulation and reproductive cycle arrest in the proestrus stage had occurred. Coadministration with DFMO did not affect the changes to the reproductive system induced by TAM. Conclusions: Coadministration of DFMO with tamoxifen did not result in toxicity unique to the combination drug regimen, but rather toxicity resulted from administration of each drug. Under the conditions of the study, the overall toxicity produced by dual administration of DFMO with tamoxifen was additive with respect to the toxicity associated with each agent alone.

Oncogenicity Evaluations of Chemopreventive Soy Components in p53(+/–) (p53 knockout) Mice

Epidemiologic data suggest that soy consumption may protect against cancer induction in several tissues in humans. Although the soy components responsible for this activity remain unidentified, isoflavones (e.g., genistein) and protease inhibitors (e.g., Bowman-Birk inhibitor complex [BBIC]) demonstrate chemopreventive activity in several animal cancer models. As part of their preclinical development for cancer prevention, PTI G-2535 (a soy isoflavone mixture containing 45% genistein, 23% daidzein, and 4% glycitein) and BBIC were evaluated for oncogenicity in p53(+/–) mice. In separate studies, groups of 25 p53(+/–) mice/sex received daily gavage exposure to PTI G-2535 (0, 250, 1000, or 2500 mg/kg/day) or BBIC (0, 500, 1000, or 2000 mg/kg/day) for 6 months. The high doses of both PTI G-2535 and BBIC were limited by viscosity. p-Cresidine (400 mg/kg/day) served as a positive-control article in both studies. PTI G-2535 induced no gross toxicity in any animal, but did induce a dose-related suppression of body weight gain in male mice. Modest hematologic alterations and increased liver and spleen weights were seen in both sexes exposed to the isoflavone mixture. BBIC had no significant effect on body weight, food consumption, clinical pathology, or organ weights in either sex. Histopathologic evaluations demonstrated no increases in the incidence of either benign or malignant tumors in any group of p53(+/–) mice exposed to PTI G-2535 or to BBIC. By contrast, the positive-control article, p-cresidine, induced urinary bladder cancers in both studies. Neither PTI G-2535 nor BBIC demonstrates any evidence of oncogenicity in the p53(+/–) mouse model.

Thirteen-week oral toxicity study of difluoromethylornithine in combination with tamoxifen citrate in female dogs

Purpose: Cancer chemoprevention is the use of pharmacologic or natural agents to inhibit the development of cancer. Difluoromethylornithine (DFMO) is an irreversible inhibitor of ornithine decarboxylase, the rate-limiting enzyme in the biosynthesis of polyamines. DFMO has demonstrated chemopreventive efficacy in animal models of tumorigenesis. Tamoxifen (TAM), a nonsteroidal antiestrogen, is approved for use in the treatment of estrogen receptor-positive breast carcinoma and has demonstrated efficacy in chemoprevention of breast cancer in women at high risk for the disease. The administration of TAM with DFMO is being considered for development by the National Cancer Institute as a potential drug regimen for the chemoprevention of breast carcinoma. Methods: The toxicity of DFMO in combination with TAM was evaluated in female Beagle dogs following 13 weeks of daily oral administration by capsule. Dose levels in milligrams per kilogram body weight per day were: 0 (vehicle control), 100 DFMO, 0.1 TAM, 1.0 TAM, 0.1 TAM + 100 DFMO and 1.0 TAM + 100 DFMO. Results: No mortalities occurred. Diarrhea was produced by TAM and vaginal discharge, due to reproductive tract lesions, was produced by both DFMO and TAM, either alone or in combination. DFMO decreased reticulocyte counts and TAM increased counts of mature neutrophils. DFMO alone resulted in lesions to the intestines and ovaries, and cornified epithelium of vagina and cervix. TAM produced cornified epithelium of vagina and cervix, and numerous lesions in the ovaries, fallopian tube, uterus, cervix and vagina which were likely due to an estrogen agonist effect. Coadministration of DFMO increased the incidence and/or severity of these reproductive tract lesions. Each compound alone produced ovarian atrophy, and antral follicles and corpora lutea were completely absent in the 1.0 TAM + 100 DFMO group. Conclusions: Coadministration of DFMO and TAM resulted in additive toxicity involving the female reproductive system.

Oral Toxicity of 1,2-Dithiole-3-Thione, a Potential Cancer Chemopreventive Agent, in the Rat

This study examined the toxicity of 1,2-dithiole-3-thione (D3T) in rats following 14 days of daily oral (gavage) administration. D3T, an extensive inducer of hepatic phase II drug-metabolizing enzymes, has demonstrated cancer preventive efficacy in rodent models of tumorigenesis and is a candidate drug for cancer prevention. Male and female CD rats (5/sex/dose group) received D3T at dose levels of 0 (corn oil vehicle control), 2, 6, 20, and 60 mg/kg/day. Oral administration of D3T for 14 days at 20 mg/kg/day resulted in decreased activity, lethargy, rough coat, and piloerection, and toxicologically significant lesions in the stomach, characterized by apoptotic necrosis and hyperplasia of the glandular mucosa. Administration of D3T at 60 mg/kg/day produced anemia in females, decreased body weight gain in males, and increased vacuolation of adrenal cortical cells. Increased liver weights, vacuolation of hepatocytes, and serum chemistry changes, indicative of altered liver function, were observed at 6 mg/kg/day, which were likely due to a pharmacologic effect of D3T on the liver and not considered to be toxicologically significant. Under the conditions of the study, the no-observed-adverse effect level (NOAEL) was 6 mg/kg/day.

Subchronic studies of pyrilamine in B6C3F1 mice

The purpose of this study was to determine the effects of subchronic exposure to the antihistamine pyrilamine maleate and to establish dosage levels to use in a 2-year chronic study. A 14-day repeated-dose study in male and female B6C3F1 mice at dietary levels of pyrilamine (as the free base) of 0, 196, 392, 783, 1563, or 3122 ppm revealed no influence on body weight gain and resulted in no deaths, nor abnormal clinical or gross necropsy observations. Parotid gland cell necrosis was more apparent in high-dosage groups than in controls of either gender. Male and female B6C3F1 mice also were administered pyrilamine for 90 days at dietary concentrations of 0, 375, 750, 1500, 3000, or 6000 ppm. Weight gain was markedly suppressed in the 6000 ppm group, but less so at 1500 and 3000 ppm. Parotid gland cell necrosis was apparent in males receiving 750 ppm pyrilamine and above and in females at 1500 ppm and above. Cytomegaly was noted in the parotid glands of both sexes at midlevel dosages but not in control, low-dosage, or high-dosage animals. It was concluded that 1500 ppm pyrilamine would not be life-threatening to B6C3F1 mice in a chronic study.

Subchronic studies of pyrilamine in Fischer 344 rats

This study was designed to determine the subchronic effects of the antihistamine, pyrilamine maleate, and to establish dose levels for a 2-year chronic study. Six male and six female F344 rats received dietary pyrilamine concentrations of 0, 392, 783, 1563, 3122, or 6276 ppm (as the free base) for 14 days. Only weight gain suppression was noted, at 3122 and 6276 ppm, especially in males. When 12 male and 12 female F344 rats were fed pyrilamine for 90 days at 0, 750, 1500, 3000, 6000, or 12,000 ppm, weight gain was suppressed and organ/body weight ratios often were higher than in controls at 3000 ppm and above. Heart/brain and thymus/brain ratios were decreased at 3000 ppm and above in males and at 6000 or 12,000 ppm in females. All rats receiving 6000 or 12,000 ppm pyrilamine exhibited severe diffuse parenchymal cell cytomegaly in the parotid salivary glands with a dose-related decrease in severity and prevalence at lower concentrations. Hepatocellular vacuolization, present only in males, increased in severity with dose, starting at 750 ppm. The severity of liver fatty metamorphosis increased with dose at the three low doses but was less apparent and/or disappeared at 6000 and 12,000 ppm. It was concluded that 3000 ppm pyrilamine would not be life threatening to F344 rats in a chronic study.

Chronic Toxicity/Oncogenicity Studies of 60 Hz Magnetic Fields in F344 Rats and B6C3F1 Mice: Final Survival, Body Weight, Clinical Observation, and Gross Pathology Data

The results of a number of epidemiology studies have suggested a positive association between exposure to power frequency magnetic fields and the incidence of several types of malignancy1–3. However, other epidemiology studies of similar design have found no such relationship between magnetic field exposure and cancer risk4,5. Because the epidemiology database for magnetic field exposure and cancer risk is contradictory, it appears unlikely that definitive assessments of human risk associated with magnetic field exposure can be developed on the basis of epidemiology data alone. In cases where the epidemiology data are inconclusive, the importance of studies in experimental animal model systems increases. Well-controlled animal studies permit evaluation of the biological effects of magnetic fields in vivo under tightly controlled exposure conditions, and in the absence of potential confounding variables. The present report provides the results of the in-life component of studies that are designed to evaluate the chronic toxicity and potential oncogenicity of chronic exposure to 60 Hz magnetic fields in F344 rats and B6C3F1 mice.