These data suggested that establishment of a suitable biomarker panel would facilitate detection of site-specific kidney injury with high sensitivity.

No abstract

Hepatocellular carcinoma which occurred in one male laboratory cynomolgus monkey at 5 years of age was investigated extensively by immunological, histopathological and electron-microscopic examinations. The animal exhibited no abnormalities in clinical laboratory tests including blood-chemistry, except for elevation of protein induced by vitamin K absence or antagonist II (PIVKA-II) in serum, which is one of the hepatic tumor markers. In virus antibody tests, the serum was positive for hepatitis A virus. On necropsy, a mass measuring 60 × 60 × 65 mm was seen in the hepatic left lobe. The tumor mass had a lobulated structure with hemorrhagic and necrotic areas, and was demarcated by thin fibrous capsules. Histopathologically, the tumor was composed of hepatocyte-like cells, having irregular trabecular structure with various thickness, lined by vascular endothelial cells. Cellular atypia such as polynucleated cells, mitotic figures, and invasion into the vascular cavity were also observed. Immunohistochemically, the tumor cells showed positive reaction for anti-PIVKA-II, anti-epithelial membrane antigen (EMA), anti-carcinoembryonic antigen (CEA), and anti-cytokeratin 18, as well as for proliferating cell nuclear antigen (PCNA). On electron-microscopic examination, the tumor cells had a number of tight junctions and formations of bile canaliculi between adjacent cells, basically resembling hepatocytes. This is the first case of hepatocellular carcinoma with PIVKA-II production in monkeys. Serological and immunohistochemical analyses for PIVKA-II are, therefore, practicable for diagnose as hepatocellular carcinoma in nonhuman primates. (J Toxicol Pathol 2002; 15: 61-68)

To facilitate development of the androgen receptor inhibitor, enzalutamide, we developed and validated methods for simultaneous determination of enzalutamide, its carboxylic acid metabolite (M1), and N-desmethyl enzalutamide (M2) in plasma from mice, rats, and dogs, and brain from mice. Following addition of stable isotope-labeled internal standards, plasma and brain samples (0.05 and 0.3 mL, respectively) were mixed with 0.1% formic acid and extracted with tert-butyl methyl ether, and then injected into a liquid chromatography-tandem mass spectrometry system. The eluent was monitored in positive electrospray ionization mode. To bracket the concentrations of the drug and metabolites in study samples, calibration curves were constructed from 20 to 50000 ng/mL for plasma and 10 to 25000 ng/g for brain. Validation data demonstrated that these methods were selective, reproducible, and accurate. Using these methods, brain-to-plasma concentration ratios in mice were determined to be 0.72 for enzalutamide, 0.048 for M1, and 1.4 for M2.

-It is well-known that indomethacin (the cyclooxygenase 1 & 2 inhibitor) and RU486 (or mifepristone, the progesterone receptor antagonist) block follicular rupture in rats. To characterize genetic alterations in unruptured follicles, gene expression profiles in ovarian follicle were analyzed in indomethacin-and RU486-treated female Sprague-Dawley rats. Ovaries are collected at 22:00 on the proestrus day and 10:00 on the following estrus day after a single dose of indomethacin and RU486. Histopathologically, changes depicting responses to LH surge were observed in ovaries, uteri and vagina. Total RNA was extracted from pre-ovulatory follicles or unruptured follicles collected by laser microdissection and analyzed by Genechip ® . Among genes showing statistically significant changes compared to control groups, following changes were considered relevant to induction of unruptured follicles. In indomethacin-treated rats, Wnt4 was down-regulated, suggesting effect on tissue integrity and steroid genesis. In RU486-treated rats, Adamts1, Adamts9, Edn2, Ednra, Lyve1, Plat, and Pparg were down-regulated. These changes suggest effects on proteolysis for extra cellular matrix or surrounding tissue (Adamts1 & 9, and Plat), constriction of smooth muscle surrounding follicles (Edn2, Ednra, and Pparg), follicular fluid (Lyve1), and angiogenesis (Pparg). Down-regulation of angiogenesis related genes (Angpt2, Hmox1, and Vegfa) was observed in both treatment groups. Here, we clarify genetic alterations induced by the inhibition of cyclooxygenase or progesterone receptor.

No abstract

Evaluation of ovarian toxicity requires an understanding of the physiological changes related to the estrous cycle in the ovary. The authors investigated the transitional gene expression profile of ovulatory follicles in rats that show normal estrous cyclicity. Ovaries were collected at 10:00 and 22:00 on the proestrus day and at 10:00 on the estrus day. Ovarian follicles or early corpora lutea were isolated using laser microdissection, and extracted total RNA was analyzed using microarray technology. Clustering analysis revealed four different expression patterns: transient up-or down-regulation only at 22:00 on the proestrus day (pattern 1), up-or down-regulation only at 10:00 on the estrus day (pattern 2), continuous increase at 22:00 on the proestrus day and at 10:00 on the estrus day (pattern 3), and up-or down-regulation at 22:00 on the proestrus day and level maintenance at 10:00 on the estrus day (pattern 4). In addition, these probe sets were functionally categorized in each pattern using the Ingenuity Pathways Analysis database. These data will aid in understanding the physiology of ovulation and may be useful in assessing ovarian toxicity and its mechanism, such as in investigations of chemical-induced ovulatory impairment.

The pharmacokinetic endpoint of a 25-fold increase in human exposure is one of the specified criteria for highdose selection for 2-year carcinogenicity studies in rodents according to ICH S1C(R2). However, this criterion is not universally accepted for 6-month carcinogenicity tests in rasH2-Tg mice. To evaluate an appropriate multiple for rasH2-Tg mice, we evaluated data for 53 compounds across five categories of rasH2-Tg mouse-positive [(1) genotoxic and (2) non-genotoxic] carcinogens and rasH2-Tg mouse-negative [(3) non-genotoxic carcinogens with clear or uncertain human relevance; (4) non-genotoxic rodent-specific carcinogens; and (5) non-carcinogens], and surveyed their tumorigenic activities and high doses in rasH2-Tg mice and 2-year rodent models.Our survey indicated that Area Under the Curve (AUC) margins (AMs) or body surface area-adjusted dose ratios (DRs) of tumorigenesis in rasH2-Tg mice to the maximum recommended human dose (MRHD) were 0.05-to 5.2fold in 6 category (1) compounds with small differences between models and 0.2-to 47-fold in 7 category (2) including three 2-year rat study-negative compounds. Among all 53 compounds, including 40 compounds of the rasH2-Tg mouse-negative category (3), (4), and (5), no histopathologic risk factors for rodent neoplasia were induced only at doses above 50-fold AM or DR in rasH2-Tg mice except for two compounds, which induced hyperplasia and had no relationship with the tumors observed in the rasH2-Tg mouse or 2-year rodent studies.From the results of these surveys, we confirmed that exceeding a high dose level of 50-fold AM in rasH2-Tg mouse carcinogenicity studies does not appear to be of value.