4-week Repeated Dose Toxicity Study Research Articles

Gene expression analysis of antioxidant and DNA methylation on the rat liver after 4-week wood preservative chromated copper arsenate exposure.

Our previous 4-week repeated dose toxicity study showed that wood preservative chromated copper arsenate (CCA) induced hepatocellular hypertrophy accompanied by biochemical hepatic dysfunction and an increase in oxidative stress marker, 8-hydroxydeoxyguanosine, in female rats. To further explore the molecular mechanisms of CCA hepatotoxicity, we analyzed 10%-buffered formalin-fixed liver samples from female rats for cell proliferation, apoptosis, and protein glutathionylation and conducted microarray analysis on frozen liver samples from female rats treated with 0 or 80 mg/kg/day of CCA. Chemical analysis revealed that dimethylated arsenical was the major metabolite in liver tissues of male and female rats. CCA increase labeling indices of proliferating cell nuclear antigen and decrease terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling accompanied with increased expression of protein glutathionylation, indicating a decrease in glutathione (GSH) in hepatocytes of female rats. Microarray analysis revealed that CCA altered gene expression of antioxidants, glutathione-S-transferase (GST), heat shock proteins and ubiquitin-proteasome pathway, cell proliferation, apoptosis, DNA methylation, cytochrome P450, and glucose and lipid metabolism in female rats. Increased expression of GSTs, including Gsta2, Gsta3, Mgst1, and Cdkn1b (p27), and decreased expression of the antioxidant Mt1, and DNA methylation Dnmt1, Dnmt3a, and Ctcf were confirmed in the liver of female rats in a dose-dependent manner. Methylation status of the promoter region of the Mt1 was not evidently changed between control and treatment groups. The results suggested that CCA decreased GSH and altered the expression of several genes, including antioxidants, GST, and DNA methylation, followed by impaired cell proliferation in the liver of female rats.

Abstract 1139: A novel HER2/4-1BB bispecific antibody, YH32367 (ABL105) exhibits the optimal efficacy and superior safety profile through tumor-directed 4-1BB agonism

Abstract Although HER2-targeted therapies such as Herceptin® (trastuzumab) have dramatically improved outcomes for solid cancer patients with HER2 overexpression, it remains a challenge to cure the HER2 positive cancer patients with resistance to current HER2-targeted therapies. Therefore, new approaches need to treat them. One approach would be to combine with immunotherapy. 4-1BB (CD137) is a key costimulatory receptor expressed on activated T-cells and natural killer (NK) cells and a promising therapeutic target in cancer. In this study, we present the results from a preclinical study on the efficacy and safety of HER2/4-1BB bispecific antibody, YH32367 (ABL105). YH32367 has been designed to overcome the challenges with HER2 resistance via tumor-directed 4-1BB agonism. Its activity was determined using cell-based 4-1BB bioassay and co-culture assay with human peripheral blood mononuclear cells (hPBMC). In vivo anti-tumor efficacy of YH32367 was assessed in both HER2-expressing tumor-bearing hPBMC humanized mice and h4-1BB knock-in mice. In addition, 4-week repeated dose toxicity study of YH32367 was conducted in cynomolgus monkeys at the dose of 10, 30 and 100 mg/kg. YH32367 stimulated IFN-γ secretion and thereby eliciting tumor cell lysis in co-culture assay with hPBMC and HER2-expressing tumor cells. YH32367 also showed a long-term tumor immunity as well as superior efficacy on tumor eradication in MC38/hHER2-bearing h4-1BB knock-in mouse models, compared to the equimolar dosing of trastuzumab and benchmark 4-1BB agonistic antibody. Even a single dosing of YH32367 at the doses of 1, 3 and 10 mg/kg elicited complete tumor regression in more than 80% of mice bearing MC38/hHER2. In GLP 4-week toxicity study in monkeys, there were no YH32367-related effects on mortality, body weight, hematology, clinical chemistry, urinalysis, microscopic observations, cardiovascular system and immunotyping/cytokine analysis in any doses, indicating a favorable safety profile. In conclusion, YH32367 exhibited potent in vitro and vivo effects via tumor immunity and it was well tolerated and safe in the 4-week toxicity study. These results suggest that YH32367 could be a promising therapeutic for HER2-positive cancer patients, especially with HER2 drug resistance as an effective HER2-targeted therapy. Phase I clinical study is expected to begin in 2022. Citation Format: Eunjung Lee, Hyejin Chung, Yangsoon Lee, Eun-jung Lee, Young Bong Park, Ju Young Park, Sujin Ahn, Junhwan Kim, Kyoung Kyu Ahn, Kyeongsu Park, Wonjun Son, Donghoon Yeom, Jaeho Jung, Jonghwa Won, Se-Woong Oh. A novel HER2/4-1BB bispecific antibody, YH32367 (ABL105) exhibits the optimal efficacy and superior safety profile through tumor-directed 4-1BB agonism [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1139.

First-in-class monoclonal antibody (mAb) PBP1510 targeting pancreatic adenocarcinoma upregulated factor (PAUF) for pancreatic cancer (PC) treatment: Preclinical perspectives.

e16274 Background: PAUF has been shown to induce PC cell proliferation, angiogenesis and evasion of immune surveillance, thereby promoting tumour progression and metastasis. PBP1510, a novel humanized IgG1 mAb developed by Prestige Biopharma Limited, Singapore, binds to and neutralizes PAUF with high specificity and affinity. With no available targeted therapies against PAUF, the first in its class, PBP1510, has been granted Orphan Drug Designation by EMA, US FDA and Korea MFDS. Methods: To assess inhibition of migration and invasion of PC in vitro, PC cell lines (CFPAC-1 and BxPC-3) with high expression levels of PAUF, and primary PC cells extracted from patients were treated with PBP1510 or human IgG (negative control). Suitability of cynomolgus monkey as animal model for pharmacokinetic (PK) and toxicity studies was verified by comparing binding affinities of PBP1510 to human and monkey PAUF. PK characteristics and immunogenicity of PBP1510 (up to 20 mg/kg), were examined in single dose studies in 6 monkeys. A 4-week repeated dose toxicity study was also conducted in 32 monkeys with PBP1510 (up to 40 mg/kg). An in vitro tissue cross-reactivity study was also conducted in a full panel of normal human tissues, to assess potential toxicity in humans. Results: PBP1510 suppressed the proliferation of the PC cell lines compared to negative control. The migration and invasion of PBP1510 treated PC cell lines as well as the primary tumour cells from patients was significantly reduced compared to the respective control groups in vitro. Similarity in binding affinity of PBP1510 to human PAUF and monkey PAUF, and amino acid sequence homology between human and monkey PAUF, suggest that monkey is an appropriate animal species for toxicity and PK studies of PBP1510. In monkey studies, a dose-proportional increase in systemic exposure was observed at low and high doses. PBP1510 at doses up to 40 mg/kg was well tolerated by study animals and no notable local or systemic toxicity was observed. Low immunogenic profile of PBP1510 was indicated by an absence of anti-PBP1510 antibodies till up to 57 days (8 doses and 28-day recovery period). These data are used to simulate human PK profile and to derive the starting dose and maximum escalated dose for the upcoming Phase 1 clinical studies. Cross-reactivity results did not reveal non-specific binding of PBP1510 in the human tissues except in the cytoplasm of acinar and duct epithelium in the salivary gland which is not expected to be of major toxicologic significance. Conclusions: The efficacy and safety data presented here, along with the data from in vivo mouse studies demonstrating superior anti-tumour activity of PBP1510 treatment, support further clinical development of PBP1510 as a novel anti-cancer agent to treat PAUF-positive PC. The pivotal step of advancing PBP1510 into initial human studies is now in progress.

Evaluation of acute and subacute toxicity of sodium taurodeoxycholate in rats

Taurodeoxycholate (TDCA) inhibits various inflammatory responses suggesting potential clinical application. However, the toxicity of TDCA has not been evaluated in detail in vivo. We investigated the acute toxicity and 4-week repeated-dose toxicity of TDCA following intravenous infusion under Good Laboratory Practice regulations. In the sighting study of acute toxicity, one of two rats (one male and one female) treated with 300 mg/kg TDCA died with hepatotoxicity, suggesting that the approximate 50% lethal dose of TDCA is 300 mg/kg. Edema and discoloration were observed at the injection sites of tails when rats were infused with 150 mg/kg or higher amount of TDCA once. In 4-week repeated-dose toxicity study, no treatment-related mortality or systemic changes in hematology and serum biochemistry, organ weights, gross pathology, or histopathology were observed. However, the tail injection site showed redness, discharge, hardening, and crust formation along with histopathological changes such as ulceration, edema, fibrosis, and thrombosis when rats were infused with 20 mg/kg TDCA. Taken together, TDCA induced no systemic toxicity or macroscopic lesions at the injection site at a dose of 10 mg/kg/day, which is 33 times higher than the median effective dose observed in a mouse sepsis model. These findings suggest that TDCA might have a favorable therapeutic index in clinical applications.

Nonclinical toxicology studies with sodium taurodeoxycholate: acute and subacute toxicity in dogs

Sodium taurodeoxycholate (TDCA) has been investigated for various inflammatory disorders such as sepsis. We recently evaluated nonclinical safety profile of TDCA using rats infused intravenously. As a series of preclinical safety investigations, we further conducted toxicity studies with TDCA delivered to dogs via intravenous administration under Good Laboratory Practice regulation in this study. In dose range-finding study (dose escalation study), dogs given with TDCA at a dose of 150 mg/kg showed marked changes in clinical signs, hematology, and serum biochemistry. And biochemical markers of liver damage and local skin lesions were observed following intravenous infusion of 100 mg/kg TDCA, suggesting that 100 mg/kg was chosen as the highest dose of TDCA for 4-week repeated-dose toxicity study using dogs. Despite no treatment-related significant changes in body weight, food consumption, ophthalmoscopy, and urinalysis, skin lesions were observed at the injection site of animals administered with higher than 50 mg/kg of TDCA along with biochemical and histopathological changes associated with liver injury. However, most of off-target effects were found to be reversible since these were recovered after stopping TDCA infusion. These findings indicate that the no-observed-adverse-effect-level (NOAEL) for TDCA in dogs was considered to be 5 mg/kg/d. Taken together, our results provide important toxicological profiles regarding the safe dose of TDCA for drug development or clinical application.

Safety assessment of Gyejibokryeong-hwan water extract: Study of acute and subacute toxicity, and influence on drug metabolizing enzymes

Ethnopharmacological relevanceGyejibokryeong-hwan is a traditional herbal medicine and is reported to have various pharmacological actions. Despite many reports of previous studies, there is limited scientific evidence concerning its safety and few drug-metabolism profiles to support the continued therapeutic application of Gyejibokryeong-hwan. Aim of the studyThe purpose of the present study was to investigate the acute and subacute toxicity profile of a Gyejibokryeong-hwan water extract (GBHW) in vivo, and its effects on the activities of drug-metabolizing enzymes in vitro. Materials and methodsAcute and subacute toxicity was evaluated by giving GBHW to rats. In a study of acute toxicity, the rats were given GBHW by single oral gavage administration at 0 and 5000 mg/kg. In a study of subacute toxicity, rats were given GBHW by oral gavage at 0, 1000, 2000, and 5000 mg/kg/day daily for 28 days. The activities of the major human microsomal cytochrome P450 (CYP450) and UDP-glucuronosyltransferase (UGT) isozymes were investigated using fluorescence- and luminescence-based enzyme assays in vitro, respectively. ResultsGBHW did not cause any mortality in the study of acute toxicity. In the study of subacute toxicity, GBHW at more than 2000 mg/kg/day was observed with minor changes in the absolute and relative organ weight, hematology, serum biochemistry and urinalysis parameters in rats of either sex. However, these changes were not considered to be important toxicologically. GBHW moderately inhibited the activities of CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2E1, CYP3A4, and UGT1A1. ConclusionsOur present data suggest that GBHW does not cause toxicologically important adverse events at doses up to 2000 mg/kg/day in the 4-week repeated dose toxicity study and provide valuable information concerning its potential to interact with conventional medicine.

Comparisons of l-cysteine and d-cysteine toxicity in 4-week repeated-dose toxicity studies of rats receiving daily oral administration.

Two 4-week repeated-dose toxicity studies were conducted to evaluate the potential toxicity of l-cysteine and d-cysteine. In one study, three groups of 6 male rats were each administered l-cysteine once daily by gavage at doses of 500, 1,000, or 2,000 mg/kg/day for 28 consecutive days. The control group was administered a 0.5% methylcellulose vehicle solution. The other study followed a similar protocol except that the experimental groups received d-cysteine. Toxicological observations showed that the l-cysteine-treated groups exhibited renal injuries such as basophilic tubules with eosinophilic material in the lumen, and there were increased numbers of basophilic tubules in all treated groups. In 1,000 or 2,000 mg/kg/day-treated groups, salivation and necropsy findings indicative of focal erosion in the stomach mucosa were found. Increases in reticulocyte counts were observed in the 2,000 mg/kg/day-treated group. Toxicological findings obtained for the d-cysteine-treated groups included anemia and renal injuries such as basophilic tubules with eosinophilic material in the lumen, increased numbers of basophilic tubules, and crystal deposition in the medulla in the 2,000 mg/kg/day-treated group. Additional findings included sperm granuloma in the epididymis, necropsy findings suggestive of focal erosion in the stomach mucosa, and salivation in the 1,000 or 2,000 mg/kg/day-treated groups. One rat in the 2,000 mg/kg/day-treated group died due to renal failure. In conclusion, the no-observed-adverse-effect levels (NOAELs) were estimated to be less than 500 mg/kg/day for l-cysteine and 500 mg/kg/day for d-cysteine under our study conditions. The toxicological profiles were similar for l-cysteine and d-cysteine; however, there were slight differences in the dose responses. The mechanisms underlying these differences remain to be determined.

Safety assessment of melinjo (Gnetum gnemon L.) seed extract: Acute and subchronic toxicity studies

Melinjo (Gnetum gnemon L.) is widely cultivated in Southeast Asia. Its fruit and seeds are common ingredients in Indonesian foods. The seeds are very rich in resveratrol dimers such as gnetin C and its glucosides, gnemonoside A and gnemonoside D, and also contain trans-resveratrol and its glucoside, trans-piceid. The safety of melinjo seeds is assured, since people in Southeast Asia have consumed them for a long time; however, their safety has not been scientifically verified. In this study, the safety of melinjo seed extract (MSE) powder was assessed in an acute oral toxicity study, a 4-week repeated dose toxicity study, and in a micronucleus test in rats. In the acute and subchronic toxicity studies, the group administered the powder did not show any toxicologically significant MSE-related changes, compared with the control group. The no observed adverse effect level (NOAEL) was determined as 1000 mg/kg/day. A genotoxicity test (rat bone marrow micronucleus test) was negative for MSE powder at levels up to 4000 mg/kg/day. These results might provide supportive evidence of safety of melinjo seeds, which has been used as food ingredients for a long time.

Spontaneous Micronucleus Formation in an Integrated Bone Marrow Micronucleus Assay in Wistar Han Rats

Multiple end-point in vivo assays can alleviate the need for multiple repeated in vivo assays, thereby reducing time, the costs involved and extensive use of laboratory animals in targeted preclinical experiments. The International conference on harmonization (Ich) guideline S2 (R1) recommends the use of the integrated micronucleus study. One option is to integrate the micronucleus end-points into a repeat dose study. a 4-week repeated dose toxicity study is one of the approaches often included in a preclinical toxicity study schedule. Thus, ten 4-week repeated dose toxicity studies were conducted in a Wistar han rat model that included integrated micronucleus end-point determinations. Those studies used common vehicles and determined different routes of administration that permitted the collection of background data of spontaneous micronucleus formation. The studies conformed to standard operation procedures (SOP), respective study protocols of the testing facility, and related regulatory guidelines. The micronucleus background data obtained from the Wistar han rat model was compared to data obtained from Sprague Dawley rats in an acute exposure design. It was concluded that micronucleus background responses were very similar between the Wistar han (repeated-dose design) and Sprague Dawley rats (acute dose design). These background data will serve as reference values for future integrated micronucleus studies. We also reported particular clinical pathology parameters (e.g., hematology, coagulation and biochemistry) and the relative organ weight data collected from the animals that participated in these studies. Journal of Environmental Immunology and Toxicology 2014; 1:159-166

A 4-week Repeated Dose Toxicity Study of Glycine in Rats by Gavage Administration

In order to examine the toxicity profile of glycine, an authorized food additive, a solution of glycine in water for injection was administered orally (via gavage) to male SD rats (Crl:CD(SD)) once daily for 4 weeks at doses of 500, 1000 and 2000 mg/kg/day in a volume of 10 mL/kg. Control animals received vehicle only. No animals died, and no glycine-related changes were observed in body weight, food consumption, water consumption, hematology, organ weight, gross pathological examination or histopathological examination. In urinalysis, daily urinary volume and urinary Cl excretion were significantly higher in the 2000 mg/kg/day dose group, and urine pH and urinary protein showed lower trends in the glycine-treated groups. However, these changes were considered to be of little toxicological significance, because there were no histopathological changes in the kidneys or urinary bladder and no changes in other urinary parameters. As regards blood chemistry, phospholipids were significantly higher in the 2000 mg/kg/day dose group. However, the increase was small and was not considered to be toxicologically significant. In conclusion, none of the animals in any of the glycine-treated groups showed changes that were considered toxicologically significant. Therefore, the no-observed-adverse-effect level of glycine was estimated to be at least 2000 mg/kg/day under the conditions of this study.

Collaborative work on evaluation of ovarian toxicity 15) Two- or four-week repeated-dose studies and fertility study of bromocriptine in female rats

The main focus of this study is to determine the optimal administration period concerning toxic effects on ovarian morphological changes in a repeated-dose toxicity study. To assess morphological and functional changes induced in the ovary by bromocriptine, the compound was administered to female rats at dose levels of 0, 0.08, 0.4 and 2 mg/kg for the 2- or 4-week repeated-dose toxicity study, and for the female fertility study from 2 weeks prior to mating to day 7 of gestation. In the 2-week repeated-dose toxicity study, increase of ovarian weights was observed at 2 mg/kg. In the 4-week repeated-dose toxicity study, ovarian weights were increased at 0.4 and 2 mg/kg. The number of corpora luteum was increased in the 0.4 and 2 mg/kg groups of the 2- and 4-week repeated-dose toxicity studies by histopathological examination of the ovaries. Bromocriptine did not affect estrous cyclicity in 2- and 4-week repeated dosing. In the female fertility study, although animals in any groups mated successfully, no females in 0.4 and 2 mg/kg groups were pregnant. There were no adverse effects on reproductive performance in the 0.08 mg/kg group. Based on these findings, the histopathological changes in the ovary are considered important parameters for evaluation of drugs including ovarian damage. We conclude that a 2-week administration period is sufficient to detect ovarian toxicity of bromocriptine in a repeated-dose toxicity study.

Collaborative work on evaluation of ovarian toxicity by repeated-dose and fertility studies in female rats

The National Institute of Health Sciences (NIHS) and 18 pharmaceutical companies of the Japan Pharmaceutical Manufacturers Association (JPMA) have conducted a validation study intended to evaluate whether a 2-week repeated general toxicity period with histopathological examination is sufficient to detect ovarian toxicity or not. The current repeated dose general toxicity study is considered to be insufficient in terms of evaluating female reproductive function due to a lack of evidence indicating that it is adequate. Evaluation of ovarian toxicity by comprehensive histopathological examination of the female reproductive organs based on the underlying morphology of a normal cycle of the reproductive tract including the ovary and additional immunohistochemical staining with proliferative cell nuclear antigen (PCNA) to identify small follicles may be a good tool to assess female reproductive function. In the collaborative study, 2- or 4-week repeated dose toxicity studies with ovarian histopathological examinations were conducted. A female fertility study was also conducted to compare the results with those of the ovarian histopathological findings. A total of 17 test substances were evaluated and categorized into hormone analogues, primordial follicle damaging agents, metabolite imbalance inducers, and endocrine imbalance inducers. Based on the results, ovarian toxicity could be detected by a careful histopatholgical examination. A 2-week dosing period may be sufficient for the evaluation of ovarian toxicity, except for cytotoxic compounds such as alkylating agents. The pathological findings of ovarian toxicity (decreases in follicles, increases in atretic follicles, increases in currently formed corpora lutea, etc) reflected the female fertility parameters (irregular estrous cycle, pre-implantation loss).

Collaborative work on evaluation of ovarian toxicity 17) Two- or four-week repeated-dose studies and fertility study of sulpiride in female rats

To find the appropriate dosing period to detect ovarian toxicity, sulpiride, a D2 antagonist was orally dosed to female rats at dose levels of 1, 10, and 100 mg/kg/day daily for 2 or 4 weeks in repeated-dose toxicity studies. In addition, sulpiride at the same dose levels was given to female rats daily during the pre-mating period, mating period, and Days 0-7 of gestation to assess its effect on fertility. In ovarian histology in the 2-week study, increases in atretic follicle were seen at 1 mg/kg or more and increases in follicular cysts at 10 mg/kg or more. In the 4-week study, these findings were seen at 1 mg/kg or more, and a decrease in large follicles was seen at 10 mg/kg or more. Increased body weight gain was observed at 10 mg/kg or more in the 2- and 4-week studies. The females in these groups exhibited development of mammary alveolus by sulpiride-induced hyperprolactinemia. In the fertility study, sulpiride-treated females showing persistent diestrus resulted in successful mating, and almost all females got pregnant. However, increased implantation loss was observed at 10 mg/kg or more, which was considered to be caused by the adverse effect of sulpiride on oocyte development. From these results, sulpiride-induced ovarian toxicity was seen at 1 mg/kg or more in the 2- and 4-week repeated-dose toxicity studies, and the observed ovarian changes were considered to be related to adverse effects on female fertility.

Collaborative work on evaluation of ovarian toxicity 3) Effects of 2- or 4- week repeated-dose toxicity and fertility studies with tamoxifen in female rats

To assess whether ovarian histopathological examination in repeated-dose rodent toxicity study could reliably anticipate toxic effects on female reproductive function and to assess whether ovarian change could be detected in a 2-week repeated-dose toxicity study, tamoxifen was administrated orally to female rats at 0.005, 0.03, or 0.2 mg/kg/day for 2 and 4 weeks in the repeated-dose toxicity studies, and for 2 weeks prior to cohabitation, during cohabitation, and through Gestation Day 7 in a female fertility study. The relationship between ovarian histopathological findings and fertility results was investigated. Findings at 0.03 and 0.2 mg/kg/day included decreases in body weight gains associated with decreases in food consumption, in 2- and 4-week repeated-dose toxicity studies and fertility study. The ovarian histopathological findings included increases in large atretic follicles, increases in interstitium cells and absence of newly-formed corpus lutea at 0.2 mg/kg/day in the 2-week study and at 0.03 and 0.2 mg/kg/day in the 4-week study. The treatment induced estrogenic and antiestrogenic reactions in the uterus, while mucinous degeneration was detected in the vagina. Effects on female fertility consisted primarily of disturbance of estrus cycle and decreases in numbers of pregnant rats which were considered to be related to ovarian histopathological changes. Based on these findings, ovarian histopathological evaluation in the repeated-dose toxicity study could anticipate the effects of tamoxifen on female fertility via ovarian dysfunction at slightly toxic doses, and 2-week treatment of tamoxifen at appropriate dose could be sufficient to detect ovarian toxicity by microscopic examination.

Collaborative work on evaluation of ovarian toxicity 1) Effects of 2- or 4-week repeated-dose administration and fertility studies with medroxyprogesterone acetate in female rats

As a part of the collaborative study to evaluate the relationship between histopathological changes of the ovary and functional changes in female fertility, 2- and 4-week repeated-dose toxicity studies and a female fertility study were conducted using female Crl:CD(SD) rats using a synthetic progestagen of medroxyprogesterone acetate (MPA) as a test compound. MPA was administered to female rats by gavage at 0, 0.4, 2.0 and 10 mg/kg/day for 2 and 4 weeks to assess the histopathological changes of ovaries and to pregnant rats at the same doses from 2 weeks prior to mating until day 7 of gestation to examine female fertility. The number of non-pregnant female rats with irregular estrous cycle increased in number and there was a decrease in weight of ovaries was observed at doses > or = 2.0 mg/kg in the 2- and 4-week-treatment groups. The histopathological examination revealed an increased number of large atretic follicles and decreases in currently formed corpora lutea and previously-formed large or small ones were observed at the same doses in the 2- and 4-week treatment groups. In female fertility study, the number of animals with an irregular estrous cycle and elongation of mean estrous cycle increased at 0.4 mg/kg, with no changes in fertility. A decreased number of copulating animals and a decreased gestation rate with low preimplantation loss were observed in the 2.0 mg/kg-treatment group and no copulation was observed in the group treated with 10 mg/kg. Based on these results, changes in fertility induced by MPA correlated well with histopathological changes of ovaries after 2 and 4 weeks of treatment, which suggests that a 2 weeks administration period is sufficient to detect ovarian toxicity of MPA with repeated dosing

General toxicity study of gadobenate dimeglumine formulation (E7155) (3)--4-week repeated dose intravenous toxicity study followed by 4-week recovery period in rats

A 4-week repeated dose toxicity study of gadobenate dimeglumine formulation (E7155) was conducted in Sprague-Dawley rats to assess its non-clinical safety. E7155 was administered intravenously at doses of 0.3, 1.0 and 3.0 mmol/kg/day to male and female rats once a day during 4 weeks. The reversibility of toxicity was evaluated during a 4-week recovery period at 3.0 mmol/kg/day. At 0.3 mmol/kg/day and higher, vacuolation of the cortical epithelium was seen in the kidneys and an increase in the incidence of local damage at the injection sites. In the 1.0 and 3.0 mmol/kg/day male and female groups, scabbing/ulceration of the tail at the injection sites, macroscopic pale/thickened fundic mucosa in the stomachs, vacuolation of the urinary bladder, and mucosal mineralization with epithelial hyperplasia of the glandular stomach were found. In the 1.0 and 3.0 mmol/kg/day male group and 3.0 mmol/kg/day female group, increases of water consumption and urinary potassium excretion, increased kidney weight and enlargement of the kidneys were observed. In the 3.0 mmol/kg/day male and female group, hepatocyte necrosis with inflammatory cells in the liver and epithelial degranulation in the interlobular ducts of the salivary glands were found. In addition, in the 3.0 mmol/kg/day male group, increases in plasma sodium and decreases of urinary sodium and chloride excretion, and degenerative changes in the testes and epididymides were observed. After the 4-week recovery period, except for an increase in urinary potassium excretion, increased kidney weights and changes in the testes and epididymides, all of the above findings had complete or partial recovery. Vacuolation of renal tubular cells was common, expected, and known as an adaptive change of treatment with hypertonic solutions, and an increase in the incidence of local damage at the injection sites was due to irritation by repeated intravenous dosing with hypertonic solutions. Therefore, these changes were not toxic changes. In conclusion, the dose level of 0.3 mmol/kg/day should be regarded as the No Observed Adverse Effect Level (NOAEL) after repeated administration of E7155 in rats.

Toxicity study of sodium N-[2-[4-(2,2-dimethylpropionyloxy) phenylsulfonylamino] benzoyl] aminoacetate tetrahydrate (ONO-5046.Na) (3). 4-week repeated dose intravenous toxicity study in dogs with 4-week recovery test

4-week repeated dose toxicity study with 4-week recovery test of sodium N-[2-[4-(2, 2-dimethylpropionyloxy) phenylsulfonylamino] benzoyl] aminoacetate tetrahydrate (ONO-5046.Na), a novel neutrophil elastase inhibitor, was conducted in beagle dogs. The dogs of both sexes were administered ONO-5046.Na intravenously at a daily dose of 0 (vehicle control), 7.5, 15 or 30 mg/kg. In the 15 mg/kg female group and the 30 mg/kg male and female groups, transient hypoactivity and ataxic gait were observed. It is considered that these symptoms were attributed to the pharmacological effect of ONO-5046.Na. Also, in the 30 mg/kg male and female groups, erythrocyte, hematocrit and hemoglobin were decreased. In the 30 mg/kg male group, lung weight was increased. However, histopathological examination revealed there were no changes in any organs including the lungs. There were no treatment-related changes in body weights, food consumption, ophthalmology, occult blood in feces, urinalysis, blood chemistry, electrocardiography, blood pressure, temperature, pulse rate, hepatic and renal function or necropsy. These results indicate that the NOAEL of ONO-5046.Na in dogs in 15 mg/kg/day for both sexes in this study.

Toxicity study of sodium N-[2-[4-(2,2-dimethylpropionyloxy) phenylsulfonylamino] benzoyl] aminoacetate tetrahydrate (ONO-5046.Na) (2). 4-week repeated dose intravenous toxicity study in rats with 4-week recovery test

A 4-week repeated dose toxicity study with 4-week recovery test of sodium N-[2-[4-(2,2-dimethylpropionyloxy) phenylsulfonylamino] benzoyl] aminoacetate tetrahydrate (ONO-5046.Na), a novel neutrophil elastase inhibitor, was conducted in Sprague-Dawley (SD) rats. The rats of both sexes were administered ONO-5046.Na intravenously at a daily dose of 0 (vehicle control and saline control), 18.75, 37.5, 75 or 150 mg/kg. ONO-5046.Na did not affect signs, body weight, food consumption, ophthalmology, urinalysis, hematology, blood chemistry, organ weights, necropsy or histopathology at any dose. These results indicate that the NOAEL of (ONO-5046.Na in rats is 150 mg/kg/day for both sexes in this study.

Toxicity studies of landiolol hydrochloride (ONO-1101) (3). 4-week repeated dose intravenous toxicity study in dogs with 4-week recovery test

4-week repeated dose toxicity study with 4-week recovery test of landiolol hydrochloride (ONO-1101), a novel ultra short acting beta-blocker, was conducted in beagle dogs. ONO-1101 was administered intravenously to dogs of both sexes at a dose level of 0 (control), 12.5, 25 or 50 mg/kg/day. No deaths occurred throughout the treatment period. Transitory licking chops, vomiting, nausea, diarrhea and soft feces were observed occasionally in both sexes dosed 25 and 50 mg/kg/day and the incidence seemed dose-dependent. However, those incidence declined in the course of the treatment period. Hematology showed a decrease in red blood cell count, hematocrit and hemoglobin value in both sexes receiving 25 and 50 mg/kg/day. ONO-1101 did not effect on body weight, food consumption, respiratory rate, pulse, rectal temperature, heart rate, blood pressure, electrocardiography, renal or hepatic function, ophthalmology, urinalysis, occult blood in feces, blood chemistry, organ weights, necropsy and microscopic findings at any doses. These results indicate that the no-adverse-effect level of ONO-1101 in dogs is 12.5 mg/kg/day for both sexes in this study.

Toxicity studies of landiolol hydrochloride (ONO-1101) (2). 4-week repeated dose intravenous toxicity study in rats with 4-week recovery test

4-week repeated dose toxicity study with 4-week recovery test of landiolol hydrochloride (ONO-1101), a novel ultra short acting beta-blocker, was conducted in Sprague-Dawley (SD) rats. ONO-1101 was administered intravenously to rats of both sexes at a dose level of 0 (control), 12.5, 25, 50 or 100 mg/kg/day. In the 100 mg/kg/day group, bradypnea or dyspnea was seen in all animals, pale in ear, eye and foot, tremor, reddish lacrimation and loss of righting reflex were also observed in some animals right after administration, and then those signs disappeared within 1 min after administration. During the treatment period, 3/20 animals of each sex in the 100 mg/kg/day showed clonic convulsion and died within 2 min after administration. No clinical changes were seen in the 50 mg/kg/day group or lower. Histopathological findings showed atrophy of the submaxillary gland in females and vessel-wall thickening and perivascular fibrosis of the injection site (tail) in both sexes at 100 mg/kg/day, however those changes were reversible. ONO-1101 did not effect on body weight, food consumption, ophthalmology, urinalysis, hematology, blood chemistry, organ weights or necropsy at any doses. These results indicate that the no-adverse-effect level of ONO-1101 in rats is 50 mg/kg/day for both sexes in this study.