Registration Dossier

Toxicological information

Developmental toxicity / teratogenicity

Currently viewing:

Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Oct 24, 2000 to Nov 24, 2000
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Cross-reference
Reason / purpose:
reference to other study

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
2002
Report Date:
2002
Reference Type:
publication
Title:
Unnamed
Year:
2004
Reference Type:
publication
Title:
Unnamed
Year:
2009

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
no
Qualifier:
according to
Guideline:
other: US FDA “Guideline on detection of toxicity to reproduction for medicinal products” (1994; 1996)
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Type:
Constituent
Test material form:
liquid

Test animals

Species:
rat
Strain:
Sprague-Dawley
Details on test animals and environmental conditions:
Crl:CD®(SD)IGS BR VAF/Plus® rats weighed 218-246 g at study assignment and were about 65 days old at arrival. Pregnant animals were individually housed in stainless steel, wire-bottomed cages at a room temperature of 18-26C with a relative humidity of 30-70%, at least 10 air changes per hour, and a 12-hour light/dark cycle. Rats were given Certified Rodent Diet #5002 (PMI Nutrition International) and reverse osmosis water ad libitum.

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
unchanged (no vehicle)
Details on exposure:
Rats were gavaged with 0 (control), 96, 240, or 480 mg/kg bw/day on gestation days 7-17 at dose volumes of 0.5, 0.1, 0.25, or 0.5 ml/kg bw, respectively. Doses were adjusted daily based on current body weights.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Samples (2 ml) of test substance were taken on first and last day of dosing for analysis. No further information available.
Details on mating procedure:
Virgin females were placed with males (1 female per male) for up to 5 days. Mating performance was evaluated daily and when spermatozoa in a vaginal smear and/or a copulatory plug were observed in females, the female was placed in individual housing and the day was designated as gestation day 0.
Duration of treatment / exposure:
Day 7-17 of gestation
Frequency of treatment:
Daily
Duration of test:
21 days
No. of animals per sex per dose:
25
Control animals:
other: 0.5 ml/kg bw/day of reverse osmosis deionized water (25 rats)
Details on study design:
Groups of pregnant rats (25/dose group) were gavaged with 0 (control), 96, 240, or 480 mg/kg bw/day on gestation days 7-17 at dosage volumes of 0.5, 0.1, 0.25, or 0.5 ml/kg bw, respectively. Viability (2x daily), abnormal clinical signs (daily), body weights (daily), abortions (daily), premature deliveries (daily), and feed consumption (gestation days 0, 7, 10, 12, 15, 18, and 21) were recorded. Mating performance was assessed. On gestation day 21, surviving rats were euthanized by carbon dioxide asphyxiation and Caesarean sections were performed to remove the foetuses.

Doses were selected based on a dose range-finding study:
Groups of pregnant rats (8/dose group) were gavaged with 0 (control), 240, 480, 960, and 1920 mg/kg bw/day on gestation days 7-17. On gestation day 21, all surviving rats were euthanized and examined for distribution of corpora lutea, implantation sites, and uterine content. Parental animals underwent gross necropsy and foetuses were weighed and examined. Body weights, body weight change, and feed consumption were generally comparable to those of controls throughout the study. Necropsy of the dams and examination of the litters revealed no differences in any of the test groups including controls. The recommended dose levels for the main developmental toxicity study were 240, 480, and 960 mg/kg bw/day. The no-observable-effect level (NOEL) for maternal and embryo-foetal toxicity was 240 mg/kg bw/day. Maternal toxicity with little or no developmental toxicity was reported at 960 mg/kg bw/day.

Examinations

Maternal examinations:
Rats underwent gross necropsy of the thoracic, abdominal and pelvic viscera. Animals found dead or moribund were examined for gross lesions, pregnancy status, and uterine contents.
Ovaries and uterine content:
The number and distribution of corpora lutea, implantation sites, live and dead foetuses, and early and late resorptions were assessed. The uteri of apparently non-pregnant rats were examined to confirm absence of implantation sites.
Fetal examinations:
Live foetuses were euthanized with an intraperitoneal injection of Beuthanasia®-D Special. Sex and body weight of each foetus were recorded and any gross alterations were noted. About half of the foetuses were examined for soft tissue alterations and fixed in Bouin’s solution. Sections were kept in alcohol. The remaining foetuses were examined for skeletal alterations by staining with alizarin red S after evisceration and clearing.
Statistics:
Clinical observation and other proportion data were analysed using the Variance Test for Homogeneity of the Binomial Distribution. Continuous data (e.g., weights, feed consumption, litter averages, etc.) were analysed using Bartlett’s Test of Homogeneity of Variances and the Analysis of Variance when appropriate. If the Analysis of Variance was significant (p≤0.05), Dunnett’s Test was used to identify the statistical significance of the individual groups. If the Analysis of Variance was not appropriate, the Kruskal-Wallis Test was used. In cases where the Kruskal-Wallis Test was statistically significant (p≤0.05), Dunn’s Method of Multiple Comparisons was used to identify the statistical significance of individual groups. If there were greater than 75% ties, Fisher’s Exact Test was used. Count data from Caesarean sections were evaluated using the Kruskal-Wallis Test.

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
Two rats found dead and one delivered prematurely in the mid-dose group but these events were not considered related to test substance because they were not dose-related and 1 death was due to gavage error. Clinical signs included a statistically significant increase in the incidence of salivation in all treated groups compared to controls and a statistically significant increase in the incidence of urine-stained abdominal fur in high-dose animals compared to controls. Red, dried or red perioral substance was noted in all treated groups. Other clinical signs noted, but not considered related to the test substance, included localized alopecia, ptosis, soft or liquid faeces, chromodacryorrhea, ungroomed coat, red perivaginal substance and missing or broken incisors. Body weight gains were significantly reduced at all dose levels during gestation days 7-10 and throughout gestation at the highest dose. Body weights were significantly reduced on gestation days 16 and 17 at the highest dose. Feed consumption was significantly reduced in all dose groups during gestation days 7-12 and continued to be reduced throughout gestation at the highest dose only.

Effect levels (maternal animals)

Key result
Dose descriptor:
NOAEL
Effect level:
240 mg/kg bw/day
Basis for effect level:
other: maternal toxicity

Maternal abnormalities

Key result
Abnormalities:
effects observed, treatment-related
Localisation:
other: Systemic effects
Description (incidence and severity):
Maternal effects were seen, see details on maternal toxic effects

Results (fetuses)

Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
Foetal body weights were reduced, but not significantly, at the highest dose level. No effects attributable to test substance administration on any litter parameters (i.e, litter averages for corpora lutea, implantations, litter sizes, live foetuses, early and late resorptions, total and percent resorbed conceptuses per litter, the numbers of dams with any resorptions, dams with viable foetuses, dams with all conceptuses), gross external, soft tissue, or skeletal malformations, or number of ossification sites per foetus per litter at any dose level.

Effect levels (fetuses)

Key result
Dose descriptor:
NOAEL
Effect level:
>= 480 mg/kg bw/day
Based on:
test mat.
Remarks on result:
not determinable due to absence of adverse toxic effects

Fetal abnormalities

Key result
Abnormalities:
no effects observed

Overall developmental toxicity

Key result
Developmental effects observed:
no

Any other information on results incl. tables

Foetal body weight reductions (non-significant) seen only at maternally toxic doses.

Applicant's summary and conclusion

Conclusions:
Based on persistent clinical observations and reduced body weights and feed consumption in the dams reported at the highest dose, the maternal NOAEL was considered to be 240 mg/kg bw/day. Reductions in foetal body weights were not statistically significant at the highest dose, and therefore, the developmental NOAEL was considered to be 480 mg/kg bw/day. It was concluded that OTNE was not a developmental toxicant.
Executive summary:

A developmental toxicity study was performed in accordance with the US FDA Guideline on detection of toxicity to reproduction for medicinal products, which is equivalent to OECD414. The study was performed under GLP conditions.

Groups of pregnant rats (25/dose group) were gavaged with 0 (control), 96, 240, or 480 mg/kg bw/day on gestation days 7-17 at dosage volumes of 0.5, 0.1, 0.25, or 0.5 ml/kg bw, respectively. Viability, abnormal clinical signs, body weights, abortions, premature deliveries, and feed consumption were recorded. Mating performance was assessed. On gestation day 21, surviving rats were euthanized by carbon dioxide asphyxiation and Caesarean sections were performed to remove the foetuses. Rats underwent gross necropsy. Foetuses were assessed for litter parameters and gross external, soft tissue, or skeletal malformations.

Maternal animals showed reduced body weights and persistent clinical signs (e.g., increased salivation and urine-stained abdominal fur) at the highest dose. There were no treatment-related effects on developmental parameters at any dose, although reduction (not statistically significant) in foetal body weight was noted at the highest dose. Based on these results, the maternal NOAEL was 240 mg/kg bw/day and the developmental NOAEL was 480 mg/kg bw/day.