1,3,5-Triazine, 2,4,6-tris([1,1'-biphenyl]-4-yl)-

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2004-12-21 - 2005-09-06

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: The GLP-compliant study is scientifically valid and is acceptable for assessment.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2005

Report date:

2005

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

other: Note for guidance on reproductive toxicology: detection of toxicity to reproduction for medicinal products. Committee for proprietary medicinal products (CPMP/ICH/386/95). European Agency for Evaluation of Medicinal Products, adopted September 1993.

Test guideline 2

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Test material form:

solid: bulk

Details on test material:

Please refer to "Confidential details on test material".

Test animals

Species:

rat

Strain:

Sprague-Dawley

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories France, L'Arbresle, France
- Age at study initiation: 10-11 weeks old
- Weight at study initiation: mean body weight of 268 g (range: 224 g to 306 g)
- Housing: individually housed in suspended wire-mesh cages (43.0 x 21.5 x 18.0 cm). A metal tray, containing autoclaved sawdust (SICSA, Alfortville, France), was placed under each cage.
- Diet: A04 C pelleted maintenance diet, (SAFE, Villemoisson, Epinay-sur-Orge, France), ad libitum
- Water: tap water, filtered with a 0.22 µm filter, ad libitum
- Acclimation period: at least 5 days before the beginning of the mating period

ENVIRONMENTAL CONDITIONS
- Temperature: 22 ± 2°C
- Humidity: 50 ± 20%
- Air changes: approximately 12 cycles/hour of filtered, non-recycled air
- Photoperiod (hrs dark / hrs light): 12 / 12

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

CMC (carboxymethyl cellulose)

Remarks:

0.5% (w/v) aqueous solution

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
The test item was administered as a suspension in the vehicle. The test item was ground to a fine powder using a mortar and pestle. It was then suspended in the vehicle in order to achieve the concentration of 20, 60 or 200 mg/mL. The suspensions were then homogenized using a magnetic stirrer. The test item dosage forms were prepared weekly, and were stored at +4°C prior to use and protected from light, according to the storage conditions of the test item.

VEHICLE
- Concentration in vehicle: 20, 60 or 200 mg/mL
- Amount of vehicle: 5 mL/kg bw

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

On the first and the last day of treatment, duplicate samples were taken at three levels of the container (top, middle and bottom) from each control and test item dosage form (except for the control dosage form on the last day of treatment) and analyzed for concentration of the test item to evaluate the homogeneity of the suspensions. The mean results of the homogeneity analyses were taken as actual values of the concentration of the test item in the dosage forms.

Details on mating procedure:

Monitoring of estrous cycle:
- during the week of mating, determined periodically from a fresh vaginal lavage

Mating:
- Impregnation procedure: cohoused
- M/F ratio per cage: 1/1
- Length of cohabitation: overnight for the duration of the mating period
- Proof of pregnancy: in situ, vaginal plug / sperm in vaginal smear referred to as day 0 of pregnancy (designated as day 0 post-coitum (p.c.))
- Verification of same strain and source of both sexes: yes, males were from the same strain and the same breeder

Allocation to groups:
- before day 3 p.c., the animals were allocated to the groups, according to a stratification procedure based on body weight, recorded on day 0 p.c. to ensure comparatively similar mean body weight among groups
- A larger number of animals than necessary was paired to permit the selection and/or the replacement of individuals before start of treatment

Duration of treatment / exposure:

Test item was administered on gestation days (GD) 6 through 19.

Frequency of treatment:

once a day

Duration of test:

sacrifice on day 20 p.c.

Doses / concentrations

Remarks:

Doses / Concentrations:
100, 300, 1000 mg/kg bw/day
Basis:
nominal conc.

No. of animals per sex per dose:

24 mated female rats (only the first 20 pregnant females were taken into consideration for fetal examinations)

Control animals:

yes, concurrent vehicle

Details on study design:

- Rationale for dose-level selection: The dose-levels (100, 300 and 1000 mg/kg bw/day) were selected based on the results of a previous preliminary embryo-fetal development toxicity study conducted in pregnant rats of the same strain, which received, orally, the test item at 300, 600 or 1000 mg/kg bw/day from day 6 to day 19 of gestation (CIT/Study No. 28342 RSR).

Examinations

Maternal examinations:

MORTALITY AND MORBIDITY: Yes
- at least twice a day during the treatment period
- at least once a day on the other days

CAGE SIDE OBSERVATIONS (CLINICAL SIGNS): Yes
- Time schedule: at least once a day

BODY WEIGHT: Yes (each female)
- Time schedule for examinations: on days 0, 3, 6, 9, 12, 15, 18 and 20 p.c.

FOOD CONSUMPTION: Yes (each female)
- Time schedule: days 0 - 3, 3 - 6, 6 - 9, 9 - 12, 12 - 15, 15 - 18 and 18 - 20 p.c.

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on day 20 p.c.
- Macroscopic post-mortem examination of the principal thoracic and abdominal organs
- Organs examined: ovaries and uterus
- Uterine horn(s) without visible implantation site was (were) immersed (when appropriate) in an aqueous solution of ammonium sulphide to reveal the presence of uterine scars.
- Gross evaluation of placentas was undertaken
- Preservation of tissues: macroscopic lesions from all females were sampled and kept preserved in appropriate buffer

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes (with at least one live fetus).
- Number of corpora lutea: Yes
- Number and distribution of dead and live fetuses: Yes
- Number and distribution of early and late resorptions: Yes
- Number and distribution of uterine scars: Yes
- Number and distribution of implantation sites: Yes

Fetal examinations:

- Body weight: Yes, all per litter
- External examinations: Yes, all per litter
- Soft tissue examinations: Yes, half per litter
- Skeletal examinations: Yes, remaining live fetuses per litter
- Head examinations: Yes, remaining live fetuses per litter
- Sex of the fetuses: determined before evisceration or at the time of serial sectioning
The fetal findings were described according to the glossary of the International Federation of Teratology Societies (IFTS) and classified as malformations or variations (Wise et al., 1997; Chahoud et al., 1999).

References:
Chahoud I, et al. (1999). Classification terms in developmental toxicology: need for harminization. Reprod Toxicol 13(1): 77-82.
Wise LD, et al. (1997). Terminology of developmental abnormalities in common laboratory mammals (version I). Teratology 55: 249-292.

Statistics:

Mean values were compared by one-way analysis of variance and the Dunnett test (mean values being considered as normally distributed and variances being considered as homogeneous). Percentage values were compared by the Fisher exact probability test.

Indices:

- Preimplantation loss (in %) [(number of corpora lutea – number of implantations)/number of corpora lutea]
- Post-implantation loss (in %) [(number of implantations - number of live fetuses)/number of implantations]
- Fetal or litter incidence (in %) [total number of fetuses or litters with a particular finding/total number of fetuses or litters examined]
- Mean proportion of affected fetuses (in %) [sum of proportion of fetuses affected in each litter/total number of litters examined]

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:

Maternal toxic effects:no effects

Effect levels (maternal animals)

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Basis for effect level:

other: Dose-levels of up to and including the high dose level of 1000 mg/kg bw/d did not evoke any test substance related maternal toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects

Effect levels (fetuses)

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: There was no test substance related developmental toxicity observed in the fetuses in any treated group.

Fetal abnormalities

Abnormalities:

not specified

Overall developmental toxicity

Developmental effects observed:

no

Any other information on results incl. tables

MATERNAL DATA:

Mortality

- There were no premature deaths during the study.

Clinical signs

- No treatment-related clinical signs were observed in any test item-treated group.

Body weight and food consumption

- The body weight, the net body weight change from day 6 p.c., the carcass weight and the mean gravid uterus weights were unaffected by treatment with the test item. Treatment had no influence on the mean food consumption in any test item-treated group.

Necropsy findings:

- No treatment-related findings were noted in any test item-treated females.

 

LITTER DATA:

- There was no effect of treatment on pregnancy parameters at any dose-level.

The numbers of corpora lutea and implantation sites were equivalent between test item-treated and control groups.

When compared to control values, the sex ratio was inverted at 100 and 1000 mg/kg/day. As this finding was not observed at 300 mg/kg/day, and as the mean values of percentage of male and female fetuses from all test item-treated animals were within or very close to the range of historical control data this was considered not to be treatment-related.

The mean fetal body weights recorded in the test item-treated groups were similar to those of the control group.

Fetal evaluation:

- Fetal external and visceral examinations: All fetuses appeared normal except one fetus from the high-dose group that showed mandibular micrognathia associated with microglossia; this fetus also presented marked dilated cerebral ventricles. As only this one fetus was affected, it was considered not to be related to treatment.

- Skeletal examination No treatment-related malformations or variations were noted

 

DOSAGE ANALYSIS:

- The results of the analyses demonstrated the homogeneity of each dosage form investigated on the first day and the last day of treatment.

- Satisfactory correspondence was noted between the nominal and the measured concentrations of the test item for each administered dosage analyzed (between -9% and - 1% of nominal values).