Registration Dossier

Diss Factsheets

Toxicological information

Developmental toxicity / teratogenicity

Currently viewing:

Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
No data. Dates of treatment were 27.12.1978 to 19.01.1979
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1979
Report date:
1979

Materials and methods

Test guideline
Guideline:
other: FDA "Guidelines for reproductive studies for evaluation of drugs for human use", segment II (teratological study)
Deviations:
not specified
Remarks:
Treatment on GD 6 - 15; no record of gravid uterine weight; number corpora lutea not recorded; no analytical confirmation of exposure levels.
Principles of method if other than guideline:
Study was used to assess the teratogenic and/or embryotoxic potential of the test substance.
GLP compliance:
no
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
Nitrilotrimethylenetris(phosphonic acid)
EC Number:
229-146-5
EC Name:
Nitrilotrimethylenetris(phosphonic acid)
Cas Number:
6419-19-8
Molecular formula:
C3H12NO9P3
IUPAC Name:
[nitrilotris(methylene)]tris(phosphonic acid)
Test material form:
liquid

Test animals

Species:
rat
Strain:
Sprague-Dawley
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River, Wilmington, Mass
- Age: 72 days at mating
- Weight at study initiation: 240 – 250 grams
- Fasting period before study: Not specified
- Housing: Individually housed in elevated stainless steel cage (however, not under the mating).
- Diet: Purina Certified Rodent Chow 5001, ad libitum
- Water: By automated water system, ad libitum
- Acclimation period: 28 November to 19 December 1978

ENVIRONMENTAL CONDITIONS
- Temperature (°C): monitored twice daily, no more details.
- Humidity (%): Not specified
- Air changes (per hr): Not specified
- Photoperiod (hrs dark / hrs light): 12 hours dark / 12 hours light

IN-LIFE DATES: From: Not specified To: 19th of January 1979

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Remarks:
Distilled
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: The test substance was mixed with water and administered at 10 mg/kg bw/day. Dosing solutions were prepared fresh daily. Individual doses were adjusted based on the most recent body weight data.

DIET PREPARATION
- Rate of preparation of diet: Daily
- Storage temperature of food: Room temperature

VEHICLE
- Justification for use and choice of vehicle: N/a
- Concentration in vehicle: 22.4%
- Amount of vehicle: 10 mg/kg bw/day
- Lot/batch no.: Not specified
- Purity: Not specified
Analytical verification of doses or concentrations:
no
Details on mating procedure:
- Impregnation procedure: cohoused
- If cohoused:
- M/F ratio per cage: 1:2
- Length of cohabitation: Overnight
- Further matings after two unsuccessful attempts: No data
- Verification of same strain and source of both sexes: No data
- Proof of pregnancy: vaginal plug referred to as day 0 of pregnancy.
Duration of treatment / exposure:
Gestation days 6 - 15
Frequency of treatment:
Daily as a single dose
Duration of test:
21 days
Doses / concentrationsopen allclose all
Dose / conc.:
100 mg/kg bw/day
Dose / conc.:
500 mg/kg bw/day
Dose / conc.:
1 000 mg/kg bw/day
No. of animals per sex per dose:
24 mated females/dose
Control animals:
yes, concurrent vehicle

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily
- Cage side observations included gross signs

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Gestation day 0, 6, 10, 15, 20 and 21 (pre-necropsy)

BODY WEIGHT: Yes
- Time schedule for examinations: Gestation day 0, 6-15, 21

FOOD CONSUMPTION AND COMPOUND INTAKE:

WATER CONSUMPTION AND COMPOUND INTAKE: No

POST-MORTEM EXAMINATIONS: Yes, all females
- Sacrifice on gestation day 21 (all surviving dams) and day 21 post-mating (all surviving non-pregnant females).
Ovaries and uterine content:
Examinations included:
- Number of corpora lutea: Yes
- Sites of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
Fetal examinations:
- External examinations: Yes, approximately 50% of foetuses subject to gross dissection

- Soft tissue examinations: Yes, approximately 50% of foetuses subject to gross dissection (followed by processing / staining (Alizarin red) for skeletal abnormalities/variations)
- Skeletal examinations: Yes, approximately 50% of foetuses subject to gross dissection
Type of skeletal malformations control: angulated ribs, cervical rib, wavy rib
- Head examinations: Not specified
Other: Approximately 1/2 of the foetuses were subject to Wilson serial sectioning for neural/visceral defects (10X or 20X magnification) after fixing in Bouin's solution. Additionally, approximately 1/2 of the foetuses were examined of skeletons anomalies and ossification variation after Alizarin red staining using the method of Crary but with the following modification: eviscerated foetuses were immediately placed into an aqueous potassium hydroxide solution followed by staining. Thereafter, stained foetuses were placed in Mall's solution to remove excess stain prior to clearing and storage in benzyl alcohol-glycerine solution. Internal sex determination occurred for both the groups.
Statistics:
Chi squared, F-test and Student's T-test (absolute data) were used to compare between control and each test substance-treated group. T-tests modified using Cochran's approximation were used when variances differed significantly. Live foetuses, resorptions, implantations and corpora lutea were compared to control by using one-tailed T-test.

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:
no effects observed
Dermal irritation (if dermal study):
not examined
Mortality:
no mortality observed
Description (incidence):
One female in the 100 mg/kg bw/day dose group was sacrificed in a moribund condition on gestation day 6 which was the first day of treatment. No other mortality occurred in any of the other dose groups or the control group.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
In the 1000 mg/kg bw/day dose group, lower weight gain compared to the control group was observed, although not statistically significant. No other significant differences in the mean body weight between the dose groups were observed.
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Description (incidence and severity):
No treatment-related effects were observed.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
Other effects:
no effects observed

Maternal developmental toxicity

Number of abortions:
not specified
Pre- and post-implantation loss:
effects observed, non-treatment-related
Description (incidence and severity):
There was a statistically significant increase in the implantation efficiency in the 100 mg/kg bw/day dose group. However, this was not considered to be treatment-related.
Total litter losses by resorption:
no effects observed
Description (incidence and severity):
The occurrence of ≥2 resorptions in each of the treated dose groups compared to the control group was higher. However, the values were within the historical control values and therefore, not considered as treatment-related.
Early or late resorptions:
effects observed, non-treatment-related
Description (incidence and severity):
The mean number of resorption observed in the 100 and 1000 mg/kg bw/day dose groups were significantly greater than the control dose group. However, the values were within the historical control range and therefore not considered as treatment-related.
Dead fetuses:
no effects observed
Description (incidence and severity):
There were no dead foetuses in any groups.
Changes in pregnancy duration:
not specified
Changes in number of pregnant:
no effects observed
Description (incidence and severity):
There were no treatment-related changes in the pregnancy rate.
Other effects:
no effects observed
Description (incidence and severity):
A statistically significant decrease in the mean number of corpora lutea were observed in the 100 mg/kg bw/day dose group, however, this was not considered as treatment-related since it was not occurring in a dose-dependent manner.

Effect levels (maternal animals)

Key result
Dose descriptor:
NOAEL
Effect level:
500 mg/kg bw/day
Basis for effect level:
body weight and weight gain

Maternal abnormalities

Abnormalities:
no effects observed

Results (fetuses)

Fetal body weight changes:
no effects observed
Description (incidence and severity):
The mean foetal weights were comparable between the treated animals and the control dose group.
Reduction in number of live offspring:
no effects observed
Description (incidence and severity):
The mean numbers of live foetuses were comparable between control and the treated groups.
Changes in sex ratio:
no effects observed
Description (incidence and severity):
Sex: males/litter 6.9/7.0/6.6/6.0; females 7.4/6.9/7.0/6.9 (no significant effect) Sex ratio (m:f): 92.4%/102.0%/94.6%/87.3% (no significant effect)
Changes in litter size and weights:
not specified
Changes in postnatal survival:
not examined
External malformations:
effects observed, non-treatment-related
Description (incidence and severity):
In the 1000 kg/kg bw/day dose group, 6 of 16 foetuses in one female had defects including flexed forepaws, shortened and thickened torso, abdominal distension and exaggerated forward flexure of the head. No other significant external malformations were observed. No other dose groups had any malformations.
Skeletal malformations:
no effects observed
Description (incidence and severity):
In the foetal skeletal examination, variations in the ossification occurred. These variations may represent delays in the ossification process or slight ossification irregularities. However, the varation were comparable between the treated groups and the control group.
Visceral malformations:
effects observed, non-treatment-related
Description (incidence and severity):
The incidence of foetuses with soft tissue malformations was considered comparable between the control group and the treatment groups. The types of soft tissue malformations, e.g. distended renal pelvis and/or distended ureter are frequently observed in the specific rat strain used. A malrotation defect of the heart was observed in two foetuses in the 1000 mg/kg bw/day. Both of these foetuses had external malformations and were from the same litter as had observed external malformations. Incidences in the 500 and 1000 mg/kg bw/day were comparable to the control group. In the 100 mg/kg bw/day, the incidence of soft tissue malformations was higher than in the control group. However, this difference was not statistically significant and there was no dose relationship. Therefore, the observed effects were not considered as treatment-related.
Other effects:
no effects observed
Description (incidence and severity):
The mean crown-rump length for both sexes was significantly greater in the 500 mg/kg bw/day compared to the control group. This difference was not considered as biologically significant and therefore, not treatment related. No other differences were observed compared to the control group.

Effect levels (fetuses)

open allclose all
Key result
Dose descriptor:
NOAEL
Effect level:
>= 1 000 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No teratogenic effects observed.
Key result
Dose descriptor:
NOAEL
Effect level:
>= 1 000 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No fetotoxic effects observed.

Fetal abnormalities

Abnormalities:
no effects observed

Overall developmental toxicity

Developmental effects observed:
no

Applicant's summary and conclusion

Conclusions:
In a developmental toxicity study, conducted prior to the adoption of OECD test guidelines and pre-GLP, ATMP-H was administered by oral gavage to pregnant Charles River SD rats (24/dose) on gestation days 6-15. The doses tested were 100, 500 or 1000 mg/kg bw/day. The maternal NOAEL was concluded to be 500 mg/kg bw/day and the NOAEL for fetotoxicity and teratogenicity were concluded to be ≥1000 mg/kg bw/day respectively. This was based on females exposed to 1000 mg/kg bw/day gained less weight compared to the controls during the dosing period and was considered as treatment related. No other treatment-related effects were observed.