1,3-diethyl-2-thiourea

Endpoint summary

Key value for chemical safety assessment

Toxic effect type:

dose-dependent

Effects on fertility

Description of key information

A Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test is currently ongoing. Testing schedule is reported in the associated RSS.

Results will be provided as soon as they will be available.

Link to relevant study records

Reference

Endpoint:

screening for reproductive / developmental toxicity

Remarks:

Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Experimental Starting Date (Animal Arrival): 10 May 2022
Audited Draft Report Date: September 2023
Final Report Date: January 2024

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Reason / purpose for cross-reference:

reference to same study

Qualifier:

according to guideline

Guideline:

OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Species:

rat

Strain:

Wistar

Details on species / strain selection:

Han Wistar rat (virgin), accepted by regulatory agencies, historical control data available.

Sex:

male/female

Details on test animals or test system and environmental conditions:

Air supply: Filtered, not recirculated.
Temperature: Maintained within the range of 20-24ºC.
Relative humidity: Maintained within the range of 40-70%.
Lighting: 12 hours light : 12 hours dark.
Diet supply: SDS VRF1 Certified (ad libitum). Pelleted diet.
Water supply: Potable water from the public supply (ad libitum).

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on exposure:

Treated at: Constant doses in mg/kg/day.
Controls (Group 1): Vehicle at the same volume dose as treated groups.
Volume dose: 5 mL/kg/day for Groups 1, 2 and 3, Group 4 up until Day 21 (30 May 2022). 3.33 mL/kg/day for Group 4 males from Day 22 (31 May 2022) onwards.
Individual dose volume: Calculated from the most recently recorded scheduled body weight.
Frequency: Once daily, at approximately the same time each day.
Sequence: Groups dosed in ascending order.
Administration: Prior to dosing of each individual animal, the dosing catheter/cannula will be dipped into a container filled with a 5% glucose solution to aid intubation.
Formulation: A daily record of the usage of formulation will be maintained based on weights before and after dosing. Formulations are stirred using a magnetic stirrer before and throughout the dosing procedure. Alternative methods may be used; these will be documented in the study records.
Storage of formulation: Refrigerated temperature (2 to 8°C).
Expected appearance of formulation: Orange opaque suspension

Details on mating procedure:

Paired for mating: After minimum of 2 weeks of treatment.
Male/female ratio: 1:1
Duration of pairing: Up to 2 weeks.
Daily checks for evidence of mating Ejected copulation plugs. Sperm within vaginal smear.
Day 0 of gestation: When positive evidence of mating detected.
Male/female separation: Day when mating evidence detected.
Pre-coital interval: Calculated for each female as time between first pairing and evidence of mating.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

3 occasions: Week 1, Week 1 of gestation and final Week.

Duration of treatment / exposure:

Males: Sacrifice during Week 5 - after at least 4 weeks of treatment
Females: Sacrifice on Day 21 of lactation

Frequency of treatment:

Once daily

Dose / conc.:

0 mg/kg bw/day (nominal)

Dose / conc.:

10 mg/kg bw/day (nominal)

Dose / conc.:

30 mg/kg bw/day (nominal)

Dose / conc.:

50 mg/kg bw/day (nominal)

No. of animals per sex per dose:

10 animals/sex/dose.

Control animals:

yes, concurrent vehicle

Details on study design:

A 14-day Dose Range Finding (DRF) study was conducted on this substance (Labcorp Study 8468465). Doses of 100, 200 or 300 mg/kg bw/day, in corn oil vehicle, were administrated to groups of 5 male and 5 female Han Wistar rats. An additional group of 3 male and 3 female Han Wistar rats was eventually added, receiving 50 mg/kg bw/day of the test item in corn oil.
Animals dosed at 200 or 300 mg/kg bw/day were terminated early, on Day 10 or 3, respectively. Severe and persistent clinical signs were observed in these animals, along with a body weight loss at 200 mg/kg bw/day associated with reduction in food consumption, and macroscopic findings in the gastrointestinal tract, especially in the stomach (dark, depressions, thickened and distended).
All animals receiving 100 or 50 mg/kg bw/day of the test item survived until completion of the study. At 100 or 50 mg/kg bw/day, limited clinical signs were reported, mostly transient and appearing shortly after dosing of the animals. At 100 mg/kg bw/day, body weight loss in males (-7 g between Days 8 and 15, -14% vs control on Day 15) and females (- 12 g between Days 11 and 15, -4% vs control on Day 15) animals was reported with an associated reduction in food intake, while water intake was increased. One female had macroscopic findings in the stomach consistent with the animals from the higher dose levels.
At 50 mg/kg bw/day, body weight loss was also apparent for males (- 4 g between Days 11 and 15, -5% vs control on Day 15) associated with low food intake, and body weight loss was evident for females (-2 g between Days 11 to 15), although their body weight on Day 15 did not differ from control animals. Water consumption was also increased in both sexes.
Considering that:
- According to the OECD Testing Guideline 422, the highest dose level should be chosen with the aim of inducing toxic effects but not death nor obvious suffering;
- The adverse effects – including local irritation effect in the stomach and reduction in bodyweight – observed in animals exposed to 100 mg/kg bw/day of test item for 14 days, can reasonably be expected to result in severe suffering and/or death when the treatment period is extended, and;
- Limited effects were reported in animals exposed to 50 mg/kg bw/day of the test item for 14 days, so this dose-level may not be sufficient to induce toxic effects.
The most appropriate top dose-level for the OECD Testing Guideline 422 is comprised between 50 and 100 mg/kg bw/day. Therefore, a dose level of 75 mg/kg bw/day is selected as the highest dose-level for the study.
30 and 10 mg/kg bw/day are selected as the intermediate and low dose-levels, respectively.
Treatment at 75 mg/kg/day was not tolerated, with 5 out of 10 females either euthanised for welfare reasons or found dead, and 2 out of 10 males euthanised for welfare reasons. Upon review of the data, the decision was taken to euthanise the remaining 5 females receiving 75 mg/kg/day in order to prevent any further undue suffering or premature deaths for these animals.
The remaining males receiving 75 mg/kg/day were not euthanised, but the dose level received by these males will be reduced to 50 mg/kg/day from Day 22 of dosing (31 May 2022) onwards.

Positive control:

Not relevant

Parental animals: Observations and examinations:

Clinical Observations: Visually inspected at least twice daily for evidence of reaction to treatment or ill-health
Body Weight: Before dosing on the day that treatment commences, weekly thereafter, twice weekly from Week 4 onwards and on the day of necropsy. Females: Days 0, 4, 7, 11, 14, 17 and 20 after mating and Days 1, 4, 7, 11, 14, 18 and 21 of lactation (the day of necropsy).
Food Consumption: Weekly during treatment.
Water consumption: Weekly during treatment. Measured for a 3-day period in each week.
Sensory Reactivity and Grip Strength / Motor Activity: Week 5 (males). Days 7-9 of lactation.
Estrous Cycles: For 14 days before treatment (all females including spares); animals that fail to exhibit 4-5 day cycles will not be allocated to study. After pairing until mating (for a maximum of 14 days). Females showing no evidence of mating: following completion of the pairing period females will be separated from the male and vaginal smearing continued for up to five days or until the first estrus smear is seen. If a female shows an estrus smear during this period, she will be killed as soon as practically possible and subject to macroscopic examination.
Hematology, Peripheral Blood: At termination.
Blood Chemistry: At termination.
Thyroid Hormone Analysis: At termination.
Duration of gestation: Time elapsing between mating and commencement of parturition.
Parturition observations
From Day 20 after mating, animals checked 3 times daily for evidence of parturition. If difficulties observed, progress of parturition process monitored. Numbers of live and dead offspring recorded.

Oestrous cyclicity (parental animals):

Wet smears: Using pipette lavage during the following phases:
• For 14 days before treatment (all females including spares); animals that fail to exhibit 4-5 day cycles will not be allocated to study.
• After pairing until mating (for a maximum of 14 days).
• Females showing no evidence of mating: following completion of the pairing period females will be separated from the male and vaginal smearing continued for up to five days or until the first estrus smear is seen. If a female shows an estrus smear during this period, she will be killed as soon as practically possible and subject to macroscopic examination. NB: The guideline states that “females showing no evidence of copulation are killed 24-26 days after the last day of the mating period”. Reading vaginal smears after separation and confirming failure to mate/non-pregnancy by continuation of cyclicity is considered an enhancement in animal welfare.
• On the day of scheduled termination.
Dry smears: For 15 days before pairing, using cotton swabs.

Sperm parameters (parental animals):

Detailed qualitative examination will be made, taking into account the tubular stages of the spermatogenic cycle. The examination will be conducted in order to identify treatment related effects such as missing germ cell layers or types, retained spermatids, multinucleate or apoptotic germ cells and sloughing of spermatogenic cells into the lumen. Any cell- or stage-specificity of testicular findings will be noted.

Litter observations:

Clinical observations: Observed approximately 24 hours after birth and then daily for evidence of ill-health or reaction to maternal treatment.
Litter size: Daily on Days 1-21 of age.
Sex ratio: Days 1, 4, 7, 14 and 21 of age.
Individual offspring body weights: Days 1, 4, 7, 14, 17 and 21 of age.
Ano-genital distance: Day 1 of age.
Nipple/areolae count: Day 13 of age - male offspring. Representative photographs may be taken if nipples are seen.

Postmortem examinations (parental animals):

Detailed macroscopic examination will be performed for evidence of adverse reaction to treatment. Organs weighed and tissues retained for light microscpy. Any abnormal tissues retained and may be weighed at the discretion of necropsy staff.
For F0 females, the number of uterine implantation sites will also be recorded.

Postmortem examinations (offspring):

F1 offspring culled on Day 4 of age: Externally normal offspring discarded without examination. Externally abnormal offspring identified on despatch to necropsy; examined externally, and retained pending possible future examination.
F1 offspring on Day 21 of age: Thyroid glands retained from two offspring – one male and one female, where possible If insufficient numbers of Day 21 offspring are available for hormone sampling and thyroid retention/preservation, priority is given to hormones, with sample for T4 given first priority. All animals will be subject to an external macroscopic examination; particular attention will be paid to the external genitalia. Abnormalities retained in appropriate fixative.

Statistics:

Included

Reproductive indices:

Fertility index
Gestation index

Offspring viability indices:

Post-implantation survival index
Live birth index
Viability index
Lactation index

Remarks on result:

not determinable

Remarks:

Conclusion will be reached once the study has been completed.

Critical effects observed:

not specified

Remarks on result:

not determinable

Remarks:

Conclusion will be reached once the study has been completed.

Critical effects observed:

not specified

Reproductive effects observed:

not specified

Conclusions:

The objective of the study is to make an assessment of the general systemic toxic potential in Han Wistar rats, including a screen for reproductive/developmental effects and assessment of endocrine disruptor relevant endpoints, with administration of 1,3-diethyl-2-thiourea by oral gavage for at least 4 weeks.

Effect on fertility: via oral route

Endpoint conclusion:

no study available (further information necessary)

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Effects on developmental toxicity

Description of key information

A reliable study on developmental toxicity study on rats in available on the analogue Dimethylthiourea (DMTU).
Foetoxicity was observed in this study at all doses but this toxicity is due to the maternal toxicity (decrease of maternal bodyweight at all doses).
The LOAEL for developmental toxicity was 15 mg/kg/day based on decrease of fetal body weight gain at all doses. No teratogenic effects were observed in this study.

A Decision on Compliance Check was received requesting a pre-natal developmental toxicity study (OECD 414) by oral route, in rats to be conducted on the registered substance. Before initiating this study, it was considered appropriate to conduct a 14-day Dose Range Finding study followed by the Screening for reproductive/developmental toxicity on rats (OECD 422) requested by the authorities as part of the same Decision, and a preliminary study in pregnant rats. The aim was to generate appropriate experimental data on the registered substance in order to select the most appropriate dose-levels to be investigated during the OECD 414, in accordance with the latest ECHA Guidance. Due to unforeseen circumstances, delays were encountered resulting in the postponment of the studies. Consequently, the regulatory deadline cannot be met. Testing schedule is reported in the associated RSS.
Results will be provided as soon as they will be available.

Link to relevant study records

Reference

Reference 1

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Experimental Starting Date (Animal Arrival): 16 February 2023
Audited Draft Report Date: December 2023
Final Report Date: April 2024

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Species:

rat

Strain:

Wistar

Details on test animals or test system and environmental conditions:

Air supply: Filtered, not recirculated.
Temperature: Maintained within the range of 20-24ºC.
Relative humidity: Maintained within the range of 40-70%.
Lighting: 12 hours light : 12 hours dark.
Diet supply: SDS VRF1 Certified, Pelleted diet (ad libitum).
Water supply: Potable water from the public supply (ad libitum).

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on exposure:

Treated at: Constant doses in mg/kg/day.
Volume dose: 4 mL/kg/day.
Individual dose volume: Calculated from the most recently recorded scheduled body weight.
Controls (Group 1): Vehicle at the same volume dose as treated groups.
Frequency: Once daily, at approximately the same time each day.
Sequence: Groups dosed in ascending order.
Method: Before administering the test substance, the required amount of dose formulation is drawn up into the syringe. After the dose has been drawn up the outside of the catheter is wiped clean of formulation residue with a disposable tissue and the end of the catheter will be lightly tapped onto clean tissue to remove any remaining droplets. The catheter will then be dipped into a container filled with 5% glucose solution.
Formulation: A record of the usage of formulation will be maintained based on weights. This balance may be compared with the expected usage as a check of correct administration if any discrepancy is suspected.
Formulations are stirred using a magnetic stirrer before and throughout the dosing procedure. Alternative methods may be used; these will be documented in the study records.
Storage of formulation: Refrigerated (2 to 8ºC).
Expected appearance of formulation: Orange opaque suspension.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

First Week and Last Week

Details on mating procedure:

Animals will be supplied time mated to identified males of the same strain and source. Arrival on GD 2 over four delivery dates.

Duration of treatment / exposure:

GD 6 to 20.

Frequency of treatment:

Once daily

Duration of test:

Termination on GD 21

No. of animals per sex per dose:

22 mated females/dose

Control animals:

yes, concurrent vehicle

Maternal examinations:

Clinical Observations: Visually inspected at least twice daily for evidence of reaction to treatment or ill-health. Physical examination: GD 3, 5, 12, 18 and 21.
Body weight: GD 3, 6-21.
Food consumption: GD 3-6, 6-9, 9-12, 12-15, 15-18 and 18-21.
Thyroid hormone analysis: All adults surviving to scheduled termination on GD 21.
Light Microscopy: Thyroid.

Ovaries and uterine content:

For females surviving to GD 21 only, the following will be recorded:
Uterus: Gravid uterine weight (including cervix and ovaries).
The following will be recorded for all animals (including those prematurely sacrificed, where possible):
Each ovary/uterine horn, number of:
Corpora lutea.
Implantation sites.
Resorption sites (assessed as early or late).
Fetuses (live and dead).

Apparently non pregnant animals: Status confirmed by appropriate staining technique for presence of implantation sites.

Blood sampling:

All adults surviving to scheduled termination on GD 21.

Fetal examinations:

Fetal Examination
All live fetuses: Fetuses and placentae dissected from the uterus and weighed individually. Fetuses sexed. Ano-genital distance recorded. External examination with particular attention paid to external genital organs of male fetuses. Individual identification within litter.
50% in each litter: Sex confirmed internally. Examined for visceral abnormalities by fresh microdissection (Modified Staples technique) and subsequently fixed in Bouins solution.
50% in each litter: Sex confirmed internally, eviscerated and fixed in Industrial Methylated Spirit.

Fetal processing
Bouin’s fixed fetuses: Head removed post-fixation and heads processed by Wilsons free-hand serial sectioning. Torsos retained in Bouins solution.
IMS fixed fetuses: Processed and stained with Alizarin Red and Alcian Blue.

Detailed Fetal Examination
Bouin’s fixed heads: Serial sections examined for visceral abnormalities.
Fetuses, stained with Alizarin Red and Alcian Blue: Skeletal development, cartilage and abnormalities assessed.

Statistics:

Included

Historical control data:

Recent HCD will be provided by the CRO.

Remarks on result:

not determinable

Remarks:

Conclusion will be reached once the study has been completed.

Abnormalities:

not specified

Remarks on result:

not determinable

Remarks:

Conclusion will be reached once the study has been completed.

Abnormalities:

not specified

Developmental effects observed:

not specified

Conclusions:

Assessment of the influence of 1,3-diethyl-2-thiourea on embryo-fetal survival and development when administered during the organogenesis and fetal growth phases of pregnancy in the Han Wistar rats.

Reference 2

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

Comparable to guideline study [OECD 414] A read-across between DMTU (dimethylthiourea) and DETU (diethylthiourea) is justified because theirs structures differ by a methyl fonction only. DETU has a molecular weight more higher than DMTU and therefore DETU is probably less toxic than DMTU.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Deviations:

no

GLP compliance:

not specified

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: From IFFA CREDO Breeding Laboratories (St Germain sur l'arbresle, France)
- Age at study initiation: no data
- Weight at study initiation: males = 350 g ; females = 200-220 g
- Fasting period before study: no data
- Housing: Bred females were individually housed in clear polycrbonate cages with hardwood shavings as bedding.
- Diet (e.g. ad libitum): UAR Alimentation Villemoisson, ad libitum
- Water (e.g. ad libitum): filetred tap water, ad libitum
- Acclimation period: 1 or 2 weeks

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- °C
- Humidity (%): 55 +/- 5%
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): a light cycle from 7AM to 7 PM

Route of administration:

oral: gavage

Vehicle:

water

Details on exposure:

Administered volume = 5 ml/kg bw
The actual volume administered was based on body weight taken on GD6.

Analytical verification of doses or concentrations:

not specified

Details on mating procedure:

- Impregnation procedure: cohoused
- If cohoused:
- M/F ratio per cage: 1M / 3 F
- Length of cohabitation: overnight
- Verification of same strain and source of both sexes: yes
- Proof of pregnancy: sperm in vaginal smear referred to as day 0 of pregnancy

Duration of treatment / exposure:

gestation days 6 to 20

Frequency of treatment:

daily, at approximately the same time each time

Duration of test:

1 month

Remarks:

Doses / Concentrations:
0, 15, 25, 50, 100 and 200 mg/kg bw/day
Basis:
actual ingested

No. of animals per sex per dose:

20-23 animals/dose

Control animals:

yes, concurrent vehicle

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes, daily
DETAILED CLINICAL: no data
BODY WEIGHT : Yes, on day 0 and every 3 days from days 6 to 21 of gestation.
POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 21
- Organs examined: uterus

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: No data
- Number of corpora lutea: No data
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Live and dead fetuses

Fetal examinations:

Fetal body weight & Fetal malformations
live fetuses were removed, weighed, sexed and examined.
- External examinations: Yes: all per litter (including those of the oral cavity)
- Soft tissue examinations: Yes: half per litter
- Skeletal examinations: Yes:half per litter
- Head examinations: Yes: No data

Statistics:

Whenever possible, the data were presented as means ± SD. Implantation sites, live fetuses, and various body weights were analyzed by the one-way analysis of variance, followed by Dunnett's test if differences were found. The frequency of nonsurviving implants, resorptions, and anomalies among litters was evaluated with the Dixon-Massey test after an arc-sine-square root transformation. Rates of pregnancy and fetal sex ratio were analyzed using Fisher's exact test. Where applicable, least-squares analysis was carried out. The reported level of statistical significance was p < 0.05.

Indices:

no data

Historical control data:

no data

Details on maternal toxic effects:

No maternal deaths, morbidity, or treatment-related clinical signs were observed during this study. Significant reduction of maternal body weight gain was observed in tha last third of gestation at 15 mg/kg bw/d and throughout treatment at higher dose levels. The maternal weight gain during days 6 to 21 of gestation and the absolute weight gain were significantly reduced at all doses (p<0.01) (table 1). Incidences of pregnancy were comparable among groups (table 2).

Dose descriptor:

NOAEL

Effect level:

< 15 mg/kg bw/day

Basis for effect level:

other: maternal toxicity

Details on embryotoxic / teratogenic effects:

No significant effect of DMTU was noted on the mean numbers of implantation sites, viable fetuses, and percentage of resorptions and on fetal sex ratio. fetal body weights were significantly reduced at all doses tested, showing a dose-depenent relationship. The 9% decrease at 15 mg/kg bw/d reached 34% at 200 mg/kg bw/d (Table 2). The decrease in maternal weight was considered severe aand the fetal effects caused by DMTU were probably secondary to the toxicity incurred by the mothers.
No animalies were seen upon external examination of the fetuses (table 3). Dilated ureter and extra lumbar ribs occurred significantly more often among litters of rats given 200 mg/kg bw/d (p<0.01 and p<0.05, respectively). No other visceral and skeletal variants were significantly affected at any tested dose. In the control group, one fetus exhibited a forelimb syndactiyly and onother a bilateral microphtalmia.

Dose descriptor:

NOAEL

Effect level:

< 15 mg/kg bw/day

Sex:

male/female

Basis for effect level:

fetal/pup body weight changes

Dose descriptor:

NOAEL

Effect level:

> 200 mg/kg bw/day

Basis for effect level:

other: teratogenicity

Abnormalities:

not specified

Developmental effects observed:

not specified

Table 1 : change in weight during gestation in SD rats treated daily by gastric intubation with DMTU on days 6 to 20 of gestation and sacrified on day 21^a

Compound	Dose	Body weight (g) on GD6	Body weight gain (g)						Absolute weight gain (g)^b
			GD 6-9	GD 9-12	GD 12/15	GD 15-18	GD 18-21	GD 6-21
Distilled water (ml/kg/d)	5	257+/-14	18+/-5	16+/-6	24+/-5	48+/-7	59+/-10	162+/-19	46+/-17
DMTU (mg/kg/d)	15	255+/-16	15+/-4	15+/-5	19+/-6	40+/-9*	46+/-11**	135+/-24**	33+/-10**
	25	255+/-14	12+/-6*	16+/-5	17+/-6**	38+/-9**	41+/-10**	126+/-24**	28+/-13**
	50	256+/-11	6+/-5**	19+/-6	15+/-5**	35+/-11**	34+/-9**	109+/-21**	19+/-10**
	100	253+/-15	3+/-10**	11+/-7*	14+/-7**	25+/-8**	35+/-9**	88+/-20**	3+/-10**
	200	257+/-16	-1+/-4**	14+/-7	8+/-6**	22+/-8**	27+/-9**	71+/-14**	-2+/-11**
Data are expressed as means +/- SD ^aIncludes all dams pregnant at sacrifice. GD = gestational day. ^b(day 21 bw) – (gravid uterus weight) – (day 6 bw) * and ** denote significant differences from the vehicle control value using Dunnett’s test, p<0.05 and p<0.01, respectively.

 

 

Table 2 : Reproductive parameters in SD rats treated daily by gastric intubation with DMTU on days 6 to 20 of gestation and sacrified on day 21^a

Compound	Dose	No. of deaths per number of treated females	% of females pregnant	Number of examined litters	Mean implantation sites per litter	Mean live fetuses per litter	Mean % nonsurviving implants per litter^b	Mean % resorption sites per litter
Distilled water (ml/kg/d)	5	0/20	95.0	19	15.26+/-1.94	14.95+/-1.96	2.02+/-3.70	2.02+/-3.70
DMTU (mg/kg/d)	15	0/21	85.7	18	14.39+/-3.47	14.06+/-3.45	2.24+/-4.08	2.24+/-4.08
	25	0/21	95.2	20	14.50+/-3.32	14.15+/-3.59	3.63+/-6.44	3.63+/-6.44
	50	0/21	90.5	19	13.79+/-3.85	13.26+/-3.75	3.49+/-5.55	3.49+/-5.55
	100	0/23	95.6	22	14.81+/-3.17	14.04+/-2.90	4.73+/-6.14	4.73+/-6.14
	200	0/23	87.0	20	14.40+/-2.85	13.70+/-2.87	4.68+/-7.19	4.27+/-7.20
Data are expressed as means +/- SD ^aIncludes all dams pregnant at sacrifice. GD = gestational day. ^bResorption plus dead fetuses * and ** denote significant differences from the vehicle control value using Dunnett’s test, p<0.05 and p<0.01, respectively.

 

Compound	Dose	Fetal sex ratio M:F (%)	Mean fetal body weight (g) per litter
			Males	Females
Distilled water (ml/kg/d)	5	0.86	6.00+/-0.30	5.68+/-0.29
DMTU (mg/kg/d)	15	1.14	5.47+/-0.33**	5.19+/-0.34**
	25	0.96	5.29+/-0.30**	5.04+/-0.25**
	50	1.15	5.20+/-0.33**	4.80+/-0.36**
	100	0.98	4.40+/-0.28**	4.26+/-0.29**
	200	0.89	3.94+/-0.28**	3.76+/-0.24**
Data are expressed as means +/- SD * and ** denote significant differences from the vehicle control value using Dunnett’s test, p<0.05 and p<0.01, respectively.

 

 

Table 3 : Incidence of anomalies in fetuses of SD rats teated daily by gastric intubation with DMTU on days 6 to 20 of gestation and sacrified on day 21

 

 

	Distilled Water (ml/kg/d)	DMTU (mg/kg/d)
	5	15	25	50	100	200
	Number of fetuses (litters) examined
External examination	284 (19)	253 (18)	283 (20)	252 (19)	309 (22)	274 (20)
Soft tissue examination	142 (19)	127 (18)	142 (20)	126 (19)	154 (22)	137 (20)
Skeletal examination	142 (19)	126 (18)	141 (20)	126 (19)	155 (22)	137 (20)
	Number of fetuses (litters) affected
Soft tissue anomalies
Microphtalmia^b	1 (1)	0	0	0	0	0
Hydroureter	0	0	0	0	0	3 (2)
Dilated renal pelvis	1 (1)	0	0	0	1 (1)	5 (2)
Dilated ureter	0	0	0	0	1 (1)	18 (11) **
Skeletal anomalies Interparietal and/or occipital not ossified	0	0	0	0	0	0
Vertebral centra dumbbell-shaped or absent	0	2 (2)	0	2 (2)	3 (3)	4 (3)
Extra lumbar ribs	9 (4)	9 (6)	8 (7)	13 (6)	14 (8)	13 (13)*
Fifth sternebrae Not ossified	4 (4)	0	1 (1)	0	0	2 (2)
Forelimb syndactilyly	1 (1)	0	0	0	0	0

 

^bConsidered to be a malformation * and ** denote significant differences from the vehicle control value, p<0.05 and p<0.01, respectively.

 

Conclusions:

DMTU was not teratogenic in rat at 200 mg/kg bw/day, which was maternally toxic. But DMTU was fetotoxic at all doses tested in the presence of maternal toxicity.

Executive summary:

A read-across between these two substances is justified for the reproduction endpoint.

Because DETU and DMTU are two thioureas having a high structural similarity: both structures differ by a carbon by chain.

Moreover DETU and DMTU have the same physicalchemical properties, and are harmful by oral route (acute toxicity).

Saillenfait,et al (1991), tested DMTU in SD rats at 0, 15, 25, 50, 100 or 200 mg/kg/day from GD 6-20.

The only maternal toxicity noted was decreased body weight gain at 15 mg/kg/day and above (NOAEL of maternal toxicity < 15 mg/kg bw/d). The LOAEL for developmental toxicity was 15 mg/kg/day based on decrease of fetal body weight gain at all doses. The decrease in maternal weight was considered severe and the fetal effects caused by DMTU were probably secondary to the toxicity incurred by the mothers. Dilated ureter and extra lumbar ribs were observed at 200 mg/kg bw/d (significant results).

No teratogenic effects related to administration of DMTU were observed at dose levels up to 200 mg/kg bw/d, a level which was toxic to the dams.

Reference 3

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Qualifier:

no guideline followed

Principles of method if other than guideline:

Developmental study was performed on rats by dermal route. Rats were exposed on day 12 of gestation only at dose levels of 450, 670, 1000, 1500 and 2250 mg/kg bw.

GLP compliance:

no

Limit test:

yes

Species:

rat

Strain:

other: Charles River-CD

Details on test animals or test system and environmental conditions:

Primigravida femelles.

Route of administration:

dermal

Vehicle:

DMSO

Details on exposure:

DETU was applied on the clipped back skin of rat.

Analytical verification of doses or concentrations:

no

Details on mating procedure:

no data

Duration of treatment / exposure:

a single exposure on gestation day 12

Frequency of treatment:

a single exposure

Remarks:

Doses / Concentrations:
450, 670, 1000, 1500 and 2250 mg/kg bw
Basis:
nominal conc.

No. of animals per sex per dose:

5 females/group

Control animals:

yes, concurrent vehicle

Details on study design:

The animals were sacrified on the twentieth day of gestation and the observations and determinations were made.

Maternal examinations:

no data

Ovaries and uterine content:

gross examinations of uterus and fetuses, number of implantation sites, number of live fetuses, number of early resorptions, number of late resorptions.

Fetal examinations:

fetal weight and fetal crown-rump length.
All fetuses were saved in appropriate fixatives for possible future determination of skeletal and visceral anomalies.

Statistics:

no data

Indices:

no data

Historical control data:

no data

Details on maternal toxic effects:

Maternal toxic effects:no effects
DETU was not toxic to pregnant rats, no mortality was observed.

Dose descriptor:

NOAEL

Effect level:

> 2 250 mg/kg bw/day

Basis for effect level:

other: maternal toxicity

Dose descriptor:

NOAEL

Effect level:

> 2 250 mg/kg bw/day

Basis for effect level:

other: developmental toxicity

Details on embryotoxic / teratogenic effects:

DETU was not toxic to pregnant rats, not embryolethal and not teratogenic under the conditions of this test at levels ranging from 450 to 2250 mg/kg bw. All animals (at all levels) delivered normal litters. All parameters used to measure the outcome of pregnancy and fetal development were similar to those of the control DMSO-treated females.

Table of results : Effects of a single topical application of DETU on the outcome of pregnancy and fetal development.

 

Compound	Number of females			Number per pregnant female				Number of fetus with anomalies
	pregnant	Not pregnant	Died	Implantation sites	Early Resorption sites	Late resorptions	Live fetuses
DMSO	4	1	0	10.8	0.3	0	10.5	None
DETU	4	1	0	10.0	1.5	0	8.5	None

Mean of all treated-animals (at all doses)

Conclusions:

DETU was not toxic to pregnant rats, not embryolethal and not teratogenic under the conditions of this test at levels ranging from 450 to 2250 mg/kg bw.

Executive summary:

The purpose of this developmental study was to determine the Appropriate lethal dose (ALD) of DETU for pregnant rats and to evaluatae embryotoxic and teratogenic potential. The test material was suspended in DMSO and a single dose was applied on the clipped back skin of primigravida Charles River-CD rats on day 12 of gestation at dose levels of 450, 670, 1000, 1500 and 2250 mg/kg bw. Five pregant females were treated with DMSO and served as controls.

The animals were sacrified on the twentieth day of gestation and the following observations and determinations were made: gross examinations of uterus and fetuses, number of implantation sites, number of live fetuses, number of early resorptions, number of late resorptions, fetal weight and fetal crown-rump length. All fetuses were saved in appropriate fixatives for possible future determination of skeletal and visceral anomalies.

DETU was not toxic to pregnant rats, not embryolethal and not teratogenic under the conditions of this test at levels ranging from 450 to 2250 mg/kg bw. All animals (at all levels) delivered normal litters. All parameters used to measure the outcome of pregnancy and fetal development were similar to those of the control DMSO-treated females.

Effect on developmental toxicity: via oral route

Endpoint conclusion:

adverse effect observed

Dose descriptor:

LOAEL

15 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

A reliable study on developmental toxicity study on rats in available on Dimethylthiourea (DMTU). The study is reliable with a klimisch score of 1.

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

2 250 mg/kg bw/day

Study duration:

subacute

Species:

rat

Additional information

Developmental study on DMTU by oral route (read-across)

Saillenfait et al. (1991), tested DMTU in SD rats at 0, 15, 25, 50, 100 or 200 mg/kg/day from GD 6-20.

The only maternal toxicity noted was decreased body weight gain at 15 mg/kg/day and above (NOAEL of maternal toxicity < 15 mg/kg bw/d). The LOAEL for developmental toxicity was 15 mg/kg/day based on decrease of fetal body weight gain at all doses. The decrease in maternal weight was considered severe and the fetal effects caused by DMTU were probably secondary to the toxicity incurred by the mothers. Dilated ureter and extra lumbar ribs were observed at 200 mg/kg bw/d (significant results).

No teratogenic effects related to administration of DMTU were observed at dose levels up to 200 mg/kg bw/d, a level which was toxic to the dams.

Developmental study on DETU by dermal route

DETU was suspended in DMSO and a single dose was applied on the clipped back skin of primigravida Charles River-CD rats on day 12 of gestation at dose levels of 450, 670, 1000, 1500 and 2250 mg/kg bw. Five pregant females were treated with DMSO and served as controls.

The animals were sacrified on the twentieth day of gestation and the following observations and determinations were made: gross examinations of uterus and fetuses, number of implantation sites, number of live fetuses, number of early resorptions, number of late resorptions, fetal weight and fetal crown-rump length. All fetuses were saved in appropriate fixatives for possible future determination of skeletal and visceral anomalies.

DETU was not toxic to pregnant rats, not embryolethal and not teratogenic under the conditions of this test at levels ranging from 450 to 2250 mg/kg bw. All animals (at all levels) delivered normal litters. All parameters used to measure the outcome of pregnancy and fetal development were similar to those of the control DMSO-treated females.

Justification for classification or non-classification

Proposed self-classification (Regulation (EC) No 1272/2008) :

Not classified.

Current conclusion based on experimental data generated on an analogue: No teratogenic effects were observed in the oral developmental study. A foetotoxicity (decrease of fetal bodyweight) was observed but is due to the maternal toxicity.

This conclusion will be revised upon receiving the results from the currently ongoing studies.