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Key value for chemical safety assessment

Genetic toxicity in vivo

Description of key information
Thiourea did not induce gene mutation in bacteria, but mixed results were obtained in assays in mammalian cells. It consistently induced chromosomal recombination in yeast and insects and induced mammalian cell transformation. Thiourea did not induce genotoxicity in an in vivo micronucleus test.
Link to relevant study records
Reference
Endpoint:
in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus
Remarks:
Type of genotoxicity: chromosome aberration
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
February 1979
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Comparable to guideline study with acceptable restrictions
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
Deviations:
yes
Remarks:
: 400 instead of 200 erythrocytes were examined
GLP compliance:
no
Type of assay:
micronucleus assay
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Central institute for the breeding of laboratory animals TNO, Zeist
- Age at study initiation: 4-5 weeks
- Assigned to test groups randomly: the animals were assigned to three test groups of 5 males and 5 females according body weight
- Fasting period before study: deprivation of food for 14-15 h
- Housing: screen bottom cages
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: yes

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 24+/-1°C
- Humidity (%): 40-60 %
- Air changes (per hr): no data. air conditioned rooms
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: gavage
Vehicle:
- Vehicle(s)/solvent(s) used: water
- Justification for choice of solvent/vehicle:
- Concentration of test material in vehicle: 350 mg/kg bw in 5 ml water per kg bw
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: freshly prepared immediately before treatment of the animals

Duration of treatment / exposure:
The test samples were administered twice with an interval of 24 h. 6 hours after the last treatment the animals were killed.
Frequency of treatment:
twice
Post exposure period:
6 hours
Remarks:
Doses / Concentrations:
350 mg/kg body weight
Basis:
nominal in water
Remarks:
Doses / Concentrations:
7 % Thiourea
Basis:
nominal in water
No. of animals per sex per dose:
5 males and 5 females
Control animals:
yes, concurrent vehicle
Positive control(s):
2,3,5-tris-ethyleneiminobenzoquinone (Trenimon; Bayer) was used as a solution in physiological saline
- Justification for choice of positive control(s): no data
- Route of administration: oral
- Doses / concentrations: 0.0625 mg/kg body weight
Tissues and cell types examined:
bone marrow of the femora; erythrocytes
Details of tissue and slide preparation:
The bone marrow of the femora was flushed into centrifuge tubescontaining foetal calf serum and centrifuged. The excess serum was removed. The cells were than resuspended by mixing gently with a pasteur pipette. A drop was placed on a slide cleaned with methanol overnight and spread with a haemocytometer cover glass. Five slides were prepared of each animal. The smears were air dried, fixed in methanol and stained according to May-Grünwald.
Sex:
male/female
Genotoxicity:
negative
Toxicity:
not examined
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
RESULTS OF RANGE-FINDING STUDY
The dose-levels used in the present study were based on acute oral LD50 of the test material for rats (CIVO letters 29/12/75)

RESULTS OF DEFINITIVE STUDY
- Induction of micronuclei (for Micronucleus assay): Oral administration of thiourea did not affect the incidence of micronucleated erythrocytes
- Ratio of PCE/NCE (for Micronucleus assay):Oral administration of thiourea did not affect the ratio of poly and normochromatic erythrocytes
- Appropriateness of dose levels and route:
- Statistical evaluation: standard deviations were determined

No mortality or abnormalities of condition or behaviour, attributable to treatment were observed in any of the animals during the exposure period.

Table 1: Average body weights of groups of 5 rats

Group No

Treatment/kg body weight

Average body weight +/- SD

males

females

390

2x5ml distilled water

99+/-3

79+/-7

392

2x5 ml 7% Thioharnstoff

96+/-4

81+/-5

Table 2: Mean numbers of micronucleated erythrocytes and percentage polychromatic erythrocytes in bone marrow of rats

Treatment

Sex

Incidence of micronucleated cells per 2000 erythrocytes per rat

Percentage polychromatic erythrocytes +/-SD

mean

range

Vehicle control

Male

4.4

2-7

74.2+/-3.5

Thioharnstoff

Male

5.4

4-8

72.0+/-3.7

Vehicle control

Female

5.0

2-8

63.8+/-5.1

Thioharnstoff

female

4.0

4-6

62.9+/-3.5

Conclusions:
Interpretation of results (migrated information): negative
Thiourea did no increase the frequency of micronucleated polychromatic erythrocytes in bone marrow. In this study thiourea did not reveal any evidence for mutagenic activity.
Executive summary:

In a Wistar rats bone marrow micronucleus assay, 5 male and 5 female animals per dose were treated by oral gavage with Thiourea (7 % a.i.) at doses of 0 and 350 mg/kg bw. Bone marrow cells were harvested at 6 hours post-treatment. The vehicle was water. There were no signs of toxicity during the study. Thiourea was tested at an adequate dose based on the results of an oral acute toxicity test. The positive control induced the appropriate response. There was not a significant increase in the frequency of micronucleated polychromatic erythrocytes in bone marrow after any treatment time. This study is classified as acceptable. This study satisfies the requirement for Test Guideline OECD 474 for in vivo cytogenetic mutagenicity data.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Additional information

Additional information from genetic toxicity in vivo:

Thiourea did not induce gene mutation in bacteria, but mixed results were obtained in mammalian cells. It consistently induced chromosomal recombination in yeast and insects and induced mammalian cell transformation. Thiourea is reported to elicit cytotoxicity in mammalian cell cultures. In several in vitro tests cytotoxicic effects have not been reported making it difficult to verify any positive result. Thiourea did not induce genotoxicity in an in vivo micronucleus test in rats.

Based on the available data the MAK commission concluded in 1990 that Thiourea is weak genotoxic. The WHO/IPCS concluded in 2003 that thiourea is not considered to be a genotoxic carcinogen and that the development of thiourea-induced thyroid tumours may involve inhibition of peroxidase in the thyroid gland, leading to decreased thyroid hormone production and increased proliferation as a result of an increase in the secretion of thyroid-stimulating hormone.

Based on the overall assessment of the available data Thiourea is not considered being genotoxic. This is in agreement with the WHO/IPCS conclusion (2003).

Andrae (1989) tested the Thiourea metabolite FASA (formamidine sulfinate) for its genotoxic effects on V79 hepatocytes of rats. FASA is formed enzymatically by FMO (FAD-dependent monooxygenase) and is inducing several genotoxic effects like induction of gene mutation, micronuclei formation and DNA repair. The author concluded that FASA would be responsible for the genotoxic effects elicited by thiourea if it would be formed in vivo, which was not verified in this study.

The results obtained in the PhD thesis of Timme Madlon (1998) on "Studies on the mechanism and significance of thiourea genotoxicity in mammalian cells in vitro and in vivo – The genotoxicity and metabolism of thiourea in human thyroid follicle cells" showed that thiourea was marginally genotoxic to human thyroid follicle cells. The genotoxicity was indicated by a slight increase in the frequency of DNA strand breaks in the cells and possibly due to slight auto oxidation of thiourea to electrophilic S oxygenated products. The investigations yielded no indication of organ-specific metabolic activation of thiourea in thyrocytes. It can therefore be assumed that human exposure to thiourea is not associated with a relevant risk of thyroid tumours due to genotoxic effects.

The following tables summarize several of the available test reports, some are not included as individual IUCLID study records, but to emphasise the amount of available, ambiguous data.

Genotoxicity and related endpoints: in vitro

Table1: summary of available studies on genotoxicity in vitro

Endpoint

Results and references

Species, strain

Result

Metabolic activation

Reference

Genotoxicity and related endpoints:in vitro

Gene mutation

Salmonella typhimuriumTA 97

Negative

+/-

Brams et al. 1987
Zeiger et al. 1988

Salmonella typhimuriumTA1535, TA100, TA1537, TA1538, TA98

Negative

+/-

Korte und Greim, 1981

Brusick, 1977

Salmonella typhimuriumTA 98

Negative

+/-

Simmon 1979a*
Dunkel et al. 1984*
Brams et al. 1987
Zeiger et al. 1988

Salmonella typhimuriumTA 100

Negative

+/-

Simmon 1979a*
Yamaguchi 1980*
Dunkel et al. 1984*
Brams et al. 1987
Zeiger et al. 1988

Salmonella typhimuriumTA1535

Negative

+/-

Simmon 1979a*
Rozenkranz and Poirier 1979*
Dunkel et al. 1984*
Zeiger et al. 1988

Salmonella typhimuriumTA1535/pSK1002 umu test

Negative

+/-

Nakamura et al. 1987

Salmonella typhimuriumTA1537

Negative

+/-

Simmon 1979°*
Dunkel et al. 1984*

Salmonella typhimuriumTA1538

Negative

+/-

Rozenkranz and Poirier 1979*
Simmon 1979a*
Dunkel et al. 1984*

Escherichia coli SOS repair

Negative

+/-

Brams et al. 1987
Kevekordes et al. 1999*

Escherichia coliWP2uvrA

Negative

+/-

Dunkel et al. 1984*

Escherichia colipol A

Negative

+

Rosenkranz and Poirier 1979*
McCarroll et al. 1981

-

McCarroll et al. 1981

Escherichia coliuvr/rec strains

Positive

-

Hellmér and Bolcsfoldi 1992

Negative

+

Hellmér and Bolcsfoldi 1992

Escherichia coliK12

Negative

+

Mamber et al. 1984*

Not tested

-

Mamber et al. 1984*

Escherichia coliRK

Negative

+/-

Hayes et al. 1984*

Aspergillus nidulans

Negative

-

Crebelli et al. 1986

Saccharamyces cerevisiae

Positive

-

Schiestl et al. 1989
Galli and Schiestl 1995, 1996, 1998*

+

Galli and Schiestl 1998*
Egilsson et al. 1979*
Wilkie and Gooneskera 1980*

ambigous

+/-

Zoushu, 1989

Saccharamyces cerevisiaetrp locus

Positive

+

Morita et al. 1989

Negative

-

Morita et al. 1989

Saccharamyces cerevisiaeD7 trp locus

Positive

Not indicated

Jiang et al. 1989*

Drosophila melanogaster

Positive

-

Batiste-Alentorn et al. 1991*

Weakly positive

-

Vogel & Nivard 1993*

Inconclusive

-

Batiste-Alentorn et al. 1994, 1995*

Negative

-

Rodriguez-Arnaiz 1997*

Mouse L5178Y cells

Weakly positive

+

Caspary 1988
Wangenheim and Bolcsfoldi 1988

-

Wangenheim and Bolcsfoldi 1988

Negative

+

Mitchell et al. 1988*

-

Mitchell et al. 1988*
Myhr and Caspary 1988*

Chinese hamster ovary cells (V79),hprtlocus

Negative

-

Bradley et al. 1982*

Positive

+/-

Ziegler-Skylakakis et al. 1985*

Chinese hamster ovary cells (V79), cell sub-line Sp5

Negative

-

Helleday et al. 1998*

Sister chromatid exchange

Chinese hamster V79 cells

Negative

-

Bradley et al. 1982*

Chinese hamster Ovary (CHO)

Positive

+/-

TNO 1978; de Raat

Unscheduled DNA synthesis

Rat liver cells

Weakly positive

-

Ziegler-Skylakakis et al. 1985*

Rat liver cells

Negative

-

Lonati-Galligani et al. 1983*
Fautz et al. 1991

Mitotic recombination

Saccharamyces cerevisiaeD3

Negative

+/-

Simmon 1979b*

Recombination

Transformed human lymphoblastoid GM6804 cells

Positive

-

Aubrecht et al. 1995*

Micronucleus formation

Syrian hamster embryo cells

Positive

-

Fritzenschaf et al. 1993

Chinese hamster V79

Weakly positive

 

Ziegler-Skylakakis et al. 1998*

DNA strand breaks

Rat hepatocytesin vitro

Positive

-

Sina et al. 1983*

Rat hepatocytesin vitro

 Negative

-

Fautz et al. 1991

Cell transformation

Syrian hamster embryo cells

Positive

-

Pienta et al. 1977*

Rauscher virus-infected rat embryo cells

Positive

-

Dunkel et al. 1981*

Bovine papilloma virus DNA-enhanced C3H10T1/2 cells

Weakly positive

-

Kowalski et al. 2000*

DNA synthesis inhibition

Human fibroblast cells

Positive

-

Painter 1977*

* studies referred to in IPCS 2003, Health Canada 2008, IARC 2001, or MAK 1990

 

Genotoxicity and related endpoints: in vivo

Table2: summary of available studies on genotoxicity in vivo

Endpoint

Results and references

Species, strain

Result

Metabolic activation

Reference

Gene mutation

Drosophila melanogaster

Positive

n.a.

Batiste-Alentorn et al. 1991

Drosophila melanogaster

Weakly Positive

n.a.

Vogel and Nivard 1993

Drosophila melanogaster

Negative

n.a.

Rodriguez-Arnaiz 1997

Drosophila melanogaster

Inconclusive

n.a.

Batiste-Alentorn et al. 1994, 1995*

Micronucleus test

Rats treated with two successive oral doses within 24 hours

Negative

n.a.

TNO 1979b



Justification for selection of genetic toxicity endpoint
mammalian in vivo micronucleus test similar to OECD 474

Justification for classification or non-classification

Thiourea did not induce gene mutation in bacteria, but mixed results were obtained in assays in mammalian cells. It consistently induced chromosomal recombination in yeast and insects and induced mammalian cell transformation. Based on the overall assessment of available data the test item is not considered a mutagen.