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Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

In a bacterial reverse mutation (Ames) test, conducted according to OECD Test Guideline 471 and to GLP, dihydrogen hexachloroplatinate, compound with 2-aminoethanol (1:2) induced reverse mutations in strains of Salmonella typhimurium and Escherichia coli both in the absence and presence of a rat liver metabolic activation system (S9) (Scarcella, 2001).

 

In an in vitro mammalian cell gene mutation assay, conducted in accordance with OECD Test Guideline 476 and to GLP, dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol (1:2) induced mutations in Chinese hamster V79 cells when tested both with and without S9 (Cinelli, 2002).

 

No in vitro mammalian cell cytogenicity data are available for dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol (1:2). Further in vitro testing in considered unnecessary in the light of the available in vivo genotoxicity test data.

Link to relevant study records
Reference
Endpoint:
in vitro gene mutation study in bacteria
Remarks:
Type of genotoxicity: gene mutation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
23 April - 25 June 2001
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
Study conducted according to GLP
Qualifier:
according to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Type of assay:
bacterial reverse mutation assay
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Species / strain / cell type:
E. coli WP2 uvr A
Metabolic activation:
with and without
Metabolic activation system:
Sprague-Dawley rat liver induced with phenobarbitone and betanaphthoflavone
Test concentrations with justification for top dose:
Two main experiments.
In the first, using the plate incorporation method, the test substance was assayed at 313, 625, 1250, 2500 or 5000 ug/plate in all five tester strains, in the absence or presence of S9. In the second, TA98, TA100, and WP2 uvrA strains were tested under the same experimental conditions as the first experiment, while for TA1535 and TA1537, a pre-incubation step was included.
Vehicle / solvent:
Sterile distilled water
Untreated negative controls:
yes
Remarks:
Dimethoxysulphoxide (DMSO)
Negative solvent / vehicle controls:
yes
Remarks:
Untreated
Positive controls:
yes
Positive control substance:
sodium azide
Remarks:
Migrated to IUCLID6: in distilled water
Positive controls:
yes
Positive control substance:
9-aminoacridine
Remarks:
Migrated to IUCLID6: in DMSO
Positive controls:
yes
Positive control substance:
2-nitrofluorene
Remarks:
Migrated to IUCLID6: in DMSO
Positive controls:
yes
Positive control substance:
other: 2-Aminoanthracene in DMSO
Positive controls:
yes
Positive control substance:
methylmethanesulfonate
Remarks:
Migrated to IUCLID6: in distilled water
Evaluation criteria:
For the test substance to be considered mutagenic, a two-fold (or more) increase in the mean revertant numbers must be observed at two-consecutive dose-levels or at the highest practiable dose-level only. In addition, there must be evidence of a dose-reponse relationship.
Statistics:
Regression analysis by the least squares method
Species / strain:
S. typhimurium, other: TA98, TA100
Metabolic activation:
with and without
Genotoxicity:
positive
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
E. coli WP2 uvr A
Metabolic activation:
with and without
Genotoxicity:
positive
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
S. typhimurium TA 1537
Metabolic activation:
with and without
Genotoxicity:
positive
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
In main experiment one, two-fold, dose-related increases in revertant colonies compared to controls were seen in TA98, TA100, and WP2 uvrA strains, with and without S9.
In main experiment two, the test substance induced reproducible, dose-related, two-fold increases in the number of revertant colonies compared to controls in TA98, TA100, and WP2 uvrA strains, with and without S9. Two-fold increases in TA1537 were also seen using the pre-incubation method, at non-toxic dose-levels, with and without S9.
Conclusions:
In an OECD Test Guideline 471 study, to GLP, dihydrogen hexachloroplatinate, compound with 2-aminoethanol (1:2) induced reverse mutations in strains of Salmonella typhimurium and Escherichia coli both with and without metabolic activation.
Executive summary:

In an OECD Test Guideline 471 study, conducted according to GLP, dihydrogen hexachloroplatinate, compound with 2-aminoethanol (1:2) was assessed for its ability to induce gene mutations in strains of S. typhimurium (TA1535, TA1537, TA98, TA100) and E. coli (WP2 uvrA). The test was performed in two independent experiments (involving the plate incorporation method, including a pre-incubation step for TA1535 and TA1537 in Experiment 2), with dose levels of up to 5000 μg/plate (determined following an initial toxicity test), both in the absence and presence of rat liver metabolic activation (S9).

In experiment 1, dose-related increases in revertant numbers, which were at least two-fold the control values, were observed in WP2 uvrA both with and without S9. The test item induced reproducible, large and dose-related increases in the number of revertant colonies, which were more than two-fold the control values, with TA98, TA100 and WP2 uvrA tester strains in the plate incorporation assays, at the higher dose levels both in the presence and absence of S9 metabolism. Increases in revertant numbers were also observed with TA1537 in the pre-incubation assay. These increases were greater than twice the control value at non-toxic dose-levels, both in the presence and absence of S9 metabolism.

It was concluded that dihydrogen hexachloroplatinate, compound with 2-aminoethanol (1:2) was mutagenic in S. typhimurium and E.coli under the reported experimental conditions.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed (positive)

Genetic toxicity in vivo

Description of key information

The in vivo genotoxicity of dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol, as evaluated by its ability to induce micronuclei in polychromatic erythrocytes and to cause DNA damage, was assessed in a combined study following OECD 474 and 489 and according to GLP. Male Wistar rats (5/group) were given gavage doses of 500, 1000 or 2000 mg/kg bw/day of the test item on three consecutive days. Comet analyses were conducted on preparations of liver, glandular stomach, duodenum and kidney tissues and micronuclei were analysed in bone marrow cells.

 

There was no evidence of an increase in the incidence of micronucleated polychromatic erythrocytes. There was no increase in % tail intensity in the liver, glandular stomach, kidney or duodenum (Eurlings, 2020). As such, and as platinum was detected in the plasma of the test animals, the test item was considered to be non-genotoxic in vivo.

Link to relevant study records

Referenceopen allclose all

Endpoint:
in vivo mammalian cell study: DNA damage and/or repair
Type of information:
experimental study
Adequacy of study:
key study
Study period:
11 Feb 2020 - 29 Jun 2020
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
An EPMF member company received an official request from the Korean authorities to perform the in vivo mutagenicity assay with hexahydroxyplatinate,compound with 2-aminoethanol(1:2) (CAS 68133-90-4), ahead of the formal testing proposal approval by ECHA. The requested experimental information is in line with the assay submitted in the TP for this substance. The deadline given by the Korean authorities was however much tighter than the anticipated date of receiving the final decision on the TP. Therefore, the in vivo mutagenicity testing with this substance has been initiated in the EU prior to receiving the final decision on the TP.

In the attached document (translation from Korean and anonymised), a justification for initiating the in vivo muta testing upon official request from an non-EU authority (i.e. request outside EU-REACH), ahead of receiving the final decision on the TP. More information is available upon request.
Reason / purpose for cross-reference:
reference to same study
Qualifier:
according to guideline
Guideline:
OECD Guideline 489 (In vivo Mammalian Alkaline Comet Assay)
Version / remarks:
29 July 2016.
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Type of assay:
mammalian comet assay
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Lot/batch number of test material:
19267COLPT.
- Expiration date of the lot/batch: 01 October 2021.
- Purity test date: CoA issued 17 December 2019.

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: At room temperature.

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: None.
- Final preparation of a solid: Test item was suspended in corn oil.

FORM AS APPLIED IN THE TEST (if different from that of starting material)
: Suspension.
Species:
rat
Strain:
Wistar
Details on species / strain selection:
The Wistar Han rat was the species and strain of choice because it is a readily available rodent which is commonly used for genotoxicity testing, with documented susceptibility to a wide range of toxic items. Moreover, historical control background data has been generated with this strain.
Sex:
male
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany.
- Age at study initiation: 6 weeks.
- Weight at study initiation: 171 ± 8.7 g (Mean body weight ± SD).
- Assigned to test groups randomly: Yes.
- Fasting period before study: No.
- Housing: Up to 5 animals of the same sex and in the same dosing group were housed together.
- Diet: Commercial pellets ad libitum, except during designated procedures.
- Water: Tap water, ad libitum.
- Acclimation period: At least 6 days.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 to 25°C.
- Humidity (%): 40 to 70%.
- Air changes (per hr): ≥ 10.
- Photoperiod: 12 hrs light/12 hrs dark, except during designated procedures.

IN-LIFE DATES:
From: Not specified.
To: 12 Mar 2020.
Route of administration:
oral: gavage
Vehicle:
- Vehicle(s)/solvent(s) used: corn oil.
- Source of vehicle: Fagron Farmaceuticals, Capelle a/d IJssel, the Netherlands.
Duration of treatment / exposure:
Three consecutive days.
Frequency of treatment:
Daily.
Post exposure period:
Tissue samples taken 3 - 4 hours after administration of final dose.
Dose / conc.:
500 mg/kg bw/day (actual dose received)
Dose / conc.:
1 000 mg/kg bw/day (actual dose received)
Dose / conc.:
2 000 mg/kg bw/day (actual dose received)
Remarks:
No treatment-related toxicity or mortality were seen in a preliminary dose range finding study in which three male and three female rats received three consecutive daily doses of 2000 mg/kg bw
No. of animals per sex per dose:
5
Control animals:
yes, concurrent vehicle
Positive control(s):
Ethyl methanesulphonate.
- Route of administration: Gavage.
- Doses / concentrations: 200 mg/kg bw, dissolved in physiological saline, administered twice.
Tissues and cell types examined:
Cells were isolated from the liver, glandular stomach, duodenum and kidney.
Details of tissue and slide preparation:
Minced liver or kidney tissue was added to collagenase and dissolved in HBSS (saline). This suspension was shaken and centrifuged. The cell pellet was resuspended in HBSS and kept on ice prior to preparation of the slides.

Tissue from the glandular stomach and duodenum was stored on ice in "mincing buffer incomplete" (HBSS + EDTA). The surface epithelium of both the glandular stomach and duodenum was discarded as it contains a high proportion of apoptotic cells which distort the comet analysis. The cells, suspended in the buffer, were filtered though a 100 µm cell strainer and stored on ice prior to preparation of the slides.

Low melting point agarose was added to the cell suspensions and layered on a comet slide, which was then incubated for 13 - 39 minutes in the refrigerator.

Slides were kept overnight in the refrigerator, immersed in pre-chilled lysis solution. After rinsing, the slides were placed in freshly-prepared alkaline solution; electrophoresis was performed for 20 minutes (stomach and duodenum) or 30 minutes (liver and kidney). Following another rinse, the slides were immersed in absolute ethanol and allowed to dry, before staining with SYBR Gold fluorescent dye.
Evaluation criteria:
A test item was considered positive if all of the following criteria were met:
a) at least one treatment group demonstrated a statistically significant increase in % tail intensity vs. control.
b) the increase was dose-related.
c) any of the results were outside the 95% confidence limits of the historical control data.

If none of the above criteria were met, and direct or indirect evidence supportive of exposure of, or toxicity to, the target tissues was demonstrated, the test item was considered negative. If the data precluded making a conclusion of clearly positive or negative, the result was concluded as equivocal.
Statistics:
ToxRat Professional v 3.2.1 (ToxRat Solutions® GmbH, Germany) was used for statistical
analysis of the comet assay data .

A test item is considered positive in the Comet assay if all of the following criteria are met:
a) At least one of the treatment groups exhibits a statistically significant (one-sided, p <
0.05) increase in percentage Tail Intensity is detected compared with the concurrent
negative control.
b) The increase is dose related when evaluated with a trend test.
c) Any of the results are outside the 95% control limits of the historical control data range.

A test item is considered negative in the Comet assay if:
a) None of the treatment groups exhibits a statistically significant (one-sided, p < 0.05)
increase in percentage Tail Intensity is detected compared with the concurrent negative
control.
b) There is no concentration-related increase when evaluated with a trend test.
c) All results are within the 95% control limits of the negative historical control data range.
Key result
Sex:
male
Genotoxicity:
negative
Remarks:
Kidney: no statistically significant increase in % tail intensity.
Toxicity:
no effects
Vehicle controls validity:
valid
Positive controls validity:
valid
Key result
Sex:
male
Genotoxicity:
negative
Remarks:
Liver: no statistically significant increase in % tail intensity.
Toxicity:
not examined
Vehicle controls validity:
valid
Positive controls validity:
valid
Key result
Sex:
male
Genotoxicity:
negative
Remarks:
Glandular stomach: no statistically significant increase in % tail intensity.
Toxicity:
not examined
Vehicle controls validity:
valid
Positive controls validity:
valid
Key result
Sex:
male
Genotoxicity:
negative
Remarks:
Duodenum: no statistically significant increase in % tail intensity.
Toxicity:
not examined
Vehicle controls validity:
valid
Positive controls validity:
valid
Additional information on results:

Platinum was quantifiable in plasma samples from high-dose (2000 mg/kg bw/day) satellite animals 1, 3, 6 and 12 hours after completing the second day of treatment. Moreover, platinum was quantifiable in plasma samples from all high-dose animals taken at necropsy approximately 3-4 hours after the third dose. Therefore it was confirmed that the target tissues were exposed to the test item. No test item was detected in the animals dosed with vehicle.

Historical data Comet assay Negative control

 

Liver
Tail Intensity (%)

Males and Females

Duodenum
Tail Intensity (%)

Males and Females

Stomach
Tail Intensity (%)

Males and Females

Kidney
Tail Intensity (%)

Males and Females

Mean

1.96

3.06

2.45

12.10

SD

0.92

1.52

1.39

8.46

n

85

45

60

30

Lower control limit

(95% control limits)

0.27

-0.86

-1.07

-1.35

Upper control limit

(95% control limits)

3.65

6.97

5.96

25.55

SD = Standard deviation

n = Number of observations

 

Kidney: Historical control data from experiments performed in Feb 2012 – July 2019

Liver, Stomach, Duodenum: Historical control data from experiments performed in Jan 2018 – July 2019

Conclusions:
When tested in the comet assay, dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol, did not induce an increase in DNA damage in the liver, kidney, glandular stomach or duodenum of rats administered up to 2000 mg/kg bw/day by gavage on three consecutive days. As such, this compound was considered to negative under the conditions of this assay.
Executive summary:

The potential for dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol, to cause DNA damage was evaluated in a study following OECD 489 and according to GLP. Male Wistar rats (5/group) were given gavage doses of 500, 1000 or 2000 mg/kg bw/day of the test item on three consecutive days, or a vehicle control. The concurrent positive control group received two doses of EMS (200 mg/kg bw/day). Comet analyses were conducted on preparations of liver, glandular stomach, duodenum and kidney tissues.

 

There was no increase in % tail intensity in the liver, kidney, glandular stomach or duodenum, indicating that the test item is not genotoxic to these tissues.

Endpoint:
in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus
Type of information:
experimental study
Adequacy of study:
key study
Study period:
11 Feb 2020 - 29 Jun 2020
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
An EPMF member company received an official request from the Korean authorities to perform the in vivo mutagenicity assay with hexahydroxyplatinate,compound with 2-aminoethanol(1:2) (CAS 68133-90-4), ahead of the formal testing proposal approval by ECHA. The requested experimental information is in line with the assay submitted in the TP for this substance. The deadline given by the Korean authorities was however much tighter than the anticipated date of receiving the final decision on the TP. Therefore, the in vivo mutagenicity testing with this substance has been initiated in the EU prior to receiving the final decision on the TP.

In the attached document (translation from Korean and anonymised), a justification for initiating the in vivo muta testing upon official request from an non-EU authority (i.e. request outside EU-REACH), ahead of receiving the final decision on the TP. More information is available upon request.
Reason / purpose for cross-reference:
reference to same study
Qualifier:
according to guideline
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
Version / remarks:
29 July 2016
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Type of assay:
mammalian erythrocyte micronucleus test
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Lot/batch number of test material:
19267COLPT.
- Expiration date of the lot/batch: 01 October 2021.
- Purity test date: CoA issued 17 December 2019.

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: At room temperature.

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: None.
- Final preparation of a solid: Test item was suspended in corn oil.

FORM AS APPLIED IN THE TEST (if different from that of starting material)
: Suspension.
Species:
rat
Strain:
Wistar
Details on species / strain selection:
The Wistar Han rat was the species and strain of choice because it is a readily available rodent which is commonly used for genotoxicity testing, with documented susceptibility to a wide range of toxic items. Moreover, historical control background data has been generated with this strain.
Sex:
male
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany.
- Age at study initiation: 6 weeks.
- Weight at study initiation: 171 ± 8.7 g (Mean body weight ± SD).
- Assigned to test groups randomly: Yes.
- Fasting period before study: No.
- Housing: Up to 5 animals of the same sex and in the same dosing group were housed together.
- Diet: Commercial pellets ad libitum, except during designated procedures.
- Water: Tap water, ad libitum.
- Acclimation period: At least 6 days.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 to 25°C.
- Humidity (%): 40 to 70%.
- Air changes (per hr): ≥ 10.
- Photoperiod: 12 hrs light/12 hrs dark, except during designated procedures.

IN-LIFE DATES:
From: Not specified.
To: 12 Mar 2020.
Route of administration:
oral: gavage
Vehicle:
- Vehicle(s)/solvent(s) used: corn oil.
- Source of vehicle: Fagron Farmaceuticals, Capelle a/d IJssel, the Netherlands.
Duration of treatment / exposure:
Three consecutive days.
Frequency of treatment:
Daily.
Post exposure period:
Tissue samples taken 3 - 4 hours after administration of final dose.
Dose / conc.:
500 mg/kg bw/day (actual dose received)
Dose / conc.:
1 000 mg/kg bw/day (actual dose received)
Dose / conc.:
2 000 mg/kg bw/day (actual dose received)
Remarks:
No treatment-related toxicity or mortality were in a preliminary dose range finding study in which three male and three female rats received three consecutive daily doses of 2000 mg/kg bw
No. of animals per sex per dose:
5
Control animals:
yes, concurrent vehicle
Positive control(s):
Cyclophosphamide.
- Route of administration: Gavage.
- Doses / concentrations: A single dose of 19 mg/kg bw, dissolved in physiological saline.
Tissues and cell types examined:
Bone marrow from the femur.
Details of tissue and slide preparation:
The femurs were flushed with foetal calf serum and the cell suspension centrifuged. The supernatant was removed and a drop of the remaining cell suspension was spread across a clean slide and fixed with methanol. The slides were automatically stained with Giemsa using the Wright Stain Procedure.
Evaluation criteria:
The test item was considered positive if all of the following criteria were met:
a) at least one treatment group showed a statistically significant increase in frequency of micronucleated polychromatic erythrocytes.
b) the increase was dose related.
c) the results were outside the 95% confidence limits of the historical control data.

If none of the above criteria were met, and bone marrow exposure to the test item occurred, the substance was considered negative.

The incidence of micronuclei was assessed in 8000 polychromatic erythrocytes per animal.
Statistics:
ToxRat Professional v 3.2.1 (ToxRat Solutions® GmbH, Germany) was used for statistical
analysis of the data.

A test item is considered positive in the micronucleus test if all of the following criteria are
met:
a) At least one of the treatment groups exhibits a statistically significant (one-sided,
p < 0.05) increase in the frequency of micronucleated polychromatic erythrocytes
compared with the concurrent negative control
b) The increase is dose related when evaluated with a trend test.
c) Any of the results are outside the 95% control limits of the historical control data range.

A test item is considered negative in the micronucleus test if:
a) None of the treatment groups exhibits a statistically significant (one-sided, p < 0.05)
increase in the frequency of micronucleated polychromatic erythrocytes compared with
the concurrent negative control.
b) There is no concentration-related increase when evaluated with a trend test.
c) All results are within the 95% control limits of the negative historical control data range.

As there were statistically significant differences between one or more of the test item groups
and the vehicle control group a Cochran Armitage trend test (p < 0.05) was performed to test
whether there is a significant trend in the induction.
Key result
Sex:
male
Genotoxicity:
negative
Toxicity:
no effects
Vehicle controls validity:
valid
Positive controls validity:
valid
Additional information on results:
Platinum was quantifiable in plasma samples from high-dose (2000 mg/kg bw/day) satellite animals 1, 3, 6 and 12 hours after completing the second day of treatment. Moreover, platinum was quantifiable in plasma samples from all high-dose animals taken at necropsy approximately 3-4 hours after the third dose. Therefore it was confirmed that the bone marrow was exposed to the test item. No test item was detected in the animals dosed with vehicle.

No statistically significant increase in the frequency of micronucleated polychromatic erythrocytes was observed.

Treated animals showed no decrease in the PCE:NCE ratio, indicating a lack of toxicity to the bone marrow.
Conclusions:
Dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol, did not induce an increase in micronucleated polychromatic erythrocytes in rats administered up to 2000 mg/kg bw/day by gavage on three consecutive days.
Executive summary:

The in vivo clastogenicity and aneugenicity of dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol, as evaluated by its ability to induce micronuclei in polychromatic erythrocytes, was assessed in a study following OECD 474 and according to GLP. Male Wistar rats (5/group) were given gavage doses of 500, 1000 or 2000 mg/kg bw/day of the test item on three consecutive days, or a vehicle control. The concurrent positive control group received a single dose of cyclophosphamide. Bone marrow was harvested from the femurs and assessed for micronuclei.

There was no increase in the number of micronucleated polychromatic erythrocytes in any treatment group. On that basis, dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol, was concluded to be non-genotoxic under the conditions of this assay.

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

Mode of Action Analysis / Human Relevance Framework

No data identified.

Additional information

In an OECD Test Guideline 471 study, conducted according to GLP, dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol (1:2) was assessed for its ability to induce gene mutations in strains of S. typhimurium (TA1535, TA1537, TA98, TA100) and E. coli (WP2 uvrA). The test was performed in two independent experiments (involving the plate incorporation method, including a pre-incubation step for TA1535 and TA1537 in Experiment 2), with dose levels of up to 5000 μg/plate (determined following an initial toxicity test), both in the absence and presence of S9 using liver fraction from rats pre-treated with phenobarbitone and beta-naphthoflavone. In experiment 1, dose-related increases in revertant numbers, which were at least two-fold the control values, were observed in WP2 uvrA both with and without S9. The test item induced reproducible, large and dose-related increases in the number of revertant colonies, which were more than two-fold the control values, with TA98, TA100 and WP2 uvrA tester strains in the plate incorporation assays, at the higher dose levels both in the presence and absence of S9 metabolism. Increases in revertant numbers were also observed with TA1537 in the pre-incubation assay. These increases were greater than twice the control value at non-toxic dose-levels, both in the presence and absence of S9 metabolism. It was concluded that dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol (1:2) was mutagenic in S. typhimurium and E.coli under the reported experimental conditions (Scarcella, 2004).

 

In an in vitro GLP study, conducted in accordance with OECD Test Guideline 476 (in vitro mammalian cell gene mutation assay), dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol (1:2) was tested for its ability to induce 6 -thioguanine resistant mutants in Chinese hamster V79 cells. Cells were exposed to test material for 3 hours, both in the presence and absence of metabolic activation using liver S9 fraction from rats pre-treated with phenobarbitone and betanaphthoflavone. On the basis of a preliminary cytotoxicity test, the maximum dose levels for the mutation assay were selected as 78.1 and 5000 μg/ml for treatment in the absence or presence of S9, respectively. Dose related and significant increases in mutant numbers or mutant frequency were observed following treatment with the test item, in the absence and presence of S9 metabolism. These increases were greater than five-fold the spontaneous mutation frequency, in replicate cultures at both expression times (days, 6 and 8). It was concluded that dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol (1:2) was mutagenic in Chinese hamster V79 cells, both in the presence and absence of S9 (Cinelli, 2002).

 

In a combined in vivo micronucleus test and Comet assay in rats, dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol, administered by gavage at doses of 500, 1000 or 2000 mg/kg bw/day for three days did not cause an increased incidence of micronucleated polychromatic erythrocytes. Treatment also gave no evidence of DNA damage in the liver, kidney, glandular stomach or duodenum when assessed by the Comet procedure. As such, the test item was considered to be non-genotoxic in vivo (Eurlings, 2020).

 

Several Expert Groups have assessed the toxicity profile of platinum, and various platinum compounds, including the assessment of CMR properties. All reviews have indicated that platinum compounds have been reported to be mutagenic in a range of in vitro studies (DECOS, 2008; EMA, 2008; SCOEL, 2011; WHO, 1991). Cisplatin and related compounds are known DNA-reactive carcinogens and, as these compounds are better investigated due to their pharmaceutical properties, this has been confirmed in vivo. As cisplatin-type substances differ in chemical reactivity (liability of ligands, number of active sites etc.) it is reasonable to expect that not all forms of platinum are carcinogenic (DECOS, 2008). Limited experimental data on carcinogenicity for other platinum compounds give no evidence of activity that would meet classification criteria (DECOS, 2008; SCOEL, 2011).

 

Following the generally positive in vitro results identified for the platinum compounds in various bacterial/mammalian cell mutagenicity assays (supported by some mammalian cell cytogenicity tests) and the unclear in vivo relevance of these in vitro findings, a combined in vivo micronucleus test and Comet assay in rats (with dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol) did not cause an increased incidence of micronucleated polychromatic erythrocytes and gave no evidence of DNA damage in the liver, kidney, glandular stomach or duodenum when assessed by the Comet procedure (Eurlings, 2020).

 

 

References

DECOS (2008). Dutch Expert Committee on Occupational Standards. Platinum and Platinum Compounds. Health-based recommended occupational exposure limit. Gezondheidsraad, 2008/12OSH. https://www.gezondheidsraad.nl/en/publications/gezonde-arbeidsomstandigheden/platinum-and-platinum-compounds-health-based-recommended

 

EMA (2008). European Medicines Agency. Guideline on the specification limits for residues of metal catalysts or metal reagents. Committee for Medicinal Products for Human Use (CHMP). EMEA/CHMP/SWP/4446/2000. http://www.ema.europa.eu/docs/en_GB/document_library/Scientific_guideline/2009/09/WC500003586.pdf

 

SCOEL (2011). Recommendation from the Scientific Committee on Occupational Exposure Limits for platinum and platinum compounds. SCOEL/SUM/150. http://ec.europa.eu/social/BlobServlet?docId=7303&langId=en

 

WHO (1991). World Health Organization. Platinum. International Programme on Chemical Safety. Environmental Health Criteria 125. http://www.inchem.org/documents/ehc/ehc/ehc125.htm#SectionNumber:7.4

Justification for classification or non-classification

Based on the existing data set, dihydrogen hexahydroxyplatinate, compound with 2-aminoethanol (1:2) does not currently meet the criteria for classification as a germ cell mutagen (category 1A/1B or 2) under EU CLP criteria (EC 1272/2008).