Registration Dossier

Diss Factsheets

Administrative data

Endpoint:
genetic toxicity in vivo, other
Remarks:
in vivo mammalian somatic cell study: combined gene mutuation and cytogenicity / micronucleus assay
Type of information:
experimental study planned
Study period:
To be determined upon receipt of ECHA final decision
Justification for type of information:
TESTING PROPOSAL ON VERTEBRATE ANIMALS
Please see attached justification.

NON-CONFIDENTIAL NAME OF SUBSTANCE:
- Name of the substance on which testing is proposed to be carried out: Hydroxybenzaldehyde polymer with phenol (EC 947-320-5).

CONSIDERATIONS THAT THE GENERAL ADAPTATION POSSIBILITIES OF ANNEX XI OF THE REACH REGULATION ARE NOT ADEQUATE TO GENERATE THE NECESSARY INFORMATION:
- Available GLP studies:
The following genotoxicity data have been generated to support an Annex VIII registration of the substance:
A bacterial reverse mutation test (OECD TG 471): Reported to be negative.
An in vitro CHO chromosome aberration test (OECD TG 473): Reported to be negative.
An in vitro mouse lymphoma assay (OECD TG 490): Reported to be positive in the presence of S9.
The REACh information requirements state that for substances demonstrated to be positive for any of the genotoxicity studies in Annex VII or VIII, appropriate in vivo mutagenicity assays shall be considered to ascertain if the genotoxicity potential observed in vitro can be expressed in vivo. Consequently, the attached testing proposal describes the applicant’s intended follow-up study, together with the justification for the choice of assay and its experimental design.
- Available non-GLP studies: There are no non-GLP studies available on the registered substance.
- Historical human data: There is no relevant historical human data available on the registered substance.
- (Q)SAR: QSAR is not appropriate for this UVCB substance.
- In vitro methods: It is not possible to conclude whether hazards identified in vitro will be expressed in vivo.
- Weight of evidence: A weight of evidence assessment of available in vitro data is presented in the attached testing proposal in order to determine the choice of the proposed in vivo study ans its experimental design.
- Grouping and read-across: No suitable surrogate substance has been identified for read-across.

CONSIDERATIONS THAT THE SPECIFIC ADAPTATION POSSIBILITIES OF ANNEXES VI TO X (AND COLUMN 2 THEREOF) OF THE REACH REGULATION ARE NOT ADEQUATE TO GENERATE THE NECESSARY INFORMATION:
Column 2 of REACH Annex VIII states that "Appropriate in vivo mutagenicity studies shall be considered in case of a positive result in any of the genotoxicity studies in Annex VII or VIII".
Testing with the registered substance produced a positive result in the Mouse Lymphoma Assay in the presence of S9, with an equivocal response also apparent in the absence of S9.
In the chromosome aberration study, the registered substance produced aberration frequencies that exceeded the laboratory’s HCD, though these findings did not display a concentration-relationship and were not reproduced. They were therefore considered biologically irrelevant and the reported negative conclusion is supported.
In order to fully evaluate the genotoxic hazard of the registered substance, it is considered justified to conduct an in vivo study.
It is proposed that a comet assay in the liver, combined with assessment of micronuclei in the bone marrow, is an appropriate and scientifically justified assay for in vivo evaluation of the genotoxicity hazard identified in vitro for the registered substance. Please see the attached for the full justification.

FURTHER INFORMATION ON TESTING PROPOSAL IN ADDITION TO INFORMATION PROVIDED IN THE MATERIALS AND METHODS SECTION:
The combined assay would be conducted as follows:
- Species: Rat (This species is more commonly used for the comet assay and most laboratories have more HCD in rat than mouse).
- Strain: Common laboratory strain, e.g., Han Wistar or Sprague Dawley. The strain will be determined by availability of suitable historical control data at the selected test facility.
- Sex: To be determined in a preliminary dose setting phase. Assuming no substantial differences in maximum tolerated dose (MTD) are identified between the sexes, the main experiment will be conducted in males only.
- Exposure route: Oral gavage (Oral exposure is a potential route of human exposure).
- Duration of treatment: 3 consecutive doses. Necropsy and tissue processing 3 hours after the final (3rd) administration, equivalent to 24 hours after the 2nd administration.
- Dose levels: 0.25x MTD, 0.5x MTD, MTD (Actual doses to be determined based on the results of a preliminary dose setting phase conducted in the same species/strain of animals and using the same dosing regimen, route and dose volume).
- Controls: Concurrent negative (vehicle) and positive controls.
- Group size: N=5-6 for all groups depending on standard practice at test facility. For the positive control N=3 would be acceptable providing the laboratory can demonstrate suitable proficiency.
- Comet analysis: Liver (Measurement of % tail intensity (% DNA in tail) of 150 cells per tissue, per animal. Hedgehog cells, excluded from comet analysis but frequency to be recorded and reported).
- Micronucleus assessment: Bone marrow smears (Number of immature and mature erythrocytes in a total of 500 erythrocytes to be determined for toxicity assessment. 4000 immature erythrocytes to be assessed for the presence of micronuclei).
- Additional assessments:
Terminal blood sample taken and analysed for clinical chemistry parameters as an indicator of liver effects.
Sample of liver from each treated rat and vehicle control to be preserved in formalin for possible histopathology assessment in the event of positive comet findings.
Terminal blood sample to be processed to plasma and retained frozen as a contingency. In the event of negative comet and micronucleus results and assuming systemic exposure cannot be confirmed from existing data, this plasma sample could be analysed for the presence of Hydroxybenzaldehyde polymer and/or its metabolites.
Please refer to the attached for the full details of the proposed study.

Data source

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 489 (In vivo Mammalian Alkaline Comet Assay)
Version / remarks:
A comet assay in the liver, combined with assessment of micronuclei in the bone marrow.
Deviations:
not applicable

Test material

Constituent 1
Chemical structure
Reference substance name:
Condensation products of phenol and salicylaldehyde
EC Number:
947-320-5
Molecular formula:
Variable as UVCB substance
IUPAC Name:
Condensation products of phenol and salicylaldehyde

Results and discussion

Applicant's summary and conclusion

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