CELEX 02008R1272 · v20250901

3.5. Germ cell mutagenicity

3.5.1.1. Germ cell mutagenicity means heritable gene mutations, including heritable structural and numerical chromosome aberrations in germ cells occurring after exposure to a substance or mixture.

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3.5.1.2. A mutation means a permanent change in the amount or structure of the genetic material in a cell. The term ‘mutation’ applies both to heritable genetic changes that may be manifested at the phenotypic level and to the underlying DNA modifications when known (including specific base pair changes and chromosomal translocations). The term ‘mutagenic’ and ‘mutagen’ will be used for agents giving rise to an increased occurrence of mutations in populations of cells and/or organisms.

3.5.1.3. The more general terms ‘genotoxic’ and ‘genotoxicity’ apply to agents or processes which alter the structure, information content, or segregation of DNA, including those which cause DNA damage by interfering with normal replication processes, or which in a non- physiological manner (temporarily) alter its replication. Genotoxicity test results are usually taken as indicators for mutagenic effects.

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3.5.2. Classification criteria for substances

3.5.2.1. This hazard class is primarily concerned with substances that may cause mutations in the germ cells of humans that can be transmitted to the progeny. However, the results from mutagenicity or genotoxicity tests in vitro and in mammalian somatic and germ cells in vivo are also considered in classifying substances and mixtures within this hazard class.

3.5.2.2. For the purpose of classification for germ cell mutagenicity, substances are allocated to one of two categories as shown in Table 3.5.1.

Table 3.5.1

Hazard categories for germ cell mutagens

Categories	Criteria
CATEGORY 1:	Substances known to induce heritable mutations or to be regarded as if they induce heritable mutations in the germ cells of humans. Substances known to induce heritable mutations in the germ cells of humans.
Category 1A:	The classification in Category 1A is based on positive evidence from human epidemiological studies. Substances to be regarded as if they induce heritable mutations in the germ cells of humans.
Category 1B:	The classification in Category 1B is based on: — positive result(s) from in vivo heritable germ cell mutagenicity tests in mammals; or — positive result(s) from in vivo somatic cell mutagenicity tests in mammals, in combination with some evidence that the substance has potential to cause mutations to germ cells. It is possible to derive this supporting evidence from mutagenicity/genotoxicity tests in germ cells in vivo, or by demonstrating the ability of the substance or its metabolite(s) to interact with the genetic material of germ cells; or — positive results from tests showing mutagenic effects in the germ cells of humans, without demonstration of transmission to progeny; for example, an increase in the frequency of aneuploidy in sperm cells of exposed people.
CATEGORY 2:	Substances which cause concern for humans owing to the possibility that they may induce heritable mutations in the germ cells of humans The classification in Category 2 is based on: — positive evidence obtained from experiments in mammals and/or in some cases from in vitro experiments, obtained from: — — somatic cell mutagenicity tests in vivo, in mammals; or — other in vivo somatic cell genotoxicity tests which are supported by positive results from in vitro mutagenicity assays. Note: Substances which are positive in in vitro mammalian mutagenicity assays, and which also show chemical structure activity relationship to known germ cell mutagens, shall be considered for classification as Category 2 mutagens.

3.5.2.3. Specific considerations for classification of substances as germ cell mutagens

3.5.2.3.1. To arrive at a classification, test results are considered from experiments determining mutagenic and/or genotoxic effects in germ and/or somatic cells of exposed animals. Mutagenic and/or genotoxic effects determined in in vitro tests shall also be considered.

3.5.2.3.2. The system is hazard based, classifying substances on the basis of their intrinsic ability to induce mutations in germ cells. The scheme is, therefore, not meant for the (quantitative) risk assessment of substances.

3.5.2.3.3. Classification for heritable effects in human germ cells is made on the basis of well conducted, sufficiently validated tests, preferably as described in Regulation (EC) No 440/2008 adopted in accordance with Article 13(3) of Regulation (EC) No 1907/2006 (‘Test Method Regulation’) such as those listed in the following paragraphs. Evaluation of the test results shall be done using expert judgement and all the available evidence shall be weighed in arriving at a classification.

3.5.2.3.4. In vivo heritable germ cell mutagenicity tests, such as:

—

rodent dominant lethal mutation test;

—

mouse heritable translocation assay.

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3.5.2.3.5. In vivo somatic cell mutagenicity tests, such as:

—

mammalian bone marrow chromosome aberration test;

—

mammalian erythrocyte micronucleus test

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3.5.2.3.6. Mutagenicity/genotoxicity tests in germ cells, such as:

(a)

mutagenicity tests:

—

mammalian spermatogonial chromosome aberration test;

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spermatid micronucleus assay;

(b)

Genotoxicity tests:

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sister chromatid exchange analysis in spermatogonia;

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unscheduled DNA synthesis test (UDS) in testicular cells.

3.5.2.3.7. Genotoxicity tests in somatic cells such as:

—

liver Unscheduled synthesis test (UDS) in vivo;

—

mammalian bone marrow Sister Chromatid Exchanges (SCE);

3.5.2.3.8. In vitro mutagenicity tests such as:

—

in vitro mammalian chromosome aberration test;

—

in vitro mammalian cell gene mutation test;

—

bacterial reverse mutation tests.

3.5.2.3.9. The classification of individual substances shall be based on the total weight of evidence available, using expert judgement (See 1.1.1). In those instances where a single well-conducted test is used for classification, it shall provide clear and unambiguously positive results. If new, well validated, tests arise these may also be used in the total weight of evidence to be considered. The relevance of the route of exposure used in the study of the substance compared to the route of human exposure shall also be taken into account.

3.5.3. Classification criteria for mixtures

3.5.3.1. Classification of mixtures when data are available for all ingredients or only for some ingredients of the mixture

3.5.3.1.1. The mixture shall be classified as a mutagen when at least one ingredient has been classified as a Category 1A, Category 1B or Category 2 mutagen and is present at or above the appropriate generic concentration limit as shown in Table 3.5.2 for Category 1A, Category 1B and Category 2 respectively.

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Table 3.5.2

Generic concentration limits of ingredients of a mixture classified as germ cell mutagens that trigger classification of the mixture

Ingredient classified as:	Concentration limits triggering classification of a mixture as:
	Category 1 mutagen		Category 2 mutagen
	Category 1A	Category 1B	Category 2 mutagen
Category 1A mutagen	≥ 0,1 %	—	—
Category 1B mutagen	—	≥ 0,1 %	—
Category 2 mutagen	—	—	≥ 1,0 %

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Note

The concentration limits in the table above apply to solids and liquids (w/w units) as well as gases (v/v units).

3.5.3.2. Classification of mixtures when data are available for the complete mixture

3.5.3.2.1. Classification of mixtures will be based on the available test data for the individual ingredients of the mixture using concentration limits for the ingredients classified as germ cell mutagens. On a case-by-case basis, test data on mixtures may be used for classification when demonstrating effects that have not been established from the evaluation based on the individual ingredients. In such cases, the test results for the mixture as a whole must be shown to be conclusive taking into account dose and other factors such as duration, observations, sensitivity and statistical analysis of germ cell mutagenicity test systems. Adequate documentation supporting the classification shall be retained and made available for review upon request.

3.5.3.3. Classification of mixtures when data are not available for the complete mixture: bridging principles

3.5.3.3.1. Where the mixture itself has not been tested to determine its germ cell mutagenicity hazard, but there are sufficient data on the individual ingredients and similar tested mixtures (subject to paragraph 3.5.3.2.1), to adequately characterise the hazards of the mixture, these data shall be used in accordance with the applicable bridging rules set out in section 1.1.3.

3.5.4. Hazard communication

3.5.4.1. Label elements shall be used in accordance with Table 3.5.3, for substances or mixtures meeting the criteria for classification in this hazard class.

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Table 3.5.3

Label elements of germ cell mutagenicity

Classification	Category 1 (Category 1A, 1B)	Category 2
GHS Pictograms
Signal Word	Danger	Warning
Hazard Statement	H340: May cause genetic defects (state route of exposure if it is conclusively proven that no other routes of exposure cause the hazard)	H341: Suspected of causing genetic defects (state route of exposure if it is conclusively proven that no other routes of exposure cause the hazard)
Precautionary Statement Prevention	P201 P202 P280	P201 P202 P280
Precautionary Statement Response	P308 + P313	P308 + P313
Precautionary Statement Storage	P405	P405
Precautionary Statement Disposal	P501	P501

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3.5.5. Additional classification considerations

It is increasingly accepted that the process of chemical-induced tumorigenesis in humans and animals involves genetic changes for example in proto-oncogenes and/or tumour suppresser genes of somatic cells. Therefore, the demonstration of mutagenic properties of substances in somatic and/or germ cells of mammals in vivo may have implications for the potential classification of these substances as carcinogens (see also Carcinogenicity, section 3.6, paragraph 3.6.2.2.6).

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3.5. Germ cell mutagenicity