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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.
3.5.2.3.5. In vivo somatic cell mutagenicicty tests, such as:
mammalian bone marrow chromosome aberration test;
mammalian erythrocyte micronucleus test.
3.5.2.3.6. Mutagenicity/genotoxicity tests in germ cells, such as:
(a) mutagenicity tests:
mammalian spermatogonial chromosome aberration test;
spermatid micronucleus assay;
(b) Genotoxicity tests:
sister chromatid exchange analysis in spermatogonia;
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.