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extended one-generation reproductive toxicity - basic test design (Cohorts 1A, and 1B without extension)
Data waiving:
other justification
Justification for data waiving:
Justification for type of information:
ZBEC – Extended one generation reproductive toxicity study
On December 12th 2017 we received the ECHA decision on ZBEC (Decision number: CCH-D-2114382075-49-01/F). ECHA requested to submit the basic extended one-generation reproductive toxicity study with the inclusion of cohorts 2A and 2B (developmental neurotoxicity). The inclusion of these cohorts was based on a particular concern on (developmental) neurotoxicity based on the results on the structurally analogous substance zinc bis dimethyldithiocarbamate (ziram, ZDMC).
We reviewed ECHAs’ request for the inclusion of cohorts 2A and 2B in light of the newly obtained OECD 408 and 414 studies for ZBEC.

OECD 408
In the OECD 408 study rats (6-7 weeks old) were exposed to doses of ZBEC up to 1000 mg/kg bw/day. Throughout the study these animals were in good health and did not show any treatment-related clinical signs. Thyroid stimulating hormone levels were slightly decreased in females of the high-dose group. This decrease in TSH levels was not corroborated by noticeable changes in T4 levels, thyroid weight or pathology, or by effects on plasma total cholesterol levels which are known to be controlled by thyroid hormone action (OECD Guideline 408, 2018). Since, moreover the values were well within the range of historical control data and this variable is known to show considerable variation, the decrease in TSH levels in high-dose females was therefore considered a chance finding. Furthermore, the results of the neurobehavioral observations and motor activity assessment did not indicate any neurotoxic potential of ZBEC in rats.
In absence of any relevant changes in any test group, the systemic NOAEL was placed at the highest level tested, namely 1000 mg/kg bw/day. The NOAEL for systemic toxicity of ZDMC is placed on 1.6 mg/kg bw/day based on a 52-week study in dogs. None of the effects reported in the ZDMC studies, which are regarded as triggers for inclusion of the 2A and 2B cohorts, were observed in animals receiving ZBEC.

OECD 414
In addition to the OECD 408 study, the draft report of an OECD 414 in rats indicates the low toxic potential of ZBEC. Unfortunately, finalisation of this draft report was not possible before the submission deadline of December 19th, consequently this data is not included in the current update of the dossier. In this study time-mated female Wister Han rats were treated with ZBEC from Day 6 to 20 post-coitum, inclusive by daily oral gavage at dose levels of 100, 300 and 1000 mg/kg bw. No treatment-related changes were noted in any of the maternal parameters investigated in this study (i.e. clinical appearance, body weight, food consumption, and macroscopic examination). No treatment-related changes were noted in any of the developmental parameters investigated in this study (i.e. litter size, post-implantation loss, sex ratio, fetal body weights, external, visceral and skeletal malformations and developmental variations). This included visceral analyzation of the brains for half of the pups. The analysis indicated that the developing brain was not affected by the test substance.
The NOAEL for maternal and developmental toxicity of ZBEC was established as being at least 1000 mg/kg bw. The NOAEL for maternal toxicity of ZDMC was placed at 6.8 mg/kg bw and for developmental toxicity on 9 mg/kg bw/day based on OECD 416 study in rats. None of the effects reported in the ZDMC studies, which are regarded as triggers for inclusion of the 2A and 2B cohorts, were observed in animals receiving ZBEC.

Cohort: developmental neurotoxicity
The request by ECHA for the inclusion of the 2A and 2B cohorts is solely based on a particular concern on (developmental) neurotoxicity of a structurally similar substance, ZDMC:
- ECHA notes that results obtained with the analogue substance zinc bis dimethyldithio-carbamate (ZDMC) show developmental neurotoxicity in the offspring, such as increased motor activity and decreased mean peak startle response in a dietary two-generation reproduction and developmental neurotoxicity study (Nemec, 1996).

We want to emphasize that we are not happy with the strategy of ECHA to first reject our read-across adaptation to ZDMC and then request an extension of the OEGRTS based on the same read-across substance without indicating the toxicological basis for this request.
In light of newly obtained repeated dose toxicity and pre-natal developmental data for ZBEC, we can conclude that the toxic potential of ZBEC is much lower than of ZDMC. This is supported by the overall toxicity overview presented in the table attached under 'Attached justification'. In addition, no triggers for the inclusion of the 2A and 2B cohorts were found in the OECD 408 and 414 studies.
As was reported in the decision, ECHA considers the developmental neurotoxicity effects found in the Nemec (1996) study to be the main trigger for the inclusion of the cohorts. In the US EPA document the validity of the developmental toxicity findings in the Nemec study (1996) were questioned. There are some major deficiencies within the study. The neurobehavioural data (motor activity, startle response and cognitive function) were not analysed statistically. Therefore, it was not indicated that the positive findings were not purely found by change.
We consider that all observed effects in this study are related to the general state of the pups and were transient (i.e. motor activity and peak startle response), while the developmental neurotoxicological endpoints with a morphological developmental basis did not show any effect. For example, no effect on learning and memory evaluations were found and brain weights and qualitative histopathological evaluation of the nervous system did not reveal any treatment-related findings. Therefore, it seems that the observed positive effects have a general-toxicity basis instead of a developmental toxicity basis.
Furthermore, we consider that the absence of adverse effects of ZBEC in the recent OECD 408 and 414 study should be the main indicator, overriding the already weak triggers from the study on ziram, to conclude that the inclusion of the 2A and 2B in the OECD 443 is not warranted. In our view, these cohorts should therefore not be included, due to a lack of particular concern and also in light of animal welfare reasons. We ask ECHA to revise their decision and agree with a basic OECD 443 study without inclusion of the developmental neurotoxicity cohort.

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Applicant's summary and conclusion