Registration Dossier

Diss Factsheets

Administrative data

Key value for chemical safety assessment

Toxic effect type:
dose-dependent

Effects on fertility

Description of key information

Short description of key information:
The toxic potential of LZ649 on basic reproductive processes was assessed in this OECD 421 screening study in which Crl:CD(SD) rats were dosed for two weeks prior to pairing, throughout gestation and lactation at dose levels of 50, 150 or 500 mg/kg/day. The administration of LZ649 was well tolerated up to and including the maximum level of 500 mg/kg/day assessed within this study. There was no adverse effect of administration with LZ649 on clinical condition, bodyweight, and food consumption, macroscopic and microscopic appearance. Reproductive performance assessments of oestrous cycles, gestation length and parturition for F0 females also showed no test article related change. Organ weights for F0 males were similar throughout the groups. The clinical condition, litter size and survival and sex ratio of offspring, exposed in utero or via the milk, were not affected by doses up to and including 500 mg/kg/day. The results observed in this screening study conclude that LZ649 has no effect on reproductive performance and offspring survival or development at doses up to and including 500 mg/kg/day.

Justification for selection of Effect on fertility via oral route:
GLP compliant study conducted in accordance with international guidelines.

Link to relevant study records
Reference
Endpoint:
screening for reproductive / developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
02 April - 16 July 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP compliant study conducted in accordance with international guidelines
Reason / purpose for cross-reference:
reference to same study
Qualifier:
according to guideline
Guideline:
OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)
Deviations:
no
Principles of method if other than guideline:
Not applicable.
GLP compliance:
yes (incl. QA statement)
Limit test:
no
Specific details on test material used for the study:
Retest Date: 23 November 2014
Species:
rat
Strain:
Sprague-Dawley
Details on species / strain selection:
Crl:CD(SD) strain was used
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Reputable supplier
- Age at study initiation: (P) Males 71 days; Females 65
- Weight at study initiation: (P) Males: 342-407 g; Females: 219-259
- Fasting period before study: No
- Housing: Polycarbonate cages, the gridded cages used during pairing were suspended over trays covered with absorbent paper which was
changed daily. For cages with solid floors, wood based material was used as bedding and was sterilised by autoclaving and changed at least twice
each week
- Use of restrainers for preventing ingestion (if dermal): no
- Diet (e.g. ad libitum): SDS VRF1 certified diet ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19-23
- Humidity (%): 40 to 70
- Air changes (per hr): Not given
- Photoperiod (hrs dark / hrs light): 12 / 12

IN-LIFE DATES: From: 18 April 2012 To: 8 June 2012
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: The LZ649 was prepared for administration as a series of graded concentrations in the vehicle. Approximately 50% of the final volume of vehicle was added to the test substance and mixed by magnetic stirring until dissolution was achieved. The volume was made up with the remaining vehicle and magnetically stirred until homogenous.

VEHICLE
- Justification for use and choice of vehicle (if other than water): Corn oil is a commonly used vehicle, test substance dissolves in this vehicle
- Concentration in vehicle: 0, 10, 30, 100 mg/mL
- Amount of vehicle (if gavage): 5 mL/kg body weight
- Lot/batch no. (if required): NA
- Purity: NA
Details on mating procedure:
- M/F ratio per cage: 1/1
- Length of cohabitation: 14 days maximum
- Proof of pregnancy: vaginal plug / sperm in vaginal smear referred to as day 0 of pregnancy
- After successful mating each pregnant female was caged (how): individually
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
A method of analysis of the test substance and formulation preparation, homogeneity and stability, was validated in this study.
The first and last preparations for dosing were analysed using the validated method to confirm the dose concentrations.
Duration of treatment / exposure:
F0 genearation only were dosed, both males and females from the start of the study until day 7 of lactation. F0 females were not dosed if parturition was in progress at the scheduled time of administration.
Frequency of treatment:
Once daily
Dose / conc.:
50 mg/kg bw/day (nominal)
Dose / conc.:
150 mg/kg bw/day (nominal)
Dose / conc.:
500 mg/kg bw/day (nominal)
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
In a previous 13 week study dose levels of 50, 150 and 500 mg/kg/day were investigated using a five day/week dosing regime and 500 mg/kg/day was found to be the No Observed Adverse Effect Level (NOAEL). Therefore dose levels were selected for this reproduction screening study taking into consideration the requirement to dose reproducing animals on a seven day/week regime, doses of no higher than those administered in the 13 week study were selected.
Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily
- Cage side observations checked in table [No.?] were included.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Weekly

BODY WEIGHT: Yes
- Time schedule for examinations: weekly, also for F0 females on Days 0, 3, 7, 10, 14, 17 and 20 after mating and days 1, 4 and 7 of lactation

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean weekly diet consumption calculated as g food/rat/day: Yes

Oestrous cyclicity (parental animals):
For 15 days before pairing, vaginal smears taken, eaxamined to establish duration and regularity of the oestrous cycle
Sperm parameters (parental animals):
Parameters examined in P male parental generation:

The following organs, taken from each male, were dissected free of adjacent fat and other contiguous tissue and the weights recorded:
Epididymides (L&R)
Testes (L&R)

L&R Bilateral organs weighed individually
Organ weights were also adjusted for terminal bodyweight, using the weight recorded before necropsy.
Litter observations:
STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: no

PARAMETERS EXAMINED
The following parameters were examined in F1 offspring:
number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain, physical or behavioural abnormalities

GROSS EXAMINATION OF DEAD PUPS:
yes, for external and internal abnormalities, possible cause of death was determined in some casesfor pups born or found dead
Missing offspring and those grossly autolysed or grossly cannibalised could not be examined.
Postmortem examinations (parental animals):
SACRIFICE
- Male animals: Surviving F0 males were killed after the first Day 7 of lactation of the females (after confirmation that a second mating was not required).
- Maternal animals: F0 females were killed on Day 7 of lactation.

GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera

HISTOPATHOLOGY / ORGAN WEIGHTS
The following tissues were prepared for microscopic examination:
Epididymides - caput, corpus and cauda
Ovaries - qualitative evaluation of one section from each ovary

Postmortem examinations (offspring):
SACRIFICE
- The F1 offspring were sacrificed at 7 days of age.
- These animals were subjected to postmortem examinations (macroscopic and/or microscopic examination) as follows:

A careful external examination was performed for gross abnormalities and externally normal offspring were discarded without further internal examination.
Externally abnormal offspring were internally examined and any abnormal tissues were retained in an appropriate fixative.
Statistics:
All statistical analyses were carried out separately for males and females. For all other adult parameters, the analyses were carried out using the individual animal as the basic experimental unit. For litter/fetal findings the litter was taken as the treated unit and the basis for statistical analysis and biological significance was assessed with relevance to the severity of the anomaly and the incidence of the finding within the background control population.
Clinical signs:
no effects observed
Description (incidence and severity):
There were no post dose signs. Clinical signs observed were of a type commonly observed in this strain of rat, at this age, in this laboratory and did not indicate any adverse reaction to the test item.
Dermal irritation (if dermal study):
not examined
Mortality:
mortality observed, non-treatment-related
Description (incidence):
One Group 3 male died following dosing on Day 30 of study. Necropsy did not give any indication of the cause of death and no evidence of dosing trauma was observed. Abnormalities at necropsy were limited to clear fluid present in the stomach. The cause of death was therefore undetermined following pathology examination. In the absence of any signs or deaths in the high dose group it was not attributed to the test material.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
Following the first week of study lower bodyweight gain in females receiving LZ649 at 500 mg/kg/day was apparent, individual responses were variable and some females at each dose level lost weight. During the second week of study the females receiving 500 mg/kg/day had similar weight gain to Control animals. There was no effect of LZ649 on bodyweight in males at any dose level.
Bodyweight on Day 0 of gestation was slightly lower in animals receiving 500 mg/kg/day but following superior bodyweight gains by Day 20 of gestation no differences were apparent.
Bodyweight gain was decreased during lactation in females receiving 150 mg/kg/day compared with Controls, there was no similar effect in the high dose group. As the 150 mg/kg/day animals started with the bodyweight values greater than controls, and ended with values similar to Controls no test article relationship was attached.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
There was no adverse effect of LZ649 on food consumption prior to pairing, during gestation or lactation.
It was apparent that food consumption was slightly low in females receiving 500 mg/kg/day after the first week of study, an effect of this degree was not considered to be adverse. The food consumption during week two of dosing was similar to Controls in both males and females at all dose levels. Food consumption during the period Days 0-2 of gestation and throughout lactation was also slightly lower than control for females receiving 500 mg/kg/day.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Description (incidence and severity):
Histopathological evaluation was performed on tissues with gross abnormalities, ovaries, testes and epididymides.
There were no microscopic findings attributable to administration with LZ649.
One control male showed slight degeneration/atrophy of the epithelium in a few seminiferous tubules, but this is not an uncommon finding in rats of this age and strain at these laboratories.
Histopathological findings: neoplastic:
not examined
Other effects:
not examined
Reproductive function: oestrous cycle:
no effects observed
Description (incidence and severity):
There was no effect of LZ649 on oestrous cycle length. All animals mated at the first oestrus opportunity following pairing.
Reproductive function: sperm measures:
no effects observed
Reproductive performance:
no effects observed
Description (incidence and severity):
Percentage mating, conception rate and fertility index was 100% in each dose group up to 500 mg/kg/day. All animals gave birth to a live litter.
Gestation length was within the expected timeframe observed at this laboratory.
SIGNS and MORTALITY:
One Group 3 male (number 29) died following dosing on Day 30 of study. Necropsy did not give any indication of the cause of death and no evidence of dosing trauma was observed. Abnormalities at necropsy were limited to clear fluid present in the stomach. The cause of death was therefore undetermined following pathology examination. In the absence of any signs or deaths in the high dose group it was not attributed to the test material.
There were no post dose signs. Clinical signs observed were of a type commonly observed in this strain of rat, at this age, in this laboratory and did not indicate any adverse reaction to LZ649.
BODYWEIGHT:
Bodyweight on Day 0 of gestation was slightly lower in animals receiving 500 mg/kg/day but following superior bodyweight gains by Day 20 of gestation no differences were apparent.
FOOD CONSUMPTION:
There was no adverse effect of LZ649 on food consumption prior to pairing, during gestation or lactation.
OESTROUS CYCLE LENGTH:
There was no effect of LZ649 on oestrous cycle length.
PRE-COITAL INTERVAL:
All animals mated at the first oestrus opportunity following pairing.
MATING PERFORMANCE and FERTILITY:
Percentage mating, conception rate and fertility index was 100% in each dose group up to 500 mg/kg/day.
GESTATION LENGTH and GESTATION INDEX:
All animals gave birth to a live litter. Gestation length was within the expected timeframe observed at this laboratory.
MACROPATHOLOGY:
There were no changes detected at macroscopic examination in any of the males or females receiving LZ649 that were related to exposure.
ORGAN WEIGHTS:
There were no effects of LZ649 on epididymides or testes weights.
HISTOPATHOLOGY:
There were no microscopic findings attributable to administration with LZ649.
One control male showed slight degeneration/atrophy of the epithelium in a few seminiferous tubules, but this is not an uncommon finding in rats of this age and strain at these laboratories.

Key result
Dose descriptor:
NOEL
Effect level:
500 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects
Dose descriptor:
NOAEL
Effect level:
500 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects
Clinical signs:
not examined
Dermal irritation (if dermal study):
not examined
Mortality:
not examined
Body weight and weight changes:
not examined
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
not examined
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
Other effects:
not examined
Reproductive function: oestrous cycle:
not examined
Reproductive function: sperm measures:
not examined
Reproductive performance:
not examined
Clinical signs:
no effects observed
Description (incidence and severity):
There was no effect of test article on the number of implantations and total litter size on Day 1, and live litter sizes on Days 1, 4 and 7 were similar to Control values.
Dermal irritation (if dermal study):
not examined
Mortality / viability:
no mortality observed
Description (incidence and severity):
The post implantation survival and offspring survival up to Day 7 was not affected by LZ649.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
Offspring bodyweight on Day 1 of age and subsequent bodyweight gain up to Day 7 of age was unaffected by parental administration with LZ649 at levels up to 150 mg/kg/day. Bodyweight gain for the period Days 1-7 of lactation of male and female offspring derived from females receiving 500 mg/kg/day was lower than Control though this was not considered adverse at the degree observed.
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Description (incidence and severity):
Macroscopic examination of offspring dying before scheduled termination or killed at scheduled termination on Day 7 of age did not reveal any findings that were attributed to parental test material exposure.
Histopathological findings:
not examined
Other effects:
not examined
Behaviour (functional findings):
not examined
Developmental immunotoxicity:
not examined
LITTER SIZE:
There was no effect of test article on the number of implantations and total litter size on Day 1, and live litter sizes on Days 1, 4 and 7 were similar to Control values.
OFFSPRING SURVIVAL INDICES:
The post implantation survival and offspring survival up to Day 7 was not affected by LZ649.
SEX RATIO:
Sex ratio was not affected by parental test material exposure.
BODYWEIGHT:
Offspring bodyweight on Day 1 of age and subsequent bodyweight gain up to Day 7 of age was unaffected by parental administration with LZ649 at levels up to 150 mg/kg/day. Bodyweight gain for the period Days 1-7 of lactation of male and female offspring derived from females receiving 500 mg/kg/day was lower than Control though this was not considered adverse at the degree observed.
OFFSPRING MACROPATHOLOGY:
Macroscopic examination of offspring dying before scheduled termination or killed at scheduled termination on Day 7 of age did not reveal any findings that were attributed to parental test material exposure.
Key result
Dose descriptor:
NOEL
Generation:
F1
Effect level:
500 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
500 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects
Clinical signs:
not examined
Dermal irritation (if dermal study):
not examined
Mortality / viability:
not examined
Body weight and weight changes:
not examined
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
not examined
Histopathological findings:
not examined
Other effects:
not examined
Behaviour (functional findings):
not examined
Developmental immunotoxicity:
not examined
Reproductive effects observed:
not specified
Conclusions:
The results observed in this screening study conclude that LZ649 has no effect on reproductive performance and offspring survival or development at doses up to 500 mg/kg/day.
Executive summary:

The toxic potential of LZ649 on basic reproductive processes was assessed in this OECD 421 screening study in which Crl:CD(SD) rats were dosed for two weeks prior to pairing, throughout gestation and lactation at dose levels of 50, 150 or 500 mg/kg/day. The administration of LZ649 was well tolerated up to and including the maximum level of 500 mg/kg/day assessed within this study. There was no adverse effect of administration with LZ649 on clinical condition, bodyweight, and food consumption, macroscopic and microscopic appearance. Reproductive performance assessments of oestrous cycles, gestation length and parturition for F0 females also showed no test article related change. Organ weights for F0 males were similar throughout the groups. The clinical condition, litter size and survival and sex ratio of offspring, exposed in utero or via the milk, were not affected by doses up to and including 500 mg/kg/day. The results observed in this screening study conclude that LZ649 has no effect on reproductive performance and offspring survival or development at doses up to and including 500 mg/kg/day.

Effect on fertility: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
500 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
Guideline study
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Additional information

According to REACH Annex IX section 8.7.3 a two-generation reproductive toxicity study was waived, as no adverse effects on reproductive organs or tissues were noted neither in an oral sub-chronic toxicity study (90-day) nor in a reproductive/developmental toxicity screening test.


Effects on developmental toxicity

Description of key information

Short description of key information:

The teratogenic potential of the test item was evaluated in female Hannover Wistar rats. The test substance was administered daily by oral gavage to pregnant rats on gestation days 6 -19, at dose levels of 100, 300 and 1000 mg/kg bw/day. The dams were sacrificed at gestation day 20 and the foetuses were examined for visceral and skeletal variations and malformations.

Up to dose level of 1000 mg/kg bw/day did not induce maternal toxicity, embryotoxicity, foetotoxicity, or teratogenicity. There were no malformations or developmental effects attributed to the test item at any dose level

The following no-observed-adverse-effect (NOAEL) levels were derived:

NOAELmaternal toxicity:                  1000 mg/kg bw/day

Based on the lack of any test-item related maternal toxicity effect in any treatment group.

NOAELembryotoxicity:                    1000 mg/kg bw/day

Based on the lack of any test-item related intrauterine effect in any treatment group.

NOAELfoetotoxicity:                               1000 mg/kg bw/day

Based on the lack of any test-item related foetotoxicity effect in any treatment group.

NOAELteratogenecity:                     1000 mg/kg bw/day

Based on the lack of any developmental effects in any treatment group.

Link to relevant study records
Reference
Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
12 March - 11 April 2019
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Version / remarks:
27 June 2018
Deviations:
no
Principles of method if other than guideline:
not applicable
GLP compliance:
yes (incl. QA statement)
Limit test:
no
Specific details on test material used for the study:
already provided
Species:
rat
Strain:
Wistar
Details on test animals or test system and environmental conditions:
Species and strain: Hannover Wistar rats (CRL:WI(Han))
Age of animals: Young adult female rats, nulliparous and non-pregnant, at least 11 weeks old at mating
Starting body weight: 189-230 g (the variation did not exceed ± 20% of the mean weight)
Acclimation period: at least 12 days
Light: 12 hours daily, from 6.00 a.m. to 6.00 p.m.
Temperature: 19.7-24.6°C (target: 22 ± 3°C)
Relative humidity: 33-56% (target: 30-70%)
Ventilation: 15-20 air exchanges/hour
Housing/Enrichment: Successfully mated animals were housed individually.

Route of administration:
oral: gavage
Vehicle:
propylene glycol
Details on exposure:
A constant volume of 5 mL/kg body weight was administered to all dose groups, including the controls. The individual volume of the treatment was based on the most recent individual body weight of the animals.
The control or test item dose formulations were administered to mated, sperm positive (assumed pregnant) female rats daily by oral gavage at approximately the same time each day, from gestation day 6 (GD 6) to gestation
day 19 (GD 19).
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Samples for test item concentration and homogeneity determination of the dosing formulations were collected four times during the treatment period. Formulation stability was established before this study. The test item concentration was analysed at the Test Site using a HPLC-UV method. The mean concentration of all measured formulations were found to be in the range of 91.8-95.6% of their nominal concentrations (20, 60 and 200 mg/mL) and were found to be homogenous.
Details on mating procedure:
The oestrus cycle of female animals was examined shortly before start of pairing. After acclimation, the females were paired according to their oestrus cycle with males in the morning for approximately 2-3 hours (1 male : 1 female) until at least 24 sperm positive females/group were attained. After the daily mating period, a vaginal smear was prepared and stained with 1% aqueous methylene blue solution. The smear was examined with a light microscope; the presence of a vaginal plug or sperm in the vaginal smear was considered as evidence of copulation (gestation day 0, GD 0). Sperm positive females were separated and caged individually.
Duration of treatment / exposure:
Days 6 - 19 of gestation
Frequency of treatment:
Once daily during exposure period
Duration of test:
Until Day 20 of gestation
Dose / conc.:
100 mg/kg bw/day
Dose / conc.:
300 mg/kg bw/day
Dose / conc.:
1 000 mg/kg bw/day
No. of animals per sex per dose:
98 female animals, 25-26 mated female animals/group, 4 groups (one control and 3 test item-treated groups)
Control animals:
yes, concurrent vehicle
Details on study design:
The sperm-positive, assumed pregnant females were allocated to the experimental groups (on each mating day) in such a way that the group averages of the body weight were as similar as possible. A computer software (SPSS PC+4.0) was used to verify homogeneity/variation among/within groups.
Maternal examinations:
Animals were inspected for signs of morbidity and mortality twice daily (at the beginning and end of each working day). Detailed clinical observations were made on all animals at the onset of treatment (GD 6) then weekly.
Parameters monitored during the study included mortality and clinical observations, body weight, body weight gain and individual food consumption. Maternal reproductive parameters associated with uterine examination were evaluated, and the foetuses were weighed and examined for external, visceral and skeletal abnormalities. Placentas were examined macroscopically.
Ovaries and uterine content:
The ovaries and uterus were removed and the pregnancy status ascertained. The uterus including the cervix was weighed and examined for early and late embryonic or foetal deaths and for the number of live foetuses.
The dams’ viscera were examined macroscopically for any structural abnormalities or pathological changes.
The number of corpora lutea in each ovary and implantation sites in each uterine horn, the number of live foetuses, early and late embryonic death and foetal death were counted and the number and percentage of pre- and postimplantation losses were calculated. The degree of resorption was described in order to estimate the relative time of death of the conceptus. The placentas were examined macroscopically.
Fetal examinations:
Each foetus was weighed individually (accuracy ±0.01 g) and subjected to external examination, plus an additional examination of the great arteries. The anogenital distance of each foetus was measured. The sex of the fetuses was assigned based on the external observations and was reconfirmed by examining the internal sex organs. All abnormalities (external, soft tissue and skeletal malformations, and variations) found during the foetal examinations were recorded; photographic records were made additionally.
Statistics:
The statistical evaluation of data was performed with the program package SAS v9.2 in case of Provantis v.9, or SPSS PC+4.0 (SPSS Hungary Kft, Budapest) in the case of data tabulated in Excel, by an appropriate statistical method.
Indices:
No information available.
Historical control data:
HC data available
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Transient signs of piloerection in some Mid and High dose animals, slightly noisy respiration in some High dose animals, and hunched back, red coloured faeces and reduced faeces for a two-days period in one
High dose animals were considered to be treatment related but not adverse.
Dermal irritation (if dermal study):
not examined
Mortality:
no mortality observed
Description (incidence):
No test item related mortality was observed in the study.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
No test item related effect on body weight was observed in the test item related groups when compared to the controls.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
No test item related effect on food consumption was observed in the test item related groups when compared to the controls.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not specified
Clinical biochemistry findings:
not specified
Urinalysis findings:
not examined
Behaviour (functional findings):
not specified
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Description (incidence and severity):
There were no treatment-related effects.
Gross pathological findings:
no effects observed
Description (incidence and severity):
There were no findings note at necropsy that were considered to be related to the treatment.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
Other effects:
no effects observed
Description (incidence and severity):
No test item related effect on the weight of the thyroid gland (with parathyroid glands) was observed in the test item related groups when compared to the controls.
No test item-related changes were observed in the thyroid hormone levels of the dams.
Number of abortions:
no effects observed
Description (incidence and severity):
The early and the late embryonic loss values of the test item treated groups, and the number of dead foetuses were comparable with the controls. There was no statistically significant difference in the postimplantation loss or
total intrauterine mortality between the treated and control groups.
Pre- and post-implantation loss:
no effects observed
Description (incidence and severity):
There was no test item related effect on the intrauterine mortality parameters.

Total litter losses by resorption:
no effects observed
Description (incidence and severity):
There was no test item related effect on the intrauterine mortality parameters.

Early or late resorptions:
no effects observed
Description (incidence and severity):
The early and the late embryonic loss values of the test item treated groups, were comparable with the controls.
Dead fetuses:
no effects observed
Description (incidence and severity):
The number of dead foetuses were comparable with the controls.
Changes in pregnancy duration:
not examined
Changes in number of pregnant:
no effects observed
Description (incidence and severity):
The number of confirmed pregnant, evaluated dams was 24 in the Control and the Mid dose groups, and 25 in the Low and High dose groups.
Other effects:
no effects observed
Description (incidence and severity):
The mean foetal weight per litter in the test item treated groups did not differ significantly from the control mean value in any treated group.
No abnormalities were observed on the placentas of any treated or control animals.
Details on maternal toxic effects:
As there were no maternal findings, it was considered that the test item had no maternal toxicity effect at dose levels up to 1000 mg/kg bw/day.
Key result
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: No maternal toxicity observed at the highest dose level
Key result
Abnormalities:
no effects observed
Description (incidence and severity):
Up to 1000 mg/kg bw/day did not induce maternal toxicity
Fetal body weight changes:
no effects observed
Description (incidence and severity):
The mean foetal weight per litter values and the number of retarded foetuses (runts) were comparable with the control in all test item treated groups.
Reduction in number of live offspring:
no effects observed
Description (incidence and severity):
The mean number of viable foetuses was comparable with the control mean in all test item treated groups.
Changes in sex ratio:
no effects observed
Description (incidence and severity):
There was no toxicologically significant difference in the sex distribution of foetuses between the control and treatment groups and no effect on anogenital distance of either sex.
Changes in litter size and weights:
no effects observed
Description (incidence and severity):
The mean foetal weight per litter values and the number of retarded foetuses (runts) were comparable with the control in all test item treated groups.
Changes in postnatal survival:
not examined
External malformations:
no effects observed
Description (incidence and severity):
Foetal malformations observed in the study were considered to be incidental. The malformations showed no dose dependency, and were not regarded as a test item related effect.
Skeletal malformations:
no effects observed
Description (incidence and severity):
There was no test item related effects on external, visceral, or skeletal development of foetuses in the study.
Visceral malformations:
no effects observed
Description (incidence and severity):
There was no test item related effects on external, visceral, or skeletal development of foetuses in the study.
Other effects:
not specified
Details on embryotoxic / teratogenic effects:
At dose levels up to 1000 mg/kg bw/day did not induce maternal toxicity, embryotoxicity, foetotoxicity, or teratogenicity.
Key result
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day
Based on:
test mat.
Sex:
female
Basis for effect level:
other: At dose levels up to 1000 mg/kg bw/day did not induce foetotoxicity
Key result
Abnormalities:
no effects observed
Description (incidence and severity):
At the highest dose levels up to 1000 mg/kg bw/day did not induce embryotoxicity or foetotoxicity.
Key result
Developmental effects observed:
no
Conclusions:
The test item, when administered daily by oral gavage to pregnant Hannover Wistar rats from gestation days GD6 to GD19 at dose levels up to 1000 mg/kg bw/day did not induce maternal toxicity, embryotoxicity, foetotoxicity, or teratogenicity. There were no malformations or developmental effects attributed to the test item at any dose level.

NOAELmaternal toxicity: 1000 mg/kg bw/day
Based on the lack of any test-item related maternal toxicity effect in any treatment group.
NOAELembryotoxicity: 1000 mg/kg bw/day
Based on the lack of any test-item related intrauterine effect in any treatment group.
NOAELfoetotoxicity: 1000 mg/kg bw/day
Based on the lack of any test-item related foetotoxicity effect in any treatment group.
NOAELteratogenecity: 1000 mg/kg bw/day
Based on the lack of any developmental effects in any treatment group. 
Executive summary:

The teratogenic potential of the test item was evaluated in female Hannover Wistar rats in year 2019 according to OECD 414 and GLP. The test substance was administered daily by oral gavage to pregnant rats on gestation days 6 -19, at dose levels of 100, 300 and 1000 mg/kg bw/day. The dams were sacrificed at gestation day 20 and the foetuses were examined for visceral and skeletal variations and malformations.

Up to dose level of 1000 mg/kg bw/day did not induce maternal toxicity, embryotoxicity, foetotoxicity, or teratogenicity. There were no malformations or developmental effects attributed to the test item at any dose level

The following no-observed-adverse-effect (NOAEL) levels were derived:

 NOAELmaternal toxicity:                  1000 mg/kg bw/day

Based on the lack of any test-item related maternal toxicity effect in any treatment group.

NOAELembryotoxicity:                    1000 mg/kg bw/day

Based on the lack of any test-item related intrauterine effect in any treatment group.

NOAELfoetotoxicity:                               1000 mg/kg bw/day

Based on the lack of any test-item related foetotoxicity effect in any treatment group.

NOAELteratogenecity:                     1000 mg/kg bw/day

Based on the lack of any developmental effects in any treatment group.

Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
1 000 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
Guideline study
Effect on developmental toxicity: via inhalation route
Endpoint conclusion:
no study available
Effect on developmental toxicity: via dermal route
Endpoint conclusion:
no study available
Additional information

The developmental toxicity study requirement according to REACH Annex IX, section 8.7 was waived as the substance did not show any toxicological activity in the available tests. A sub-chronic oral toxicity study in the rat was performed according to OECD Guideline 408. The results of this oral gavage study indicate that the test item has no potential to produce toxic effects when administered to rats at dosage of 500 mg/kg/day. Based on the lack of treatment-related effects in clinical signs, ophthalmic examinations, feed consumption, weight gain, clinical pathology, organ weights, gross pathology, microscopic pathology, and FOB results, the no-observed-effect level (NOEL) for subchronic exposure to AAEM was considered to be 500 mg/kg/day for both male and female rats when administered 5 days per week for 13 weeks. The toxic potential of LZ649 on basic reproductive processes was assessed in a OECD 421 screening study in which rats were dosed for two weeks prior to pairing, throughout gestation and lactation at dose levels of 50, 150 or 500 mg/kg/day. There was no adverse effect of administration with LZ649 on clinical condition, bodyweight, and food consumption, macroscopic and microscopic appearance. Reproductive performance assessments of oestrous cycles, gestation length and parturition for F0 females also showed no test article related change. Organ weights for F0 males were similar throughout the groups. The clinical condition, litter size and survival and sex ratio of offspring, exposed in utero or via the milk, were not affected by doses up to and including 500 mg/kg/day. The results observed in this screening study conclude that LZ649 has no effect on reproductive performance and offspring survival or development at doses up to and including 500 mg/kg/day.

The teratogenic potential of the test item was evaluated in female Hannover Wistar rats according to OECD 414. The test substance was administered daily by oral gavage to pregnant rats on gestation days 6 -19, at dose levels of 100, 300 and 1000 mg/kg bw/day. The dams were sacrificed at gestation day 20 and the foetuses were examined for visceral and skeletal variations and malformations.

Up to dose level of 1000 mg/kg bw/day did not induce maternal toxicity, embryotoxicity, foetotoxicity, or teratogenicity. There were no malformations or developmental effects attributed to the test item at any dose level. The NOAEL is 1000 mg/kg bw/day.

Justification for classification or non-classification

Based on the data available the substance is not classified and labeled according to Regulation 1272/2008/EEC (CLP).

Additional information