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Toxicological information

Repeated dose toxicity: oral

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Administrative data

Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2021-2022
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2022

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)
Version / remarks:
25.06.2018
GLP compliance:
yes (incl. QA statement)
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
3-[[3-[[(2-cyanoethyl)amino]methyl]-3,5,5-trimethylcyclohexyl]amino]propiononitrile
EC Number:
300-496-1
EC Name:
3-[[3-[[(2-cyanoethyl)amino]methyl]-3,5,5-trimethylcyclohexyl]amino]propiononitrile
Cas Number:
93940-97-7
Molecular formula:
C16H28N4
IUPAC Name:
3-[(3-{[(2-cyanoethyl)amino]methyl}-3,5,5-trimethylcyclohexyl)amino]propanenitrile
Test material form:
liquid
Specific details on test material used for the study:
Identity: 3-[[3-[[(2-cyanoethyl)amino]methyl]-3,5,5-trimethylcyclohexyl]amino]propiononitrile (13/0241-5)
Label name: Baxxodur(TM) PC 136
Alternative name: MIPDA
CAS number: 93940-97-7 (Batch no. 18315014)
Retest date: 21 July 2022
Appearance: clear colourless liquid (purity 97.0 area%)
Storage conditions: room temperature

Test animals

Species:
rat
Strain:
Wistar
Remarks:
Wistar Hannover rats
Sex:
male/female
Details on test animals or test system and environmental conditions:
A total of 90 Wistar Hannover rats (45 males and 45 females), 27-29 days old and with body weight of approximately 75-99 g, were ordered from Envigo RMS srl. After arrival, the weight range for each sex was determined (84 - 107 g for males, 86 - 102 for females) and the animals were temporarily identified within the cage by means of a coloured mark on the tail. A health check was then performed by a veterinarian. An acclimatisation period of 19 days was allowed before the start of treatment, during which time the health status of the animals was assessed by thorough observations.

The animals were housed in a limited access rodent facility. Animal room controls were set to maintain temperature and relative humidity at 22 °C ± 2 °C and 55 % ± 15 %, respectively; actual conditions were monitored, recorded and the records retained. No relevant deviations from these ranges were recorded during the study. There were approximately 15 to 20 air changes per hour and the rooms were lit by artificial light for 12 hours each day. The animals were housed up to 5 of one sex to a cage, in clear polysulfone solid bottomed cages as indicated in the relevant SOP. Nesting material was provided inside suitable bedding bags and changed at least twice a week. Drinking water was supplied ad libitum to each cage via water bottles. A commercially available laboratory rodent diet was offered ad libitum throughout the study. There was no information available to indicate that any non-nutrient substance likely to influence the effect of the test item was present in the drinking water or the diet. Records of analysis of water, diet and bedding material are kept on file. Dated and signed records of activities relating to the day to day running and maintenance of the study in the animal house were recorded.

Allocation to groups
On the day of allocation (7 days prior to the start of treatment) all animals were weighed. Animals at the extremes of the weight distribution were excluded to leave the required number of animals. The rats were allocated to the 4 groups by computerised stratified randomisation to give approximately equal initial group mean body weights. Individuals were uniquely identified within the study by sex, tattoo on the hind feet and ear notch and housed up to 3 of one sex per cage. The cages were identified by a label recording the study number, animal numbers and details of treatment. The arrangement of cages in batteries was such that cages from each treatment group were evenly distributed across the battery to minimise possible environmental effects. Two males initially allocated to Groups 2 (no. A4326024) and 4 (no. A4326078) respectively, and 2 females allocated to Groups 3 (no. A4326045) and 4 (no. A4326071) respectively, showing ocular lesions, were replaced with spare animals showing no ocular abnormality, from the batch initially ordered for the study. No replacements occurred after the first dose was administered.

Deviation
The body weights of the animals at arrival were slightly outside the range indicated in the study protocol (84 - 107 g for males, 86 - 102 for females instead of 75 to 99 g). For one female of Group 4 (no. A4326069) the thyroid weight was inadvertently not recorded. These deviations were not considered to have compromised the purpose or conduct of the study. No other deviations occurred during the study.

Administration / exposure

Route of administration:
oral: gavage
Details on route of administration:
The test item was administered orally by gavage at a dose volume of 10 mL/kg body weight/day. Control animals received the vehicle alone at the same dose volume. The dose was administered to each animal on the basis of the most recently recorded body weight and the volume administered was recorded for each animal.
Vehicle:
CMC (carboxymethyl cellulose)
Remarks:
0.5% Sodium carboxymethyl cellulose (CMC) suspension in deionized water (with 10 mg/100 mL Cremophor). Preparations were made weekly (concentrations of 1, 3 and 10 mg/mL), unless specified otherwise.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The analytical method was validated in ERBC Study no. A4325 in the range from 1 to 10 mg/mL. Linearity, accuracy and precision were within the limits stated in the validation protocol (r > 0.995; accuracy 85-115%; precision CV < 10%).
A 28 hour stability at room temperature and an 8 day stabiliy at 2-8°C was verified in the range from 1 to 10 mg/mL. Suspensions are considered to be stable if concentration and homogeneity, after the defined period of storage, are still acceptable (85%-115% for concentration and CV < 10% for homogeneity). The proposed preparation procedure for the test item was checked in the range from 1 to 10 mg/mL by chemical analysis (concentration and homogeneity) to confirm that the method was suitable. Final results for all levels were within the acceptability limits for concentration (85-115%) and homogeneity (CV < 10%). Samples of the formulations prepared on Week 1 and Week 13 were analysed to check the homogeneity and concentration. Results of these analyses were within the acceptability limits for suspensions (85-115% for concentration and CV < 10% for homogeneity). The validated software used for this activity was Chromeleon.
Duration of treatment / exposure:
All animals were dosed once a day, 7 days a week, for a minimum of 13 consecutive weeks. All animals were dosed up until the day before necropsy.
Doses / concentrationsopen allclose all
Dose / conc.:
10 mg/kg bw/day (nominal)
Remarks:
low dose
Dose / conc.:
30 mg/kg bw/day (nominal)
Remarks:
medium dose
Dose / conc.:
100 mg/kg bw/day (nominal)
Remarks:
high dose
No. of animals per sex per dose:
Each group comprised 10 male and 10 female rats.
Control animals:
yes, concurrent vehicle
Details on study design:
dose level rational
Dose levels were selected in consultation with the Sponsor based on information from a preliminary OECD TG 422 study (BASF Project No.: 85R0241/13R184). In this study, dose levels of 15, 50, 150 mg/kg/day were used for dosing Wistar Hannover rats, by oral gavage, with a dose volume of 10 mL/kg. The results obtained showed a reduction in body weight and food consumption in animals treated at 150 mg/kg/day as well as adverse changes in clinical pathology parameters and histopathology (the findings were mainly focus on brain, eyes with optic nerves, liver, skeletal muscle, axillary and mesenteric lymph nodes, kidney and spleen).
In detail, in the high dose group 150 mg/kg bw/day, histopathology revealed in the liver a centrilobular microvesicular vacuolation in all males and females accompanied by single cell necrosis. It cannot be excluded if other hepatic findings like multifocal necrosis, multinuclear hepatocytes and karyomegaly, affecting very few animals were treatment-related and were, therefore, assumed to be possibly treatment-related and adverse. In the skeletal muscle of males and females, the fiber vacuolation was accompanied by necrosis and regeneration of fibers. In the axillary lymph node of females, the vacuolation was present in the high endothelial venules (HEV) and was accompanied by apoptosis. The mesenteric lymph nodes of females showed vacuolation in the HEV, lymphocytes of medullary cords and macrophages of medullary sinuses, apoptosis and macrophage aggregates in the periphery of paracortical areas, and decreased cellularity in paracortical regions (T-lymphocytes). In the kidneys of females, a tubular cytoplasmic macro- and microvesicular vacuolation affecting all animals was accompanied by tubular degeneration/regeneration and granulomatous inflammation. Both types of vacuolation affected predominantly the cortical tubular epithelium but the microvesicular vacuolation extended also into the outer stripe of the outer medulla. The spleen of females displayed a vacuolation of the B-lymphocytes (follicular and marginal zone) and a decreased cellularity of T-lymphocytes in the periarteriolar lymphocyte sheath (PALS). Finally, a microvesicular vacuolation without signs of cytotoxicity was detected in the plexus choroideus of the brain, and in the stromal iris and choroid of the eyes of males and females.
According to the OECD TG 408, the highest dose level should be chosen with the aim to induce toxicity but not death or severe suffering. Considering all above mentioned effects, the dose of 150 mg/kg bw/day was considered as severe suffering. Since the treatment period of the OECD TG 408 is much longer than the treatment in the OECD TG 422 and the no observed adverse effect level (NOAEL) for general systemic toxicity was set at 50 mg/kg/day for male and female Wistar Hannover rats, a dose level of 100 mg/kg/day was considered suitable to be the highest dose level for this 13-week treatment study.

Examinations

Observations and examinations performed and frequency:
In vivo observations
Full records were maintained for all measurements and observations.
Mortality
Throughout the study, all animals were checked early in each working day and again in the afternoon. At weekends and Public Holiday a similar procedure was followed except that the final check was carried out at approximately mid-day.

Clinical signs and neurotoxicity assessment
All clinical signs were recorded for individual animals. Once before commencement of treatment and once daily during the study, each animal was observed and any clinical signs was recorded. These data (clinical signs) will be archived with all raw data. Observations were performed at the same time interval each day (approximately 30 minutes, 2 and 4 hours up to Day 3, 30 minutes post-dose from Day 4), the interval was selected taking into consideration the presence of post-dose reactions. Once before commencement of treatment and at least once per week during the study from
the start of treatment, each animal was given a detailed clinical examination. Each animal was observed in an open arena. The test included observation and record of changes in gait and posture, reactivity to handling, presence of clonic or tonic movements, stereotypies or bizarre behaviour and effects on the autonomic nervous system (e.g. lachrymation, piloerection, unusual respiratory pattern). Changes in fur, skin, eyes, mucous membranes, occurrences of secretions and excretions were also recorded. Once during Week 13 of treatment an evaluation of sensory reactivity to stimuli of different modalities (e.g. auditory, visual and proprioceptive stimuli) and an assessment of grip strength were also performed.

Motor activity assessment (MA)
The motor activity (MA) of all animals was measured once during Week 12 of treatment by an automated activity recording. Measurements were performed using a computer generated random order.

Body weight
Each animal was weighed on the day of allocation to treatment group, on the day that treatment commenced, weekly thereafter and just prior to necropsy.

Food consumption
The weight of food consumed by each cage of rats was recorded at weekly intervals following allocation. The group mean daily intake per rat was calculated.

Ophthalmoscopy
Both eyes of all animals were examined prior to the commencement of treatment by means of an ophthalmoscope, and by a slit-lamp microscope, after the instillation of 0.5% Tropicamide (Visumidriatic®, Visufarma, Rome, Italy). Two males and 2 females showing ocular lesions, were replaced with spare animals showing no ocular abnormality, from the batch initially ordered for the study. The eyes of all animals from high-dose and control groups were re-examined during Week 13 of treatment.

Oestrous cycle
At the end of the treatment and just prior to necropsy, vaginal smears were taken from all animals.

Clinical pathology investigations
During the last week of treatment, blood samples of approximately 1.1 mL for thyroid hormone determination (T3, T4, TSH) were collected from the sublingual vein of all animals, under light isoflurane anaesthesia and condition of food deprivation. Samples were transferred into tubes containing no anticoagulant and centrifuged at room temperature. The serum obtained was divided in two aliquots, the first (Aliquot A) with at least 250 µL of serum and the second (Aliquot B) with the remaining amount. Samples were stored at -20°C pending despatch to the test site. Prior to necropsy, blood samples were collected from all animals under isoflurane anaesthesia from the abdominal vena cava for haematology, coagulation and clinical chemistry, under conditions of food deprivation. The study day indicated on Tables/Appendices refers to the first scheduled day (Day 92 for males; Day 93 for females). Blood samples were collected and analysed in the same order. The blood samples collected were divided into tubes as follows:
– EDTA anticoagulant for haematological investigations
– Heparin anticoagulant for biochemical tests
– Citrate anticoagulant for coagulation tests
– No anticoagulant for hormone determination

The measurements performed on blood samples are listed below:
Haematology
– Haematocrit
– Haemoglobin
– Red blood cell count
– Reticulocyte count
– Mean red blood cell volume
– Mean corpuscular haemoglobin
– Mean corpuscular haemoglobin concentration
– White blood cell count
– Differential leucocyte count
· Neutrophils
· Lymphocytes
· Eosinophils
· Basophils
· Monocytes
· Large unstained cells
– Platelets
These parameters were analysed by Siemens Advia 120. No haematological data are available for one Group 2 female (no. A4326021) since the sample was not analysed (clotted samples).

Coagulation
– Prothrombin time
This parameter was analysed by Instrumentation Laboratory ACL Elite PRO. No coagulation data are available for one Group 2 male (no. A4326030) and for one Group 3 female (no. A4326057) since the samples were not analysed (samples with microfilaments).

Clinical chemistry
– Alkaline phosphatase
– Alanine aminotransferase
– Aspartate aminotransferase
– Gamma-glutamyltransferase
– Urea
– Blood urea nitrogen
– Creatinine
– Glucose
– Triglycerides
– Inorganic phosphorus
– Total bilirubin
– Total cholesterol
– HDL
– LDL
– Total protein
– Albumin
– Globulin
– A/G Ratio
– Sodium
– Potassium
– Calcium
– Chloride
These parameters were analysed by Siemens Advia 1800.

Immunoanalysis -Thyroid hormone determination (T3, T4 and TSH) (delegated phase)
Immunoanalysis was carried out according to immunoanalytical methods validated at the Test Site:
– RV-T3/R/S-BKM/RIA-002 for T3
– RV-T4/R/S-BKM/RIA-002 for T4
– RV-TSH/R/S-IZO/RIA-002 for TSH
Samples from all animals were assayed to determine the serum levels of Total triiodothyronine (total T3), Total thyroxine (total T4) and Thyroid stimulating hormone (TSH) by RadioImmunoAssay (RIA).
Total number of samples: 80 × 2 aliquots. The aliquots were maintained at -20°C until transfer to the Test Site indicated below, where the immunoanalysis was carried out.

Samples were labelled as follows: study number, animal number, group and sex, sampling date. Samples were transported in dry ice, by a dedicated courier and were subject to the latter’s standard insurance policy. In the first instance, only Aliquots A were despatched to the Test Site. The backup samples (aliquots B) were not despatched since not requested by the Test Site. These samples were
destroyed with the Study Director’s authorisation and following Sponsor’s approval.

A Final Delegated Phase Report with the results of these analyses was sent for inclusion in the Final Study Report. The results were presented as individual data (mean of singlicate or duplicates), group mean and group standard deviation. Evaluation of the data and the statistical analysis was performed by the testing lab.
Sacrifice and pathology:
Terminal studies
Euthanasia
Animals that had completed the scheduled test period were killed by exsanguination under isoflurane anaesthesia. All animals were subjected to necropsy, supervised by a pathologist, as detailed below.

Necropsy
The clinical history of the animals was studied and a detailed post mortem examination was conducted (including examination of the external surface and orifices). Changes were noted, the requisite organs weighed and the required tissue samples preserved in fixative.

Organ weights
From all animals completing the scheduled test period, all organs were dissected free of fat and weighed. The ratios of organ weight to body weight were calculated for each animal. For one female of Group 4 (no. A4326069), the thyroid weight was inadvertently not recorded.

Tissues fixed and preserved
Samples of all the tissues listed in section 4.5.9 were fixed and preserved in 10% neutral buffered formalin (except eyes, optic nerves, Harderian glands, testes and epididymides which were fixed in Modified Davidson’s fluid and preserved in 70% ethyl alcohol).

Despatch of tissues and organs (Delegated phase)
The wet tissues required for histopathological examination were shipped at room temperature by dedicated courier to TPL Path Labs GmbH. Prior to shipment, the Principal Investigator was notified by e-mail. A detailed list of samples accompanying the shipment was included for control at receipt. Moreover, individual data of macroscopic observations, the history, the histopathology check list and scheme for males and females were sent to TPL Path Labs GmbH.

Paraffin blocks preparation and despatch - Histotechnology - (Delegated phase)
The preparation of paraffin blocks, following the ERBC embedding scheme for male and female, was performed at TPL Path Labs GmbH. In first instance, only the paraffin blocks were returned to Test Facility for the preparation of the slides and histopathological examination. Prior to shipment, the Study Director was notified by e-mail. In the second instance, the wet tissues were returned to Test Facility, following Study Director communication. A detailed list of samples accompanying the shipment was included for control at receipt.

Histopathological examination
The tissues required for histopathological examinations were cut at 5 micrometre thickness and stained with haematoxylin and eosin. In addition, the testes and epididymides of all animals were cut at 2-3 micrometre thickness and stained with Periodic Acid Schiff (PAS). The morphological evaluation of the seminiferous epithelium (staging of spermatogenic cycle) was performed initially in all animals in the control and high dose groups killed at the end of the 13 weeks of treatment. Sections from the vagina, uterus and ovaries from the control and high dose females killed at the end of the 13 weeks of treatment were examined for evaluation of oestrous cycle.
The examination was as detailed below:
i Tissues specified in the guideline from all animals in the control and high dose groups.
ii All abnormalities in all groups.
On the basis of the histopathological differences observed between control and high dose groups, the examination was extended to the epidydimides of males in Groups 2 and 3.

Histological processing from wet tissue to H&E stained slide - Histotechnology - (Delegated phase)
Sections of the epidydimides of males in Groups 2 and 3 were cut at 5 micrometre thickness and stained with haematoxylin and eosin. The slides were returned to Test Facility for the histopathological examination. A detailed list of samples accompanying the shipment was included for control at receipt.
Statistics:
Statistical analysis
Standard deviations were calculated as considered appropriate. For continuous variables the significance of the differences amongst groups was assessed by analysis of variance. Differences between each treated group and the control group were assessed by Dunnett’s test using a pooled error variance. The homogeneity of the data was verified by Bartlett’s test before Dunnett’s test. If the data were found to be inhomogeneous a Modified t test (Cochran and Cox) was applied.
The mean values, standard deviations and statistical analysis were calculated from the actual values in the computer without rounding off. Statistical analysis of histopathological finding was carried out by means of a nonparametric Kolmogorov-Smirnov test.

Results and discussion

Results of examinations

Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
No treatment-related clinical signs were observed during the study.
Hard abdomen was observed for some days in a single female dosed at 30 mg/kg/day (no. 43), while piloerection was occasionally noted in 2 males dosed at 100 mg/kg/day (nos. 78 and 80). These signs were not considered to be treatment-related.
Mortality:
no mortality observed
Description (incidence):
No mortality occurred during the study.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
When compared to control animals, no treatment-related changes were noted in mean body weights and body weight gain in both genders during the study. There were no treatment-related terminal body weight changes.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
No treatment-related changes were observed in food consumption in male and female animals, during the study.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
no effects observed
Description (incidence and severity):
Animals with no ocular abnormality were selected for the study by an ophthalmoscopic examination performed before the start of treatment. No findings were detected in both eyes of all animals, from high dose and control groups, when they were re-examined during Week 13 of treatment.
Haematological findings:
effects observed, non-treatment-related
Description (incidence and severity):
No treatment-related changes were recorded. The statistically significant differences recorded between control and treated animals (lymphocytes and large unstained cells in males, reticulocytes and monocytes in females) were within the range of expected biological variation and/or not dose-related, therefore they were considered to be incidental.
Clinical biochemistry findings:
effects observed, non-treatment-related
Description (incidence and severity):
Females dosed at 100 mg/kg/day showed decreases of LDL cholesterol (-37%) and an increase of triglycerides (62%). LDL cholesterol was also decreased in females receiving 30 mg/kg/day (-28%). Due to the slight severity, these changes were not considered to be adverse, even though their relation with the test item cannot be definitively excluded. The other statistically significant differences recorded between control and treated animals (alkaline phosphatase, sodium and potassium in males, alanine aminotransferase, aspartate aminotransferase, creatinine, chloride and calcium in females), were within the range of expected biological variation and/or not dose-related, therefore they were considered to be unrelated to treatment.
Endocrine findings:
effects observed, non-treatment-related
Description (incidence and severity):
Coagulation
No changes were recorded.

Oestrous cycle
No treatment-related anomalies were recorded in the oestrous cycle of the treated females, when compared to controls, at the end of the study.

Thyroid hormone determination (T3, T4 and TSH) (delegated phase)
TSH was lower than control in males dosed at 100 mg/kg/day (45% to 47%). Changes were mostly due to the high values of some control animals (nos. A4326002, A4326006, A4326014 and A4326018), which showed values outside the range of historical data (ULOQ = 12.2 ng/mL). In addition, no other related changes were recorded (i.e. T3/T4 increases and thyroid weight changes) and no dose-relation were observed, therefore the finding recorded was considered to be incidental.
Urinalysis findings:
not specified
Behaviour (functional findings):
effects observed, non-treatment-related
Description (incidence and severity):
Observation of treated animals at removal from the cage and in an open arena did not reveal significant changes when compared to controls, when examined at the weekly interval.
Some differences often associated to significance, at statistical analysis were noted between control and treated groups during the study:
– on Day 38 of the study a decrease in defecation numbers at 30 and 100 mg/kg/day in males;
– on day 66 an increase in rearing values at 30 and 100 mg/kg/day in males;
– on Day 32 an increase in rearing values at 30 and 100 mg/kg/day in females;
– on Day 60 an increase in rearing values at all doses in females.
These temporary changes were considered to be incidental.

No relevant differences that could be considered treatment-related were observed at functional tests (sensory reaction to stimuli, landing footsplay, grip strength) performed at the end of the treatment period. Increase in grip strength measurement (first, second trials and mean values), at the dose level of 30 and/or 100 mg/kg/day was recorded in treated males, when compared to control. Motor activity measurements performed at the end of the treatment was similar among treated animals and controls for both sexes although a slight increase in the value was noted in females receiving 100 mg/kg/day compared to the control group.
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, non-treatment-related
Description (incidence and severity):
There were no treatment-related organ weight changes when compared to the controls. Any organ weight variations, including the increase in relative and absolute mean liver weight in high dose males (+ 14%
for liver relative weight to body weight) were not correlated to any histopathological changes, in the range of expected spontaneous changes in rats of the same age and considered unrelated to treatment.
Gross pathological findings:
effects observed, non-treatment-related
Description (incidence and severity):
There were no treatment-related macroscopic observations at the end of the treatment period. Any macroscopic observations had a comparable incidence in control and treated groups and/or are characteristically seen in untreated rats
of the same age and were considered incidental and unrelated to treatment.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, non-treatment-related
Description (incidence and severity):
Treatment-related microscopic observations, associated with the oral administration of 3-[[3-[[(2-cyanoethyl)amino]methyl]-3,5,5-trimethyl
cyclohexyl]amino]propiononitrile were present only in the epididymides of all high dose treated males (10/10) when compared to the controls. Findings consisted in mild to moderate vacuolation of the epithelium. Vacuolation was of microvesicular type (multiple fine to small vacuoles, that were no larger than the nuclei of the respective cell types) and mostly present in the caudal part of the corpus of the epididymides. Since there were no treatment-related changes in organ weight or cell morphological changes leading to cytotoxicity (cell death, degeneration and/or inflammation), vacuolisation was considered treatment-related but not adverse.
There were no test item-related microscopic observations in the testis (stage aware evaluation on PAS-stained slides). There was no test item-related effect on estrous cycle stages. Any other microscopic observations than those listed above had a comparable incidence in control and treated groups and/or are characteristically seen in untreated rats of the same age and were considered incidental and unrelated to treatment.
Histopathological findings: neoplastic:
no effects observed
Other effects:
not specified

Effect levels

Key result
Dose descriptor:
NOAEL
Effect level:
100 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects

Target system / organ toxicity

Key result
Critical effects observed:
no

Applicant's summary and conclusion

Conclusions:
With reference to the results reported, it can it can be concluded that the No Observed Adverse Effect Level (NOAEL) for males and females was 100 mg/kg/day.
Executive summary:

The oral toxicity of 3-[[3-[[(2-cyanoethyl)amino]methyl]-3,5,5-trimethylcyclohexyl]amino]propiononitrile in Wistar Hannover rats, following daily oral administration at dose levels of 10, 30 and 100 mg/kg/day for 13 consecutive weeks, was investigated in this study according to OECD 408.


No mortality occurred and no relevant clinical signs were recorded during the study.


No signs indicating neurotoxic effects were seen during the in vivo phase of the study.


Body weight, food consumption and oestrous cycle were not affected by treatment.


No lesions were recorded at ophthalmological examination.


No treatment-related changes were recorded at the end of treatment period in haematological parameters and coagulation. Decreases in LDL cholesterol were observed in females dosed at 30 and 100 mg/kg/day, while an increase in triglycerides was seen in females receiving 100 mg/kg/day. Due to the slight severity, these changes were not considered to be adverse, even though their relation with the test item cannot be definitively excluded.


No treatment-related changes were recorded for thyroid hormones levels.


No treatment-related changes were observed in terminal body weight and absolute and relative organ weights of treated animals, when compared to the controls.


No treatment-related post mortem macroscopic observations were recorded in animals of sexes when compared to the control. Treatment-related microscopic but not adverse findings were present in the epididymides of all high dose treated males (100 mg/kg/day) when compared to the controls. There were no test item-related microscopic observations in the testis (stage aware evaluation on PAS-stained slides).


There was no test item-related effect on estrous cycle stages.


With reference to the above mentioned results it can it can be concluded that the No Observed Adverse Effect Level (NOAEL) for males and females was 100 mg/kg/day.