Registration Dossier

Administrative data

Key value for chemical safety assessment

Effects on fertility

Link to relevant study records
Reference
Endpoint:
screening for reproductive / developmental toxicity
Remarks:
based on test type (migrated information)
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Read-across from GLP-compliant guideline study performed with similar substance.
Reason / purpose:
reference to same study
Reference:
Composition 0
Qualifier:
according to
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Version / remarks:
of 1996
Deviations:
no
GLP compliance:
yes (incl. certificate)
Limit test:
no
Test material information:
Composition 1
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Sprague Dawley rats, strain: Crl:CD(SD) with appropriate range of bodyweight at study start.
- Source: Charles River (UK) Ltd.
- Age at treatment start: 61-68 days.
- Weight at treatment start: Males: minimum 332 g, maximum 397 g,
Females: minimum 210 g, maximum 255 g.
- Housing Inside a barriered rodent facility:
all animals pre-pairing + toxicity subgroups: In groups up to 5 by sex in solid floor polycarbonate cages.
during pairing (1 male+1 female/cage): In RB3 modified polypropylene cages with stainless steel grid-floor over absorbent paper-lined trays.
males after pairing: In groups of up to 5 in solid floor polycarbonate cages.
females during gestation and lactation: Females housed individually (+litter) in solid floor polycarbonate cages.
- Bedding material (in solid floor cages): Wood based bedding.
- Cage enrichment: Aspen chew block + plastic shelter (except during pairing or post gestation day 20).
- Diet (ad libitum): Standard rodent diet (SDS VRF1 Certified) without antibiotic, chemotherapeutic or prophylactic agent.
- Fasting (diet withheld): Main phase males and Toxicity phase females overnight before blood sampling for clinical pathology.
- Acclimation period: 5 days before treatment start, under laboratory conditions.

Routine analysis of the batch of diet used and water, chew blocks and bedding material did not provide evidence of contamination that might have prejudiced the study.

IN-LIFE DATES:
- Duration of test, males & toxicity phase females: Five weeks
Duration of test, main phase females (i.e. reproductive subgroup): From 14 days prior to pairing to day 7 of lactation.
Duration of test, offspring: From birth to day 7 of lactation.

ENVIRONMENTAL CONDITIONS

Air conditioned room kept at positve pressure without re-circulation of the filtered fresh air supplied to the room.
Controlled environment, environmental conditions were set at:
- Temperature (°C): 21 ± 2°C
- Relative Humidity (%): 40 to 70%
- Photoperiod (artificial lighting): 12 hrs day / 12 hrs night
Deviations from the target ranges for temperature and relative humidity were not evident.
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
- Concentration in vehicle: The concentration of the test material in vehicle was 20, 60, and 200 mg/ml.
- Amount (dose volume by gavage): 5 mL/kg bw/day..
Actual dose volumes were calculated at about weekly or shorter intervals accounting for the latest body weight. Litter animals were not dosed.
Details on mating procedure:
- Male/female ratio per cage: 1/1
- Length of cohabitation: At the most 14 days, until proof of pregnancy was confirmed. 
- Proof of successful mating: Formation of at least one copulation plug and a sperm positive vaginal smear.
The day this was found was referred to as day 0 of gestation.
(During cohabitation, females were checked every morning for pregnancy).

Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
- Chemical analysis of test material formulations by high performance liquid chromatography coupled with mass spectrometry (HPLC-MS/MS).
- Concentrations (verified at first and last treatment week) of the test material formulations were confirmed at each dose level.
- Chemical analysis confirmed that the mean concentrations of WS405777 in prepared formulations were 100% to 113% of the corresponding
nominal concentration, thus confirming accuracy of formulation.
Duration of treatment / exposure:
- Treatment period, males & toxicity phase females: Daily, for five consecutive weeks, in males commencing 14 days prior to mating
- Treatment period, main phase females (i.e. reproductive subgroup): 43 to 46 days (from 14 days prior to pairing to day 6 of lactation)
- Offspring were not dosed
Frequency of treatment:
Daily, 7 days/week (during parturition, dosing omitted as appropriate)
Details on study schedule:
- Age at mating of the mated animals in the study: 11 to 12 weeks
Remarks:
Doses / Concentrations:
0 (vehicle control), 100, 300, 1000 mg/kg bw/day
Basis:
actual ingested
No. of animals per sex per dose:
Main phase animals (i.e. reproductive subgroups): 10 males / 10 females
Control animals:
yes, concurrent vehicle
Details on study design:
This study was conducted to examine both repeated dose toxicity and  reproductive/developmental toxicity as an OECD screening combined study
(OECD 422 test guideline).  Therefore, animals initially entering the study were divided into toxicity subgroup animals (toxicity phase) and reproductive subgroup animals (main phase), whereby 5 of the 10 F0 males (used for pairing) per dose group formed the toxicity male subgroups.

Dose selection was based on the results of a 7-day preliminary oral toxicity study in the rat in which dose levels of 100, 300 or 1000 mg/kg/day did not have any overt treatment-related effects on young adult animals (females nulliparous and non-pregnant). Accordingly the same dose levels were used in the present OECD 422 combined repeat dose toxicity and reprotoxic/develpmental toxicity screening study.
Positive control:
Not included in the study.
Parental animals: Observations and examinations:
Clinical observations performed and frequency:
- Clinical signs : At least twice a day (before and after administration)
- Detailed physical examination
and arena observations: Before treatment start and at least once a treatment week.
- Body weight, all males: Weekly throughout the study.
Body weight, Repro. Females: Weekly for pre-pairing period; on gestation days 0, 6, 13, 20; on lactation days 1,4 and 7.
- Food consumption, all males: Weekly for pre-pairing period and for the period after mating.
Food cons., Repro. Females: Weekly for pre-pairing period, during gestation for days 0-5, 6-12, 13-19, during lactation for days 1-3 & 4-6.
Estrous cyclicity (parental animals):
Frequency of vaginal oestrus was determined by examination of vaginal smears taken daily from all main phase (i.e. reproductive subgroup) females from the beginning of the treatment period to the day of confirmed copulation.
- Regular: All observed cycles of 4 or 5 days
- Irregular: At least one cycle of 2, 3 or 6 to 10 days
- Acyclic: At least 10 days without oestrus
Sperm parameters (parental animals):
Parameters examined in male parental animals:
- testis weight,
- epididymis weight
- detailed qualitative histopathology examination of the testes taking into account the tubular stages of the spermatogenic cycle. This was to identify treatment related effects such as missing germ cell layers or types, retained spermatids, multinucleate or apoptotic germ cells and sloughing of spermatogenic cells in the lumen. Any cell- or stage-specificity of testicular findings was noted.
Litter observations:
STANDARDISATION OF LITTERS: Not performed. The study ended on lactation day 6.

LITTER PARAMETERS EXAMINED
- From Day 20 post copulation 3 times a day checks for evidence of parturition, any difficulties and numbers of live and dead offspring.
- Total litter size on day 1 of age and mortality/live litter size on each day until 7 days after littering.
- Sex ratio expressed as percentage males and calculated for total offspring on Day 1 and for live offspring on Days 1, 4 and 7
(No. of male pups in litter/No. of offspring in litter) x 100
- Gestation index (No. of live litters born on day 0/No. of living pregnant females) x 100
- Clinical signs, recorded daily
- Individual body weight of live pups (on days 1, 4 and 7 after littering) and weight change from days 1-4, 4-7 and 1-7.



Postmortem examinations (parental animals):
GROSS PATHOLOGY: Yes, see below
WEIGHING OF ORGANS: Yes, see below
HISTOPATHOLOGY: Yes, see below

Terminal sacrifice
- all males and toxicity subgr. females: Killed in Week 6, after completion of the Treatment Week 5 investigations.
- reproductive subgr. females & offspring: Killed on Day 7 post partum.

Gross pathology:
- adult/parental animals: Full macroscopic examination with tissue collection.

Organ Weights:
- main phase and tox. subgr. adults: Adrenals, brain, epididymides, heart, kidneys, liver, ovaries, pituitary, prostate, seminal
vesicles & coagulation gland, spleen, testes, thymus, uterus with cervix & oviducts

Histopathology:
- reproductive subgroups Gross lesions from all adult animals from all dose groups were examined by light microscopy.
Postmortem examinations (offspring):
Pup survivors were killed on Day 7 post partum.
Full macroscopic examination of decedent and surviving pups including assessment of the presence of milk in the stomach, where possible.
(Missing or grossly autolysed or cannibalised pups could not be examined).
Statistics:
As detailed in Endpoint study record "7.5.1 Repeated dose toxicity: oral - Repeat dose tox combined_gavage_rat_HLS_GAH0213"
Reproductive indices:
- Pre-coital interval (pairing days until detection of mating)
- No. of animals mating (evidence of successful copulation, i.e. at least one copulation plug and a sperm positive vaginal smear)
- No. of animals achieving pregnancy
- Percentage mating (No. of animals mating/No. of animals paired) x 100
- Fertility index (No. of animals achieving pregnancy/ No. or animals paired) x 100
- Conception rate (No. of animals achieving pregnancy/No. of animals mated) x 100
- Gestation length (time elapsing between detection of mating and commencement of parturition)
- No. of living pregnant females
- For further reproductive parameters, see also the above section "Litter observations" and section "Offspring viability indices" below.
Offspring viability indices:
- Post-implantation survival index  (Total no. of pups born/Total no. of uterine implantation sites) x 100
- Live birth index (No. of live pups on day 1 after littering/Total no. of pups born) x 100
- Viability index (No. of live pups on day 4 after littering /No. of live pups on day 1 after littering) x 100
- Lactation index (No. of live pups on day 7 after littering /No. of live pups on day 1 after littering) x 100
- For further parameters indicative of the viability of the offspring, see also the above section "Litter observations"
Clinical signs:
no effects observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Description (incidence and severity):
in toxicity phase females and parental males. In parental females only gross lesions were histopathologically examined.
Other effects:
no effects observed
Reproductive function: estrous cycle:
no effects observed
Reproductive function: sperm measures:
no effects observed
Reproductive performance:
no effects observed
Dose descriptor:
NOAEL
Effect level:
>= 1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: NOAEL = highest dose tested.
Clinical signs:
no effects observed
Mortality / viability:
no mortality observed
Body weight and weight changes:
no effects observed
Sexual maturation:
not specified
Organ weight findings including organ / body weight ratios:
not specified
Gross pathological findings:
no effects observed
Histopathological findings:
not examined
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
>= 1 000 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: NOAEL = highest dose tested. Offspring development up to Day 7 of age.
Reproductive effects observed:
not specified

As outlined in the „Validity Assessment Report“ for the read-across approach (see IUCLID Section 13) read-across from testing data obtained with the UVCB substance WS405777 is considered appropriate for the safety evaluation as well as classification and labelling of the mono-constituent substance WS406663 based on the close chemical similarity between the two substances.

Executive summary:

In this screening study, administration of the read-across source substance WS405777 to rats at dose levels of 100, 300 and 1000 mg/kg/day was well tolerated with no effects to parental animals and to reproductive endpoints. The NOAEL was derived at the highest dose, i.e. 1000 mg/kg/day.

As outlined in the „Validity Assessment Report“ for the read-across approach (see IUCLID Section 13) read-across from testing data obtained with the UVCB substance WS405777 is considered appropriate for the safety evaluation as well as classification and labelling of the mono-constituent substance WS406663 based on the close chemical similarity between the two substances.

The NOAEL of 1000 mg/kg/day as derived with the read-across source substance WS405777 in this screening study also can be used for the read-across target substance WS406663 for the regulatory purposes of REACH.

Effect on fertility: via oral route
Endpoint conclusion:
no adverse effect observed
Study duration:
subacute
Species:
rat
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Additional information
Justification for selection of Effect on fertility via oral route:
Justification for use of the results obtained with the read-across source substance WS405777 for classification of WS406663 is provided in the “Validity Assessment Report for the read-across approach” attached in IUCLID Section 13.

Effects on developmental toxicity

Link to relevant study records
Reference
Endpoint:
developmental toxicity
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Read-across from GLP-compliant guideline study performed with similar substance.
Reason / purpose:
reference to same study
Reference:
Composition 0
Qualifier:
according to
Guideline:
other: OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test) of 1996
Deviations:
no
GLP compliance:
yes (incl. certificate)
Limit test:
no
Test material information:
Composition 1
Species:
rat
Strain:
Sprague-Dawley
Details on test animals and environmental conditions:
TEST ANIMALS
- Sprague Dawley rats, strain: Crl:CD(SD) with appropriate range of bodyweight at study start.
- Source: Charles River (UK) Ltd.
- Age at treatment start: 61-68 days.
- Weight at treatment start: Males: minimum 332 g, maximum 397 g,
Females: minimum 210 g, maximum 255 g.
- Housing Inside a barriered rodent facility:
all animals pre-pairing + toxicity subgroups: In groups up to 5 by sex in solid floor polycarbonate cages.
during pairing (1 male+1 female/cage): In RB3 modified polypropylene cages with stainless steel grid-floor over absorbent paper-lined trays.
males after pairing: In groups of up to 5 in solid floor polycarbonate cages.
females during gestation and lactation: Females housed individually (+litter) in solid floor polycarbonate cages.
- Bedding material (in solid floor cages): Wood based bedding.
- Cage enrichment: Aspen chew block + plastic shelter (except during pairing or post gestation day 20).
- Diet (ad libitum): Standard rodent diet (SDS VRF1 Certified) without antibiotic, chemotherapeutic or prophylactic agent.
- Water (ad libitum): Potable drinking water from the public supply.
- Acclimation period: 5 days before treatment start, under laboratory conditions.

Routine analysis of the batch of diet used and water, chew blocks and bedding material did not provide evidence of contamination that might have prejudiced the study.

IN-LIFE DATES:
- Duration of test, males & toxicity phase females: Five weeks
Duration of test, main phase females (i.e. reproductive subgroup): From 14 days prior to pairing to day 7 of lactation.
Duration of test, offspring: From birth to day 7 of lactation.

ENVIRONMENTAL CONDITIONS

Air conditioned room kept at positve pressure without re-circulation of the filtered fresh air supplied to the room.
Controlled environment, environmental conditions were set at:
- Temperature (°C): 21 ± 2°C
- Relative Humidity (%): 40 to 70%
- Photoperiod (artificial lighting): 12 hrs day / 12 hrs night
Deviations from the target ranges for temperature and relative humidity were not evident.
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
Treatment of parental animals by oral gavage administration. Test substance was not directly administered to F1 animals.

- Concentration in vehicle: 20, 60 and 200 mg/ml
- Amount (dose volume by gavage): 5 mL/kg bw/day.
Actual dose volumes were calculated at about weekly or shorter intervals accounting for the latest body weight.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
- Chemical analysis of test material formulations by high performance liquid chromatography coupled with mass spectrometry (HPLC-MS/MS).
- Concentrations (verified at first and last treatment week) of the test material formulations were confirmed at each dose level.
- Chemical analysis confirmed that the mean concentrations of WS405777 in prepared formulations were 100% to 113% of the corresponding
nominal concentration, thus confirming accuracy of formulation.
Details on mating procedure:
- Male/female ratio per cage: 1/1
- Length of cohabitation: At the most 14 days, until proof of pregnancy was confirmed. 
- Proof of successful mating: Formation of at least one copulation plug and a sperm positive vaginal smear.
The day this was found was referred to as day 0 of gestation.
(During cohabitation, females were checked every morning for pregnancy).

Duration of treatment / exposure:
- Treatment period, males & toxicity phase females: Daily, for five consecutive weeks, in males commencing 14 days prior to mating
- Treatment period, main phase females (i.e. reproductive subgroup): 43 to 46 days (from 14 days prior to pairing to day 7 of lactation)
- Offspring were not dosed
Frequency of treatment:
Daily, 7 days/week (during parturition, dosing omitted as appropriate)
Duration of test:
- Duration of test, all F0 males & toxicity subgroup females: Five weeks
Duration of test, reproductive subgroup females (F0): From 14 days prior to mating to day 7 of lactation.
Duration of test, offspring: From birth to day 7 of lactation.
Remarks:
Doses / Concentrations:
0 (vehicle control), 100, 300, 1000 mg/kg bw/day
Basis:
actual ingested
No. of animals per sex per dose:
Main phase animals (i.e. reproductive subgroups): 10 males / 10 females
Control animals:
yes, concurrent vehicle
Details on study design:
This study was conducted to examine both repeated dose toxicity and  reproductive/developmental toxicity as an OECD screening combined study
(OECD 422 test guideline).  Therefore, animals initially entering the study were divided into toxicity subgroup animals (toxicity phase) and reproductive subgroup animals (main phase), whereby 5 of the 10 F0 males (used for pairing) per dose group formed the toxicity male subgroups.

Dose selection was based on the results of a 7-day preliminary oral toxicity study in the rat in which dose levels of 100, 300 or 1000 mg/kg/day did not have any overt treatment-related effects on young adult animals (females nulliparous and non-pregnant). Accordingly the same dose levels were used in the present OECD 422 combined repeat dose toxicity and reprotoxic/develpmental toxicity screening study.
Maternal examinations:
Clinical observations performed and frequency:
- Clinical signs : At least twice a day (before and after administration)
- Detailed physical examination
and arena observations: Before treatment start and at least once a treatment week.
- Body weight: Weekly for pre-pairing period; on gestation days 0, 6, 13, 20; on lactation days 1 & 7.
- Food consumption: Weekly for pre-pairing period, during gestation for days 0-6, 6-13, 13-20, during lactation for days 1-4 & 4-7.
- Frequency of vaginal estrus: Daily by examination of vaginal smears taken from the beginning of the treatment period to the day of confirmed copulation.
(During the pairing period, females were checked every morning for pregnancy).

Additional parameters examined:
- No. of animals mating (evidence of successful copulation, i.e. sperm positive vaginal smear and at least one copulation plug)
- Pre-coital interval (pairing days until detection of mating)
- No. of animals achieving pregnancy
- No. of living pregnant females
- From Day 20 post copulation 3 times a day checks for evidence of parturition, any difficulties and numbers of live and dead offspring.
- Gross pathology
- Organ weights
adrenals, brain, heart, kidneys, liver,ovaries, pituitary, spleen, thymus, uterus with cervix & oviducts.
- Histopathology (gross lesions).





Ovaries and uterine content:
The ovaries and uterine content were macroscopically examined after termination on day 7 of lactation, i.e. day 7 post partum.
Numbers of uterine implantation sites were recorded and post implantation survival determined.
In addition, gestation length (time elapsing between detection of mating and commencement of parturition) was recorded.
Fetal examinations:
Litters were examined post partum as follows (day of birth = day 0):
- Number:  Daily until day 7 post partum.
- Sex: In total litter: 1st day; in live litter on days 1, 4 and 7
- Live births/mortality: Daily until day 7 post partum.
- Clinical signs: Daily until day 7 post partum
- Body weight of live pups: 1st, 4th, 7th day and weight change from days 1-4, 4-7 and 1-7.
- Necropsy:  7th day full macroscopic examination of all pups including assessment of the presence of milk in the stomach, where possible.
(Missing or grossly autolysed or cannibalised pups could not be examined)
Statistics:
As detailed in Endpoint study record "7.5.1 Repeated dose toxicity: oral - Repeat dose tox combined_gavage_rat_HLS_GAH0213"
Indices:
- Percentage mating (No. of animals mating/No. of animals paired) x 100
- Conception rate (No. of animals achieving pregnancy/No. of animals mated) x 100
- Fertility index (No. of animals achieving pregnancy/ No. of animals paired) x 100
- Gestation index (No. of live litters born on day 0/No. of living pregnant females) x 100
- Post-implantation survival index  (Total no. of pups born/Total no. of uterine implantation sites) x 100
- Live birth index (No. of live pups on day 1 after littering/Total no. of pups born) x 100
- Sex ratio expressed as percentage males and calculated for total offspring on Day 1 and for live offspring on Days 1, 4 and 7
(No. of male pups in litter/No. of offspring in litter) x 100
- Viability index (No. of live pups on day 7 after littering /No. of live pups on day 1 after littering) x 100
Historical control data:
Not included in the study report.
Details on maternal toxic effects:
Maternal toxic effects:no effects
Dose descriptor:
NOAEL
Effect level:
>= 1 000 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: maternal toxicity
Dose descriptor:
NOAEL
Effect level:
>= 1 000 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: developmental toxicity
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects
Abnormalities:
not specified
Developmental effects observed:
not specified

As outlined in the „Validity Assessment Report“ for the read-across approach (see IUCLID Section 13) read-across from testing data obtained with the UVCB substance WS405777 is considered appropriate for the safety evaluation as well as classification and labelling of the mono-constituent substance WS406663 based on the close chemical similarity between the two substances.

Executive summary:

In this screening study, administration of the read-across substance WS405777 to rats at dose levels of 100, 300 and 1000 mg/kg/day was well tolerated with no effects to parental animals, to reproductive endpoints and to offspring. The NOAEL was derived at the highest dose, i.e. 1000 mg/kg/day.

As outlined in the „Validity Assessment Report“ for the read-across approach (see IUCLID Section 13) read-across from testing data obtained with the UVCB substance WS405777 is considered appropriate for the safety evaluation as well as classification and labelling of the mono-constituent substance WS406663 based on the close chemical similarity between the two substances.

The NOAEL of 1000 mg/kg/day as derived with the read-across source substance WS405777 in this sscreening study also can be used for the read-across target substance WS406663 for the regulatory purposes of REACH.

Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Study duration:
subacute
Species:
rat
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
Justification for selection of Effect on developmental toxicity: via oral route:
Justification for use of the results obtained with the read-across source substance WS405777 for classification of WS406663 is provided in the “Validity Assessment Report for the read-across approach” attached in IUCLID Section 13.

Justification for classification or non-classification

There were no findings or adverse effects in the Combined Repeated Dose Oral (Gavage) Toxicity Study with the Reproduction/Developmental Toxicity Screening Test (OECD 422) warranting the classification of the read-across source substance WS405777 regarding reproductive or developmental toxicity according to European classification rules [REGULATION (EC) 1272/2008].

The same conclusion of non-classification regarding reproductive or developmental toxicity can be drawn for the read-across target substance WS406663 based on the close chemical similarity between the two substances (see IUCLID Section 13 „Validity Assessment Report“ for the read-across approach).