cis-2-(2,4-dichlorophenyl)-2-(1H-imidazol-1-ylmethyl)-1,3-dioxolane-4-ylmethyl methanesulphonate monohydrochloride

Endpoint summary

Description of key information

Acute toxicity: oral:

In an acute oral toxicity study in female Crl:WI (Han) (SPF) rats, following the acute toxic class method in accordance with the OECD Guideline 423 and EU Method B.1 tris, the LD50 was established to be within the range of 300 - 2000 mg eq/kg. According to the OECD 423 guideline, the LD50 cut-off value was considered to be 500 mg eq/kg body weight (Latour, 2015).

Acute toxicity: inhalation:

In addition to the oral route of exposure, for substances other than gases, the information mentioned under REACH section 8.5.2 to 8.5.3 shall be provided for at least one other exposure route (REACH Regulation, column 2 adaptation of Annex VIII). For T001202, a key study is available for the dermal route of exposure. Therefore, an acute inhalation toxicity study should not be performed.

Acute toxicity: dermal:

In an acute dermal toxicity study in male and female Crl:WI (Han) (SPF) rats, following the standard acute method according to OECD Guideline 402 and EC method B.3, the LD50 was established to exceed 2000 mg eq/kg body weight (Latour, 2016).

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records

Reference

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 2015-06-09 to 2015-07-03

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.1100 (Acute Oral Toxicity)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

acute toxic class method

Limit test:

no

Specific details on test material used for the study:

Test Material SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Janssen Pharmaceutica N.V., I14KB4717
- Expiration date of the lot/batch: 25 November 2016 (retest date)
- Purity correction factor: 1.09

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature
- Solubility and stability of the test substance in the solvent/vehicle: Analysis of test item in vehicle for
concentration, stability, homogeneity was not performed, however preparation was performed with
approved procedure and documented in detail. Homogeneity was visually inspected prior to use.

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: Test preparation material was kept at room temperature no more than 4 hours before animals were dosed. Preparations were stirred on magnetic stirrer during dosing.

Species:

rat

Strain:

Wistar

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Wistar strain Crl:WI (Han) (outbred, SPF-Quality); Charles River Deutschland, Sulzfeld, Germany
- Age at study initiation: approximately 8 weeks old
- Weight at study initiation (day 1): 149 - 173 grams
- Fasting period before study: animals were deprived of food overnight prior to dosing and until 3-4 hours after administration of the test item. Water was available ad libitum.
- Housing: group housing of 3 animals per cage in labeled Makrolon cages (MIV type; height 18 cm.) containing sterilized sawdust as bedding material (Lignocel S 8-15, JRS - J.Rettenmaier & Söhne GmbH + CO. KG, Rosenberg, Germany) and paper as cage-enrichment (Enviro-dri, Wm. Lillico & Son (Wonham Mill Ltd), Surrey, United Kingdom).
- Diet (e.g. ad libitum): ad libitum, free access to pelleted rodent diet (SM R/M-Z from SSNIFF® Spezialdiäten GmbH, Soest, Germany).
- Water (e.g. ad libitum): ad libitum, free access to tap water.
- Acclimation period: at least 5 days before start of treatment under laboratory conditions.
- Diet, water, bedding and cage enrichment evaluation for contaminants and/or nutrients was performed according to facility standard procedures. There were no findings that could interfere with the study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18-24 °C
- Humidity (%): 40-70%
- Air changes (per hr): at least 10
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:

oral: gavage

Vehicle:

propylene glycol

Details on oral exposure:

VEHICLE
- Concentration in vehicle: 30 mg/ml and 200 mg/ml
- Amount of vehicle (if gavage): 10 ml/kg
- Justification for choice of vehicle: the vehicle was selected based on trial preparations performed at WIL Research Europe and on test item data supplied by the Sponsor. The vehicle was chosen from (in order of preference): water (Elix) (test item did not dissolve), 1% aq. carboxymethyl cellulose (test item did not dissolve), propylene glycol (spec.gravity 1.036), polyethylene glycol 400 (spec. gravity 1.125) (turbid solution) and corn oil (spec. gravity 0.92).

MAXIMUM DOSE VOLUME APPLIED: 10 ml/kg

DOSAGE PREPARATION (if unusual):
- The preparations (w/w) were kept at room temperature and were dosed within 4 hours after adding the vehicle to the test item. Homogeneity was assessed by visual inspection of the solutions and the formulations were stirred during dosing, which ensures homogeneity sufficient for these kinds of studies.
- Adjustment was made for specific gravity of the vehicle. A correction was made for the purity of the test item.
- The concentration of the test item in vehicle was varied to allow constant dosage volume in terms of ml/kg body weight.

CLASS METHOD (if applicable)
- Rationale for the selection of the starting dose: the dose level was selected based on toxicity data (e.g. existing human and animal data, literature, substance data supplied by the Sponsor, analysis of structure activity relationships (SAR) and in vitro, ex-vivo and in vivo tests) of the test item (specified and approved by the Study Director in the study files).

Doses:

300 mg eq/kg bw and 2000 mg eq/kg bw

No. of animals per sex per dose:

3 females per dose

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days (until day 15)
- Frequency of observations and weighing:
Mortality/viability: twice daily. Animals showing pain, distress or discomfort, which was considered not transient in nature or was likely to become more severe, were sacrificed for humane reasons based on OECD guidance document on humane endpoints (ENV/JM/MONO/ 2000/7). The time of death was recorded as precisely as possible;
Body weights: days 1 (pre-administration), 8 and 15 and at death;
Clinical signs: at periodic intervals on the day of dosing (day 1) and once daily thereafter, until day 15. The signs were graded according to fixed scales and the time of onset, degree and duration were recorded: maximum grade 4: grading slight (1) to very severe (4); maximum grade 3: grading slight (1) to severe (3); maximum grade 1: presence is scored (1).
- Necropsy of survivors performed: yes, the moribund animals and animals surviving to the end of the observation period were sacrificed by oxygen/carbon dioxide procedure. All animals assigned to the study were subjected to necropsy and descriptions of all internal macroscopic abnormalities recorded.

Statistics:

No statistical analysis was performed (the method used is not intended to allow the calculation of a precise LD50 value).

Key result

Sex:

female

Dose descriptor:

LD50

Effect level:

300 - 2 000 other: mg eq/kg bw

Based on:

test mat.

Mortality:

At 300 mg eq/kg bw, no mortality occurred.
At 2000 mg eq/kg bw, two animals were found dead and one animal was sacrificed for humane reasons, on day 1.

Clinical signs:

At 300 mg eq/kg bw, lethargy, hunched posture, uncoordinated movements, piloerection and/or ptosis were noted for the animals on days 1 and/or 2.
At 2000 mg eq/kg bw, lethargy, flat- and hunched posture, uncoordinated movements, slow breathing, rales, shallow respiration, piloerection, watery discharge from the eyes, hypothermia and/or ptosis were noted for the animals on day 1.

Body weight:

The body weight gain shown by the animals over the study period was considered to be similar to that expected for normal untreated animals of the same age and strain.

Gross pathology:

No abnormalities were found at macroscopic post mortem examination of all animals.

Interpretation of results:

Category 4 based on GHS criteria

Conclusions:

The oral LD50 value of T001202 in Wistar rats was established to be within the range of 300-2000 mg eq/kg body weight.
According to the OECD 423 test guideline, the LD50 cut-off value was considered to be 500 mg eq/kg body weight.

Based on these results:
- according to the Globally Harmonized System of Classification and Labelling of Chemicals (GHS) of the United Nations (2011) (including all amendments), T001202 should be classified as: harmful if swallowed (Category 4);
- according to the Regulation (EC) No 1272/2008 on classification, labelling and packaging of substances and mixtures (including all amendments), T001202 should be classified as Category 4 and should be labeled as H302: Harmful if swallowed.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

LD50

Value:

500 mg/kg bw

Acute toxicity: via inhalation route

Link to relevant study records

Reference

Endpoint:

acute toxicity: inhalation

Data waiving:

other justification

Justification for data waiving:

other:

Endpoint conclusion

Endpoint conclusion:

no study available

Acute toxicity: via dermal route

Link to relevant study records

Reference

Endpoint:

acute toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 2016-01-07 to 2016-01-21

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 402 (Acute Dermal Toxicity)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.3 (Acute Toxicity (Dermal))

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.1200 (Acute Dermal Toxicity)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

standard acute method

Limit test:

yes

Specific details on test material used for the study:

Test Material SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Janssen Pharmaceutica N.V., I14KB4717
- Expiration date of the lot/batch: 25 November 2016 (retest date)
- Purity correction factor: 1.09

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature
- Solubility and stability of the test substance in the solvent/vehicle: Analysis of test item in vehicle for
concentration, stability, homogeneity was not performed, however preparation was performed with
approved procedure and documented in detail. Homogeneity was visually inspected prior to use.

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: Test preparation material was kept at room temperature no more than 4 hours before animals were dosed. Preparations were stirred on magnetic stirrer during dosing.

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Wistar strain Crl:WI (Han) (outbred, SPF-Quality); Charles River Deutschland, Sulzfeld, Germany
- Age at study initiation: Young adult animals (approx. 10 weeks old)
- Weight at study initiation: 181 - 303 grams
- Housing: group housed in Makrolon cages (MIV type, height 18 cm) during acclimatization period; Individually housed in labeled Makrolon cages (MIII type, height 18 cm.) containing sterilized sawdust as bedding material (Lignocel S 8-15, JRS - J.Rettenmaier & Söhne GmbH + CO. KG, Rosenberg, Germany) and paper as cage-enrichment (Enviro-dri, Wm. Lillico & Son (Wonham Mill Ltd), Surrey, United Kingdom).
- Diet (e.g. ad libitum): ad libitum, free access to pelleted rodent diet.
- Water (e.g. ad libitum): ad libitum, free access to tap water
- Acclimation period: at least 5 days before start of treatment under laboratory conditions

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18-24 °C
- Humidity (%): 40-70%
- Air changes (per hr): at least 10
- Photoperiod (hrs dark / hrs light): 12/12

Type of coverage:

not specified

Vehicle:

propylene glycol

Details on dermal exposure:

TEST SITE
- Area of exposure: approx. 10% of the total body surface, i.e. approx. 25 cm² for males and 18 cm² for females
- Type of wrap if used: surgical gauze patch (Surgy 1D), successively covered with aluminum foil and Coban elastic bandage. A piece of Micropore tape was additionally used for fixation of the bandages in females only.

REMOVAL OF TEST SUBSTANCE
- Washing (if done): after removal of dressing, the skin was cleaned of residual test item using tap water
- Time after start of exposure: 24 h

Duration of exposure:

24 h

Doses:

2000 mg eq/kg (single dosing)

No. of animals per sex per dose:

5

Control animals:

no

Details on study design:

Preparation of test item:
The preparation (w/w) was kept at room temperature and dosed within 4 hours after adding the vehicle to the test item. Homogeneity was assessed by visual inspection of the solutions and the formulations were stirred during dosing, which ensures homogeneity sufficient for these kinds of studies.
Adjustment was made for specific gravity of the vehicle. A factor of 1.09 was used to correct for the purity/composition of the test item.

Treatment of animals and application of test item:
Method: Dermal application. Test preparation was stirred on a magnetic stirrer during application.
Clipping: One day before exposure (Day -1) an area of approximately 5x7 cm on the back of each animal was clipped.

Frequency of dosing: Single dosage, on Day 1.

Observations:
Observation period: until day 15 after treatment
- Mortality/Viability: Twice daily.
- Body weights: Days 1 (pre-administration), 8 and 15.
- Clinical signs: At periodic intervals on the day of dosing (Day 1) and once daily thereafter, until Day 15. The time of onset, degree and duration were recorded and the symptoms graded according to fixed scales:
Maximum grade 4: grading slight (1) to very severe (4)
Maximum grade 3: grading slight (1) to severe (3)
Maximum grade 1: presence is scored (1).
- Necropsy: At the end of the observation period, all animals were sacrificed by oxygen/carbon dioxide procedure and subjected to necropsy. Descriptions of all internal macroscopic abnormalities were recorded.

Statistics:

No statistical analysis was performed (the method used is not intended to allow the calculation of a precise LD50 value).

Key result

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 2 000 other: mg eq/kg bw

Based on:

test mat.

Mortality:

No mortality occurred.

Clinical signs:

Flat posture, ptosis, piloerection and/or chromodacryorrhoea were noted for the animals on Days 1 and/or 2. White discolouration, erythema maculate, scales and/or scabs were seen in the treated skin-area or left flank of the animals during the observation period. These local effects were considered not to have affected the conclusion of the study.

Body weight:

The changes noted in body weight gain in males and females were within the range expected for rats used in this type of study and were therefore considered not indicative of toxicity.

Gross pathology:

Abnormalities of the lungs (dark red discolouration) were noted for two male animals at macroscopic post mortem examination. No abnormalities were found at macroscopic examination of the other animals.

Interpretation of results:

GHS criteria not met

Conclusions:

The dermal LD50 value of T001202 in Wistar rats was established to exceed 2000 mg eq/kg body weight. Based on these results, the substance does not have to be classified and has no obligatory labelling requirement for acute dermal toxicity according to the Globally Harmonized System of Classification and Labelling of Chemicals (GHS) of the United Nations (2015) (including all amendments) and Regulation (EC) No 1272/2008 on classification, labelling and packaging of substances and mixtures (including all amendments).

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

2 000 mg/kg bw

Additional information

• Acute oral toxicity:

An acute oral toxicity study with T001202, according to the acute toxic class method (OECD guideline 423 and EU Method B.1 tris) in female Crl:WI(Han) (SPF) rats was performed (Latour, 2015). Initially, the substance was administered to three female Wistar rats at 300 mg eq/kg body weight. In a stepwise procedure two addition groups of three females were dosed at 2000 and 300 mg eq/kg body weight. The vehicle was propylene glycol. The rats received a single oral dose of test item, and were observed during 14 days following administration. At 300 mg eq/kg bw, no mortality occurred. At 2000 mg eq/kg bw, two animals were found dead and one animal was sacrificed for humane reasons, on day 1. At 300 mg eq/kg bw, lethargy, hunched posture, uncoordinated movements, piloerection and/or ptosis were noted for the animals on days 1 and/or 2. Animals dosed at 2000 mg eq/kg bw, were observed to show signs of lethargy, flat- and hunched posture, uncoordinated movements, slow breathing, rales, shallow respiration, piloerection, watery discharge from the eyes, hypothermia and/or ptosis on day 1only. The body weight gain shown by the animals over the study period was considered to be similar to that expected for normal untreated animals of the same age and strain. No abnormalities were found at macroscopic post mortem examination of all animals.

The oral LD50 value of T001202 was established to be within the range of 300-2000 mg eq/kg body weight. According to the OECD 423 test guideline, the LD50 cut-off value was considered to be 500 mg eq/kg body weight.

This reliable guideline study is assigned as key study.

In addition, an acute oral LD50 value in rats of 226.3 (173.1 - 295.7) mg/kg body weight was established in a K4 supporting study (Schellekens, 2004). Although the LD50 value established in this study is lower than the LD50 value determined in the K1 study of Latour (2015), the result of the K4 study is not used for classification nor for risk assessment as it is not clear at which doses lower than 640 and 320 mg/kg body weight the substance was tested.

• Acute inhalation toxicity:

In addition to the oral route of exposure, for substances other than gases, the information mentioned under REACH section 8.5.2 to 8.5.3 shall be provided for at least one other exposure route (REACH Regulation, column 2 adaptation of Annex VIII). For this substance a key study is available for the dermal route of exposure. Therefore, an acute inhalation toxicity study should not be performed.

• Acute dermal Toxicity:

An acute dermal toxicity study with T001202, according to OECD guideline 402 and EU Method B.3, in male and female Crl:WI (Han) (SPF) rats was performed (Latour, 2016). The vehicle was propylene glycol and the test item formulation was applied on a clipped area on the back at a dose of 2000 mg/kg bw. After 24 hours of exposure, the test item was washed-off using tap water. The animals were observed over 14 days.

During the study no mortality occurred. Clinical signs observed were: flat posture, ptosis, piloerection and/or chromodacryorrhoea were noted for the animals on Days 1 and/or 2. White discolouration, erythema maculate, scales and/or scabs were seen in the treated skin area or left flank of the animals during the observation period. These local effects were considered not to have affected the conclusion of the study. The changes noted in body weight gain in males and females were within the range expected for rats used in this type of study and were therefore considered not indicative of toxicity. Abnormalities of the lungs (dark red discolouration) were noted for two male animals at macroscopic post mortem examination. No abnormalities were found at macroscopic examination of the other animals. The dermal LD50 value of T001202 in Wistar rats was established to exceed 2000 mg/kg body weight.

Justification for classification or non-classification

Based on the results showing an LD50 between 300 - 2000 mg/kg eq bw and according to the criteria laid down in Regulation (EC) No 1272/2008 on classification, labelling and packaging of substances and mixtures, T001202 should be classified as acute oral toxic category 4 and should be labelled as H302: Harmful if swallowed.

No data were available to decide on the classification for the inhalation route.

Based on the dermal LD50 exceeding 2000 mg/kg bw, T001202 does not have to be classified and has no obligatory labelling requirement for acute dermal toxicity according to the criteria laid down in Regulation (EC) No 1272/2008.