4-(6-(3,6-diazabicyclo[3.1.1]heptan-3-yl)pyridin-3-yl)-6-(2-hydroxy-2-methylpropoxy)pyrazolo[1,5-a]pyridine-3-carbonitrile bis sulphate

Acute Toxicity: oral

Administrative data

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

20 August - 9 September 2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2019

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

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

GLP compliance:

not specified

Test type:

acute toxic class method

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

4-(6-{3,6-diazabicyclo[3.1.1]heptan-3-yl}pyridin-3-yl)-6-(2-hydroxy-2-methylpropoxy)pyrazolo[1,5-a]pyridine-3-carbonitrile

Molecular formula:

C22H24N6O2.[2]H2O4S

IUPAC Name:

4-(6-{3,6-diazabicyclo[3.1.1]heptan-3-yl}pyridin-3-yl)-6-(2-hydroxy-2-methylpropoxy)pyrazolo[1,5-a]pyridine-3-carbonitrile

Test material form:

solid: particulate/powder

Details on test material:

Batch (Lot) Number: 18-547.25-002
Physical Description: White powder
Storage Conditions: Kept in a room temperature area, protected from light

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

Justification for Test System and number of animals:
The Sprague Dawley rat was chosen as the animal model for this study as it is an accepted rodent species for preclinical toxicity testing by regulatory agencies. A total of 13 male and 13 female Sprague Dawley rats were received on 20 Aug 2019.

Six days were allowed between receipt of the animals and the start of treatment to accustom the animals to the laboratory environment. Each animal was uniquely identified using a subcutaneously implanted electronic identification chip.

Housing:
On arrival, animals were individually housed until randomization. Following, randomization, animals were socially housed in polycarbonate cages containing appropriate bedding equipped with an automatic watering valve.

Selection, Assignment, Replacement, and Disposition of Animals:
Prior to randomization procedures, the animals were weighed. Animals determined to be suitable as test subjects were assigned to groups by a stratified randomization scheme designed to achieve similar group mean body weights. Males and females were randomized separately.

Initial age: 8 weeks

Weight at the Initiation of Dosing: Males: 206 to 245 g.
Females: 149 to 167 g.

Environmental Conditions:
The animal room environment and photoperiod were controlled (targeted conditions: temperature 20C to 26C, humidity 30% to 70%, 12 hours light and 12 hours dark, except during designated periods). The overall average temperature and relative humidity ranges during the study were 21°C and 53% to 56%, respectively.

Food:
PMI Nutrition International Certified Rodent Chow No. 5CR4 was provided ad libitum throughout the study, except during designated procedures.
Maximum allowable concentrations of contaminants in the diet were controlled and routinely analyzed by the manufacturers.

Water:
Municipal tap water, purified by reverse osmosis and exposed to ultraviolet irradiation, was freely available. Periodic analysis of the water was subcontracted to management-authorized analytical laboratories. The analytical results are retained in the archives of CR-SPV .It was considered that there were no known contaminants in the dietary materials that could interfere with the objectives of the study.

Veterinary Care:
Veterinary care was available throughout the study; however, no veterinary examinations or medicinal treatments were administered during the study.

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

other: Reverse Osmosis Deionized (RODI) water

Details on oral exposure:

Administration of Test item:
On the day prior to dosing the animals chosen for use on the study were weighed and fasted. On Day 0, the test article was administered orally as a single dose (5, 50, 300, and 2000 mg/kg, 10 mL/kg) to rats (3/sex/group) using a syringe attached to a gavage cannula. Individual doses were calculated based on the animal's fasted (Day 0) body weight.

Doses:

5 mg/kg (group 1), 50 mg/kg (group 2), 300 mg/kg (group 3), 2,000 mg/kg (group 4)

No. of animals per sex per dose:

3/sex/group

Control animals:

no

Details on study design:

In-life Procedures, Observations, and Measurements
Mortality/Moribundity Checks:
The animals were observed for general health/mortality and moribundity at least twice daily, at least once in the morning and at least once in the afternoon, throughout the study.

Clinical observations:
Each animal was removed from the cage and observed in detail a minimum of 2 times on Day 0 (postdose), with the first observation within approximately 30 minutes after dosing, and daily thereafter (Days 1 to 14).

Body Weights:
Individual body weights were recorded for all animals prior to fasting, prior to dosing (Day 0), and on Days 7 and 14. Animals found dead were also weighed.

Terminal Procedures:
For the animals that died on study, a necropsy was conducted. The animals were refrigerated to minimize autolysis. Animals surviving until scheduled euthanasia were weighed, euthanized by carbon dioxide inhalation, and discarded without examination. Animals that died on study were subjected to a complete necropsy examination, which included evaluation of the carcass and musculoskeletal system; all external surfaces and orifices; cranial cavity and external surfaces of the brain; and thoracic, abdominal, and pelvic cavities with their associated organs and tissues. No tissues were retained.

Statistics:

Body weight means and standard deviations were calculated separately for males and females for each limit level administered.
The LD50 was estimated as indicated below:
< 50% mortality: LD50 was estimated as greater than the administered dose.
= 50% mortality: LD50 was estimated as equal to administered dose.
> 50% mortality: LD50 was estimated as less than the administered dose.

Results and discussion

Effect levels

Key result

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 300 - < 2 000 mg/kg bw

Based on:

test mat.

Mortality:

Mortality occurred in animals administered 2000 mg/kg. Following dose administration of 2000 mg/kg on Day 0, 1/3 males was found dead and 2/3 females were found dead; on Day 3, 1/3 males were found dead. The remaining male and female administered 2000 mg/kg and all animals administered ≤ 300 mg/kg survived to scheduled euthanasia.

Clinical signs:

other: Test article-related clinical observations in animals administered 2000 mg/kg included abnormal breathing, erected fur, partially closed eyes, and fur staining. No test articlerelated clinical observations were noted in animals administered ≤300 mg/kg. I

Gross pathology:

A gross necropsy examination was performed for all found dead animals that were administered 2000 mg/kg. There were no gross necropsy findings noted for the animals.

Applicant's summary and conclusion

Interpretation of results:

Category 4 based on GHS criteria

Conclusions:

The oral LD50 value of Intermediate 3532670 in Sprague-Dawley rats was established to be within the range of 300-2,000 mg/kg/bw.