2-Propenamide, N-[5-[[4-[4-[(dimethylamino)methyl]-3-phenyl-1H-pyrazol-1-yl]-2-pyrimidinyl]amino]-4-methoxy-2-(4-morpholinyl)phenyl]

Basic toxicokinetics

Administrative data

Endpoint:

basic toxicokinetics in vivo

Remarks:

Short term (7 days) repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2016-06-16 to 2016-06-29

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

test procedure in accordance with national standard methods with acceptable restrictions

Justification for type of information:

Considered bridging study - reference section 13

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2016

Report date:

2016

Materials and methods

Objective of study:

absorption

other: accumulation

Test guideline

Qualifier:

no guideline followed

Principles of method if other than guideline:

The objective of study was to investigate the pharmacokinetics of YH25448 following repeated oral administrations of free form and mesylate salt of YH25448 to rats.

GLP compliance:

not specified

Test material

Test material information

Constituent 1

Reference substance name:

N-{5-[(4-{4-[(dimethylamino)methyl]-3-phenyl-1H-pyrazol-1-yl}pyrimidin-2-yl)amino]-4-methoxy-2-(morpholin-4-yl)phenyl}prop-2-enamide

EC Number:

841-500-6

Cas Number:

1903008-80-9

Molecular formula:

C30H34N8O3

IUPAC Name:

N-{5-[(4-{4-[(dimethylamino)methyl]-3-phenyl-1H-pyrazol-1-yl}pyrimidin-2-yl)amino]-4-methoxy-2-(morpholin-4-yl)phenyl}prop-2-enamide

Test material form:

solid: particulate/powder

Reference

Reference 1

Constituent 1

Reference substance name:

N-{5-[(4-{4-[(dimethylamino)methyl]-3-phenyl-1H-pyrazol-1-yl}pyrimidin-2-yl)amino]-4-methoxy-2-(morpholin-4-yl)phenyl}prop-2-enamide

EC Number:

841-500-6

Cas Number:

1903008-80-9

Molecular formula:

C30H34N8O3

IUPAC Name:

N-{5-[(4-{4-[(dimethylamino)methyl]-3-phenyl-1H-pyrazol-1-yl}pyrimidin-2-yl)amino]-4-methoxy-2-(morpholin-4-yl)phenyl}prop-2-enamide

Test material form:

solid: particulate/powder

Reference 2

Constituent 1

Test material form:

solid: particulate/powder

Specific details on test material used for the study:

*Test article 1
SOURCE OF TEST MATERIAL
- YH25448 (T003925, Lazertinib free base)
- lot/batch number of test material: 5138-117
- Appearance: Light yellow solid
- Purity: 98.7%

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: at 4°C
- Stability under storage conditions: not indicated
- Stability under test conditions: not indicated
- Solubility and stability of the test substance in the solvent/dispersant/vehicle/test medium: not indicated

FORM AS APPLIED IN THE TEST: liquid


*Test article 2
SOURCE OF TEST MATERIAL
- YH25448A (YH25448, Lazertinib mesylate )
- lot/batch number of test material: 5138-156
- Appearance: Off -white solid
- Purity: 99.3%

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: at 4°C
- Stability under storage conditions: not indicated
- Stability under test conditions: not indicated
- Solubility and stability of the test substance in the solvent/dispersant/vehicle/test medium: not indicated

FORM AS APPLIED IN THE TEST: liquid

Radiolabelling:

no

Test animals

Species:

rat

Strain:

Sprague-Dawley

Remarks:

Crl:CD (SD)

Details on species / strain selection:

Rationale for species selection: SD rats were commonly used species in pre-clinical studies with large historical scientific data and easy to handle.

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: 34 male Crl:CD (SD) rats from Orient Bio Inc.
- Age at study initiation: 7 weeks
- Weight at study initiation: 0.260 - 0.280 kg
- Fasting period before study: not indicated
- Housing: Three or four rats per cage were housed in polycarbonate cages (240 W x 400 L x 180 H mm) for acclimation.

- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 5 days

DETAILS OF FOOD AND WATER QUALITY: not indicated

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23±2°C
- Humidity (%): 40 - 60 %
- Air changes (per hr): 10 times/hr
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 2016-06-17. To: 2016-06-28

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

other: 50 mM citric acid/0.5% methyl cellulose and 0.5% methyl cellulose

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
- To prepare the oral dosing formulation, appropriate amounts of test item were weighed. The test items were dissolved in vehicle on the day of dosing to achieve the final concentrations.
Group 1: YH25448 (test item), 50 mg/kg (dose), 10 mL/kg (dose volume), 1.50 g (Weighed amount), 300 mL (Volume of vehicle added), 5 mg/mL (Final concentration)
Group 2: YH25448A (test item), 50 mg/kg (dose), 10 mL/kg (dose volume), 1.76 g (Weighed amount), 300 mL (Volume of vehicle added), 5 mg/mL* (Final concentration)
Group 3: YH25448A (test item), 50 mg/kg (dose), 10 mL/kg (dose volume), 1.76 g (Weighed amount), 300 mL (Volume of vehicle added), 5 mg/mL* (Final concentration)
*: corrected as free form with salt factor

VEHICLE
Citric acid/Methyl cellulose:
- Justification for use and choice of vehicle (if other than water): not indicated
- Concentration in vehicle: 50 mM (citric acid), 0.5% (methyl cellulose)
- Amount of vehicle (if gavage): 300 mL
- Lot/batch no. (if required): 104K0117
- Purity: not indicated

Methyl cellulose:
- Justification for use and choice of vehicle (if other than water): not indicated
- Concentration in vehicle: 0.5 %
- Amount of vehicle (if gavage): 300 mL
- Lot/batch no. (if required): SLBJ8516V
- Purity: not indicated

Vehicle preparation:
1) 50 mM citric acid/0.5 % methyl cellulose
The vehicle of 50 mM citric acid/0.5 % methyl cellulose was made by following procedure.
- 5 g of methyl cellulose and 10.51 g of citric acid were added to 1000 mL of bottle.
- 750 mL of distilled water was added to the bottle and the mixture was stirred overnight to dissolve completely.
- Distilled water was added up to 1000 mL of final volume.

2) 0.5 % methyl cellulose
The vehicle of 0.5 % methyl cellulose was made by following procedure.
- 5 g of methyl cellulose was added to 1000 mL of bottle.
- 750 mL of distilled water was added to the bottle and the mixture was stirred overnight to dissolve
completely.
- Distilled water was added up to 1000 mL of final volume.

Duration and frequency of treatment / exposure:

7 days, daily

Doses / concentrations

Dose / conc.:

50 mg/kg bw/day (nominal)

Remarks:

Group 1, 2 and 3

No. of animals per sex per dose / concentration:

6 males/group (3 groups), the animals in each group were divided into 2 subgroups again (n=3/time point). Dosing of 50 mg/kg in each group.

Control animals:

no

Positive control reference chemical:

yes, YH25448-d5 (Internal standard)

Details on study design:

- Dose selection rationale: The dose level was determined as 50 mg/kg for the study considering a dose level in 4-week repeated oral dose toxicity study in rats (COVANCE Study No.: 83337361)).
- Fasting period before blood sampling for clinical biochemistry: not indicated

Details on dosing and sampling:

TOXICOKINETIC / PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled: blood, plasma
- Time and frequency of sampling: Blood sampling were collected alternately from the 2 subgroups via jugular vein using 1 mL of disposable syringe at predose, 0.5, 1, 2, 4, 8, 12 and 24 hr post-dose on Day 1 and Day 7.
- Anaesthetic used for blood collection: no data
- Animals fasted: no data
- How many animals: 18
- Other: Plasma samples were obtained from blood samples after centrifugation at 13,000 rpm for 2 min and aliquoted 50 µL and residual plasma. All plasma samples were stored in deep freezer at approximately -70°C before analysis.


ANALYTICAL METHOD
- Plasma samples were quantitated following ‘Bioanalytical method validation for determination of YH25448 in mouse, rat, dog, monkey and human plasma by LC-MS/MS (RDTE152442))’. Using triple quadrupole mass spectrometer, YH25448 (reference standard, m/z 555.256 → 510.100) and YH25448-d5 (internal standard, m/z 560.377 → 515.200) were detected by MRM (multiple reaction monitoring) mode.
- Protein precipitation method was used for determination of YH25448 in plasma samples. 10 µL of internal standard solution (YH25448-d5, 1 µg/mL in 50% acetonitrile) and 500 µL of methanol for protein precipitation were added to 50 µL of plasma samples and vortexed for 1 min. The mixtures were centrifuged at 13,000 rpm for 10 min. 200 µL of deionized water was added to 200 µL of supernatants and vortexed for 10 sec. The mixtures were injected (2 µL) into LC-MS/MS system.

Statistics:

no statistics available

Results and discussion

Toxicokinetic / pharmacokinetic studies

Toxicokinetic parameters

Toxicokinetic parameters 1

Key result

Test no.:

#2

Toxicokinetic parameters:

AUC: 30504.8 ng·hr/mL (group 1), 27408.6 ng·hr/mL (group 2) and 25391.4 ng·hr/mL (group 3)

Toxicokinetic parameters 2

Key result

Test no.:

#2

Toxicokinetic parameters:

Cmax: 1873.2 ng/mL (group 1), 1741.7 ng/mL (group 2) and 1583.2 ng/mL (group 3)

Toxicokinetic parameters 3

Key result

Test no.:

#1

Toxicokinetic parameters:

AUC: 21180.2 ng·hr/mL (group 1), 13267.3 ng·hr/mL (group 2) and 19059.4 ng·hr/mL (group 3)

Toxicokinetic parameters 4

Key result

Test no.:

#1

Toxicokinetic parameters:

Tmax: 2.0 to 8.0 hr

Toxicokinetic parameters 5

Key result

Test no.:

#1

Toxicokinetic parameters:

Cmax: 1169.1 ng/mL (group 1), 1061.5 ng/mL (group 2) and 1170.9 ng/mL (group 3)

Any other information on results incl. tables

After single oral administrations of YH25448 or YH25448A to rats at 50 mg/kg, Cmax for YH25448 was generally similar among groups. Although AUClast for YH25448 in Group 2 (YH25448A in 50 mM citric acid/0.5 % methyl cellulose) was slightly low compared with other groups, the difference was not significant (less than 2 fold).
After repeated oral administrations of YH25448 or YH25448A to rats for 7 days at 50 mg/kg, mean plasma concentration-time profiles and the values of Cmax and AUClast for YH25448 were generally similar among groups.
Accumulation was observed after repeated administration for 7 days with accumulation ratio of 1.3 to 2.1.

Applicant's summary and conclusion

Conclusions:

In conclusion, pharmacokinetics of YH25448 was generally similar after single and repeated dose of YH25448 (free base) or YH25448A (mesylate salt) to rats and moderate accumulation was observed after repeated administration.