(isopropyl (1s,3s)-3-(methyl(7H-pyrrolo[2,3-d]pyrimidin-4-yl)amino)cyclobutane-1-carboxylate)

Skin irritation / corrosion

Administrative data

Endpoint:

skin corrosion: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 12 Dec 2018 to 21 Dec 2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Report date:

2019

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 431 (In Vitro Skin Corrosion: Reconstructed Human Epidermis (RHE) Test Method)

Version / remarks:

OECD Guideline 431. “In Vitro Skin Corrosion: reconstructed human epidermis (RHE) test method”, (adopted 29 July 2016).

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.40 (In Vitro Skin Corrosion: Transcutaneous Electrical Resistance Test (TER))

Version / remarks:

EC Guideline No. 440/2008. Part B: Methods for the Determination of Toxicity and
other health effects, Guideline B.40 BIS: "In Vitro Skin Corrosion: Human Skin Model Test". Official Journal of the European Union No. L142, 31 May 2008.

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

(isopropyl (1s,3s)-3-(methyl(7H-pyrrolo[2,3-d]pyrimidin-4-yl)amino)cyclobutane-1-carboxylate)

EC Number:

950-860-4

Molecular formula:

C15H20N4O2

IUPAC Name:

(isopropyl (1s,3s)-3-(methyl(7H-pyrrolo[2,3-d]pyrimidin-4-yl)amino)cyclobutane-1-carboxylate)

Test material form:

solid: bulk

Details on test material:

White Crystalline Solid

Specific details on test material used for the study:

Batch GR13419 ofPF-07094402 was a white solid with a purity of 100%.

In vitro test system

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Details on animal used as source of test system:

The model consists of normal, human-derived epidermal keratinocytes which have been cultured to form a multilayered, highly differentiated model of the human epidermis. It consists of organized basal, spinous and granular layers, and a multi-layered stratum corneum containing intercellular lamellar lipid layers arranged in patterns analogous to those found in vivo. The EpiDerm tissues (surface 0.6 cm2) were cultured on polycarbonate membranes of 10 mm cell culture inserts.

Justification for test system used:

Recommended test system in international guidelines (OECD and EC).

Vehicle:

unchanged (no vehicle)

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

The skin was moistened with 25 μL Milli-Q water (Millipore Corp., Bedford, Mass., USA) to ensure close contact of the test item to the tissue and 27.3 to 29.3 mg of the solid test item was added into the 6-well plates on top of the skin tissues.

Duration of treatment / exposure:

Two tissues were used for a 3-minute exposure to PF-07094402 and two for a 1-hour exposure.

Number of replicates:

2 each group

Results and discussion

In vitro

Results

Results 1

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

3-minute application

Value:

91

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Results 2

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

1 hour application

Value:

96

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

In conclusion, PF-07094402 is not corrosive in the in vitro skin corrosion test under the experimental conditions described in this report.

Executive summary:

PF-07094402 was checked for color interference in aqueous conditions and possible direct MTT reduction by adding the test item to MTT medium. Because the solutions did not turn blue / purple nor a blue / purple precipitate was observed it was concluded that the test item did not interfere with the MTT endpoint.

Skin corrosion is expressed as the remaining cell viability after exposure to the test item. The relative mean tissue viability obtained after the 3-minute and 1-hour treatments with PF-07094402 compared to the

negative control tissues was 91% and 96% respectively. Because the mean relative tissue viability for PF-07094402 was not below 50% after 3 minutes treatment and not below 15% after 1 hour treatment PF-07094402 is considered to be not corrosive. The absolute mean OD570 (optical density at 570 nm) of the negative control tissues was within the acceptance limits of OECD 431 (lower acceptance limit ³0.8 and upper acceptance limit <_2.8) and the laboratory historical control data range. The mean relative tissue viability following the 1-hour exposure to the positive control was 6.6%.

In the range of 20 - 100% viability the Coefficient of Variation between tissue replicates was <_ 6.7%, indicating that the test system functioned properly.