N-((3R,6S)-6-methylpiperidin-3-yl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

31st January 2019 to 24th February 2020

Reliability:

1 (reliable without restriction)

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2020

Report date:

2020

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

1

Reference substance name:

Reference substance 001

Cas Number:

2374827-92-4

Test material form:

solid: particulate/powder

Method

Target gene:

Histidine locus in in several strains of Salmonella typhimurium (S. typhimurium; TA98, TA100, TA1535, and TA1537), and base-pair substitutions of Escherichia coli (E. coli) strain WP2uvrA

Species / strain

Species / strain 1

Species / strain / cell type:

E. coli WP2 uvr A

Species / strain 2

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Metabolic activation:

with and without

Metabolic activation system:

S9

Test concentrations with justification for top dose:

Selection of an adequate range of doses was based on a dose-range finding test with the strains TA100 and WP2uvrA, both with and without 5% (v/v) S9-mix. Eight concentrations, 1.7, 5.4, 17, 52, 164, 512, 1600 and 5000 µg/plate were tested in triplicate. The highest concentration of the test item used in the subsequent mutation assays was 5000 µg/plate.
Based on the results of the dose-range finding test, the following dose-range was selected for the first mutation experiment with the tester strains, TA1535, TA1537 and TA98 in the absence and presence of S9-mix: 52, 164, 512, 1600 and 5000 µg/plate.

Vehicle / solvent:

The vehicle of the test item was dimethyl sulfoxide (DMSO, Merck, Darmstadt, Germany).

Controls

Controls 1

Negative solvent / vehicle controls:

yes

Remarks:

Dimethyl sulfoxide

Positive controls:

yes

Remarks:

without metabolic action

Positive control substance:

4-nitroquinoline-N-oxide

2-nitrofluorene

sodium azide

methylmethanesulfonate

other: ICR-191

Controls 2

Negative solvent / vehicle controls:

yes

Remarks:

Dimethyl sulfoxide

Positive controls:

yes

Remarks:

with metabolic action

Positive control substance:

other:

Results and discussion

Test results

Test results 1

Key result

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Key result

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 3

Key result

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 4

Key result

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 5

Key result

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other:

Remarks:

PF-06715298 is not mutagenic in either the Salmonella typhimurium reverse mutation assay or the Escherichia coli reverse mutation assay.

Applicant's summary and conclusion

Conclusions:

Based on the results of this study it is concluded that PF-06715298 is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichiacoli reverse mutation assay.

Executive summary:

The objective of this study was to determine the potential of PF-06715298 and/or its metabolites to induce reverse mutations at the histidinelocus in several strains of Salmonella typhimurium (S. typhimurium; TA98, TA100, TA1535, and TA1537), and at the tryptophan locus of Escherichia coli (E. coli) strain WP2uvrA in the presence or absence of an exogenous mammalian metabolic activation system (S9).
The study procedures described in this report were based on the most recent OECD and EC guidelines.
Batch 902/6651600/C/21/1 of the test item was a white powder. A correction factor of 1.08 was used to correct for the purity. The vehicle of the test item was dimethyl sulfoxide. In the dose-range finding study, the test item was initially tested up to concentrations of 5000 µg/plate in the strains TA100 and WP2uvrA in the absence and presence of 5% (v/v) S9-mix. In the first mutation experiment, the test item was again tested up to concentrations of 5000 µg/plate in the strains TA1535, TA1537 and TA98 in the absence and presence of 5% (v/v) S9-mix. In a follow-up experiment of the assay with additional parameters, the test item
was tested at a concentration range of 492 to 5000 µg/plate in the absence and presence of 10% (v/v) S9-mix in the tester strains TA1535, TA1537, TA98, TA100 and WP2uvrA. In all three experiments the test item did not precipitate on the plates at this dose level. The bacterial background lawn was not reduced at any of the concentrations tested and no biologically relevant decrease in the number of revertants was observed.
The test item did not induce a significant dose-related increase in the number of revertant (His+) colonies in each of the four tester strains (TA1535, TA1537, TA98 and TA100) and in the number of revertant (Trp+) colonies in the tester strain WP2uvrA both in the absence and presence of S9-metabolic activation. These results were confirmed in a follow-up experiment.
The negative and strain-specific positive control values were within the laboratory historical control data ranges indicating that the test conditions were adequate and that the metabolic activation system functioned properly.
In conclusion, based on the results of this study it is concluded that PF-06715298 is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichiacoli reverse mutation assay.