2-Pyridinecarboxamide, 4-(4-amino-3-fluorophenoxy)-N-methyl-

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

experimental study

Adequacy of study:

other information

Study period:

Nov 2009 to Mar 2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2011

Report date:

2011

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

GLP compliance:

no

Type of assay:

micronucleus assay

Test material

Test material information

Constituent 1

Reference substance name:

4-(4-amino-3-fluorophenoxy)-N-methylpyridine-2-carboxamide

EC Number:

692-721-8

Cas Number:

757251-39-1

Molecular formula:

C13 H12 F N3 O2

IUPAC Name:

4-(4-amino-3-fluorophenoxy)-N-methylpyridine-2-carboxamide

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

Vehicle composition: 0.7 mL Polyethylenglycol + 0.05 mL Ethanol abs. + 0.25 mL Solutol HS15

Details on exposure:

- Application volume: 5 mL/kg bw

- Rationale for the selection of the starting dose:
In order to select an appropriate dose range, male and female rats were treated in a pre-experiment with BAY 74-2329 and observed over a period of three days from start of treatment. After a single oral treatment of three male and female rats with 2000 mg/kg BAY 74-2329, all three female animals died or were sacrificed in moribund state on the day after treatment. Following single oral treatment of 2 male and 2 female rats with 1000 mg/kg, one male and one female animal died on the day after treatment. Daily treatment (i.g.) of 2 male and 2 female rats with 500 mg/kg BAY 74-2329 over three days resulted in some clinical signs (atactic gait) in animals of both sex but was tolerated without mortality.

Based on the findings of the pre-experiment, 500 mg/kg BAY 74-2329 was chosen as the highest dose in the bone marrow Micronucleus test. Additionally, 250 and 125 mg/kg were chosen as mid and low dose.

Duration of treatment / exposure:

3 days

Frequency of treatment:

once daily

Post exposure period:

none

Doses / concentrations

Remarks:

Doses / Concentrations:
125, 250, 500 mg/kg
Basis:
actual ingested

No. of animals per sex per dose:

6

Control animals:

yes, concurrent vehicle

Positive control(s):

Ethyl methanesulfonate (200 mg/kg bw)

Examinations

Tissues and cell types examined:

micronuclei in rat bone marrow

Details of tissue and slide preparation:

The preparation and microscopic analysis of micronuclei in rat bone marrow was performed according the pertinent SOP, which complies with internationally accepted guidelines and recommendations. Bone marrow was collected from all animals after sacrifice on study day 3. The femurs were dissected out from each animal and the bone marrow was flushed/ aspirated into fetal calf serum. The resulting cell suspensions were filtered through a cellulose column, centrifuged and smears were prepared from drops of the cell pellets which had been resuspended in a few drops of remained serum. The slides were air-dried and stained using May-Gruenwald and Giemsa solutions. The slides were coded and analyzed "blind" in random order. The stained smears were examined using oil immersion high power magnification in regions where cells were well spread and stained. The slides were examined for the incidence of micronucleated cells per 2000 polychromatic (PCE) and 1000 normochromatic (NCE) erythrocytes per animal. The ratio of polychromatic to normochromatic erythrocytes was calculated on the basis of 1000 NCE scored.

Statistics:

The statistical evaluation for was performed by Global Drug Discovery Statistics using the software SAS version 9.1.3 SP4. Each of the following variables were statistical analysed:

p1 proportion of micronucleated PCE (bone marrow micronucleus assay) or micronucleated RET (peripheral blood micronucleus assay)

p2 proportion of micronucleated NCE

p3 ratio of PCE / NCE (bone marrow micronucleus assay) or % RET (peripheral blood micronucleus assay)

For investigation of treatment differences the variables p1 and p2 were arc sin ( pi ) transformed; the analyses were conducted separately for each variable. One-sided t-tests were performed to assess the difference between negative and positive controls, the latter was then excluded from further analysis. Thereafter, in an analysis of variance it was investigated whether any treatment effect was present. If a treatment effect could be shown, contrasts were used for group comparisons between each dose group and the respective vehicle control. Contrasts were analyzed on the 5 % level.

Results and discussion

Test results

Sex:

male/female

Genotoxicity:

negative

Toxicity:

yes

Vehicle controls validity:

valid

Negative controls validity:

not examined

Positive controls validity:

valid

Any other information on results incl. tables

Following treatment with 500 mg/kg BAY 74-2329, one male animal died on the second treatment day and 5 (out of 11 remaining) animals of this dose group showed adverse clinical signs, demonstrating that 500 mg/kg is the maximum tolerated dose (MTD) under these treatment conditions.

 

Results obtained in the Micronucleus test showed that male and female rats treated with BAY 74-2329 showed at all 3 dose levels (125, 250 and 500 mg/kg body weight) neither a biologically relevant nor statistically significant increase (p < 0.05) in micronucleated PCE and NCE as compared to the vehicle control. BAY 74-2329 induced a statistically significant decrease (p < 0.05) in the ratio of PCE to NCE in male animals, indicating that the test item reached the target cells at cytotoxic concentration. The positive control compound EMS gave the expected statistically significant (p < 0.05) increase in the micronucleated PCE and NCE cell counts as compared to the vehicle control, thus demonstrating the ability of the test system to detect cytogenetic damage.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): negative

Executive summary:

Evaluation of the Micronucleus test data did not show any evidence for a mutagenic potential of BAY 74-2329 in male and female rats when administered intragastrically up to the toxic dose level of 500 mg/kg bw. It is therefore concluded that BAY 74-2329 is non-mutagenic in the rat bone

marrow Micronucleus test according to OECD TG 474.