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Administrative data

Endpoint:
in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From February 27, 2020 to September 1, 2020
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2020
Report date:
2020

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
GLP compliance:
yes
Type of assay:
mammalian erythrocyte micronucleus test

Test material

Constituent 1
Chemical structure
Reference substance name:
5-({4-chloro-6-[ethyl(phenyl)amino]-1,3,5-triazin-2-yl}amino)-4-hydroxy-3-[(1-sulfo-2-naphthyl)diazenyl]naphthalene-2,7-disulfonic acid, lithium sodium salts
EC Number:
942-803-7
Molecular formula:
Not applicable; this UVCB substance contains: C31H21ClN7O10S3.xLi.yNa, (x + y) = 3; 0 < (x,y) < 3 with 804.0 < MW < 852.1 g/mol (UVCB substance), C31H22N7O11S3.xLi.yNa, (x + y) = 3; 0 < (x,y) < 3 with 785.5 < MW < 833.7 g/mol (UVCB substnace), and traces of NaCl and Na2SO4.
IUPAC Name:
5-({4-chloro-6-[ethyl(phenyl)amino]-1,3,5-triazin-2-yl}amino)-4-hydroxy-3-[(1-sulfo-2-naphthyl)diazenyl]naphthalene-2,7-disulfonic acid, lithium sodium salts
Test material form:
solid: particulate/powder
Remarks:
migrated information: powder

Test animals

Species:
mouse
Strain:
ICR
Sex:
male/female
Details on test animals or test system and environmental conditions:
- Source: SPF (Beijing) biotechnology Co., Ltd.
- Age at study initiation: 51-61 days old
- Weight at study initiation: Males: 35.50-38.87 g; Females: 29.12-31.83 g
- Housing: The male mice were housed one per cage and the female mice were housed 3 or 5 per cage.
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 7 days
- Temperature (°C): 21.7-23.6 °C
- Humidity (%): 56-62 %
- Photoperiod: 12-hrs dark / 12-hrs light

Administration / exposure

Route of administration:
intravenous
Vehicle:
Ultra-pure water (CAS No.: 7732-18-5)
Frequency of treatment:
Mice were administered twice by tail vein injection, with an interval of approximately 24 hours.
Doses / concentrationsopen allclose all
Dose / conc.:
25 mg/kg bw/day (nominal)
Remarks:
for females
Dose / conc.:
50 mg/kg bw/day (nominal)
Remarks:
for males and females
Dose / conc.:
100 mg/kg bw/day (nominal)
Remarks:
for males and females
Dose / conc.:
200 mg/kg bw/day (nominal)
Remarks:
for males
No. of animals per sex per dose:
five males and five females
Control animals:
yes, concurrent vehicle
Positive control(s):
Cyclophosphamide monohydrate (CP, CAS No.: 6055-19-2)

Results and discussion

Test results
Sex:
male/female
Genotoxicity:
negative
Toxicity:
no effects
Vehicle controls validity:
valid
Positive controls validity:
valid

Any other information on results incl. tables

Table 1. Statistics results of the body weights in male and female mice

Sex

Group

Dose

(mg/kg.bw)

Animal number

First Dosing (g)

Mean±SD

Last Dosing (g)

Mean±SD

Sacrifice (g)

Mean±SD

Male

Negative control

0

1000-1004

37.09 ± 0.98

36.08 ± 0.40

36.78 ± 0.57

Treated Group

50

1100-1104

37.22 ± 0.91

36.31 ± 1.97

37.81*± 0.55

100

1200-1204

37.23 ± 0.83

37.02 ± 0.97

36.90 ± 0.95

200

1300-1304

37.30 ± 0.88

37.34 ± 0.36

37.01 ± 1.84

CP

50

1400-1404

37.31 ± 1.07

36.01 ± 0.93

36.36 ± 1.44

Female

Negative control

0

2000-2004

30.35 ± 0.79

29.52 ± 1.12

30.06 ± 1.39

Treated Group

25

2100-2104

30.47 ± 0.64

29.80 ± 0.71

28.75 ± 0.95

50

2200-2204

30.48 ± 0.69

29.48 ± 1.23

29.83 ± 1.10

100

2300-2304

30.48 ± 0.75

29.84 ± 0.70

29.59 ± 1.35

CP

50

2400-2404

30.52 ± 0.84

29.14 ± 1.25

29.41 ± 1.47

Note: Statistically significant difference compared to negative control (*: P<0.05).

 

Table 2. Statistics results of microscopic analysis in mice

Sex

Group

Animal number

Number of PCE

MNPCE/PCE (‰)

Mean±SD

PCE/RBC

Mean±SD

Male

Negative control

1000-1004

20000

1.7± 0.7

0.60 ± 0.05

50

1100-1104

20000

1.7 ± 0.6

0.56#± 0.03

100

1200-1204

20000

1.8 ± 0.8

0.57#± 0.04

200

1300-1304

20000

1.6 ± 0.7

0.57#± 0.10

CP (50)

1400-1404

20000

25.6**± 3.2

0.58 ± 0.05

Female

Negative control

2000-2004

20000

1.8 ± 0.6

0.60 ± 0.02

25

2100-2104

20000

1.6 ± 0.6

0.57#± 0.09

50

2200-2204

20000

1.6 ± 0.7

0.56#± 0.09

100

2300-2304

20000

1.7 ± 0.4

0.54#± 0.07

CP (50)

2400-2404

20000

25.0**± 3.5

0.55 ± 0.05

Note: Statistically significant difference compared to negative control (**: P<0.01).

#: the ratio was more than 20 percent of the ratio in the negative control group.

Applicant's summary and conclusion

Conclusions:
According to OECD 474 test method, CJ306 was negative effect under the condition of in vivo mammalian somatic cell-erythrocyte micronucleus test.
Executive summary:

This test using the procedures outlined in the SYRICI Study for G1980C0060 which is based on OECD 474 (OECD, 2016). The results of this OECD 474 test for CJ306 show that test validity criteria was met.

According to the results of the preliminary test, the maximum tolerated dose (MTD) for male mice were 200 mg/kg.bw/day and for female mice were 100 mg/kg.bw/day. Therefore, the three dose levels in female mice were 25, 50 and 100 mg/kg.bw/day, and the three dose levels in the male mice were 50, 100 and 200 mg/kg.bw/day. Comparing with the concurrent negative control group, the incidence of micronucleated PCE for the treated groups had no ststistically significant difference (P>0.05) and the incidence of micronucleated PCE for CP group had ststistically significant difference (P<0.01). Furthermoer, the ratios betweeen PCE and RBC in all treated groups were more than 20 percent of the ratio in the concurrent negative control group. Under the conditions of this study, the results of micronucleus test were negative. Therefore, CJ306 was not to induce the increase of the incidence of micronucleated PCE in mice.