STI571 F8

Repeated dose toxicity: oral

Administrative data

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2005

Reliability:

1 (reliable without restriction)

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2005

Report date:

2005

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Test material form:

solid: particulate/powder

Details on test material:

Batch number: 022401

Test animals

Species:

rat

Strain:

Fischer 344

Details on species / strain selection:

Rats are recommended by the test guideline.
The strain chosen (F344) is commonly used in our laboratory for subacute toxicity studies.
Background data are therefore available, if necessary.

Sex:

male/female

Details on test animals or test system and environmental conditions:

Species, strain: Rats, Fischer, F344/NHsd, SPF.
Supplier: Harlan Winkelman GmbH, D-33176 Borchen.
Number of animals and sex: 30 males and 30 females. Females were nulliparous and non-pregnant.
All animals were within ±20 % of the mean weight per sex at the time of the allocation to the groups.
Age at delivery: About 6 weeks.
Age at time of the first administration: About 7 weeks.
Health status: The animals were supplied in an SPF-status and subjected to a health check prior to first dosing.
All animals were in a good state of health at this term.

Hygiene: Optimal hygienic conditions.
Room temperature: Average of 22.3 °C (continuous control and recording).
Relative humidity: Average of 56.1% (continuous control and recording).
Air exchange: 12 per hour.
Light :Artificial light from 6 a.m. to 6 p.m.
Cages: Single caging
Makrolon cages: Type III, high version (39 cm x 23 cm ground area, 18 cm height).
Bedding material: Aspen wood chips (ABEDD Dominik Mayr KEG, A-8580 Köflach), autoclaved. Bedding material was changed weekly. Acceptable contaminants are identical with the limits for contaminants in the feed.
Feed: Altromin 1324 forte, gamma irradiated with 25 kGy60Co, ad libitum. Exception: Feed was withdrawn on the day before blood sampling at about 5 p.m. and was offered again immediately
thereafter. Random samples of the feed are analysed for contaminants by Altromin GmbH, D-32791 Lage. One sample was additionally analysed by Agrar- und Umweltanalytik GmbH, D-07749 Jena. Acceptable limits of contaminants were taken from the animal feed regulation of the
Federal Republic of Germany.
Water: Tap water, acidified with HCl to pH ≥3, from an automatic watering system, ad libitum. Random samples of the water are analysed by the "Bundesanstalt für Lebensmitteluntersuchung
und –forschung", A-1090 Vienna, to check, if the water fulfils the requirements for drinking water for humans (exception: the pH).
Environmental enrichment: Nesting material and nibbling wood bricks (sized 10 cm x 2 cm x
2 cm), same material and source as the bedding material, were offered freshly once week.
Identification: By tattooing the individual animal number into the right pinna and cage tag.
Acclimatisation: 5 days.

Administration / exposure

Route of administration:

oral: gavage

Details on route of administration:

An oral administration was performed by stomach intubation using a metal gavage once a
day in the morning on 7 days per week.
The individual dose volumes were calculated using the last determined body weights.
The oral administration is a common route in toxicity testing of chemicals.

Vehicle:

other: Water. An aqueous solution of the test substance was used in the dosed groups.

Details on oral exposure:

Method of administration:
Stomach intubation.

Analytical verification of doses or concentrations:

yes

Remarks:

Determinations of the test substance in some selected preparations for stability and concentration were performed by HPLC.

Details on analytical verification of doses or concentrations:

System: Thermo Separation Instruments Thermoquest 1.
Software: Chrom Quest Chromatography Data System; version 4.2.
Column: Aquasil C18 5 μM 100*2.1 mm; SN0451685X, Lot. P5A02 Fa.Thermo Electron Corporation.
Column temperature: 30 °C.
Autosampler temperature: 20 °C.
Flow: 0.4 mL/min.
Mobile phase: 40 % methanol, 60 % 50 mM KH2PO4 + 2.3 mM KHSO4 + 5 mM heptane 1 -
sulfonic acid Na-salt.
Injection volume: 10 μL.
Detection: UV-absorption at 268 nm.

The following analytical method was developed to determine the concentration of the test
substance (as it is, in aqueous solutions) using high performance liquid chromatography.
The method was validated according to:
• "Validation of analytical procedures": Methodology ICH Q2B, December 1996 (as far as
applicable)
Calculation of linearity and variance homogeneity was performed according to:
• "Calibration of analytical methods": DIN 38402 part 51, May 1986
Sample Pre-treatment
All samples were diluted with water/methanol 60/40 (v/v), to obtain concentrations within the
calibration range
Test substance
Name: STI571 F8
Batch No.: 022401
Reference substance
The test substance "as it is" was used as reference substance.

Solutions of the test substance: Water/acetonitrile 60/40 (v/v).
HPLC equipment and conditions:
System: Thermo Separation Instruments Thermoquest 1, SCM1000 Vacuum Membrane Degasser, SpectraSYSTEM® Gradient pump P2000, SpectraSYSTEM® autosampler AS3000, SpectraSYSTEM® UV6000LP Photodiode Array Detector with 50mm flowcell
Software: ChromQuest Chromatography Data System version 4.2
Column: Aquasil C18, 5 μm 100*2.1 mm, SN 0451685X, Lot No. P5A02 Fa.Thermo Electron Corporation.
Column temperature: 30 °C
Autosampler temperature: 20 °C
Flow: 0.4 mL/min.
Injection volume: 10 μL
Detection: UV-absorption at 268 nm
Mobile phase: 60 % 50 mmol/L KH2PO4 + 2.2 mmol/L KHSO4 + 5 mmol/L, Heptane 1-sulfonic acid sodium salt, 40 % methanol LiChrosolv

Duration of treatment / exposure:

Test duration: 28 days

Frequency of treatment:

Dosing regime: 7 days/week

No. of animals per sex per dose:

Male: 5 animals at 0 mg/kg bw/day
Male: 5 animals at 20 mg/kg bw/day
Male: 5 animals at 65 mg/kg bw/day
Male: 5 animals at 200 mg/kg bw/day
Female: 5 animals at 0 mg/kg bw/day
Female: 5 animals at 20 mg/kg bw/day
Female: 5 animals at 65 mg/kg bw/day
Female: 5 animals at 200 mg/kg bw/day

Control animals:

yes, concurrent vehicle

Details on study design:

Test substance administration:
The test substance was administered as a freshly prepared aqueous solution, orally by
gavage to 3 groups of 5 male and 5 female F344 rats each, once a day for 28°consecutive
days (Days 1 to 28).
Doses used:
• Group A (low dose): 20 mg "STI571 F8"
per kg body weight and day,
• Group B (mid dose): 65 mg "STI571 F8"
per kg body weight and day,
• Group C (high dose): 200 mg "STI571 F8"
per kg body weight and day.
An equally sized negative control group (group K) received the vehicle for the test substance.
The dose volume was uniformly 10 mL per kg body weight.
In addition, two groups of 5 males and 5 females each, i.e. one high dose satellite group
(group CS) and one control satellite group (group KS), were treated in the same way as their
corresponding groups, but were kept for further 14 days without test substance
administration in an attempt to observe the reversibility or persistence of test substance
induced lesions.

Examinations

Observations and examinations performed and frequency:

Investigations:
• Animal observations: all animals, once a day (plus a daily check for viability).
• Detailed clinical observations: all animals, once a week.
• Functional observations: all animals, once, in the last week of the dosing period.
• Body weights: all animals, once a week.
• Feed consumption: all animals, for each week.
• Haematology: all animals of groups K, A, B and C, on Day 29 and all animals of groups KS and CS on Day 43.
• Clinical biochemistry: all animals of groups K, A, B and C on Day 29 and all animals of groups KS and CS on Day 43.
• Necropsy with gross pathological examination: all animals of groups K, A, B and C on Day 29 and all animals of groups KS and CS on Day 43.
• Organ weight determination: all animals at necropsy.
• Histopathological examination: all animals of groups K and C.

Sacrifice and pathology:

GROSS PATHOLOGY: Yes

HISTOPATHOLOGY: Yes

Statistics:

Analysis of variance followed by the Scheffé-test: all data with means and standard deviations determined, comparison of more than two groups
t-test: all data with means and standard deviations determined, for comparison of two groups only
H-test of Kruskal and Wallis followed by the test of Nemenyi: counted events with scoring or in cases where the requirements for the analysis of variance were not fulfilled
Chi2-test: counted events
Fisher's exact test counted: events, if the Chi2-Test was not applicable.

Results and discussion

Results of examinations

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Water consumption and compound intake (if drinking water study):

not specified

Haematological findings:

effects observed, non-treatment-related

Clinical biochemistry findings:

no effects observed

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Details on results:

Clinical observations:
Mortality: none in any group.

Observations in life, clinical and functional observations:
There were no test substance related effects.

Body weight gain: No relevant changes were observed,
compared to the control.

Feed consumption: No relevant changes were observed,
compared to the control.

Laboratory findings:
Haematology: No relevant changes were observed, compared to
the control.

Blood chemistry: No relevant changes were observed, compared
to the control.

Effects in organs:
Necropsy: no relevant findings.

Organ weights: no relevant findings.

Histopathology: no relevant findings.

Effect levels

1

Key result

Dose descriptor:

NOAEL

Effect level:

200 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

body weight and weight gain

gross pathology

histopathology: non-neoplastic

2

Dose descriptor:

NOEL

Effect level:

200 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

body weight and weight gain

clinical signs

gross pathology

haematology

histopathology: non-neoplastic

other: original NCD unit is mg/kg/day.

Target system / organ toxicity

Critical effects observed:

no

Applicant's summary and conclusion

Conclusions:

Classified as: Not classified

No target organ or specfic toxic effect could be elucidated.
According to EU-Directive 2001/59/EC, the application of "R48" is not considered as necessary for " STI571 F8", as no toxic effects were noted at doses of 200 mg/kg and below.