4-[(3,5-Dichlorobenzoyl)amino]-3-hydroxybenzoic acid

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:

April to September 2014

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2015

Report date:

2015

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.7 (Repeated Dose (28 Days) Toxicity (Oral))

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

4-(3,5-dichlorobenzamido)-3-hydroxybenzoic acid

EC Number:

832-365-4

Cas Number:

1184581-58-5

Molecular formula:

C14H9Cl2NO4

IUPAC Name:

4-(3,5-dichlorobenzamido)-3-hydroxybenzoic acid

Test material form:

solid: particulate/powder

Details on test material:

Batch E010014831, re-test date: 30 June 2015

Specific details on test material used for the study:

Purity/composition correction factor (required) - No
Hygroscopic - No
Volatile - No
pH (1% in water, indicative range) - 5.3 – 5.0 (determined by WIL Research Europe B.V. )
Stability in vehicle:
• 1% Aq. Carboxymethyl cellulose - Unknown
Solubility in vehicle:
• 1% Aq. Carboxymethyl cellulose - Not indicated

Analysis of stability, homogeneity and concentration of the test substance under test conditions were not performed as part of this study.

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

Test system: Rat: Crl:WI(Han) (outbred, SPF-Quality).
Rationale: Recognized by international guidelines as the recommended test system (e.g. EPA, FDA, OECD and EC).
Source: Charles River Deutschland, Sulzfeld, Germany.
Total number of animals: 12 males and 12 females (females were nulliparous and non-pregnant).
Age at start of treatment: Approximately 9 weeks.
Identification: Tailmark (markerpen) and earmark.
Randomization: By computer-generated random algorithm according to body weight, with all animals within ± 20% of the sex mean.
Acclimatization period: At least 5 days before the start of treatment under laboratory conditions.
Health inspection Upon receipt of the animals.

Environmental controls for the animal room were set to maintain 18 to 24°C, a relative humidity of 40 to 70%, at least 10 air changes/hour, and a 12-hour light/12-hour dark cycle. Any variations to these conditions were maintained in the raw data and had no effect on the outcome of the study.
Accomodateion: Group housing of 3 animals per sex in Makrolon cages (MIV type, height 18 cm) with sterilized sawdust as bedding material (Lignocel S 8-15, JRS - J.Rettenmaier &Söhne GmbH + CO. KG, Rosenberg, Germany) and paper as cage-enrichment (Enviro-dri, Wm. Lilico & Son (Wonham Mill Ltd), Surrey, United Kingdom).

Administration / exposure

Route of administration:

oral: gavage

Details on route of administration:

Oral gavage, using a plastic feeding tube.
Formulations were placed on a magnetic stirrer during dosing.

Vehicle:

CMC (carboxymethyl cellulose)

Details on oral exposure:

10 mL/kg body weight. Actual dose volumes were calculated according to body weights determined on Days 1 and 4.
Once daily for up to 7 consecutive days, approximately the same time each day with a maximum of 6 hours difference between the earliest and latest dose. Animals were dosed up to the day prior to necropsy.

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

7 days

Frequency of treatment:

Once daily for up to 7 consecutive days, approximately the same time each day with a maximum of 6 hours difference between the earliest and latest dose. Animals were dosed up to the day prior to necropsy.

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/kg bw/day (nominal)

Doses / concentrations 2

Dose / conc.:

150 mg/kg bw/day (nominal)

Doses / concentrations 3

Dose / conc.:

300 mg/kg bw/day (nominal)

Doses / concentrations 4

Dose / conc.:

1 000 mg/kg bw/day (nominal)

No. of animals per sex per dose:

3

Control animals:

yes, concurrent no treatment

Details on study design:

Male and female Wistar Han rats, approximately 9 weeks of age on study Day 1, were administered PF-06410251 via oral gavagedaily for 7 consecutive days

Examinations

Sacrifice and pathology:

At least twice daily. Animals showing pain, distress or discomfort, which is considered not transient in nature or is likely to become more severe, will be sacrificed for humane reasons based on OECD guidance document on humane endpoints (ENV/JM/MONO/ 2000/7). The time of death will be recorded as precisely as possible.

Other examinations:

Clinical signs: At least once daily from start of treatment onwards, detailed clinical observations will be made in all animals. The time of onset, grade and duration of any observed signs will be recorded. Signs will be graded for severity and the maximum grade will be predefined at 3 or 4. Grades will be coded as slight (grade 1), moderate (grade 2), severe (grade 3) andvery severe (grade 4). For certain signs, only its presence (grade 1) or absence (grade 0) will be scored. In the data tables, the scored grades will be reported, as well as the percentage of animals affected in summary tables.
Body weights: On Days 1, 4 and 7. In order to monitor the health status animals may be weighed more often.
Food consumption: Over Days 1-4 and 4-7.
Water consumption: Subjective appraisal will be maintained during the study and a quantitative assessment introduced in the event of a suspected treatment related effect.

Results and discussion

Results of examinations

Clinical signs:

no effects observed

Description (incidence and severity):

No clinical signs of toxicity were noted during the observation period.

Mortality:

no mortality observed

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

One male and one female at 1000 mg/kg showed body weight loss between Days 4 and 7. In addition, one control female showed body weight loss between Days 4 and 7. And at 300 mg/kg body weight loss was noted in one female between Days 1-4 and recovered thereafter. Reduced body weight gain was also noted in individual females at 300 and 1000 mg/kg. No toxicologically significant changes in body weights and body weight gain were noted at 150 mg/kg (both sexes) and 300 mg/kg (males).

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

A slight trend of lower food consumption was observed in females at 300 and 1000 mg/kg, predominantly between Days 1-4 and recovered at 300 mg/kg between Days 4-7. No toxicologically significant changes in food consumption before or after allowance for body weight were noted in males.

Food efficiency:

not examined

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

no effects observed

Ophthalmological findings:

not examined

Clinical biochemistry findings:

no effects observed

Description (incidence and severity):

No toxicologically relevant changes occurred in clinical biochemistry parameters of treated rats. Slightly lower calcium was noted in males at 1000 mg/kg and slightly higher protein and albumin were noted in females at 1000 mg/kg. However all these changes remained within the range considered normal for rats of this age and strain and/or occurred in absence of supportive morphological changes. Therefore these changes were considered not toxicologically significant. Slight variation in the other parameters were considered to be of no toxicological significance as they occurred in the absence of a treatment-related distribution and remained within the range considered normal for rats of this age and strain.

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

no effects observed

Description (incidence and severity):

No toxicologically significant changes were noted in organ weights and organ to body weight ratios. Higher absolute and relative epididymides weights were noted at 1000 mg/kg. Since these values remained within the range considered normal for rats of this age and strain and no supportive morphological changes were noted these changes were not considered toxicologically significant. A single animal showed a higher ovaries weight, which was supported by the cysts noted at macroand microscopic examination. This finding is occasionally seen in animals of this age and strain and therefore not considered to be toxicologically relevant. Slight variation in the other organs among the dose groups occurred well within the range considered
normal for rats of this age and strain and in absence of supportive morphological changes. Therefore these changes were not considered toxicologically significant.

Gross pathological findings:

no effects observed

Description (incidence and severity):

Necropsy did not reveal any toxicologically relevant alterations. Other necropsy findings, among all treated groups, were considered to be of no toxicological significance since they occurred in the absence of a treatment-related distribution, and are occasionally seen among rats used in these types of studies.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

not specified

Histopathological findings: neoplastic:

not specified

Other effects:

effects observed, treatment-related

Description (incidence and severity):

An increased incidence and/or severity of lymphogranulocytic inflammation of the glandular stomach, hyperkeratosis of the forestomach and/or vacuolation of the limiting ridge was recorded in the stomach of male rats at 1000 mg/kg/day. All other microscopic findings were within the range of background pathology encountered in Wistar
rats of this age and strain and occurred at similar incidences and severity in both control and treated rats.

Details on results:

Wistar rats were treated withPF-06410251 for 7 consecutive days by daily oralgavageat dose levels up to 300 mg/kg. Slight body weight loss and/or reduced body weight gain was noted for individual animals at 300 and 1000 mg/kg, which was accompanied by a slight trend for reduced food consumption in females. At microscopic examination, lymphogranulocytic inflammation of the glandular stomach, hyperkeratosos of the forestomach and/or vacuolation of the limiting ridge were noted in males treated at 1000 mg/kg. No toxicologically relevant microscopic findings were observed for females. All of the remaining parameters investigated in this study (i.e. clinical appearance, clinical laboratory investigations, macroscopic examination and organ weights) were considered to be normal for rats of this age and strain. Based on the microscopic findings in the stomach of males at 1000 mg/kg, it was concluded thatPF06410251
has no toxic potential when administered to rats by daily oralgavagefor a period of up to 7 consecutive days up to 300 mg/kg.

Effect levels

Dose descriptor:

NOAEL

Effect level:

> 300 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

food consumption and compound intake

Target system / organ toxicity

Critical effects observed:

no

Lowest effective dose / conc.:

300 mg/kg bw/day (nominal)

System:

gastrointestinal tract

Organ:

not specified

Applicant's summary and conclusion

Conclusions:

Based on the microscopic findings in the stomach of males at 1000 mg/kg, it was concluded thatPF06410251 has no toxic potential when administered to rats by daily oralgavagefor a period of up to 7 consecutive days up to 300 mg/kg.

Executive summary:

Title:

7-Day repeated dose study with PF-06410251 by daily gavage in the rat.

Guidelines:

The study was based on the following guidelines: - EC No 440/2008, B.7 Repeated Dose (28 days) Toxicity (oral), 2008. - OECD 407, Repeated Dose 28-day Oral Toxicity Study in Rodents, 2008.

Rationale for dose levels

Dose levels for this 7-day oralgavagestudy were selected to be 0, 150, 300 and 1000 mg/kg, based on the results of the acute oral toxicity study (Project 504696) and on information from the Sponsor.

Study outline

The test substance, formulated in 1% aqueous carboxymethyl cellulose, was administered daily for 7 days by oralgavageto SPF-bred Wistar rats. One control group and three treated groups were tested, each consisting of 3 males and 3 females.

Evaluated parameters

The following parameters were evaluated: clinical signs daily; body weight on Days 1, 4 and 7 and food consumption over Days 1-4 and 4-7; clinical pathology and macroscopy at termination; organ weights and histopathology on a selection of tissues.

Results

Body weight loss and/or reduced body weight gain was noted at 300 and 1000 mg/kg, which was accompanied by a slight trend for reduced food consumption in females only. At microscopic examination, lymphogranulocytic inflammation of the glandular stomach, hyperkeratosos of the forestomach and/or vacuolation of the limiting ridge were noted in males treated at 1000 mg/kg. No toxicologically relevant microscopic findings were observed for females. All of the remaining parameters investigated in this study (i.e. clinical appearance, clinical laboratory investigations, macroscopic examination and organ weights) were considered to be normal for rats of this age and strain.