Registration Dossier

Administrative data

Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
18 March 2016 to 21 July 2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2017
Report Date:
2017

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity in Rodents)
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid
Specific details on test material used for the study:
Identity POLYOL R2490
EC name Poly[oxy(methyl-1,2-ethanediyl)],α,α’-(2,2-dimethyl-1,3-propanediyl) bis[ω-hydroxy-]
Batch number 150602165
EC number 610-848-9
CAS number 52479-58-0
Expiry date 20 December 2016
Appearance Clear colourless solution
Storage conditions Room temperature

Test animals

Species:
rat
Strain:
Sprague-Dawley
Details on species / strain selection:
Standard species/strain use for toxicity studies
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Envigo RMS srl, San Pietro al Natisone (UD), Italy

- Age at study initiation: Seven to eight weeks
- Weight at study initiation: Group mean of male rats (195 to 197g); Group mean of female rats (160 to 162g)
- Fasting period before study: No
- Housing: in groups of up to five by sex in solid floor polysulphone cages with bedding bags.
- Diet: ad libitum 4 RF 21
- Water: ad libitum
- Acclimation period: two weeks

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2
- Humidity (%): 55 ± 15
- Air changes (per hr): approximately fifteen to twenty air changes per hour
- Photoperiod (hrs dark / hrs light): twelve hours continuous light and twelve hours darkness cycle

IN-LIFE DATES: 18 March 2016 (allocation of animals) to 21 July 2016 (final day of necropsy)

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS:
The test item was prepared at appropriate concentrations in softened water. The stability of the test item formulations at the lowest and highest concentrations formulated (10 and 75 mg/mL) was determined and these formulations were found to be stable for up to 8 days when stored at room temperature. Formulations were prepared on a weekly or daily basis.
VEHICLE
- Concentration in vehicle: 0, 10, 27.5, 75 mg/mL (main phase) and 0, 75 mg/mL (recovery phase)
-Dose volume: 10 mL/kg bw
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Samples of the formulations prepared on Weeks 1 and 13 were analysed to confirm the test item concentration for all dosing solutions prepared. The results indicate that the prepared formulations were within 90 to 110% of the nominal concentration.
Duration of treatment / exposure:
13 weeks
Frequency of treatment:
Once daily (oral gavage) for all animals.
Doses / concentrationsopen allclose all
Dose / conc.:
0 mg/kg bw/day (actual dose received)
Remarks:
Control
Dose / conc.:
100 mg/kg bw/day (actual dose received)
Dose / conc.:
275 mg/kg bw/day (actual dose received)
Dose / conc.:
750 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
10 males and 10 females per dose (main study phase); 5 males and 5 females were used for the recovery phase (control and high dose groups)
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The dose levels were chosen based on information from a preliminary non GLP compliant study (RTC Study No. E0022). This oral gavage preliminary study was carried out at dose levels of 100, 300 and 1000 mg/kg bw/d. The highest dose level of 1000 mg/kg bw/d induced some slight changes (increased liver weight and size). Therefore, the highest dose of 1000 mg/kg bw/d was not considered to be safe (in terms of potential liver toxicity) in a subsequent toxicity study of longer duration. Since the changes at this dose level were very slight, a high dose of 750 mg/kg/day was assuemd to be reasonably safe. Dosages of 100 and 275 mg/kg bw/d were selected for low and mid-dose levels.

Rationale for animal assignment: The animals were randomly allocated to treatment groups using a stratified randomisation procedure to give approximately equal initial group.
Positive control:
Not required for this study type.

Examinations

Observations and examinations performed and frequency:
DETAILED CLINICAL OBSERVATIONS
All clinical signs were recorded for individual animals. Once before commencement of treatment and at least once daily during the study, each animal was observed and any clinical sign was recorded. Observations were performed at the same time interval each day, the interval was selected taking into consideration the presence of post-dose reactions.

Once before commencement of treatment and at least once per week from the start of treatment, each animal was given a detailed clinical examination. Each animal was observed in an open arena. The test included observation of changes in gait and posture, reactivity to handling, presence of clonic or tonic movements, stereotypies or bizarre behaviour and effects on the autonomic nervous system (e.g. lachrymation, piloerection, unusual respiratory pattern). Changes in fur, skin, eyes, mucous membranes, occurrences of secretions and excretions were also recorded.

BODY WEIGHT
Individual body weights were recorded on the day of allocation to treatment groups, the day before treatment commenced and at weekly intervals thereafter. Body weights were also recorded just prior to necropsy.

FOOD CONSUMPTION
The weight of food consumed by each cage of rats was recorded at weekly intervals following allocation. The interval between allocation and treatment initiation was less than a full week. The group mean daily intake per rat was calculated.

OPHTHALMOSCOPIC EXAMINATION
Both eyes of all animals were examined prior to the commencement of treatment by means of an ophthalmoscope, and by a slit-lamp microscope, after the instillation of 0.5% Tropicamide (Visumidriatic®, Visufarma, Rome, Italy). The eyes of all animals from high dose and control groups were re-examined duringWeek 13 of treatment.

CLINICAL PATHOLOGY
Prior to necropsy, samples of blood were withdrawn under isofluorane anaesthesia from the abdominal vena cava of 10 male and 10 female animals from each group, following overnight food deprivation (approximately 17-18 hours). Blood samples were collected and analysed in the same order. The blood samples collected were divided into tubes as follows:
– EDTA anticoagulant for haematological investigations
– Heparin anticoagulant for biochemical tests
– Citrate anticoagulant for coagulation tests

HAEMATOLOGY
Parameters checked included: Red blood cell count, Haematocrit, Haemoglobin, Mean corpuscular haemoglobin, Mean red blood cell volume, Mean corpuscular hemoglobin concentration, White blood cell count, Differential leukocyte count (neutrophils, lymphocytes, monocytes, eosinophils, basophils, large unstained cells), Platelets, Reticulocyte count. Samples from two animals (one male 100 mg/mg bw/d and one female 750 mg/kg bw/d) were unable to be analysed.

CLINICAL CHEMISTRY
Parameters checked included: Urea, Glucose, Total protein, Albumin, Globulin, Albumin/Globulin ratio, Sodium, Potassium, Chloride, Calcium, Phosphorus, Aspartate aminotransferase, Alanine aminotransferase, Alkaline phosphatase, Gamma-glutamyltransferase, Creatinine, Total cholesterol, Total bilirubin, Triglycerides. Samples from one animal (one female 750 mg/kg bw/d) was unable to be analysed.

COAGULATION
Prothrombin time was analysed. Samples from four animals (one male 100 mg/mg bw/d; one male, 275 mg/kg bw/d and two females, 750 mg/kg bw/d) were unable to be analysed.

NEUROTOXICITY
Once during Week 12 of treatment and once during Week 4 of recovery, an evaluation of sensory reactivity to stimuli of different modalities (e.g. auditory, visual and proprioceptive stimuli) and an assessment of grip strength were also performed.

MOTOR ACTIVITY
The motor activity (MA) of all animals was measured once during Week 13 of treatment and once during Week 4 of recovery by an automated activity recording. Measurements were performed using a computer generated random order.
Sacrifice and pathology:
GROSS PATHOLOGY
One animal in extremis and those that had completed the scheduled test period were killed by exsanguination under isofluorane anaesthesia. All animals were subjected to necropsy,
supervised by a pathologist.

ORGAN WEIGHTS
The following organs, removed from animals that were killed at the test period, were dissected free from fat and weighed before fixation:
Adrenals, Brain (cerebrum, cerebellum, medulla/pons), Epididymides, Heart, Kidneys, Liver, Ovaries (including oviducts), Spleen, Testes, Thymus, Cervix,
The ratios of organ weight to body weight were calculated for each animal.

HISTOPATHOLOGY
Samples of the following tissues were removed from all animals and preserved in buffered 10% formalin, Davidson’s fluid or modified Davidson’s fluid as appropriate:
Abnormalities, Adrenals, Aorta, Bone marrow (from sternum), Bone & bone marrow (sternum), Brain (cerebrum, cerebellum and medulla/pons), Caecum, Colon, Duodenum, Epididymides, Eyes, Femur with joint, Heart, Ileum, Jejunum including Peyer’s patches, Kidneys, Liver, Lungs (including mainstem bronchi), Lymph nodes (cervical and mesenteric), Mammary area, Oesophagus, Ovaries, Oviducts. Pancreas, Pituitary gland, Prostate gland, Rectum, Salivary glands, Sciatic nerve, Seminal vesicles, Skeletal muscle, Skin, Spinal column, Spinal cord, Spleen, Stomach, Testes , Thymus, Thyroid/Parathyroid, Trachea, Urinary bladder, Uterus (with cervix).

All tissues (with the exception of the eyes, femur, seminal vesicles, skeletal muscle and spinal column) were prepared as paraffin blocks, sectioned at a nominal thickness of 5 μm and stained with Hematoxylin and Eosin for subsequent microscopic examination. Examination was conducted on tissues specified from all animals in the control and high dose groups dying during the treatment period or killed at the end of the 13 weeks of treatment, tissues from one animal killed during the treatment period and all abnormalities in all main phase groups.
Other examinations:
No further
Statistics:
Standard deviations were calculated as considered appropriate. For continuous variables the significance of the differences amongst groups was assessed by analysis of variance.
Differences between each treated group and the control group were assessed by Dunnett’s test using a pooled error variance. The homogeneity of the data was verified by Bartlett’s test before Dunnett’s test. If the data were found to be inhomogeneous, a Modified t test (Cochran and Cox) was applied.

The mean values, standard deviations and statistical analysis were calculated from the actual
values in the computer without rounding off. Statistical analysis of histopathological finding
was carried out by means of a nonparametric Kolmogorov-Smirnov test.

Results and discussion

Results of examinations

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Convulsions and salivation were observed in all males and females of the high dose group from Weeks 5 and 6, respectively. These signs occurred initially occasionally in individual animals, becoming gradually more frequent and severe from Week 9 up to the end of treatment period. They were no longer observed during the recovery period. A palpable mass was seen in a single female animal of the high dose group starting from the last week of treatment and during the recovery period. No changes of toxicological significance were found at the weekly clinical examination, which included an evaluation of neurotoxicity.
Mortality:
mortality observed, treatment-related
Description (incidence):
One female animal from the high dose group (receiving 750 mg/kg bw/d) was sacrificed for humane reasons on Day 36 of treatment period. Piloerection, decreased activity, semi-closed eyes, dyspnoea and hunched posture were observed in this animal prior to sacrifice. In addition, convulsions and salivation were observed in this animal during the week before death. At necropsy, this animal showed multiple dark abnormal areas in the lungs, showing no correlation at histopathology, and abnormal dark areas in the mucosa of the glandular stomach, that correlated at histopathology with focal mucosal erosion. No treatment-induced changes were seen at microscopic examination. All reported microscopic observations were considered to be an expression of spontaneous and/or incidental pathology. In particular, focal erosion of gastric mucosa and mild lymphocytolysis in the thymus are considered most likely stress-related and could be the contributory factors related to the poor health conditions of this animal.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Slight but statistically significant increases in body weights (from +6% to +15%) and absolute body weight changes (from +13% to +37%) were observed in the high dose females when compared to controls during treatment period. Body weight was also slightly increased in the high dose males on Day 92 of the treatment period (+6%). These increases gradually disappeared during recovery period.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Slight but statistically significant increases in food consumption (from +8% to +31%) were observed from Day 22 of treatment period in the high dose females. These increases were also observed with minor extent in the high dose males (from +12% to +14%) from Day 78 of treatment. These increases were no longer observed during recovery period.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
no effects observed
Description (incidence and severity):
Animals with no ocular abnormality were selected for the study by an ophthalmoscopic examination performed before the start of treatment. No findings were detected in both eyes of all surviving animals, from high dose and control groups, when they were re-examined during Week 13 of treatment.
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
Leucocytosis was recorded in animals of both sexes dosed at 750 mg/kg bw/d (54%) and in two males receiving 275 mg/kg bw/d (mean value were 29% above controls). Due to the slight severity and the absence of other related changes, the above findings were not considered adverse. The other statistically significant changes recorded (mean corpuscular volume, mean corpuscular haemoglobin concentration and reticulocytes in males, erythrocytes and haemoglobin in females) were of minimal severity and/or not dose-related, therefore considered of no toxicological relevance. No changes in coagulation were recorded.
Clinical biochemistry findings:
effects observed, non-treatment-related
Description (incidence and severity):
Statistically significant fluctuations of some biochemical parameters, mainly metabolic markers, were recorded in treated animals. Aspartate aminotransferase and total bilirubin alterations were not attributed to treatment, since they were not dose-related. Concerning the other findings observed, their severity was not considered to be suggestive of tissue/organ injury.
Urinalysis findings:
not examined
Behaviour (functional findings):
effects observed, treatment-related
Description (incidence and severity):
No differences between treated animals and controls, which could be considered treatment related, were observed at functional tests (sensory reactivity, landing footsplay, grip strength) performed at the end of treatment and recovery periods. A slight but statistically significant increase in motor activity (+36% and +27% in males and females, respectively) was observed in the high dose animals (dosed at 750mg/kg/day) when compared to controls at measurements performed at the end of treatment. No significant differences were seen at the end of recovery.
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
Terminal body weights showed slight increases (statistically significant in females only) in the high dose animals when compared to controls at the end of the treatment period (+6% and +15% in males and females, respectively). Dose-related, statistically significant increases in absolute liver weights were noted in the high dose animals (+22% and 37% in males and females, respectively) and in the mid-dose females (+12%). The relative liverweight was also significantly increased at statistical analysis in the the mid- and high dose males (+10% and +16% greater than controls, mid- and high dose groups, respectively) and in the mid- and high dose females (+11% and +19% greater than controls, mid- and high dose groups, respectively) at the end of the treatment period. The relative liver weight increases were no longer present at the end of the recovery period. No toxicological relevance was attributed to the remaining statistically significant increases of the absolute weights of brain and kidneys seen in the high dose males and of heart and kidneys seen in the high dose females, since they were not associated to a correspondent increase of the relative weights. They were therefore ascribed to the increased body weight observed in these animals. The statistically significant reduction of the relative weight of the uterus seen in the mid-dose females was considered to be incidental.
Gross pathological findings:
no effects observed
Description (incidence and severity):
No treatment-related changes were noted following gross pathology examination in both the main and recovery phases. All observed changes are suggested to be incidental, having a comparable incidence in control and treated groups, and/or are characteristically seen in untreated Sprague Dawley SD rats of the same age.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, non-treatment-related
Description (incidence and severity):
No treatment-related changes were noted. A minor range of changes was seen in control and treated animals, having a relatively comparable incidence, or is characteristically seen in untreated Sprague Dawley rats of the same age, administered with the test compound by the oral route.
Histopathological findings: neoplastic:
no effects observed
Other effects:
no effects observed

Effect levels

open allclose all
Key result
Dose descriptor:
LOAEL
Effect level:
750 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
clinical signs
mortality
body weight and weight gain
haematology
clinical biochemistry
Key result
Dose descriptor:
NOAEL
Effect level:
275 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
clinical signs
mortality
haematology
clinical biochemistry
organ weights and organ / body weight ratios
Dose descriptor:
NOEL
Effect level:
100 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
clinical biochemistry
organ weights and organ / body weight ratios

Target system / organ toxicity

Key result
Critical effects observed:
no

Any other information on results incl. tables

SUMMARY OF RELEVENT FINDINGS

 

Male

Female

Dose level (mg/kg bw/d)

Control

100

275

750

Control

100

275

750

Convulsions

0/15

0/15

0/15

15/15

0/15

0/15

0/15

15/15

Salivation

0/15

0/15

0/15

15/15

0/15

0/15

0/15

15/15

Motor activitya

1043.9

957.3

1104.4

1422.7+D

1068.7

1285.1*D

1059.4

1354.7+D

Motor activityb

884.4

N/A

N/A

1123.2

999.4

N/A

N/A

1041.2

Terminal body weights (g)

461.37

459.34

463.22

490.50*D

285.53

279.27

291.65

329.15+D

Body weight gain (g)

263.99

263.22

268.25

294.36+D

123.75

118.47

129.65

169.31+D

Food consumption (Day 92) (g)

21.91

21.29

21.89

25.08+D

17.00

16.55

17.14

20.85+D

Liver weight (g)c

12.002

12.381

13.223

14.694+D

6.084

6.136

6.840*D

8.333+D

Liver weight to terminal body weight (%)c

2.6113

2.7188

2.8805+D

3.250+D

2.2516

2.3234

2.4969*D

2.684+D

Liver weight (g)d

12.120

N/A

N/A

13.138

6.593

N/A

N/A

7.922*D

Liver weight to terminal body weight (%)d

2.5904

N/A

N/A

2.7633

2.2728

 

N/A

N/A

2.5696

a           Mean counter display for dosing animals on Day 79
b           Mean counter display for recovery animals on Recovery Day 26
c           Dosing group
d           Recovery group
N/A       Not applicable
*D         Statistically significant at 0.05 level with Dunnett LSD Test
+D         Statistically significant at 0.01 level with Dunnett LSD Test

CLINICAL CHEMISTRY

 

Control data are expressed in absolute values and data from treated groups as a percentage difference with the controls.

Male animals

Dose (mg/kg bw/d)

AST (U/L)

TBIL (mg/dL)

CHOL (mg/dL)

TRIG (mg/dL)

GLU

(mg/dL)

UREA (mg/dL)

CREA (mg/dL)

Cl (mmol/L)

Ca (mmol/L)

Na (mmol/L)

IP (mg/dL)

GLO (g/dL)

0 (Control)

66.1

0.022

82.67

75.93

124.27

37.19

0.305

102.52

2.496

144.63

5.219

2.49

100

-6

64

1

-30*D

7

16*D

-7

0

4*D

0

12

2

275

-9

114

20*D

-28

20

14

-2

-1

5+D

0

11

2

750

-1

32

23*D

-33*D

12

30+D

-4

0

5+D

-1

35+D

0

 

Female animals

Dose (mg/kg bw/d)

AST (U/L)

TBIL (mg/dL)

CHOL (mg/dL)

TRIG (mg/dL)

GLU

(mg/dL)

UREA (mg/dL)

CREA (mg/dL)

Cl (mmol/L)

Ca (mmol/L)

Na (mmol/L)

IP (mg/dL)

GLO (g/dL)

0 (Control)

68.27

0.079

86.82

47.58

97.7

35.62

0.44

105.32

2.452

143.91

4.324

2.36

100

-12

-3

2

27

14

9

-2

-1

-1

-1

26+C

-1

275

-15+D

-41+D

15

32

24*D

20

8

-1

1

-1

23+C

3

750

-6

-27

21*D

79*C

39+D

39+D

17D

-3+D

7*C

-2+D

58+C

8*C

 

 

+C = Cochran and Cox Test Significant at the 0.01 level

*C = Cochran and Cox Test Significant at the 0.05 level

+D = Dunnett LSD Test Significant at the 0.01 level

*D = Dunnett LSD Test Significant at the 0.05 level

AST: Aspartate aminotransferase
TBIL: Total bilirubin
CHOL: Total cholesterol
TRIG: Trigyycerides
GLU: Glucose
Cl: Chloride
Ca: Calcium
Na: Sodium
IP: Inorganic phosphorus
GLO: Globulin

 

HAEMATOLOGY

 

Male animals

Dose (mg/kg bw/d)

WBC

(x103/µL)

NEU

(x103/µL)

LYM

(x103/µL)

MON

(x103/µL)

EOS

(x103/µL)

BAS

(x103/µL)

LUC

(x103/µL)

NEUR (%)

LYMR (%)

MONR (%)

EOSR (%)

BASR

(%)

LUCR

(%)

0 (Control)

6.859

0.775

5.710

0.151

0.136

0.052

0.037

11.38

83.06

2.23

2.00

0.75

0.55

100

7.320

0.923

6.063

0.150

0.096

0.047

0.037

12.94

82.54

2.08

1.30+D

0.64

0.49

275

8.845*c

0.844

7.561*D

0.182

0.132

0.071

0.056

9.54

85.53

2.06

1.48+D

0.78

0.60

750

10530#C

1.230

8.661+D

0.346#C

0.129

0.099+D

0.066+C

11.91

81.99

3.33*C

1.22+D

0.94*D

0.61

 

Female animals

Dose

(mg/kg bw/d)

WBC

(x103/µL)

NEU

(x103/µL)

LYM

(x103/µL)

MON

(x103/µL)

EOS

(x103/µL)

BAS

(x103/µL)

LUC

(x103/µL)

NEUR (%)

LYMR (%)

MONR (%)

EOSR (%)

BASR

(%)

LUCR

(%)

0 (Control)

4.744

0.345

4.173

0.106

0.076

0.020

0.023

7.60

87.49

2.31

1.65

0.43

0.56

100

4.900

0.352

4.304

0.098

0.083

0.026

0.042

7.40

87.65

2.07

1.67

0.48

0.73

275

5.784

0.419

5.095

0.109

0.097

0.030*C

0.036

7.46

87.84

1.88

1.69

0.52

0.61

750

7.525+D

0.575+D

6.533+D

0.188+D

0.114

0.058+C

0.056

7.91

86.54

2.56

1.53

0.75+D

0.70

 

+C = Cochran and Cox Test Significant at the 0.01 level

*C = Cochran and Cox Test Significant at the 0.05 level

#C = Cochran and Cox Test Significant at 0.001 level

+D = Dunnett LSD Test Significant at the 0.01 level

*D = Dunnett LSD Test Significant at the 0.05 level

WBC: White blood cell count

NEUR/NEU: Neutrophils (relative or absolute)

LYMR/LYM: Lymphocytes (relative or absolute)

MONR/MON: Monocytes (relative or absolute)

EOSR/EOS: Eosinophils (relative or absolute)

BASR/BAS: Basophils (relative or absolute)

LUCR/LUC: Large unstained cells (relative or absolute)

MEAN BODY WEIGHTS

 

Mean body weights are presented in grams (g).

Male animals

 

Quarantine

Dosing

Dose
(mg/kg/bw)

Day 9

Day 1

Day 8

Day 15

Day 22

Day 29

Day 36

Day 43

Day 50

Day 57

Day 64

Day 71

Day 78

Day 85

Day 92

0 (Control)

147.08

197.37

246.36

290.19

325.87

355.03

380.30

402.10

415.93

428.80

439.72

450.61

458.87

470.91

461.37

100

147.16

196.12

246.03

288.22

372.21

354.97

381.05

402.62

419.43

429.19

442.96

452.82

458.82

473.99

459.34

275

147.18

194.97

246.46

292.17

331.49

358.86

358.86

406.63

420.75

431.88

443.39

457.44

66.43

478.33

463.22

750

146.85

196.14

247.41

391.44

329.73

354.99

384.11

406.59

419.85

437.88

452.63

466.21

477.44

493.39

490.50*D

*D = Dunnett LSD Test Significant at the 0.05 level

 

Female animals

 

Quarantine

Dosing

Dose
(mg/kg/bw)

Day 9

Day 1

Day 8

Day 15

Day 22

Day 29

Day 36

Day 43

Day 50

Day 57

Day 64

Day 71

Day 78

Day 85

Day 92

0 (Control)

131.99

161.78

182.93

204.75

222.08

234.55

245.05

258.01

264.00

267.35

274.32

275.19

279.49

283.87

285.53

100

132.13

160.80

181.82

204.49

217.95

231.96

240.73

251.55

262.08

58.90

62.42

266.69

269.99

275.49

279.27

275

132.05

162.00

184.20

208.94

225.96

238.12

249.51

275.61

266.67

271.21

276.30

278.36

281.14

290.34

291.65

750

132.13

160.21

183.02

205.20

226.01

242.68

259.10*D

273.24*D

283.56+D

292.82+D

300.56+D

309.64+D

315.29+D

326.33+D

329.15+D

+D = Dunnett LSD Test Significant at the 0.01 level

*D = Dunnett LSD Test Significant at the 0.05 level

Applicant's summary and conclusion

Conclusions:
Signs of effects related to treatment with POLYOL R2490 were observed in animals at the high dose level of 750 mg/kg bw/d, when administered by oral gavage for 13 consecutive weeks at dose levels of 0, 100, 275 and 750mg/kg bw/d.
Executive summary:

In a 90-day study performed to OECD 408, Polyol R2490 was administered by oral gavage to groups of Sprague Dawley rats (10/sex) on ninety-two consecutive days at dose levels of 0 (controls) 100, 275 and 750 mg/kg bw/d. Dose levels were based on a range-finding study in which enlarged liver was observed at 1000 mg/kg bw/d. A control group was administered with the vehicle (softened water) by oral gavage. Additional animals (5/sex) were treated with Polyol R2490 for ninety-two consecutive days at a dose level of 750 mg/kg bw/d and allowed a 4 -week recovery period. A control recovery group was administered with the vehicle by oral gavage. Clinical signs, functional observations, body weight change, dietary intake, ophthalmoscopy, were monitored during the study. Haematology and blood chemistry were evaluated for all animals at the end of the study. All animals were subjected to gross necropsy examination and histopathological evaluation of selected tissues was performed. There was one unscheduled death during the study. One female animal from the high dose group (receiving 750 mg/kg bw/d) was sacrificed for humane reasons on Day 36. Convulsions and salivation were observed for all animals of the high dose group from Week 5 (males) and from Week 6 (females). These signs were occasional and occurred in individual animals initially, but increased in frequency and severity from Week 9 to the end of treatment. These signs were not seen during the 4-week recovery period. Statistically significant increases in motor activity were observed in high dose animals at the end of the treatment period when compared to the concurrent control animals, however no significant changes were noted at the end of the recovery phase. In addition, no differences were noted in sensory reactivity, landing footsplay or grip strength. Slight statistically significant increases in bodyweights (from +6% to +15%) and absolute body weight changes (from +13% to +37%) were observed in females treated with 750 mg/kg bw/d when compared to the concurrent control animals. A statistically significant increase in the high dose males was observed on Day 92 of treatment. Increases in food consumption were also observed during the treatment period in high dose females from Day 22 (from +8% to +31%) and in high dose males from Day 78 (from +12% to +14%). Increases in bodyweights and food consumption were not seen during the recovery period. Increases in terminal body weight were observed in animals treated with 750 mg/mg bw/d in both males (+6%) and females (+15%), although this was statistically significant in females only.

Statistically significant increases in absolute liver weight were observed in high dose males and females and in mid-dose females. These were dose-related in both sexes. Relative liver weights were also increased in high and mid-dose males and females (up to 16% in males and up to 19% in females). These changes were not seen in the recovery animals. Leucyocytosis was recorded in males and females dosed with 750 mg/kg bw/d and in two males dosed with 275 mg/kg bw/d. These findings were considered not to be adverse due to the slight severity and the absence of other related findings. Some changes in clinical chemistry were noted. Increases in aspartate aminotransferase and total bilirubin were not treatment related. Changes in cholesterol, triglycerides, glucose, urea, creatantine, chloride, calcium, sodium, inorganic phosphorus and globulin were considered not to indicate tissue or organ injury. The majority of findings were attributed to severe stress which was caused by treatments with Polyol R2490. Due to the post-dose convulsions and poor health of the animals, these were considered to be adverse. A NOAEL of 275 mg/kg bw/d can be determined for this study. Only very slight changes in clinical chemistry parameters and slight increase in liver weight were observed at this dose level.