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Repeated dose toxicity: oral

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Administrative data

Endpoint:
chronic toxicity: oral
Remarks:
combined repeated dose and carcinogenicity
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Study noted as being performed according to OECD453 and to GLP principles with a focus on urinary and bladder effects. No information was provided on hematology and clinical chemistry, with limited information on histopathology of other organs.

Data source

Reference
Reference Type:
publication
Title:
Unnamed
Year:
2002

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 453 (Combined Chronic Toxicity / Carcinogenicity Studies)
Version / remarks:
1981
Deviations:
not specified
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): biphenyl
- Analytical purity: >98%
- Stability under test conditions: concentrations in the diet were confirmed by gas chromatography and found within the range of 86.2 and 101% of target concentrations.
- Other: test substance obtained from Wako Pure Chemical Industries, Ltd. (Tokyo, Japan)

Test animals

Species:
rat
Strain:
Fischer 344/DuCrj
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Japan Inc. (Kanagawa, Japan)
- Age at study initiation: 6 weeks
- Weight at study initiation: mean body weights read from figure: roughly 95-130 g. The weight variation for each sex did not seem to exceed ± 20 % of the mean weight.
- Fasting period before study: no
- Housing: individually in stainless-steel wire-mesh hanging cages (170 x 294 x 176 mm)
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 1 week quarantine followed by 1 week acclimation

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 24 +/- 2
- Humidity (%): 55 +/- 10
- Air changes (per hr): 15-17
- Photoperiod (hrs dark / hrs light): 12 / 12 (fluorescent lighting)

Administration / exposure

Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS: no information

DIET PREPARATION
- Rate of preparation of diet (frequency): no information;
the diet containing appropriate concentrations of test substance was prepared by mixing in a spiral mixer for 20 min.
- Mixing appropriate amounts with (Type of food): gamma-irridiation sterilized CRF-1 powdered diet (Oriental Yeast Co., Chiba Japan)
- Storage temperature of food: 4 degC

Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Concentrations in the diet were confirmed by gas chromatography and found within the range of 86.2 and 101% of target concentrations.
Duration of treatment / exposure:
105 weeks
Frequency of treatment:
continuous
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
500 ppm
Basis:
nominal in diet
Remarks:
Doses / Concentrations:
1500 ppm
Basis:
nominal in diet
Remarks:
Doses / Concentrations:
4500 ppm
Basis:
nominal in diet
No. of animals per sex per dose:
50
Control animals:
yes, plain diet
Details on study design:
- Dose selection rationale: not indicated
- Rationale for animal assignment (if not random): not indicated
- Section schedule rationale (if not random): not indicated
Positive control:
no positive control tested

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily
- Cage side observations included: clinical signs, behavioral changes and mortality

BODY WEIGHT: Yes
- Time schedule for examinations: once a week for the first 14 weeks, and every 4 weeks thereafter

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): food consumption was measured once a week for the first 14 weeks, and every 4 weeks thereafter.
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No data
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No data

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No data

OPHTHALMOSCOPIC EXAMINATION: Yes, as anemia-colored eyes were noticed.
- Time schedule for examinations: no data
- Dose groups that were examined: at least the high dose group males (showing hematuria)

HAEMATOLOGY: No data

CLINICAL CHEMISTRY: No data

URINALYSIS: Yes
- Time schedule for collection of urine: final week (week 105)
- Metabolism cages used for collection of urine: No data
- Animals fasted: No data
- Parameters examined: urinary parameters including pH and occult blood using Urolabsix (Diagnostic Diutsior, bayer, Elkhart, Germany)

NEUROBEHAVIOURAL EXAMINATION: No data
Sacrifice and pathology:
All organs were examined macroscopically, the organs weighed and the tissues for microscopic examination included the ones specified in the OECD 453 test guideline.
Statistics:
Chi-square test (incidence non-neoplastic lesions, urinary data) and Fisher's exact test (incidence neoplastic lesions)

Results and discussion

Results of examinations

Clinical signs:
effects observed, treatment-related
Mortality:
mortality observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
not specified
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
effects observed, treatment-related
Haematological findings:
not specified
Clinical biochemistry findings:
not specified
Urinalysis findings:
effects observed, treatment-related
Behaviour (functional findings):
not specified
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
effects observed, treatment-related
Details on results:
CLINICAL SIGNS AND MORTALITY:
Mortality males: control 13/50; 500 ppm 9/50; 1500 ppm 9/50; 4500 ppm 19/50 (statistically significant)
Mortality females: control 5/50; 500 ppm 12/50; 1500 ppm 5/50; 4500 ppm 13/50
Clinical hematuria was observed from the 40th week onward in 32 males of the 4500 ppm group, of which 14 had anemia-colored skin and/or eyes.
No clinical signs in females.

BODY WEIGHT AND WEIGHT GAIN
Significant body weight decrease in 4500 ppm males. Other dose groups and females were normal.

OPHTHALMOSCOPIC EXAMINATION
Anemia colored eyes in 4500 ppm males (number unknown) with clinical hematuria.

URINALYSIS
4500 ppm males: significant increase in urinary pH (ca. 104%) and incidences of positive occult blood (23/50), compared to controls (1/50).
4500 ppm females: significant increase in incidences of positive occult blood (10/50) compared to controls (1/50).

ORGAN WEIGHTS
Relative kidney weight: statistically significant increase in 1500 and 4500 ppm rats (males and females)
Absolute kidney weight: statistically significant increase in 4500 ppm males.

GROSS PATHOLOGY
Bladder calculi: 43/50 in the 4500 ppm males (40th week onwards), 8/50 in the 4500 ppm females. No calculi were observed in other dose groups.
Male calculi varied in color, shape and sizes. The male calculi were triangular, pyrimidal and cubical in shape, with sizes ranging form 0.3-1.0 cm
Female calculi were white/yellow, spheroidal and of same size.
4/8 in the 4500 ppm females with calculi had thickening of bladder wall.
41/50 in the 4500 ppm males had polyp-like or papillary nodules which protruded from the bladder wall into the lumen. Of these 41, 38 had also bladder calculi.

HISTOPATHOLOGY: NON-NEOPLASTIC
Other organs than bladder: no tumor-related lesions.

500 ppm and 1500 ppm groups: There were no significant lesions of the urinary bladder, ureter or kidney. In the 1500 ppm females there was simple transitional cell hyperplasia of renal pelvis (12/50) and deposit of hemosiderin (22/50).

4500 ppm dose groups:
Significant lesion of the bladder in 4500 ppm males :
- transitional cell hyperplasia: simple (12/50), nodular (40/50), papillary (17/50), total (45/50).
- Squamous metaplasia: (19/50)
- Squamous cell hyperplasia (13/50)
- Inflammatory polyp (10/50).

In 4500 ppm females some bladder lesions identified as significant in the males dosed with 4500 ppm were also found, but they were not statistically significant.

Significant other lesions in 4500 ppm males:
- Ureter transitional cell hyperplasia: simple (8/50)
- Ureter dilatation (14/50)
- Renal pelvis transitional cell hyperplasia: simple (19/50) or nodular (21/50)
- Pelvis desquamation (11/50)
- Pelvis calculi (13/50)
- Mineralization of cortico-medullary junction: (10/50)
- Mineralization of papilla (23/50).

Significant other findings/lesions in 4500 ppm females:
- Renal pelvis transitional cell hyperplasia: simple (25/50) or nodular (12/50)
- Mineralization of pelvis (27/50)
- Mineralization of papilla (12/50)
- Papillary necrosis (23/50) or infarct (8/50)
- deposit of hemosiderin (25/50)

Significant other findings/lesions in 1500 ppm females:
- Renal pelvis transitional cell hyperplasia: simple (12/50)
- deposit of hemosiderin (22/50)

A few other none statisitically significant lesions were also identified in both males and females dosed with 4500 ppm of the test substance.

HISTOPATHOLOGY: NEOPLASTIC (if applicable)
Other organs than bladder: no tumor or tumor-related lesions.
Urinary bladder, ureter, kidney: no lesions in control, 500 ppm and 1500 ppm groups (m/f).
Significant neoplastic lesions in 4500 ppm males:
- Transitional cell papilloma (10/50)
- Carcinoma (24/50).
- Total bladder tumors (31/50)(3 rats had both transitional cell papilloma and carcinoma). One rat showed squamous cell papilloma and carcinoma (not significant).

In 4500 ppm females no neoplastic lesions were found.

HISTORICAL CONTROL DATA: Neither squamous cell papilloma nor squamous cell carcinoma have been observed in the F344 male rats historically either in the laboratory performing the study nor by NTP.

Effect levels

open allclose all
Dose descriptor:
LOAEL
Effect level:
4 500 ppm
Based on:
test mat.
Sex:
male
Basis for effect level:
other: see 'Remark'
Dose descriptor:
NOAEL
Effect level:
1 500 ppm
Based on:
test mat.
Sex:
male
Basis for effect level:
food consumption and compound intake
other: none
Dose descriptor:
LOAEL
Effect level:
1 500 ppm
Based on:
test mat.
Sex:
female
Basis for effect level:
other: A statistically significant (and dose-dependent) increase in simple transitional cell hyperplasia and deposit of hemosiderin was observed at levels at and above 1500 ppm.
Dose descriptor:
NOAEL
Effect level:
500 ppm
Based on:
test mat.
Sex:
female
Basis for effect level:
body weight and weight gain

Target system / organ toxicity

Critical effects observed:
not specified

Any other information on results incl. tables

Tumors in male rats dosed with 4500 ppm biphenyl were induced in high association with calculus formation and hematuria, together with pre-neoplastic, regenerative lesions in the urinary system.

In the male rat, the transitional cell hyperplasia and the squamous cell hyperplasia were considered to represent a regenerative, pre-neoplastic response to the epithelial damage caused by bladder calculi rather than to administration of biphenyl. The hyperplasia can be therefore categorized as a pre-neoplastic lesion with possible progression into bladder tumors.

The sustained mechanical damage caused by the bladder calculi plays an important role in tumorigenesis. The physical characteristics of the bladder calculi such as size and shape can be considered to affect the degree and/or frequency of pre-neoplastic and neoplastic lesions occurring in response to mechanical damage to the bladder epithelium caused by calculi.

The authors indicated that a sex-difference in the metabolic pathways of biphenyl plays an important role in the formation of bladder calculi and the development of bladder tumors.

Applicant's summary and conclusion

Conclusions:
Mortality was significantly increased in males dosed with 4500 ppm over two years. In addition to mortality, animals in this dose group also exhibited statistically significant decreases in body weight, increases in urinary pH and presence of occult blood in the urine. Males dosed with 4500 ppm also exhibited significant increases in non-neoplastic and neoplastic lesions of the bladder and urinary system.
In females statistically significant (and dose-dependent) increases of simple transitional cell hyperplasia of renal pelvis and deposit of hemosiderin was observed at levels at and above 1500 ppm.