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Toxicological information

Carcinogenicity

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Description of key information

No study is available for MIPN. Bis(isopropyl)naphthalene (DIPN), a closely structure-related analogue, is used to make read-across: DIPN was not carcinogenic in a 24-months feeding study in rats.

Key value for chemical safety assessment

Carcinogenicity: via oral route

Link to relevant study records
Reference
Endpoint:
carcinogenicity: oral
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Reliability:
3 (not reliable)
Rationale for reliability incl. deficiencies:
other: Comparable to guideline study with acceptable restrictions, but guideline validity criteria for a negative test are not met.
Reason / purpose:
reference to same study
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 453 (Combined Chronic Toxicity / Carcinogenicity Studies)
Deviations:
yes
Remarks:
: group sizes for interim kill low; interference with pulmonary disease/pneumonia associated with poor survival with progressing age
Principles of method if other than guideline:
Combined chronic toxicity carcinogenicity study; no guideline stated, but largely in compliance with OECD guideline 453
GLP compliance:
no
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Shizuoka Experimental Animal Co.
- Age at study initiation: 5 weeks
- Weight at study initiation: males 100 - 146 g, females 74 - 117 g
- Housing: two per cage
- Diet (e.g. ad libitum): CE-2 from Japan Pelleted Feed Co., made available from feeding baskets, ad libitum
- Water (e.g. ad libitum): from automatic drinking aparatus, ad libitum
- Acclimation period: 1 week

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 - 25
Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
Pellets containing DIPN were prepared using CE-2 feed

DIET PREPARATION
- Rate of preparation of diet (frequency): every six months
- Storage temperature of food: no data
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
no further information reported
Duration of treatment / exposure:
24 months
Frequency of treatment:
continuous
Post exposure period:
no
Remarks:
Doses / Concentrations:
0, 96, 240, 600, 1500 mg/kg diet [= ca. 0, 5, 12, 30, 85 mg/(kg*d)]
Basis:

No. of animals per sex per dose:
60 (included are 13 animals for interim sacrifice at 6, 12, and 18 months - 3, 5, and 5 animals, respectively)
Control animals:
yes, concurrent no treatment
Positive control:
not required
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: daily

BODY WEIGHT: Yes
- Time schedule for examinations: once every week until week 27; thereafter once biweekly

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption: weekly determinations, calculation of mean daily food and test substance intake over 24 months (g food/day/rat and mg/kg body weight/day respectively): Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No data

FOOD EFFICIENCY: No

WATER CONSUMPTIO: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at 6, 12, 18, and 24 months
- Anaesthetic used for blood collection: no data, blood samples were collected from the coccygeal vein.
- Animals fasted: no data
- How many animals: all surviving animals
- Parameter sexamined: erythrocyte count (RBC), leucocyte count (WBC), hematocrit (Ht), hemoglobin (Hb)

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at 6, 12, 18, and 24 months
- Animals fasted: no data
- How many animals: 3, 5, 5, and all surviving animals; total blood was collected by incission of the carotid artery.
- Parameters examined: blood sugar (BS), total cholesterol (Total chol.), total protein (T.p), albumin/globulin ratio (A/G), bilirubin, urea nitrogen (BUN), transaminases (GOT, GPT), alkaline phosphatase (AlP), lactic acid dehydrogenase (LDH), leucine aminopeptidase

URINALYSIS: Yes
- Time schedule for collection of urine: at 6, 12, 18, and 24 months
- Metabolism cages used for collection of urine: no data
- Animals fasted: no data
- Parameters examined: protein, sugar, pH, and occult blood

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
On rats sacrificed and those that died or became moribund during the experimental period, an autopsy was performed (sacrifice of 3, 5, 5, and all surviving animals at 6, 12, 18, and 24 months, respectively). Each organ was examined macroscopically for pathological changes.

HISTOPATHOLOGY: Yes
Histological specimens were collected from organs and tissues of rats sacrificed at 24 months and of those which died or became moribund and were sacrificed during the experimental period. They were imbedded in paraffin, cut into sections, stained with hematoxylin and eosin, and then observed microscopically.

Organs and tissues examined: liver, kidney, spleen, brain, lung, heart, adrenal gland, thymus, testis or ovary, esophagus, stomach, duodenum, large intestine, thyroid gland, pancreas, submaxillary gland, hypophysis, urinary bladder, mesenteric lymphatic node, skeletal muscle, bone marrow (of femur), eyeball, palpebral skin, skin of the back, inferior vena cava, and other organs (e.g., mammary gland and uterus) affected with tumor.
Statistics:
Means and statistical significance (p-values) were calculated. Method not reported.
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):
no effects observed
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
effects observed, treatment-related
Urinalysis findings:
no effects observed
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed
Details on results:
CLINICAL SIGNS AND MORTALITY
Clinical signs started to became manifest 6 months after the beginning of treatment. At about 9 months, some rats exhibited rhinorrhea, epistaxis, lacrimation , palpebral seborrhea, pilar erection, anorexia, and symptoms suggesting pneumonia (decrease in voluntary locomotion, loss of pilar luster, rough hair coat, hyposthenia). These signs were observed throughout the study period. But there was no difference between the control group and the treatments groups on general symptoms, occurrence of pneumonia, and loss of pilar luster.

Mortality increased with age and was partly combined with symptoms of pneumonia. The increase in mortality at later time points was higher in treated animals compared to controls with lowest survival rates in the highest dose group (group IV: 38.3 and 21.3% survival at 24 months for males and females, respectively, controls 72.3 and 53.2%). At 18 months, survival was still distinctly of the order of 85 - 90% for males and 81 - 87% for females, i.e. most deaths occurred at high age, mainly due to increased frequency of pneumonia and nephromegaly occurring in all groups. There was a dose-related increase in mortality, which became evident not until 18 months of treatment (Tab. 1, Fig. A-1 and A-2). At 24 months, the mortality was just below 50% in the three lower dose groups (survival 43 to 49%), however, in the highest dose group clearly higher than 50 % (see above).

BODY WEIGHT AND WEIGHT GAIN
Body weight gain was not appreciably affected in surviving animals until 18 months, but a trend was seen already after 12 months in the group IV females (94% of control, Tab. 2, Fig. A-3). After 18 months, the body weight gain in group IV females was 91% (p >0.01). At 24 months, mean body weight gain for group III and group IV females had decreased to 83% and 75% respectively (p <0.01). At this time point, group III and IV males also showed a decreased body weight gain of 94% and 87% of controls, respectively (p <0.01).

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study)
No significant difference in food intake was observed between the control and treatment groups. Administration of DIPN exerted no influence on food consumption.

Food intake as mg/kg bw/day decreased over time due to the increasing weight of the test animals. Calculated over the total exposure period, mean compound intake was 0, 5.1, 12.6, 31.6, and 79.2 mg/kg bw/day for males and 0, 6.1, 13.8, 36.3, and 91.9 mg/kg bw/day for females (controls, group I, II, III, and IV respectively).

HAEMATOLOGY
No significant differences in any test parameter at 6, 12, 18, and 24 months were observed between control and treatment groups.


CLINICAL CHEMISTRY
At 6, 12, and 18 months of the study, no significant differences were noticed in the clinical chemistry findings. At 24 months, GOT was significantly increased in the group III (p <0.05) and group IV (p < 0.01) males (+64% and +130%) and females (+26% and +95%). GPT values were significantly higher as well but only in the group IV males (+50%, p <0.05).

URINALYSIS
Some abnormalities in the values of sugar and occult blood were found infrequently. However, no dose response was observed indicating that this effect was not caused by the administration of DIPN, but seemed to be spontaneous.

ORGAN WEIGHTS
At 6, 12, and 18 months of the study, some variance in absolute organ weights were observed. However for relative organ weights (ratio organ weight to body weight), no significant differences between controls and treatment groups were noticed.

At 24 months, the absolute organ weights of treatment groups were not significantly different from controls with the exception of the liver weight of the group IV females (+14%, p <0.05). But significant differences were seen in relative organ weights for the livers and kidneys of group IV male and females (liver: males +50%, p <0.05, females +23%, p <0.01; kidney: males/females +17%, p <0.01). These results are related to the decreased body weights of group III and group IV animals which are presumed to have been brought about by the effect of DIPN administration.

For spleen, apparent mean relative weight increases (group II and III, both sexes) and an apparent mean weight decrease (group IV, males) could be derived. However, obviously due to individual variability, the variance was very high. No statistical significance was achieved (Tab. 6).

GROSS PATHOLOGY
Up to 18 months, increasing incidences of pneumonic changes and abscess formation in the lung, yellow or white area in the testis, and age involution were the predominant changes. Particular changes were rarely seen, and there was little difference in the frequency of occurrence of such changes between any experimental group and the control group (only 5 animals examined).

At 24 months, predominant changes were caused by age involution, and there were no particular changes in any experimental group (sacrificed animals). However, pulmonary changes were outstanding in every group. Nodules, considered to be tumors, were recognized in every group. There was, however, no difference in the frequency of occurrence of these nodules between any experimental group and the control group. It was impossible to find gross pathological changes which could be assumed to have been induced by administration of DIPN.

In rats which died during the experimental period or were sacrificed in a moribund stage, predominant changes were noted as enlargement of the liver, kidney, and spleen. Age involution of the testis, pneumonic changes, and abscess formation were also observed. Hematomegaly and nephromegaly were specific changes, but no distinct dose-relationship was observed.

The same effects were seen in sacrificed survivors at 24 months, but overall, incidences in liver and kidney were somewhat lower. In contrast, apparently higher incidences were observed for changes of the lung.

Hypertrophy of the liver and kidneys is considered as related to treatment, yet no clear dose-response relation is evident. Hypertrophic phenomena for these organs are accompanied by a significant mean increase in the relative organ weight (see above). For the spleen, variable relative organ weights were observed. But no morphologically significant effects are addressed.

Gross pathology findings for tumor incidence are displayed in the following table:
control groups: 1 male and 4 females
Group I: 2 males and 1 female
Group II: 1 male and 1 female
Group III: 1 male and 2 females
Group IV: 0 male and 2 females.


HISTOPATHOLOGY: NON-NEOPLASTIC
Due to post mortem changes, 34 rats could not be inspected histopathologically. All other animals that died or were sacrificed were examined.

Five main organs were identified with an elevated incidence of histopathological alterations: liver, kidney, lungs, heart, and testis. Focal necrosis and fatty degeneration were important morphological indicators of hepatic disorder, as well as congestion of the kidney indicated kidney hypertrophy. Other changes observed in kidney, lung, heart and testis (amongst others, dilatation of uriniferous tubules, fibrosis of cardiac muscle, and atrophy and fibrosis of some other organs) were assumed to have been induced by age involution.

There was no microscopic evidence of any substance-related lesion in the spleen (Tab. A-27). Therefore, the high variability of the spleen sizes appeared to be of no toxicological relevance.

Incidences of congestion of the liver and spleen, hyperplasia of bile ducts, partial fibrosis of cardiac muscle and the formation of protein cylinders in uriniferous tubules of the kidney indicated a sex difference (higher incidences in males). In addition, there was a difference in the incidence of pulmonary congestion and suppurative pneumonopathy between surviving and dead rats. Degeneration of the liver also seemed to be slightly more frequent in dead rats than in surviving ones.

But overall, there was no difference in the incidence of any disorder between control groups and the treatment groups. Histopathology did not reveal any morphological abnormalities in any organ that could be related to DIPN administration.


HISTOPATHOLOGY: NEOPLASTIC
Most of the tumors were mammary tumors, fibrosarcomas, keratoacanthomas and inflammatory tumors.

Excluding testicular Leydig cell tumors, the overall tumor incidence for all rats was 18.9 % (p. 15). It was highest ( 26.4%) in the control group, but less than 20 % in any treated group. Frequency of malignant tumors overall was 6.9 %, highest (10.3%) in group II versus 7.7% in the control, and 5.7% or less in all other groups.

The overall rate of leukemia was 4.6%. In the control group it was 4.4 % compared with slightly lower and higher incidences in the treated groups (group I 3.4%, group II 6.9%, group III 4.6%, and group IV 3.6%).

Extremely high was the frequency of Leydig-cell tumors in male rats, overall 87.5%, in the control 87.2, and 81.8 - 91.3% in the experimental groups.
(Note according to authors: The spontaneous, age-related frequency of this kind of tumor was reportedly common and very high in ageing male rats of this Wistar strain at that time in Japan (reference 2: Taniguchi et al., 1977) (reference 3: Nishibe et al., 1974]

Overall, there was no significant difference in the incidence of tumors between the control and experimental groups. The incidence of both benignant and malignant types of tumors was even higher in the control than in some experimental groups. Therefore it is assumed that DIPN is free from carcinogenicity.
Relevance of carcinogenic effects / potential:
DIPN did not exhibit any carcinogenic potential even at the highest dose tested (1500 ppm (nomimal), 79.2 mg/kg bw/day males, 91.9 mg/kg bw/day females). Due to high mortality at 24 months, the guideline criterion for a test result to be accepted as negative was not met.
Dose descriptor:
NOEC
Effect level:
1 500 ppm (nominal)
Sex:
male/female
Basis for effect level:
other: incidence of tumours / highest concentration tested in diet = approx. 79 and 92 mg/(kg bw*d) in male and female rats, respectively
Remarks on result:
other: Effect type: carcinogenicity (migrated information)
Conclusions:
In this study, DIPN did not show any carcinogenic potential up to the highest dose tested. Due to pneumonia, survival rates especially at 24 months were poor. The authors concluded that this issue did not limit the reliability of the study results.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Study duration:
chronic
Species:
rat

Additional information

Justification for selection of carcinogenicity via oral route endpoint:
acceptable for assessment, read-across to a chemical structure (bis(isopropyl)naphthalene) closely related to that of the target substance; no effects seen at the highest dose tested (approx. 79 and 92 mg/kg bw/day in male and female rats, respectively).

Justification for classification or non-classification

Not found carcinogenic, classification not warranted.