m-bis(2,3-epoxypropoxy)benzene

Carcinogenicity

Administrative data

Endpoint:

carcinogenicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

The mice studies were started in April 1980 and completed in April 1982.

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1986

Materials and methods

Test guideline

Qualifier:

no guideline available

Principles of method if other than guideline:

Toxicology and carcinogenesis studies of technical grade diglycidyl resorcinol ether (81%pure) were conducted by administering the chemical in corn oil by gavage to groups of 50 male and 50 female F344/Nrats at doses of 25 or 50 mg/kg. A supplemental study of similar design in male and female rats (0 or 12 mg/kg)was started approximately 12 months later because of high mortality in the 50 mg/kg dose groups. Doses were administered five times per week for 103 weeks. Groups of 50 rats of each sex received corn oil by gavage on the same dosing schedule and served as vehicle controls.
All animals were observed twice daily for signs of morbidity or mortality. Clinical signs were recorded monthly. Body weights by cage were recorded every week for the first 12 weeks and monthly thereafter. The mean body weight of each group was calculated by dividing the total weight of all animals in the group by the number of surviving animals in the group. Moribund animals and animals that survived to the end of the study were killed with carbon dioxide and necropsies were performed. Examinations for grossly visible lesions were performed on major tissues or organs. Tissues were preserved in 10% neutral buffered formalin, embedded in paraffin, sectioned, and stained with hematoxylin and eosin. The following were examined microscopically: tissue masses, abnormal lymph nodes, skin, mandibular or mesenteric lymph nodes, mammary gland, salivary gland, sternebrae, bone marrow, thymus, larynx, trachea, lungs and bronchi, heart, thyroid, parathyroid, esophagus, stomach, small intestine, colon, liver, gallbladder (mice), pancreas, spleen, kidneys, adrenals, urinary bladder, prostate/ testes or ovaries/uterus, brain, and pituitary. Necropsies were performed on all animals unless precluded in whole or in part by autolysis or cannibalization.
The classification of neoplastic nodules in the liver was done according to the recommendations of Squire and Levitt (1975) and the National Academy of Sciences (1980).
When the pathology examination was completed, the slides, individual animal data records, and summary tables were sent to an independent quality assurance laboratory. Individual animal records and tables were compared for accuracy, slides and tissues were verified, and histotechniques were evaluated. All tumor diagnoses, target tissues, and tissues from a randomly selected 10% of the animals were evaluated by a pathologist. Slides of all target tissues, neoplasms, and other slides about which the original and quality assurance pathologists disagreed were submitted to the Chairperson of the Pathology Working Group (PWG) for evaluation. Representative slides selected by the Chairperson were reviewed blindly by PWG pathologists, who reached a consensus and compared their findings with the original diagnoses. When disagreements were found, the PWG sent the appropriate slides and their comments to the original pathologist for review. The final diagnosis represents a consensus of contractor pathologists and the NTP Pathology Working Group.
Data on this experiment were recorded in the Carcinogenesis Bioassay Data System (Linhart et al., 1974). The data elements include descriptive information on the chemicals, animals, experimental design, clinical observations, survival, body weight, and individual pathologic results, as recommended by the lnternational Union Against Cancer (Berenblum, 1969).

GLP compliance:

no

Test material

Test material information

Constituent 1

Reference substance name:

m-bis(2,3-epoxypropoxy)benzene

EC Number:

202-987-5

EC Name:

m-bis(2,3-epoxypropoxy)benzene

Cas Number:

101-90-6

Molecular formula:

C12H14O4

IUPAC Name:

2-({3-[(oxiran-2-yl)methoxy]phenoxy}methyl)oxirane

Test material form:

liquid: viscous

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material:
Ciba-Geigy Corporation (Ardsley, NY), Araldite ERE 1359 in a single lot (No. P-60002)
- Purity test date:
81.2%

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material:
in the dark at 23 °C in its original container
- Stability under test conditions:
stable

FORM AS APPLIED IN THE TEST: suspended in corn oil (dissolved in acetone before being added to corn oil)

Test animals

Species:

mouse

Strain:

B6C3F1

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source:
Charles River Breeding Laboratories, Portage, MI
- Age at study initiation:
8 to 9 weeks
- Weight at study initiation:
NA
- Fasting period before study:
NA
- Housing:
five animals/cage
- Diet:
ad libitum
- Water:
ad libitum
- Acclimation period:
3 weeks

DETAILS OF FOOD AND WATER QUALITY:
Feed: Wayne Laboratory Blox (r), Allied Mills (Chicago, IL)
Water: Edstrom automatic watering system, Edstrom Industries (Waterford, WI)

ENVIRONMENTAL CONDITIONS
- Temperature (°C):
17-32
- Humidity (%):
20-81
- Air changes (per hr):
12 changes of room air per hour
- Photoperiod (hrs dark / hrs light):
12/12

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on exposure:

Using the observations and results from the 13-week study, groups of 50 mice of each sex were administered diglycidyl resorcinol ether in corn oil by gavage 5 days per week for 103 weeks at doses of 0,50, or 100 mg/ kg body weight.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

One-milliliter aliquots of the sample vials were extracted with 10 mL of methanol containing 0.7 g/mL of dibutyl phthalate as an internal standard. A reference calibration plot was prepared from spiked samples which were extracted in the same manner. The supernatant solutions were analyzed by VPC-FID at 210° on a 6 ft x 1/4 in x 2 mm ID glass column packed with SP2250 on 100/120 Supelcoport.

Duration of treatment / exposure:

103 weeks

Frequency of treatment:

Daily, 5 days per week

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/kg bw/day (actual dose received)

Doses / concentrations 2

Dose / conc.:

25 mg/kg bw/day (actual dose received)

Doses / concentrations 3

Dose / conc.:

50 mg/kg bw/day (actual dose received)

No. of animals per sex per dose:

50 Males and 50 Males

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: based on 13-week study observation and results
- Rationale for animal assignment (if not random): random

Examinations

Observations and examinations performed and frequency:

All animals were observed twice daily for signs of morbidity or mortality. Clinical signs were recorded monthly. Body weights by cage were recorded every week for the first 12 weeks and monthly thereafter. The mean body weight of each group was calculated by dividing the total weight of all animals in the group by the number of surviving animals in the group. Moribund animals and animals that survived to the end of the study were killed with carbon dioxide and necropsies were performed. Examinations for grossly visible lesions were performed on major tissues or organs. Tissues were preserved in 10% neutral buffered formalin, embedded in paraffin, sectioned, and stained with hematoxylin and eosin. The following were examined microscopically: tissue masses, abnormal lymph nodes, skin, mandibular or mesenteric lymph nodes, mammary gland, salivary gland, sternebrae, bone marrow, thymus, larynx, trachea, lungs and bronchi, heart, thyroid, parathyroid, esophagus, stomach, small intestine, colon, liver, gallbladder (mice), pancreas, spleen, kidneys, adrenals, urinary bladder, prostate/ testes or ovaries/ uterus, brain, and pituitary.

Sacrifice and pathology:

Necropsies were performed on all animals unless precluded in whole or in part by autolysis or cannibalization. Thus, the number of animals from which particular organs or tissues were examined microscopically varies and is not necessarily equal to the number of animals that were placed on study in each group. The classification of neoplastic nodules in the liver was done according to the recommendations of Squire and Levitt (1975) and the National Academy of Sciences (1980). When the pathology examination was completed, the slides, individual animal data records, and summary tables were sent to an independent quality assurance laboratory. Individual animal records and tables were compared for accuracy, slides and tissues were verified, and histotechniques were evaluated. All tumor diagnoses, target tissues, and tissues from a randomly selected 10% of the animals were evaluated by a pathologist. Slides of all target tissues, neoplasms, and other slides about which the original and quality assurance pathologists disagreed were submitted to the Chairperson of the Pathology Working Group (PWG) for evaluation. Representative slides selected by the Chairperson were reviewed blindly by PWG pathologists, who reached a consensus and compared their findings with the original diagnoses. When disagreements were found, the PWG sent the appropriate slides and their comments to the original pathologist for review. (This procedure has been described by Maronpot and Boorman, 1982.) The final diagnosis represents a consensus of contractor pathologists and the NTP Pathology Working Group.

Statistics:

Probabilities of survival were estimated by the product-limit procedure of Kaplan and Meier (1958).
Animals were statistically censored as of the time that they died of other than natural causes or were found to be missing; animals dying from natural causes were not statistically censored. Statistical analyses for a possible dose-related effect on survival used the method of Cox (1972) for testing two groups for equality and Tarone’s (1975) extensions of Cox’s methods for testing for a dose-related trend. All reported P values for the survival analyses are two-sided.
For the statistical analysis of tumor incidence data, two different methods of adjusting for intercurrent mortality were employed. Each used the classical method for combining contingency tables developed by Mantel and Haenszel (1959). Tests of significance included pairwise comparisons of high and low dose groups with controls and tests for overall dose-response trends.

Results and discussion

Results of examinations

Clinical signs:

no effects observed

Dermal irritation (if dermal study):

not examined

Mortality:

no mortality observed

Description (incidence):

No significant differences in survival were observed between the dosed and control groups.
In male mice, 30/50 (60%) of the controls, 26/50 (52%) of the low dose, and 34/50 (68%) of the high dose group lived to the end of the study (104-105 weeks). In female mice, 20/50 (40%) of the controls, 13/50 (26%) of the low dose, and lOj50 (20%) of the high dose group lived to the end of the study. The survival data include one control and one low dose male that died during the termination period of the study. For statistical purposes, these animals have been pooled with those killed at the end of the study. The major cause of death in dosed female mice was a necrosuppurative lesion of the ovary which spread to other areas of the abdominal cavity.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Mean body weights of high dose female mice were lower than those of the controls after week 20 of the study. Mean body weights of high and low dose male mice arid of low dose female mice were comparable with those of the controls. No compound-related clinical signs were observed.

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Ovary: The ovaries were enlarged and filled with a viscous yellow exudate in 17/50 control, 12/50 low dose, and 15/50 high dose females. Ovarian tissue was not macroscopically recognizable in many of these masses. Microscopically, a mantle of neutrophils, macrophages, and fibrosis surrounded the multiple abscesses. Extensive adhesions were present between the mass and the omentum. Neutrophils, lymphocytes, and plasma cells were present in the adjacent adipose tissue. Fibrinoid exudate was disseminated both in the abdominal and thoracic cavities. Overall, necrotizing inflammation was found in the abdominal cavity, ovary, uterus, or multiple organs in 18/30 vehicle control, 18/36 low dose, and 16/40 high dose females that died before the end of the study. Although microbiologic examinations were not performed on mice in this study, Klebsiella oxyroca has been isolated from mice that had similar lesions in other studies performed at the same laboratory.

Histopathological findings: neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Stomach: Hyperplastic and neoplastic lesions were observed at increased incidences in male and female mice. Squamous cell papillomas and carcinomas and papillomatosis occurred in male and female mice with statistically significant positive trends and the incidences in the high dose groups were significantly higher than those in the controls. The squamous cell papillomas were papillary growths of the epithelium and were supported by a narrow or broad fibrovascular stalk. They were covered by markedly thickened epithelium which was often heavily keratinized. Multiple lesions in the stomach of a single animal were referred to as papillomatosis. Squamous cell carcinomas were characterized by infiltrative growth into the submucosa and muscularis. The component cells varied in size and shape and many had indistinct margins. The cytoplasm was more eosinophilic than normal and, in some cells in the superficial layers, it contained keratohyalin granules. Nuclei were enlarged and contained coarse or stippled chromatin and one or two prominent nucleoli. Mitotic figures were present but not numerous. Keratin pearls were present in many of the carcinomas of different sizes. The lumina of the large pearls were filled with desquamated material, inflammatory cells, and necrotic debris. Nonkeratinizing squamous cell carcinomas were seen in the forestomach of a few mice. Areas of necrosis and hemorrhage were common in the large tumors. The morphology of the gastric neoplasms in mice was comparable to that obtained in rats. In 4 low dose males, 10 high dose males, 1 low dose female, and 9 high dose females, the squamous cell carcinomas of the stomach had disseminated onto the serosal surfaces of the abdominal cavity and in some mice had metastasized to the lung (most common site), liver, lymph nodes, spleen, adrenal glands, heart, and kidneys.
Liver: While pairwise comparisons were not significant (P>0.05), positive trends occurred in the incidences of female mice with hepatocellular carcinomas. The incidences in the high dose group were significantly higher by the life table test than those in the controls. The incidences of females with either adenomas or carcinomas had a significant positive trend; and the pairwise comparison between the high dose group and the controls was significant by the life table test.
Kidney: Mineralization was found in the kidneys of 8/50 control males, 18/50 low dose males, and 30/50 high dose males. The mineralization was minimal, and the distribution was multifocal, being located primarily in the cortex. The foci were small, ranging from the size of one or two renal tubular epithelial cells to the size of a tubule.
Hematopoietic System: Malignant lymphocytic lymphomas, malignant lymphomas of mixed types, and malignant lymphomas of all types occurred in female mice with negative dose-related trends. The incidence of all types of malignant lymphomas was significantly lower in the high dose group than in the controls.
Subcutaneous Tissue: Fibroma, fibrosarcoma, or sarcoma, NOS (combined) occurred in male mice with a statistically significant negative trend.

Effect levels

Key result

Dose descriptor:

LOAEL

Effect level:

50 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

histopathology: neoplastic

Applicant's summary and conclusion

Conclusions:

Under the conditions of the study, technical grade diglycidyl resorcinol ether (DGRE) caused hyperkeratosis and hyperplasia of the forestomach in mice. DGRE was carcinogenic for male and female mice, causing both benign and malingnant neoplasms of the forestomach.
Under the conditions of the study, the LOAEL may be assigned to 50 mg/kg bw/day dose. It is however not excluded, that the real LOAEL value may be lower as under given testing conditions. On the other hand, the chronic toxicity of DGRE via oral exposure is not relevant to humans. There was also an opinion, that the observed forestomach tumors were likely to have resulted from an indirect or local toxic effect of DGRE. Therefore by assessing the substance hazard for humans it is essential to identify the effects of toxicity from the other than oral exposure routes.

Executive summary:

Toxicology and carcinogenesis study of technical grade diglycidyl resorcinol ether (81% pure) was conducted by administering the chemical in corn oil by gavage to groups of 50 male and 50 female B6C3F1 mice at doses of 50 or 100 mg/kg. Doses were administered five times per week for 103 weeks. Groups of 50 mice of each sex received corn oil by gavage on the same dosing schedule and served as vehicle controls.

Throughout most of the primary study, mean body weights of high dose male and female mice were unaffected by administration of DGRE. Survival of dosed mice of each sex in the primary study was also mostly unaffected. The major cause of death in dosed female mice was a necrosuppurative lesion of the ovary which spread to other areas of the abdominal cavity. There was no significant difference in survival between dosed and control female mice in the supplemental study.

Hyperplastic and neoplastic lesions were observed at increased incidences in male and female mice. Squamous cell papillomas and carcinomas and papillomatosis occurred in male and female mice with statistically significant positive trends and the incidences in the high dose groups were significantly higher than those in the controls.

An audit of the experimental data was conducted for the 2-year studies of diglycidyl resorcinol ether. No data discrepancies were found that influenced the final interpretations.

The authors concluded, that under the condition of the 2-year gavage study, technical grade diglycidyl resorcinol ether caused hyperkeratosis and hyperplasia of the forestomach in mice. DGRE was carcinogenic for male and female B6C3F1 mice, causing both benign and malignant neoplasms of the forestomach.

Under the conditions of the study, the LOAEL may be assigned to 50 mg/kg bw/day dose. It is however not excluded, that the real LOAEL value may be lower as under given testing conditions. On the other hand, the chronic toxicity of DGRE via oral exposure is not relevant to humans. There was also an opinion, that the observed forestomach tumors were likely to have resulted from an indirect or local toxic effect of DGRE. Therefore, by assessing the substance hazard for humans is essential to identify the effects of toxicity from the other than oral exposure routes.