Amines, polyethylenepoly-, triethylenetetramine fraction

Carcinogenicity

Description of key information

Amines, polyethylenepoly-, triethylenetetramine fraction was not locally or systemically carcinogenic when applied to the skin of mice up to a level of 5%.

Key value for chemical safety assessment

Carcinogenicity: via oral route

Endpoint conclusion

Endpoint conclusion:

no study available

Carcinogenicity: via inhalation route

Endpoint conclusion

Endpoint conclusion:

no study available

Carcinogenicity: via dermal route

Link to relevant study records

Reference

Reference 1

Endpoint:

carcinogenicity: dermal

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

January 27, 1982 - January 30, 1984

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

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

Remarks:

Only male animals were used, no details on exposure and a dose can only be estimated, limited investigations and reporting.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 451 (Carcinogenicity Studies)

Deviations:

yes

Remarks:

only males tested; animals were treated 3 times a week; no details on exposure are given; dose can only be estimated; food consumption and food efficiency were not measured

Principles of method if other than guideline:

In the case of dermal administration, animals are normally treated with the test substance for at least 6 hours per day, 7 days per week. In this study animals were treated 3 times a week. However workers may also be exposed 3 times/week on the skin. This exposure route and frequency is appriopriate.
- Mice that died during the first 2 months on study and were replaced with extra mice from the original shipment.

GLP compliance:

yes

Species:

mouse

Strain:

C3H

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Jackson Laboratories, Bar Harbor, ME
- Age at study initiation: 4 weeks
- Weight at study initiation: 22.1-22.4 avarage for the three groups
- Fasting period before study: not applicable
- Housing: single housing in stainless steel wire-bottom suspended cages.
- Diet (e.g. ad libitum): well water, ad libitum
- Water (e.g. ad libitum): Purina Certified Rodent Chow #5002, ad libitum
- Acclimation period: at least 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22.2 ± 2 (72 ± 3 F)
- Humidity (%): 40-60
- Air changes (per hr): 6
- Photoperiod (hrs dark / hrs light): 12/12


IN-LIFE DATES: January 27, 1982 - January 30, 1984

Route of administration:

dermal

Vehicle:

ethanol

Details on exposure:

TEST SITE
- Area of exposure: back
- % coverage: no data
- Type of wrap if used: no data
- Time intervals for shavings or clipplings: The hair was clipped from the application site on an "as needed" basis, generally at 3-week intervals.

REMOVAL OF TEST SUBSTANCE
- Washing (if done): no data
- Time after start of exposure: no data

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 25 µl
- Concentration (if solution): 0, 0.2, 2%
- Constant volume or concentration used: no data

VEHICLE
- Justification for use and choice of vehicle (if other than water): Ethanol was chosen as the application vehicle after preliminary studies indicated that aqueous solutions did not produce acceptable wetting of the test area, and the test material did not appear to be stable in acetone.
- Amount(s) applied (volume or weight with unit): no data
- Concentration (if solution): no data
- Lot/batch no. (if required): no data
- Purity: no data


USE OF RESTRAINERS FOR PREVENTING INGESTION: yes/no

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Solutions of the test material in ethanol (w/w %) were prepared at approximately 8-12 week intervals. (Analyses of selected samples indicated no decrease in test material concentration over this time interval.) Observed concentrations were generally equal to or higher than the targeted dose level concentrations indicative of some evaporation
of the ethanol vehicle.

Duration of treatment / exposure:

104 weeks

Frequency of treatment:

3 times a week

Post exposure period:

none

Dose / conc.:

0.2 other: %

Remarks:

Basis: nominal in vehicle

Dose / conc.:

2 other: %

Remarks:

Basis: nominal in vehicle

No. of animals per sex per dose:

50 males

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: range fining study
- Rationale for animal assignment (if not random): random
- Section schedule rationale (if not random): no data

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: No

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: All mice were observed at least daily for signs of toxicity and for mortality. 3 times per week all mice were examined for changes in demeanor, general appearance, and evidence of cutaneous alterations at the application site. At approximately monthly intervals, the mice were palpated for cutaneous growths or other masses.

DERMAL IRRITATION (if dermal study): No

BODY WEIGHT: Yes
- Time schedule for examinations: All mice were weighed at 2-week intervals for the first 3 months and monthly thereafter until termination.

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

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: No

CLINICAL CHEMISTRY: No

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes

Other examinations:

None

Statistics:

see below

Clinical signs:

effects observed, treatment-related

Dermal irritation (if dermal study):

no effects observed

Mortality:

mortality observed, treatment-related

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Gross pathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Histopathological findings: neoplastic:

effects observed, treatment-related

Details on results:

CLINICAL SIGNS AND MORTALITY
There were no statistically identified differences in the mortality pattern of either treatment group relative to the control group. Five mice died during the first two months and were replaced by extra mice (3 control and 2 low dose rnice)(These animals are not included in the cumulative mortality table or in any subsequent data tables.)
The major apparent causes of death were liver tumors and urogenital tract infections, and these occurred in approximately equal percentages in all experimental groups. The remaining causes of death were varied, and there was no particular trend or clustering of specific causes that would suggest an association with administration of the test ma terial.
Although the incidence of urogential tract infections may be slightly higher in treated groups than controls, the condition is relatively common in male mice and is interpreted as being related to hygiene rather than to application of the test material.

BODY WEIGHT AND WEIGHT GAIN
No effects

GROSS PATHOLOGY
No treatment related effects

HISTOPATHOLOGY: NON-NEOPLASTIC
A few non-metastatic neoplasms of low grade malignancy were observed at other cutaneous locations in both treated and control mice. These were most common on the pinna of the ear, probably associated with chronic irritation by the metal ear tag. One fibrosarcoma was located on the tail of a high dose animal. None of these tumors was considered related to treatment.

Relevance of carcinogenic effects / potential:

No substance related tumors were obeserved after 2 year dermal application up to 2%.

Dose descriptor:

NOAEL

Effect level:

>= 20 mg/kg bw/day

Based on:

test mat.

Remarks:

corresponds to 2%

Sex:

male

Basis for effect level:

other: no adverse effects observed at highest dose tested

Critical effects observed:

no

Cummulative mortality

Note: mortality rate is very high towards the last 6 months of the study. With this strain of mice it would have been more appropriate to let the study run for 18 months instead of 24 months. 

Dose %	0	0.2	2.0
Number of mice/group	50	50	50
Months on test
1	0	0	0
2	0	0	0
3	0	0	1
4	0	0	2
5	0	0	2
6	0	0	2
7	1	1	3
8	1	2	3
9	2	2	3
10	2	2	3
11	2	2	4
12	2	3	4
13	2	3	5
14	2	4	5
15	2	4	5
16	2	6	6
17	4	8	8
18	7	8	10
19	11	8	10
20	13	10	17
21	14	14	18
22	16	23	20
23	21	29	21
24	27	32	26

Conclusions:

The test substance is not carcinogenic after 2-year dermal application up to 2%.

Executive summary:

The test substance was applied to the skin of male C3H/HeJ mice (50/group) at concentrations of 0, 0.2, or 2% (w/w) in ethanol, 3 times/week, for up to 2 years. The parameters evaluated during the study included mortality, clinical observations and palpable masses, body weights, gross pathology, and histopathologic examinations of complete sets of tissues from all mice. Dermal application of the tes substance was not associated with any increased incidence of neoplasms of the skin or internal organ systems, nor with any systemic organ toxicity . The treatment regimen had no adverse effects on survivability or any other study parameter evaluated. The primary causes of mortality among all groups, including controls, were liver tumors and urogenital tract infections. A few malignant cutaneous tumors were noted in both control and treated groups, however, none were located at the site of application of the test material. Four of the five observed cutaneous tumors were on the ear associated with the metal ear tag, and one fibrosarcoma was present on the tail of a high dose mouse. None o f these tumors was interpreted as related to dermal administration of the test substance.

Reference 2

Endpoint:

carcinogenicity: dermal

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

January 31, 1979 - May 23, 1981

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

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

Remarks:

Only male animals were used, limited investigations and reporting. An exact dose can only be estimated.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 451 (Carcinogenicity Studies)

Deviations:

yes

Remarks:

only males used; only one dose used; animals were treated treated 3 times a week and continuously; food consumption and food efficiency were not measured.

Principles of method if other than guideline:

In the case of dermal administration, animals are normally treated with the test substance for at least 6 hours per day, 7 days per week. In this study animals were treated 3 times a week. However workers may also be exposed 3 times/week on the skin. This exposure route and frequency is appropriate.

GLP compliance:

no

Remarks:

inhouse QA in place

Species:

mouse

Strain:

C3H

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Jackson Laboratories, Bar Harbor, ME
- Age at study initiation: 46-77 days
- Weight at study initiation: The mice assigned to the TETA group had body weights from 17.6 t o 26.6 grams and the deionized water group weighed from 17.2 t o 27.8 grams on the day of randomization.
- Fasting period before study: not applicable
- Housing: no data
- Diet (e.g. ad libitum): no data
- Water (e.g. ad libitum): no data
- Acclimation period: ± 2 weeks to 1 month


ENVIRONMENTAL CONDITIONS
- Temperature (°C): no data
- Humidity (%): no data
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): no data


IN-LIFE DATES: January 31, 1979 - May 23, 1981

Route of administration:

dermal

Vehicle:

water

Details on exposure:

TEST SITE
- Area of exposure: back
- % coverage: no data
- Type of wrap if used: no data
- Time intervals for shavings or clipplings: On Tuesday or Thursday of each week, the fur was clipped from the back of each mouse.

REMOVAL OF TEST SUBSTANCE
- Washing (if done): no data
- Time after start of exposure: no data

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 25 µl
- Concentration (if solution): 5%
- Constant volume or concentration used: yes, concentration

VEHICLE
- Justification for use and choice of vehicle (if other than water): water

USE OF RESTRAINERS FOR PREVENTING INGESTION: no

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The reports of analyses performed on the parent sample at 6, 18 and 24 months of dosing have not yet been received. The stability analyses of diluted test substance using NMR spectroscopy and titration analysis showed that the test substance in water was stable for at least one month. Monthly samples of the diluted test substance used for dosing were sent to UCC, South Charleston, WV, for titration and NMR analyses to determine that concentrations used for dosing were accurate. Reports received for these analyses indicated slight dilution differences in the samples over the dosing period. Titration analysis showed values ranging from 4.29 to 7.06 wt % with a mean value of 5.39 + 0.45 wt %. The solvent for this study was water (CAS #7732-18-5) that was deionized in a Milli-Q reagent-grade water system (#zo2011574, Millipore Corp., Bedford, MA)

Duration of treatment / exposure:

Lifetime

Frequency of treatment:

Mice were treated three times weekly, following a Monday, Wednesday, and Friday treatment schedule.

Post exposure period:

None

Dose / conc.:

5 other: %

Remarks:

Basis:
nominal in vehicle

No. of animals per sex per dose:

50 males

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: range finding: highest dose which is non-irritant and non-toxic
- Rationale for animal assignment (if not random): random
- Rationale for selecting satellite groups: not provided
- Section schedule rationale (if not random): no data

Positive control:

The ability of the C3H/HeJ mice to produce dermal carcinomas under similar experimental conditions was verified with mice treated with 3-methylcholanthrene in the laboratory.

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: No

DETAILED CLINICAL OBSERVATIONS: No
Mice were observed daily for mortality

DERMAL IRRITATION (if dermal study): Yes
- Time schedule for examinations: Monthly

BODY WEIGHT: No

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

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: No

CLINICAL CHEMISTRY: No

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes
Necropsy included the careful examination of the skin and body cavities, and the recording of observations. All suspect tumors and the dorsal skin of all mice, with or without tumors, were fixed in 10% neutral buffered formalin (NBF). In addition, all livers, kidneys and lungs, unless autolyzed, were fixed in NBF for possible his topathologic examination. Sections were prepared from the dorsal skin of all mice and any suspect internal tumors. Histopathologic examinations were performed.

Other examinations:

none

Statistics:

Mortality incidences were assessed by the product - limit method (Kaplan and Meier, 1958). The Mantel-Cox and Breslow statistics were used for testing the equality of the survival curves (Mantel, 1966 ; Breslow, 1970).

Clinical signs:

no effects observed

Dermal irritation (if dermal study):

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

not examined

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

no effects observed

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Other effects:

not examined

Details on results:

In the group that received the test substance as a 5% (v/v) dilution in deionized water the mean survival time was not statistically different from that of the deionized water control group (627 versus 626 days, respectively) . No papillomas or carcinomas were observed on the skin of any mice in either the test substance treated group or the deionized water controls at the site of application. One papilloma was diagnosed on the upper lip of a test substance-treated mouse, but this finding was considered unrelated to the treatment because of its location. A sebaceous adenoma of the skin of the thorax was observed in a mouse in the deionized water control group. The last surviving mouse in the test substance-treated group died on May 25, 1981. The last mouse in the deionized water control group died on May 23, 1981.

Relevance of carcinogenic effects / potential:

No substance related tumors were obeserved after a lifetime dermal application of 5% (v/v) solution in deionized water until the death of the animals.

Dose descriptor:

NOAEL

Effect level:

>= 50 mg/kg bw/day (nominal)

Based on:

test mat.

Remarks:

corresponding to 5% test substance solution

Sex:

male

Basis for effect level:

clinical signs

gross pathology

histopathology: neoplastic

histopathology: non-neoplastic

mortality

Critical effects observed:

no

25 µL x 0.05 mg/µL= 1.25 mg mouse, or the NOAEL >= 50 mg/kg bw

Conclusions:

The results of the present study indicated that the test substance was not carcinogenic to the skin of C3H/HeJ male mice when applied as a 5% (v/v) solution in deionized water until the death of the animals.

Executive summary:

The dermal carcinogenic potential of the test substance was assessed by applying 25 µL of a 5% (v/v) solution in deionized water to the backs of 50 male C3H/HeJ mice. A negative control group was dosed with deionized water. Both applications were performed three times a week until the death of the animals. No treatment-related skin tumors were observed in the groups treated with the test substance or deionized water. A papilloma was observed on the upper lip of a test substance-treated mouse, but this was considered unrelated to treatment because of its location. One negative control mouse had a sebaceous adenoma of the skin of the thorax. No statistically significant difference in mortality rates was observed between the treated group and the water control group. The test substance was not locally carcinogenic when applied to the skin of C3H/HeJ mice.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

50 mg/kg bw/day

Study duration:

chronic

Species:

mouse

Quality of whole database:

Similar to OECD 451 with acceptable restrictions.

Justification for classification or non-classification

Reliable data with the registration substance does not meet the criteria for classification according to Regulation (EC) No. 1272/2008, and the available data are therefore conclusive but not sufficient for classification.

Additional information

Two reliable studies similar to OECD guideline 451 are available with Amines, polyethylenepoly-, triethylenetetramine fraction.

Amines, polyethylenepoly-, triethylenetetramine fraction was applied to the skin of male C3H/HeJ mice (50/group) at concentrations of 0, 0.2, or 2% (w/w) in ethanol, 3 times/week, for up to 2 years (Young, 1986). The parameters evaluated during the study included mortality, clinical observations and palpable masses, body weights, gross pathology, and histopathologic examinations of complete sets of tissues from all mice. Dermal application of the test substance was not associated with any increased incidence of neoplasms of the skin or internal organ systems, nor with any systemic organ toxicity. The treatment regimen had no adverse effects on survivability or any other study parameter evaluated. The primary causes of mortality among all groups, including controls, were liver tumors and urogenital tract infections. A few malignant cutaneous tumors were noted in both control and treated groups, however, none were located at the site of application of the test material. Four of the five observed cutaneous tumors were on the ear associated with the metal ear tag, and one fibrosarcoma was present on the tail of a high dose mouse. None of these tumors was interpreted as related to dermal administration of the test substance.

In a second study, the dermal carcinogenic potential of Amines, polyethylenepoly-, triethylenetetramine fraction was assessed by applying 25 µL of a 5% (v/v) solution in deionized water to the backs of 50 male C3H/HeJ mice (Guzzie, 1982). A negative control group was dosed with deionized water. Both applications were performed three times a week until the death of the animals. No treatment-related skin tumors were observed in the groups treated with the test substance or deionized water. A papilloma was observed on the upper lip of a test substance-treated mouse, but this was considered unrelated to treatment because of its location. One negative control mouse had a sebaceous adenoma of the skin of the thorax. No statistically significant difference in mortality rates was observed between the treated group and the water control group. Amines, polyethylenepoly-, triethylenetetramine fraction was not locally carcinogenic when applied to the skin of C3H/HeJ mice.

In conclusion, Amines, polyethylenepoly-, triethylenetetramine fraction was not locally or systemically carcinogenic when applied to the skin of mice up to a level of 5%. This concentration can be recalculated to about 50 mg/kg bw/day assuming a mean mice weight of 25 g.