1978 

National Cancer Institute 

CARCINOGENESIS 

Technical Report Series 

No. 143 

BIOASSAY OF 

1,5-NAPHTHALENEDIAMINE 

FOR POSSIBLE CARCINOGENICITY 

CAS No. 2243-62-1 

NCI-CG-TR-143 

U.S. DEPARTMENT OF HEALTH, EDUCATION, AND WELFARE 

Public Health Service 

National Institutes of Health 

BIOASSAY OF
 

1,5-NAPHTHALENEDIAMINE
 

FOR POSSIBLE CARCINOGENICITY
 

Carcinogenesis Testing Program
 

Division of Cancer Cause and Prevention
 

National Cancer Institute
 

National Institutes of Health
 

Bethesda, Maryland 20014
 

U.S. DEPARTMENT OF HEALTH, EDUCATION, AND WELFARE
 

Public Health Service
 

National Institutes of Health
 

DHEW Publication No. (NIH) 78-1398
 

REPORT ON THE BIOASSAY OF 1,5-NAPHTHALENEDIAMINE
 

FOR POSSIBLE CARCINOGENICITY
 

CARCINOGENESIS TESTING PROGRAM
 

DIVISION OF CANCER CAUSE AND PREVENTION
 

NATIONAL CANCER INSTITUTE, NATIONAL INSTITUTES OF HEALTH
 

FOREWORD: This report presents the results of the bioassay of
 

1,5-naphthalenediamine conducted for the Carcinogenesis Testing Program, Division of Cancer Cause and Prevention, National Cancer Institute (NCI), National Institutes of Health, Bethesda, Maryland. This
 

is one of a series of experiments designed to determine whether selected chemicals have the capacity to produce cancer in animals. Negative results, in which the test animals do not have a significantly
 

greater incidence of cancer than control animals, do not necessarily
 

mean the test chemical is not a carcinogen because the experiments
 

are conducted under a limited set of circumstances. Positive results
 

demonstrate that the test chemical is carcinogenic for animals under
 

the conditions of the test and indicate a potential risk to man. The
 

actual determination of the risk to man from animal carcinogens requires a wider analysis.
 

CONTRIBUTORS: This bioassay of 1,5-naphthalenediamine was conducted
 

by Mason Research Institute, Worcester, Massachusetts, initially
 

under direct contract to the NCI and currently under a subcontract
 

to Tracor Jitco, Inc., prime contractor for the NCI Carcinogenesis
 

Testing Program.
 

The experimental design was determined by the NCI Project Officers, Dr. J. H. Weisburger (1,2) and Dr. E. K. Weisburger (1). The
 

principal investigators for the contract were Dr. E. Smith (3) and
 

Dr. A. Handler (3). Animal treatment and observation were supervised
 

by Mr. G. Wade (3) and Ms. E. Zepp (3). Chemical analysis was performed by Midwest Research Institute (4) and the analytical results
 

were reviewed by Dr. N. Zimmerman (5).
 

Histopathologic examinations were performed by Dr. A. S. Krishna
 

Murthy (3), Dr. A. Russfield (3) and Dr. D. S. Wyand (3) at the Mason
 

Research Institute, the pathology narratives were written by Dr. A.
 

Russfield (3) and Dr. D. S. Wyand (3), and the diagnoses included in
 

this report represent the interpretation of these pathologists. Histopathology findings and reports were reviewed by Dr. R. L. Schueler
 

(6).
 

Compilation of individual animal survival, pathology, and summary tables was performed by EG&G Mason Research Institute (7); the
 

iii
 
statistical analysis was performed by Mr. W. W. Belew (5,8) and Mr.
 

R. M. Helfand (5), using methods selected for the Carcinogenesis
 

Testing Program by Dr. J. J. Gart (9).
 

This report was prepared at METREK, a Division of The MITRE Corporation (5) under the direction of the NCI. Those responsible for
 

this report at METREK are the project coordinator, Dr. L. W. Thomas
 

(5), task leader Dr. M. R. Kornreich (5,10), senior biologist Ms. P.
 

Walker (5), biochemist Mr. S. C. Drill (5), and technical editor Ms.
 

P. A. Miller (5). The final report was reviewed by members of the
 

participating organizations.
 

The following other scientists at the National Cancer Institute
 

were responsible for evaluating the bioassay experiment, interpreting
 

the results, and reporting the findings: Dr. K. C. Chu (1), Dr. C.
 

Cueto, Jr. (1), Dr. J. F. Douglas (1), Dr. D. G. Goodman (1,10), Dr.
 

R. A. Griesemer (1), Dr. M. H. Levitt (1), Dr. H. A. Milman (1), Dr.
 

T. W. Orme (1), Dr. R. A. Squire (1,11), Dr. S. F. Stinson (1), Dr.
 

J. M. Ward (1), and Dr. C. E. Whitmire (1).
 

1.	 Carcinogenesis Testing Program, Division of Cancer Cause and
 

Prevention, National Cancer Institute, National Institutes of
 

Health, Bethesda, Maryland.
 

2.	 No wit w h the Naylor Dana Institute for Disease Prevention, American Health Foundation, Hammon House Road, Valhalla, New York.
 

3.	 Mason Research Institute, 57 Union Street, Worcester, Massachusetts.
 

4.	 Midwest Research Institute, 425 Volker Boulevard, Kansas City,
 

Missouri.
 

5.	 The MITRE Corporation, METREK Division, 1820 Dolley Madison
 

Boulevard, McLean, Virginia.
 

6.	 Tracer Jitco, Inc., 1776 East Jefferson Street, Rockville,
 

Maryland.
 

7.	 EG&G Mason Research Institute, 1530 East Jefferson Street,
 

Rockville, Maryland.
 

8.	 No wit w h the Solar Energy Research Institute, Cole Boulevard,
 

Golden, Colorado.
 

IV
 
9.	 Mathematical Statistics and Applied Mathematics Section, Biometry
 

Branch, Field Studies and Statistics Program, Division of Cancer
 

Cause and Prevention, National Cancer Institute, National Institutes of Health, Bethesda, Maryland.
 

10.	 No wit w h Clement Associates, Inc., 1010 Wisconsin Avenue, N.W.,
 

Washington, B.C.
 

11.	 Now with the Division of Comparative Medicine, Johns Hopkins
 

University, School of Medicine, Traylor Building, Baltimore,
 

Maryland.
 

SUMMARY
 

A bioassay of 1,5-naphthalenediamine for possible carcinogenicity
 

was conducted using Fischer 344 rats and B6C3F1 mice. 1,5-Naphthalenediamine was administered in the feed, at either of two concentrations,
 

to groups of 50 male and 50 female animals of each species. The high
 

and low dietary concentrations utilized in the chronic bioassay were,
 

respectively, 0.1 and 0.05 percent for rats and 0.2 and 0.1 percent
 

for mice. The compound was administered in the diet for 103 weeks,
 

followed by up to 4 weeks of observation. Fifty mice of each sex and
 

25 rats of each sex were placed on test as controls. These animals
 

were observed for up to 110 weeks.
 

There were no significant positive associations between the administered concentrations of 1,5-naphthalenediamine and mortality in
 

either sex of rats or mice. In all groups adequate numbers of animals
 

survived sufficiently long to be at risk from late-developing tumors.
 

Among dosed female rats, a statistically significant increase
 

in endometrial stromal polyps was observed. Several of these tumors
 

underwent malignant transformation to endometrial stromal sarcomas.
 

The incidence of female rats having either adenoma or carcinoma of
 

the clitoral gland was statistically significant. No neoplasms were
 

observed at significantly increased incidences in dosed male rats.
 

Based on lack of clinical signs or weight loss, the male rats may
 

have been able to withstand a higher dose.
 

In mice, dose-related increases in thyroid neoplasms were observed
 

in both sexes. The incidence of thyroid C-cell carcinomas was significant for high dose female mice. The combined incidences of papillary
 

adenomas, follicular-cell adenomas and papillary cystadenomas of the
 

thyroid were significant for mice of both sexes. The incidence of
 

hepatocellular carcinomas and the incidence of alveolar/bronchiolar
 

adenomas were each significant for dosed female mice.
 

Under the conditions of this bioassay, 1,5-naphthalenediamine was
 

carcinogenic in female Fischer 344 rats, causing clitoral and uterine
 

neoplasms. 1,5-Naphthalenediamine was also carcinogenic for B6C3F1
 

mice, producing thyroid neoplasms in males and neoplasms of the thyroid,
 

liver, and lung in females. Insufficient evidence was provided for the
 

carcinogenicity of the compound in male Fischer 344 rats.
 

VI1
 

TABLE OF CONTENTS
 

Page
 

I. INTRODUCTION 1
 

II. MATERIALS AND METHODS 4
 

A. Chemicals 4
 

B. Dietary Preparation 5
 

C. Animals 5
 

D. Animal Maintenance 6
 

E. Selection of Initial Concentrations 9
 

F. Experimental Design 10
 

G. Clinical and Histopathologic Examinations 13
 

H. Data Recording and Statistical Analyses 14
 

III. CHRONIC TESTING RESULTS: RATS 20
 

A. Body Weights and Clinical Observations 20
 

B. Survival 20
 

C. Pathology 23
 

D. Statistical Analyses of Results 26
 

IV. CHRONIC TESTING RESULTS: MICE 39
 

A. Body Weights and Clinical Observations 39
 

B. Survival 39
 

C. Pathology 42
 

D. Statistical Analyses of Results 44
 

V. DISCUSSION 54
 

VI. BIBLIOGRAPHY 56
 

APPENDIX A SUMMARY OF THE INCIDENCE OF NEOPLASMS IN
 

RATS TREATED WITH 1,5-NAPHTHALENEDIAMINE A-l
 

APPENDIX B SUMMARY OF THE INCIDENCE OF NEOPLASMS IN
 

MICE TREATED WITH 1,5-NAPHTHALENEDIAMINE B-l
 

APPENDIX C SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC
 

LESIONS IN RATS TREATED WITH 1,5-NAPHTHALENEDIAMINE C-l
 

APPENDIX D SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC
 

LESIONS IN MICE TREATED WITH 1,5-NAPHTHALENEDIAMINE D-l
 

ix
 
LIST OF ILLUSTRATIONS 

Figure Number Page 

1 CHEMICAL STRUCTURE OF 1,5-NAPHTHALENEDIAMINE 

2 

GROWTH CURVES FOR 1,5-NAPHTHALENEDIAMINE 

CHRONIC STUDY RATS 21 

SURVIVAL COMPARISONS OF 1,5-NAPHTHALENEDIAMINE CHRONIC STUDY RATS 22 

GROWTH CURVES FOR 1,5-NAPHTHALENEDIAMINE 

CHRONIC STUDY MICE 40 

SURVIVAL COMPARISONS OF 1,5-NAPHTHALENEDIAMINE CHRONIC STUDY MICE 41 

LIST OF TABLES 

Table Number Page 

DESIGN SUMMARY FOR FISCHER 344 RATS— 

1,5-NAPHTHALENEDIAMINE FEEDING EXPERIMENT 11 

DESIGN SUMMARY FOR B6C3F1 MICE—1,5-NAPHTHALENEDIAMINE FEEDING EXPERIMENT 12 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS 

AT SPECIFIC SITES IN MALE RATS TREATED WITH 

1,5-NAPHTHALENEDIAMINE 27 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS 

AT SPECIFIC SITES IN FEMALE RATS TREATED 

WITH 1,5-NAPHTHALENEDIAMINE 31 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS 

AT SPECIFIC SITES IN MALE MICE TREATED WITH 

1,5-NAPHTHALENEDIAMINE 45 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS 

AT SPECIFIC SITES IN FEMALE MICE TREATED 

WITH 1,5-NAPHTHALENEDIAMINE 48 
LIST OF TABLES (Concluded) 

Table Number Page 

Al SUMMARY OF THE INCIDENCE OF NEOPLASMS IN 

MALE RATS TREATED WITH 1,5-NAPHTHALENEDIAMINE A-3 

A2 SUMMARY OF THE INCIDENCE OF NEOPLASMS IN 

FEMALE RATS TREATED WITH 1,5-NAPHTHALENEDIAMINE A-8 

Bl SUMMARY OF THE INCIDENCE OF NEOPLASMS IN 

MALE MICE TREATED WITH 1,5-NAPHTHALENEDIAMINE B-3 

B2 SUMMARY OF THE INCIDENCE OF NEOPLASMS IN 

FEMALE MICE TREATED WITH 1,5-NAPHTHALENEDIAMINE B-7 

Cl SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC 

LESIONS IN MALE RATS TREATED WITH 1,5­

NAPHTHALENED LAMINE C-3 

C2 SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC 

LESIONS IN FEMALE RATS TREATED WITH 1,5­

NAPHTHALENEDIAMINE C-8 

Dl SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC 

LESIONS IN MALE MICE TREATED WITH 1,5­

NAPHTHALENEDIAMINE D-3 

D2 SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC 

LESIONS IN FEMALE MICE TREATED WITH 1,5­

NAPHTHALENED LAMINE D-7 

XI
 

I. INTRODUCTION
 

1,5-Naphthalenediamine (Figure 1) (NCI No. C03021), a bicyclic
 

aromatic amine used in the dye industry, was selected for bioassay by
 

the National Cancer Institute because of the high incidence of bladder
 

cancer reported among dye manufacturing industry workers (Anthony and
 

Thomas, 1970; Wynder et al., 1963). Aromatic amines are one class of
 

chemicals believed to contribute to the increased cancer risk in this
 

industry (Wynder et al., 1963). The structural similarity of 1,5­

naphthalenediamine to both the human bladder carcinogen 2-naphthylamine (International Agency for Research on Cancer [IARC], 1974) and
 

the suspected carcinogen 1-naphthy1amine (IARC, 1974) was an additional factor in its selection for testing.
 

The Chemical Abstracts Service (CAS) Ninth Collective Index
 

*
 

(1977) name for this compound is 1,5-naphthalenediamine. It is also
 

known as 1,5-diaminonaphthalene.
 

1,5-Naphthalenediamine can be used as an oxidation base (Colour
 

Index [C.I.] 76595), an intermediate in the synthesis of the dye
 

Naphthylene Red (C.I. 21650) (Society of Dyers and Colourists, 1956),
 

and in the production of a black trisazo dye for cotton (Taube, 1973).
 

1,5-Naphthalenediamine has also been used as a precursor for 1,5-naphthalenediisocyanate (Hirai and Yamamoto, 1975); as an intermediate
 

in the synthesis of drugs for the symptomatic treatment of asthma or
 

The CAS registry number is 2243-62-1
 
NH 

FIGURE 1 

CHEMICAL STRUCTURE OF 1,5-IMAPHTHALENEDIAMINE 
rhinitis (Hall, 1976); as a component of piperazine-modified aromatic
 

polyamides (Fujiwara et al., 1974); and as a modifier for phenolic
 

resins used in rapid curing compounds (Freeman et al. , 1974); however,
 

these uses appear to be purely experimental.
 

Specific production data for 1,5-naphthalenediamine are not
 

available; however, the exclusion of this compound from the 1977
 

Directory of Chemical Producers, U.S.A. (Stanford Research Institute,
 

1977) implies that it is not produced in commercial quantities (in
 

excess of 1000 pounds or $1000 in value annually).
 

The potential for exposure to 1,5-naphthalenediamine may be
 

greatest for workers in the dye industry and persons engaged in
 

chemical research with this compound.
 
II. MATERIALS AND METHODS
 

A. Chemicals
 

1,5-Naphthalenediamine was purchased from Carroll Products, Wood
 

River Junction, Rhode Island by the NCI for Mason Research Institute,
 

Worcester, Massachusetts, and chemical analysis was performed by Midwest Research Institute, Kansas City, Missouri. The experimentally
 

determined melting point of 190° to 191°C suggested a compound of
 

high purity based on its narrow range and its close proximity to the
 

value (190°C) reported in the literature (Pollock and Stevens, 1965).
 

Elemental analysis was consistent with cir.H N » the molecular formula
 

for 1,5-naphthalenediamine. However, nonaqueous amine group titration
 

was approximately 89 to 90 percent of that expected on a theoretical
 

basis. Vapor-phase chromatography revealed one homogeneous peak, but
 

thin-layer chromatography utilizing two solvent systems (acetone:ammonium hydroxide and methylethylketone:formic acid), each visualized with
 

254 nm and 367 nm light, indicated the presence of one nonmotile impurity. Nuclear magnetic and infrared analyses were consistent with
 

the structure of the compound. Ultraviolet analysis showed \ at
 

max
 

232, 328 and 498 nm with e values of 62,800, 10,640 and 9, respectively. The literature (Sadtler Standard Spectra) indicates a X.
 1 max
 

at 328.5 nm with e = 10,000 for 1,5-naphthalenediamine. The observed
 

e at 328 nm was 10,640 (6 percent greater than expected).
 

Throughout this report the term 1,5-naphthalenediamine is used
 

to represent this compound.
 

4
 
B. Dietary Preparation
 

The basal laboratory diet for both dosed and control animals
 

consisted of Wayne Lab-Blox (Allied Mills, Inc., Chicago, Illinois).
 

1,5-Naphthalenediamine was administered to the dosed animals as a
 

component of the diet. Under an exhaust hood, proper amounts of the
 

chemical were removed from the stock bottle. The compound was
 

blended in an aluminum bowl with an aliquot of the ground feed.
 

Once visual homogeneity was attained, the mixture was placed into a
 

6 kg capacity Patterson-Kelley twin-shell stainless steel V-blender,
 

along with the remainder of the meal and blended for 20 minutes.
 

Prepared diets were placed in double plastic bags and stored in the
 

dark at 4°C. The mixture was used for 1 week only.
 

C. Animals
 

Two animal species, rats and mice, were used in the carcinogenicity bioassay. Fischer 344 rats and B6C3F1 mice were obtained through
 

contracts of the Division of Cancer Treatment, National Cancer Institute. All animals were obtained from Charles River Breeding Laboratories, Inc., Wilmington, Massachusetts. Dosed and control animals
 

were received in separate shipments. Upon arrival, a sample of animals was examined for parasites and other signs of disease. All animals appeared to have parasites. They were treated with 3.0 gm of
 

piperazine adipate per liter of drinking water, a^i libitum, for 3
 

days, followed by 3 days of plain tapwater and 3 subsequent days of
 

piperazine adipate administration. During this period, new cages
 
with fresh bedding were provided daily. Animals were held in quarantine by species for 2 weeks prior to initiation of test. Animals
 

were assigned to groups and distributed among cages so that average
 

body weight per cage was approximately equal for a given sex and
 

species.
 

D.	 Animal Maintenance
 

All animals were housed by species in rooms having a temperature
 

(R)
 

range of 23° to 34°C. Incoming air was filtered through Tri-Dek
 

®
 15/40 denier Dacron filters (Tri-Dim Filter Corp., Hawthorne, New
 

Jersey) providing six changes of room air per hour. Fluorescent
 

lighting was provided on a 12-hour-daily cycle.
 

Rats were housed five per cage by sex. During quarantine and
 

for the first 14 months of study rats were housed in galvanized-steel
 

wire-mesh cages suspended over newspapers. Newspapers under cages
 

were replaced daily and cages and racks washed weekly. For the remainder of the study, rats were held in suspended polycarbonate cages
 

equipped with disposable nonwoven fiber filter sheets. Clean bedding
 

®
 

and cages were provided twice weekly. SAN-I-CEL corncob bedding
 

(Paxton Processing Company, Paxton, Illinois) was used for the first
 

2 months that rats were housed in polycarbonate cages. For the remainder of the study, Aspen hardwood chip bedding (American Excelsior
 

Company, Baltimore, Maryland) was used. Stainless steel cage racks
 

were cleaned once every 2 weeks, and disposable filters were replaced
 

at that time.
 
Mice were housed by sex in polycarbonate shoe box type cages.
 

Cages were fitted with perforated stainless steel lids (Lab Products,
 

Inc., Garfield, New Jersey). Nonwoven fiber filter bonnets were used
 

over cage lids. Control mice were housed ten per cage for the first
 

month of study and five per cage thereafter. Dosed mice were held
 

five per cage throughout the study. Clean cages, lids, and bedding
 

®


were provided twice per week. SAN-I-CEL was used during the first 9
 

®
 months of study. A second corncob bedding (Bed-o-Cobs , The Andersons
 

Cob Division, Maumee, Ohio) was used for the next 8 months. Aspen
 

bedding was used for the remainder of the study. Reusable filter
 

bonnets and pipe racks were sanitized every 2 weeks throughout the
 

s tudy.
 

Water was available from 250 ml polycarbonate water bottles
 

equipped with rubber stoppers and stainless steel sipper tubes.
 

Bottles were replaced twice weekly and, for rats only, water was
 

supplied as needed between changes. Food and water were available
 

ad libitum.
 

®


Wayne Lab-Blox meal was supplied to rats for 12 months and mice
 

(R)
 for 11 months from Alpine aluminum feed cups (Curtin Matheson Scientific, Inc., Woburn, Massachusetts) containing stainless steel baffles.
 

After that period, meal was supplied from stainless steel gangstyle
 

food hoppers (Scientific Cages, Inc., Bryan, Texas). During the
 

2-year period of chemical administration, dosed animals were supplied
 
with meal containing the appropriate concentrations of 1,5-naphthalenediamine. Control animals had untreated meal available. Food hoppers
 

were changed on the same schedule as were cages. Food was replenished
 

(R)
 daily in Alpine^ feed cups.
 

All rats utilized in the 1,5-naphthalenediamine bioassay were
 

*
 

housed in a room with other rats receiving diets containing acetylaminofluorene (53-96-3); sodium nitrite (76-32-00-0); L-arginine glutamate (4320-30-3); N-butylurea (592-31-4); N,N-dimethyl-p-nitrosoaniline
 

(138-89-6); 2,5-toluenediamine sulfate (6369-59-1); 2,4-dinitrotoluene
 

(121-14-2); 4-nitroanthranilic acid (619-17-0); N-(l-naphthyl)ethylenediamine dihydrochloride (1465-25-4); 2-chloro-p-phenylenediamine sulfate (61702-44-1); aniline hydrochloride (142-04-1); and p-anisidine
 

hydrochloride (20265-97-8).
 

Dosed mice were in a room with mice intubated with m-cresidine
 

(102-50-1); and with other mice receiving diets containing N-(l-naphthyDethylenediamine dihydrochloride (1465-25-4) and IH-benzotriazole
 

(95-14-7). Control mice were in a room with other mice receiving
 

diets containing hydrazobenzene (530-50-7); 2,3,5,6-tetrachloro-4­

nitroanisole (2438-88-2); tris(2,3-dibromopropyl)phosphate (126-72-7);
 

N-(l-naphthyl)ethylenediamine dihydrochloride (1465-25-4); aniline
 

hydrochloride (142-04-1); and 2-chloro-o-phenylenediamine sulfate.
 

*
 

CAS registry numbers are given in parentheses.
 
E. Selection of Initial Concentrations
 

In order to establish the maximum tolerated concentrations of
 

1,5-naphthalenediatnine for administration to dosed animals in the
 

chronic studies, subchronic toxicity studies were conducted with both
 

rats and mice. Animals of each species were distributed among six
 

groups, each consisting of five males and five females. 1,5-Naphthalenediamine was incorporated into the basal laboratory diet and supplied ad libitum to five of the six rat groups and five of the six
 

mouse groups in concentrations of 0.03, 0.1, 0.3, 1.0, and 3.0
 

percent. The sixth group of each species served as a control group,
 

receiving only the basal laboratory diet. The dosed dietary preparations were administered for 8 weeks.
 

The highest concentration causing no deaths, no compound-related
 

gross abnormalities, and no mean body weight depression in excess of
 

20 percent relative to controls was selected as the high concentration
 

for the chronic bioassay.
 

Deaths were recorded for all groups of rats receiving concentrations of 0.3 percent or more. Mean body weight depression was
 

approximately 19 and 9 percent, respectively, in males and females
 

dosed with 0.1 percent 1,5-naphthalenediamine. The concentration
 

of 1,5-naphthalenediamine selected for administration as the high
 

dose in the rat chronic bioassay was 0.1 percent.
 

Deaths were recorded for all groups of mice receiving concentrations of 0.3 percent or more and in the group of female mice
 
receiving 0.03 percent. Mean body weight depression was approximately
 

22 and 3 percent, respectively, in males and females dosed with 0.3
 

percent. Males receiving 0.1 percent experienced mean body weight
 

depression of approximately 3 percent, while females receiving the
 

same concentration had a greater mean body weight than the controls.
 

The concentration of 1,5-naphthalenediamine selected for administration as the high dose in the mouse chronic bioassay was 0.2 percent.
 

F. Experimental Design
 

The experimental design parameters for the chronic study (species, sex, group size, concentrations administered, and duration of
 

treated and untreated observation periods) are summarized in Tables
 

1 and 2.
 

Rats were all approximately 7 weeks old at the time they were
 

placed on test. Dosed rats were born approximately 1 month earlier
 

than controls and were started on test 1 month earlier than controls.
 

The dietary concentrations of 1,5-naphthalenediamine administered
 

were 0.10 and 0.05 percent. Throughout this report those rats
 

receiving the former concentration are referred to as the high dose
 

groups and those receiving the latter concentration are referred to
 

as the low dose groups. The dosed rats were supplied with feed containing 1,5-naphthalenediamine for a total of 103 weeks, followed by
 

a 3- to 4-week observation period.
 

All mice were approximately 7 weeks old at the time they were
 

placed on test. Dosed mice were born approximately 1 month earlier
 

10
 
TABLE 1 

DESIGN SUMMARY FOR FISCHER 344 RATS 

1,5-NAPHTHALENEDIAMINE FEEDING EXPERIMENT 

INITIAL

GROUP

SIZE

1,5-NAPHTHALENE­

 DIAMINE 

 CONCENTRATION 

 (PERCENT) 

OBSERVATION PERIOD 

TREATED UNTREATED 

(WEEKS) (WEEKS) 

MALE 

CONTROL 25 0 0 109 

LOW DOSE 50 0.05 

0 

103 

3 

HIGH DOSE 50 0.10 

0 

103 

3 

FEMALE 

CONTROL 25 0 0 110 

LOW DOSE 50 0.05 

0 

103 

3 

HIGH DOSE 50 0.10 

0 

103 

4 

11
 
TABLE 2
 

DESIGN SUMMARY FOR B6C3F1 MICE
 

1,5-NAPHTHALENEDIAMINE FEEDING EXPERIMENT
 

1,5-NAPHTHALENEINITIAL DIAMINE OBSERVATION PERIOD 

GROUP CONCENTRATION TREATED UNTREATED 

SIZE (PERCENT) (WEEKS) (WEEKS) 

MALE 

CONTROL 50 0 0 109 

LOW DOSE 50 0.1 103 

0 2 

HIGH DOSE 50 0.2 103 

0 2 

FEMALE 

CONTROL 50 0 0 109 

LOW DOSE 50 0.1 103 

0 2 

HIGH DOSE 50 0.2 103 

0 3 

12
 
than controls and were started on test 1 month earlier than controls.
 

The dietary concentrations of 1,5-naphthalenediamine administered
 

were 0.2 and 0.1 percent. Throughout this report those mice receiving the former concentration are referred to as the high dose groups
 

and those receiving the latter concentration are referred to as the
 

low dose groups. The dosed mice were supplied with feed containing
 

1,5-naphthalenediamine for a total of 103 weeks, followed by a 2- to
 

3-week observation period.
 

G. Clinical and Histopathologic Examinations
 

Animals were weighed immediately prior to initiation of the
 

experiment. Body weights were recorded twice weekly for the first
 

12 weeks of the study and at monthly intervals thereafter. From the
 

first day, all animals were inspected twice daily for mortality.
 

Food consumption, for two cages from each group, was monitored for
 

seven consecutive days once a month for the first nine months of the
 

bioassay and for three consecutive days each month thereafter. The
 

presence of tissue masses and lesions was determined by monthly observation and palpation of each animal.
 

A necropsy was performed on each animal regardless of whether it
 

died, was killed when moribund, or was sacrificed at the end of the
 

bioassay. The animals were euthanized by carbon dioxide inhalation,
 

and were immediately necropsied. The histopathologic examination consisted of gross and microscopic examination of major tissues, organs,
 

13
 
and gross lesions taken from sacrificed animals and, whenever possible, from animals found dead.
 

Tissues were preserved in 10 percent buffered formalin, embedded
 

in paraffin, sectioned, and stained with hematoxylin and eosin prior
 

to microscopic examination. An occasional section was subjected to
 

special staining techniques for more definitive diagnosis.
 

Slides were prepared from the following tissues: skin, subcutaneous tissue, larynx, lungs and bronchi, trachea, bone marrow, spleen,
 

lymph nodes, thymus, heart, salivary gland, liver, gallbladder (mice),
 

pancreas, esophagus, stomach, small intestine, large intestine, kidney, urinary bladder, pituitary, adrenal, thyroid, parathyroid, ear,
 

brain, testis, prostate, uterus, mammary gland, and ovary.
 

A few tissues were not examined for some animals, particularly
 

for those that died early. Also, some animals were missing, cannibalized, or judged to be in such an advanced state of autolysis as to
 

preclude histopathologic interpretation. Thus, the number of animals
 

for which particular organs, tissues, or lesions were examined microscopically varies and does not necessarily represent the number of
 

animals that were placed on experiment in each group.
 

H. Data Recording and Statistical Analyses
 

Pertinent data on this experiment have been recorded in an automatic data processing system, the Carcinogenesis Bioassay Data System
 

(Linhart et al., 1974). The data elements include descriptive information on the chemicals, animals, experimental design, clinical
 

14
 
observations, survival, body weight, and individual pathologic results,
 

as recommended by the International Union Against Cancer (Berenblum,
 

1969). Data tables were generated for verification of data transcription and for statistical review.
 

These data were analyzed using the statistical techniques described in this section. Those analyses of the experimental results
 

that bear on the possibility of carcinogenicity are discussed in the
 

statistical narrative sections.
 

Probabilities of survival were estimated by the product-limit
 

procedure of Kaplan and Meier (1958) and are presented in this report
 

in the form of graphs. 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) when testing two groups for
 

equality and used Tarone's (1975) extensions of Cox's methods when
 

testing a dose-related trend. One-tailed P-values have been reported
 

for all tests except the departure from linearity test, which is only
 

reported when its two-tailed P-value is less than 0.05.
 

The incidence of neoplastic or nonneoplastic lesions has been
 

given as the ratio of the number of animals bearing such lesions at a
 

specific anatomic site (numerator) to the number of animals in which
 

that site was examined (denominator). In most instances, the denominators included only those animals for which that site was examined
 

15
 
histologically. However, when macroscopic examination was required
 

to detect lesions prior to histologic sampling (e.g., skin or mammary
 

tumors), or when lesions could have appeared at multiple sites (e.g.,
 

lymphomas), the denominators consist of the numbers of animals necropsied.
 

The purpose of the statistical analyses of tumor incidence is to
 

determine whether animals receiving the test chemical developed a significantly higher proportion of tumors than did the control animals.
 

As a part of these analyses, the one-tailed Fisher exact test (Cox,
 

1970, pp. 48-52) was used to compare the tumor incidence of a control
 

group to that of a group of treated animals at each dose level. When
 

results for a number of treated groups, k, are compared simultaneously
 

with those for a control group, a correction to ensure an overall
 

significance level of 0.05 may be made. The Bonferroni inequality
 

(Miller, 1966, pp. 6-10) requires that the P-value for any comparison
 

be less than or equal to 0.05/k. In cases where this correction was
 

used, it is discussed in the narrative section. It is not, however,
 

presented in the tables, where the Fisher exact P-values are shown.
 

The Cochran-Armitage test for linear trend in proportions, with
 

continuity correction (Armitage, 1971, pp. 362-365), was also used
 

when appropriate. Under the assumption of a linear trend, this test
 

determined if the slope of the dose-response curve is different from
 

zero at the one-tailed 0.05 level of significance. Unless otherwise
 

16
 
noted, the direction of the significant trend was a positive dose relationship. This method also provides a two-tailed test of departure
 

from linear trend.
 

A time-adjusted analysis was applied when numerous early deaths
 

resulted from causes that were not associated with the formation of
 

tumors. In this analysis, deaths that occurred before the first
 

tumor was observed were excluded by basing the statistical tests on
 

animals that survived at least 52 weeks, unless a tumor was found at
 

the anatomic site of interest before week 52. When such an early
 

tumor was found, comparisons were based exclusively on animals that
 

survived at least as long as the animal in which the first tumor was
 

found. Once this reduced set of data was obtained, the standard procedures for analyses of the incidence of tumors (Fisher exact tests,
 

Cochran-Armitage tests, etc.) were followed.
 

When appropriate, life-table methods were used to analyze the
 

incidence of tumors. Curves of the proportions surviving without an
 

observed tumor were computed as in Saffiotti et al. (1972). The week
 

during which animals died naturally or were sacrificed was entered as
 

the time point of tumor observation. Cox's methods of comparing
 

these curves were used for two groups; Tarone's extension to testing
 

for linear trend was used for three groups. The statistical tests for
 

the incidence of tumors which used life-table methods were one-tailed
 

and, unless otherwise noted, in the direction of a positive dose
 

17
 
relationship. Significant departures from linearity (P < 0.05, two-


tailed test) were also noted.
 

The approximate 95 percent confidence interval for the relative
 

risk of each dosed group compared to its control was calculated from
 

the exact interval on the odds ratio (Gart, 1971). The relative risk
 

is defined as p /p where p is the true binomial probability of the
 

t c t
 

incidence of a specific type of tumor in a treated group of animals
 

and p is the true probability of the spontaneous incidence of the
 

same type of tumor in a control group. The hypothesis of equality
 

between the true proportion of a specific tumor in a treated group
 

and the proportion in a control group corresponds to a relative risk
 

of unity. Values in excess of unity represent the condition of a
 

larger proportion in the treated group than in the control.
 

The lower and upper limits of the confidence interval of the
 

relative risk have been included in the tables of statistical analyses. The interpretation of the limits is that in approximately 95
 

percent of a large number of identical experiments, the true ratio
 

of the risk in a treated group of animals to that in a control group
 

would be within the interval calculated from the experiment. When
 

the lower limi't of the confidence interval is greater than one, it
 

can be inferred that a statistically significant result (a P < 0.025
 

one-tailed test when the control incidence is not zero, P < 0.050
 

when the control incidence is zero) has occurred. When the lower
 

limit is less than unity but the upper limit is greater than unity,
 

18
 
the lower limit indicates the absence of a significant result while
 

the upper limit indicates that there is a theoretical possibility
 

of the induction of tumors by the test chemical which could not be
 

detected under the conditions of this test.
 

19
 
III. CHRONIC TESTING RESULTS: RATS
 

A. Body Weights and Clinical Observations
 

There was no appreciable depression in mean body weight when
 

dosed rats were compared with their respective controls (Figure 2).
 

Subcutaneous masses were observed in 2 high dose, 3 low dose,
 

and 1 control males, and in 12 high dose, 3 low dose, and 2 control
 

females. Crusted cutaneous masses occurred in 4 high dose males, 1
 

low dose male, 2 low dose females, and 1 control female, while firm
 

nodular growths were detected in 1 high dose, 2 low dose, and 2
 

control males, and in 1 low dose female. Swelling of the eyes was
 

exhibited by 2 high dose males, 2 high dose females, and 2 low dose
 

females and swelling of the nose by 1 low dose male. Only 1 control
 

female experienced crusted lesions in the vaginal area while 4 low
 

dose and 9 high dose females were so effected. Alopecia was recorded
 

for 1 low dose female, emaciation was observed in 1 male and 1 female
 

control, and 1 female control exhibited abdominal distention.
 

B. Survival
 

The estimated probabilities of survival for male and female rats
 

in the control and 1,5-naphthalenediamine-dosed groups are shown in
 

Figure 3. There was no significant positive association between
 

dosage and mortality for either male or female rats.
 

Adequate numbers of male rats were at risk from late-developing
 

tumors with 74 percent (37/50) of the high dose, 80 percent (40/50)
 

of the low dose and 68 percent (17/25) of the control surviving on
 

20
 
750 •750 

-600 

-450 

-300 

CONTROL 

-150 

LOW DOSE 

HIGH DOSE 

T~ T T~ 1 

' 30 45 60 75 90 105 120 

TIME ON TEST (WEEKS' 

750­ •750 

600­ -600 

I- 450 — -450 

I 

C3 

HI 

g 

g300­ -300 

CD 

150­

FEMALE RATS 

I 

15 30 15 60 75 90 105 120 

TIME ON TEST (WEEKS) 

FIGURE 2 

GROWTH CURVES FOR 1,5-NAPHTHALENEDIAMINE CHRONIC STUDY RATS 

21 
PROBABILITY OF SURVIVAL PROBABILITY OF SURVI

o 

o 

V) 

30 

> o 

o 3 "O

 

> 

30 

! .D O O I u) Z. ro 

ro 

m" 

m 

O 

n 8­

1 

a 

30 

i 

D 

I I I I O O o o 

01 
test until the termination of the study. No lesions were reported
 

for the 4 control rats that died in week 55.
 

With 76 percent (38/50) of the high dose, 76 percent (38/50) of
 

the low dose and 64 percent (16/25) of the control rats surviving on
 

test until the termination of the study, adequate numbers of females
 

were at risk from late-developing tumors.
 

C. Pathology
 

Histopathologic findings on neoplasms in rats are summarized in
 

Appendix A (Tables Al and A2); findings on nonneoplastic lesions are
 

summarized in Appendix C (Tables Cl and C2).
 

The incidence of liver neoplasms in male and female rats administered 1,5-naphthalenediamine in the diet appeared to be increased
 

relative to controls. In female rats, tumors of the clitoral gland,
 

uterus, and C-cell neoplasms of the thyroid appeared to be related
 

to compound administration. The incidences of these tutiors are as
 

follows:
 

MALES FEMALES 

Con­ Low High Con- Low High 

trol Dose Dose trol Dose Dose 

LIVER
 

(Number of animals with tissues
 

examined histopathologically) (25) (49) (49) (24) (50) (49)
 

Neoplastic Nodule 1 3 0 4
 

Hepatocellular Carcinoma 0 4 0 0
 

PREPUTIAL/CLITORAL GLAND
 

(Number of animals necropsied) (25) (49) (50) (24) (50) (50)
 

Carcinoma 0 0 1 3 8
 

Adenoma 0 0 0 0 5
 

23
 
Control

MALES

 Low High

 Dose Dose

 FEMALES 

 Con­ Low High 

 trol Dose Dose 

UTERUS AND ENDOMETRIUM 

(Number of animals with tissues 

examined histopathologically) ­ (24) (49) (48) 

Adenocarcinoma

Endometrial Stromal Polyp

Endometrial Stromal Sarcoma

 1 

2

 1 

2 

 14

2 

4 

 20 

2 

THYROID
 

(Number of animals with tissues
 

examined histopathologically) (21) (47) (47) (21) (49) (48)
 

C-Cell Adenoma 0 2 5 0 7 3
 

C-Cell Carcinoma 2 3 3 1 5 1
 

Neoplasms of the clitoral (preputial) gland were presented
 

grossly as round, fluctuant cystic subcutaneous lesions in the genital area, which on section were filled with pasty green material. On
 

microscopic examination, the cyst contents consisted of desquamated
 

epithelial cells, frequently mixed with leukocytes from secondary
 

inflammation. The inner portion of the cyst wall was lined by hyperkeratinized squamous epithelium often thrown into papillary folds.
 

Peripheral to this was a zone of large, round glandular cells at least
 

a few of which had coarse, brightly eosinophilic cytoplasmic granules.
 

If the peripheral border appeared smooth and intact, the lesion was
 

classified as an adenoma. If there was disorganization of the glandular structure and invasion into the surrounding stroma, the tumor
 

was called a carcinoma.
 

24
 
Thyroid C-cell tumors were observed in dosed female rats at
 

incidences increased relative to controls (4/48 [8 percent] high
 

dose, 12/49 [24 percent] low dose, 1/21 [5 percent] controls). C-


cell adenomas were discrete masses of these cells, often containing
 

small cysts lined by flat epithelium and containing colloid-like
 

material. In C-cell carcinomas, the tumor cells often assumed a
 

spindle shape and tended to invade surrounding tissue.
 

Uterine horns containing neoplasms were usually grossly enlarged.
 

The neoplasms themselves were varicolored, polypoid, frequently
 

gelatinous masses projecting into the uterine cavity. Endometrial
 

stromal polyps had a fibrous connective tissue core richly supplied
 

with large vessels. The surface of the polyps was covered with well-


differentiated endometrium which often formed glands in the superficial portion of the polyps. These tumors frequently became necrotic
 

at the tip and exhibited hemorrhage and secondary inflammation. In
 

a few rats, the connective tissue stroma of these lesions underwent
 

malignant transformation characterized by increased cellularity,
 

mitoses, and formation of plump, pleomorphic nuclei. Such tumors
 

were classified as stromal sarcomas. A uterine adenocarcinoma was a
 

collection of fairly well-differentiated glands arranged back-to-back
 

with no obvious intervening stroma. Nuclei of the glands were
 

markedly pleomorphic with frequent mitoses. There was invasion into
 

the myometrium and sometimes into extra uterine structures.
 

25
 
There were instances in this study, as noted in the summary
 

tables, where neoplastic lesions occurred only in dosed animals,
 

or with increased frequency when compared to the control group. No
 

pulmonary neoplasms were found in the controls; alveolar/bronchiolar
 

tumors were seen in dosed rats of both sexes. There was only one
 

urinary tract neoplasm in a female control; a few more occurred in
 

dosed rats, both male and female. No gliomas of the brain were seen
 

in controls; a few gliomas were found in dosed rats of both sexes.
 

These neoplasms occurred in such small numbers that a conclusive
 

interpretation as to their significance is not possible.
 

Rats in all groups exhibited a variety of nonneoplastic inflammatory and degenerative changes, and none were associated with administration of the compound.
 

Based upon the results of this pathologic examination, 1,5-naphthalenediamine was carcinogenic to female Fischer 344 rats since
 

feeding of the compound was associated with adenomas and carcinomas
 

of the clitoral gland. In addition, 1,5-naphthalenediamine feeding
 

appeared to be associated with increased incidences of thyroid, liver
 

and uterine neoplasms in female rats and liver neoplasms in male
 

rats.
 

B. Statistical Analyses of Results
 

The results of the statistical analyses of tumor incidence in
 

rats are summarized in Tables 3 and 4. The analysis is included for
 

26
 
TABLE 3
 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS AT
 

SPECIFIC SITES IN MALE RATS TREATED WITH 1,5-NAPHTHALENEDIAMINE£
 

HIGH
 

DOSE
 

2/50(0.04)
 

N.S.
 

1.000
 

0.056
 

56.712
 

102
 

1/50(0.02)
 

N.S.
 

0.250
 

0.004
 

4.616
 

106
 

4/47(0.09)
 

N.S.
 

Infinite
 

0.508
 

Infinite
 

106
 

TOPOGRAPHY : MORPHOLOGY
 

Subcutaneous Tissue: Fibroma
 

P Values
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

NJ
 

Skin: Squamous-Cell Papilloma
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Lung: Alveolar/Bronchiolar Adenoma or
 

Alveolar/Bronchiolar Carcinoma'3
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

LOW 

CONTROL DOSE 

1/25(0.04) 3/49(0.06) 

N.S. N.S. 

1.531 

0.133 

78.493 

99 106 

2/25(0.08) 1/49(0.02) 

N.S. N.S. 

0.255 

0.005 

4.707 

109 106 

0/25(0.00) 3/49(0.06) 

N.S. N.S. 

__.» Infinite 

0.315 

Infinite 

104 
___ 

TOPOGRAPHY: MORPHOLOGY
 

Hematopoietic System: Leukemia or
 

Malignant Lymphoma"
 

P Values
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Liver: Hepatocellular Carcinoma or
 

Neoplastic Nodule"
 

00 P Values0 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Pituitary: Adenoma NOS, Chromophobe
 

Adenoma, Acidophil Adenoma, or
 

Basophil Adenoma'3
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

TABLE 3 (CONTINUED) 

CONTROL
 

1/25(0.04)
 

N.S.
 

109
 

1/25(0.04)
 

N.S.
 

_—
 

109
 

2/22(0.09)
 

N.S,
 

98
 

LOW
 

DOSE
 

10/49(0.20)
 

N.S.
 

5.102
 

0.801
 

212.137
 

100
 

7/49(0.14)
 

N.S.
 

3.571
 

0.503
 

156.046
 

106
 

7/44(0.16)
 

N.S.
 

1.750
 

0.376
 

16.365
 

96
 

HIGH
 

DOSE
 

10/50(0.20)
 

N.S.
 

5.000
 

0.787
 

213.351
 

97
 

4/49(0.08)
 

N.S.
 

2.041
 

0.218
 

96.949
 

104
 

11/44(0.25)
 

N.S.
 

2.750
 

0.683
 

24.081
 

65
 
___ 

___ 

TABLE 3 (CONTINUED)
 

TOPOGRAPHY : MORPHOLOGY
 

Adrenal: Pheochromocytoma or Malignant
 

Pheochromocytoma^
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Thyroid: C-Cell Carcinoma
 

P Values0
 

VO
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Thyroid: C-Cell Adenoma or C-Cell
 

Carcinoma^5
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

CONTROL
 

2/24(0.08)
 

N.S.
 

109
 

2/21(0.10)
 

N.S.
 

97
 

2/21(0.10)
 

N.S.
 

97
 

LOW
 

DOSE
 

4/48(0.08)
 

N.S.
 

1.000
 

0.157
 

10.563
 

106
 

3/47(0.06)
 

N.S.
 

0.670
 

0.084
 

7.650
 

100
 

5/47(0.11)
 

N.S.
 

1.117
 

0.205
 

11.249
 

100
 

HIGH
 

DOSE
 

5/48(0.10)
 

N.S.
 

1.250
 

0.226
 

12.529
 

102
 

3/47(0.06)
 

N.S.
 

0.670
 

0.084
 

7.650
 

8/47(0.17)
 

N.S.
 

1.787
 

0.405
 

16.445
 

106 

104 
TABLE 3 (CONCLUDED)
 

TOPOGRAPHY : MORPHOLOGY CONTROL 

LOW 

DOSE 

HIGH 

DOSE 

Pancreatic Islets: Islet-Cell Adenoma 

or Islet-Cell Carcinoma15 

P Values0 

1/25(0.04) 

N.S. 

2/48(0.04) 

N.S. 

5/45(0.11) 

N.S. 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

1.042 

0.058 

60.184 

2.778 

0.340 

128.213 

Weeks to First Observed Tumor 98 106 104 

Testis: Interstitial-Cell Tumor 

P Values0 

21/25(0.84) 

N.S. 

44/49(0.90) 

N.S. 

45/49(0.92) 

N.S. 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

1.069 

0.890 

1.325 

1.093 

0.912 

1.324 

Weeks to First Observed Tumor 97 94 65 

Treated groups received doses of 0.05 or 0.10 percent in feed.
 

Number of tumor-bearing animals/number of animals examined at site (proportion).
 

C

The probability level for the Cochran-Armitage test is given beneath the incidence of tumors in
 

the control group when P < 0.05; otherwise, not significant (N.S.) is indicated. The probability
 

level for the Fisher exact test for the comparison of a treated group with the control group is
 

given beneath the incidence of tumors in the treated group when P < 0.05; otherwise, not significant (N.S.) is indicated. For both Cochran-Armitage and Fisher exact tests a negative designation (N) indicates a lower incidence in the treated group(s) than in the control group.
 

The 95% confidence interval on the relative risk of the treated group to the control group.
 
___ 

TABLE 4
 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS AT
 

SPECIFIC SITES IN FEMALE RATS TREATED WITH 1,5-NAPHTHALENEDIAMINE*
 

TOPOGRAPHY: MORPHOLOGY
 

Hematopoietic System: Leukemia or
 

Malignant Lymphoma-1 0
 

c
 

P Values
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Liver: Hepatocellular Carcinoma or
 

Neoplastic Nodule°
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Pituitary: Adenoma NOS, Chromophobe
 

Adenoma, Acidophil Adenoma, or Basophil Adenoma'3
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

CONTROL
 

3/24(0.13)
 

N.S.
 

94
 

0/24(0.00)
 

N.S.
 

6/21(0.29)
 

N.S.
 

91
 

LOW
 

DOSE
 

7/50(0.14)
 

N.S.
 

1.120
 

0.287
 

6.292
 

76
 

4/50(0.08)
 

N.S.
 

Infinite
 

0.458
 

Infinite
 

102
 

10/50(0.20)
 

N.S.
 

0.700
 

0.275
 

2.090
 

98
 

HIGH
 

DOSE
 

1/50(0.02)
 

N.S.
 

0.160
 

0.003
 

1.890
 

103
 

4/49(0.08)
 

N.S.
 

Infinite
 

0.467
 

Infinite
 

106
 

17/47(0.36)
 

N.S.
 

1.266
 

0.577
 

3.426
 

98
 
TABLE 4 (CONTINUED) 

TOPOGRAPHY :MORPHOLOGY CONTROL
 

Pituitary: Carcinoma NOS, Adenoma NOS,
 

Chromophobe Adenoma, Chromophobe Carcinoma, Acidophil Adenoma, or Basophil
 

Adenoma° 6/21(0.29)
 

P Values0 N.S.
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor 91
 

to 

Adrenal: Cortical Adenoma or Cortical
 

Carcinoma" 0/24(0.00)
 

P Values0 N.S.
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Adrenal: Pheochromocytoma 1/24(0.04)
 

P Values0 N.S.
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor 110
 

LOW
 

DOSE
 

11/50(0.22)
 

N.S.
 

0.770
 

0.312
 

2.262
 

88
 

3/50(0.06)
 

N.S.
 

Infinite
 

0.297
 

Infinite
 

106
 

0/50(0.00)
 

N.S.
 

0.000
 

0.000
 

8.966
 

HIGH
 

DOSE
 

18/47(0.38)
 

N.S.
 

1.340
 

0.618
 

3.606
 

98
 

1/49(0.02)
 

N.S.
 

Infinite
 

0.027
 

Infinite
 

106
 

3/49(0.06)
 

N.S.
 

1.469
 

0.127
 

75.534
 

106
 
TABLE 4 (CONTINUED)
 

TOPOGRAPHY : MORPHOLOGY CONTROL
 

Thyroid: C-Cell Carcinoma 1/21(0.05)
 

P Values0 N.S.
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor 109
 

Thyroid: C-Cell Adenoma or C-Cell
 

Carcinoma" 1/21(0.05)
 

LO P Values0 N.S.
 

Departure from Linear Trend P = 0.009
 

Relative Risk (Control) —_
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor 109
 

Thyroid: Papillary Carcinoma, Follicular-


Cell Carcinoma, or Papillary
 

Cystadenocarcinoma NOS^ 1/21(0.05)
 

P Values0 N.S.
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor 110
 

LOW
 

DOSE
 

5/49(0.10)
 

N.S.
 

2.143
 

0.266
 

99.147
 

106
 

12/49(0.24)
 

P = 0.046
 

—_
 

5.143
 

0.855
 

215.370
 

104
 

1/49(0.02)
 

N.S.
 

0.429
 

0.006
 

32.983
 

106
 

HIGH
 

DOSE
 

1/48(0.02)
 

N.S.
 

0.438
 

0.006
 

33.659
 

106
 

4/48(0.08)
 

N.S.
 

1.750
 

0.192
 

83.548
 

103
 

3/48(0.06)
 

N.S.
 

1.313
 

0.115
 

67.452
 

99
 
TABLE

TOPOGRAPHY : MORPHOLOGY
 

Thyroid: Papillary Carcinoma, Follicular-


Cell Carcinoma, Papillary Cystadenocarcinoma NOS, or Papillary Cystadenoma^
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Mammary Gland: Fibroadenoma
 

u> 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Mammary Gland: Fibroadenoma, Adenocarcinoma NOS, or Papillary Adenocarcinoma
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

4 (CONTINUED) 

CONTROL
 

1/21(0.05)
 

N.S.
 

110
 

4/24(0.17)
 

N.S.
 

109
 

4/24(0.17)
 

N.S.
 

109
 

LOW
 

DOSE
 

2/49(0.04)
 

N.S.
 

0.857
 

0.648
 

49.555
 

106
 

5/50(0.10)
 

N.S.
 

0.600
 

0.145
 

2.812
 

102
 

5/50(0.10)
 

N.S.
 

0.600
 

0.145
 

2.807
 

102
 

HIGH
 

DOSE
 

4/48(0.08)
 

N.S.
 

1.750
 

0.191
 

84.310
 

81
 

13/50(0.26)
 

N.S.
 

1.560
 

0.556
 

6.019
 

98
 

14/50(0.28)
 

N.S.
 

1.680
 

0.609
 

6.412
 

98
 
___ 

TOPOGRAPHY: MORPHOLOGY
 

Clitoral Gland: Carcinoma NOS
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Clitoral Gland: Adenoma NOS or 

u> Carcinoma NOSb 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Uterus: Endometrial Stromal Polyp
 

P Values
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

TABLE 4 (CONTINUED) 

CONTROL
 

1/24(0.04)
 

N.S.
 

110
 

1/24(0.04)
 

P = 0.003
 

110
 

2/24(0.08)
 

P = 0.003
 

102
 

LOW
 

DOSE
 

3/50(0.06)
 

N.S.
 

1.440
 

0.125
 

75.487
 

106
 

3/50(0.06)
 

N.S.
 

1.440
 

0.125
 

74.077
 

106
 

14/49(0.29)
 

P = 0.043
 

3.429
 

0.892
 

29.588
 

88
 

HIGH
 

DOSE
 

8/50(0.16)
 

N.S.
 

3.840
 

0.566
 

168.221
 

69
 

13/50(0.26)
 

P = 0.021
 

6.240
 

1.043
 

258.268
 

69
 

20/48(0.42)
 

P = 0.003
 

5.000
 

1.385
 

41.202
 

96
 
TABLE 4 (CONCLUDED)
 

TOPOGRAPHY: MORPHOLOGY CONTROL 

LOW 

DOSE 

HIGH 

DOSE 

Uterus and Endometrium: Adenocarcin

NOSb 

oma 

1/24(0.04) 2/49(0.04) 4/48(0.08) 

P Values0 N.S. N.S. N.S. 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

0.980 

0.054 

56.627 

2.000 

0.216 

96.367 

Weeks to First Observed Tumor 110 104 106 

Zymbal's Gland: Sebaceous Adenocarcinoma'5 0/24(0.00) 0/50(0.00) 3/50(0.06)
 

P Values0 N.S. N.S. N.S.
 

Relative Risk (Control) Infinite
 

Lower Limit 0.296
 

Upper Limit Infinite
 

Weeks to First Observed Tumor 89
 

Treated groups received doses of 0.05 or 0.10 percent in feed.
 

Number of tumor-bearing animals/number of animals examined at site (proportion).
 

"The probability level for the Cochran-Armitage test is given beneath the incidence of tumors in
 

the control group when P < 0.05; otherwise, not significant (N.S.) is indicated. The probability
 

level for the Fisher exact test for the comparison of a treated group with the control group is
 

given beneath the incidence of tumors in the treated group when P < 0.05; otherwise, not significant (N.S.) is indicated. For both Cochran-Armitage and Fisher exact tests a negative designation (N) indicates a lower incidence in the treated group(s) than in the control group.
 ;J
 

The 95% confidence interval on the relative risk of the treated group to the control group.
 

"The probability level of the test for departure from linear trend is given beneath the control
 

group when P < 0.05.
 

3 
every type of tumor in either sex where at least two such tumors were
 

observed in at least one of the control or 1,5-naphthalenediaminedosed groups and where such tumors were observed in at least 5 percent of the group.
 

For female rats an increased incidence of endometrial stromal
 

polyps was observed in both the high and low dose groups compared to
 

the control group. The Cochran-Armitage test indicated a significant
 

(P = 0.003) positive association between compound administration and
 

tumor incidence. The Fisher exact tests supported this result with a
 

significant (P = 0.003) comparison of the high dose group to the control; for the low dose comparison the probability level was P = 0.043,
 

a marginal result which was not significant under the Bonferroni criterion. Based on these results, the administration of 1,5-naphthalenediamine was associated with an elevated incidence of endometrial
 

stromal polyps in female rats.
 

A number of adenomas NOS and carcinomas NOS of the clitoral gland
 

were observed in female rats. The Cochran-Armitage test indicated a
 

significant (P = 0.003) positive association between dose and the
 

combined incidence of adenomas NOS or carcinomas NOS of the clitoral
 

gland. The Fisher exact test comparing high dose to control was also
 

significant (P = 0.021). In historical data collected by this laboratory for the NCI Carcinogenesis Testing Program, 4/249 (2 percent) of
 

the untreated female Fischer 344 rats had one of these tumors, compared to the 13/50 (26 percent) observed in the high dose group in
 

37
 
this bioassay. Based upon these statistical results, the administration of 1,5-naphthalenediamine was associated with an elevated incidence of clitoral gland neoplasms in female rats.
 

For females the Fisher exact test comparing control to low dose
 

for the combined incidence of C-cell adenomas or C-cell carcinomas of
 

the thyroid had a probability level of P = 0.046, a marginal result
 

which was not significant under the Bonferroni criterion.
 

Based on these statistical tests, it is concluded that 1,5-naphthalenediamine was carcinogenic for female rats, producing tumors of
 

the clitoral gland and uterus.
 

38
 
IV. CHRONIC TESTING RESULTS: MICE
 

A. Body Weights and Clinical Observations
 

Mean body weight depression was readily apparent in dosed male
 

mice when compared to controls. A similar but less pronounced trend
 

was evident in dosed females (Figure 4).
 

One low dose male had a soft subcutaneous mass on the leg and
 

two males in this group had palpable abdominal masses. Firm nodular
 

growths developed in one low dose male and two high dose females.
 

Alopecia was observed in 27 control males, 16 low dose males, 4 high
 

dose males, 25 control females, and 3 low dose females. Two low dose
 

and two high dose males experienced noticeable swelling of the eyes.
 

Abdominal distention was observed in one control male and one control
 

female mouse.
 

B. Survival
 

The estimated probabilities of survival for male and female mice
 

in the control and 1,5-naphthalenediamine-dosed groups are shown in
 

Figure 5. There was no significant positive association between dosage and mortality for either male or female mice.
 

Adequate numbers of male mice were at risk from late-developing
 

tumors with 58 percent (29/50) of the high dose, 78 percent (39/50)
 

of the low dose and 66 percent (33/50) of the controls surviving on
 

test until the termination of the study. The 6 control male mice
 

that died in week 11 were autolyzed, as were 2 of the 4 high dose
 

male mice that died in week 41.
 

39
 
45 60 75 120

 

TIME ON TEST (WEEKS) 

TIME ON TEST (WEEKS) 

FIGURE 4 

GROWTH CURVES FOR 1,5-NAPHTHALENEDIAMINE CHRONIC STUDY MICE 

40 

120 
PROBABILITY OF SURVIVAL PROBABILITY OF SURVI

o

I 

V) 

3D 

ro o o (71 

1 

3D 

CO 

O 

m 

O 

30 >m 

2 

m 

o 

3D 

O 

D 

° § 

T" o o o o 01 
For female mice, with 68 percent (34/50) of the high dose, 82
 

percent (41/50) of the low dose and 60 percent (30/50) of the control
 

mice surviving on test until the termination of the study, adequate
 

numbers were at risk from late-developing tumors.
 

C. Pathology
 

Histopathologic findings on neoplasms in mice are summarized in
 

Appendix B (Tables Bl and B2); findings on nonneoplastic lesions are
 

summarized in Appendix D (Tables Dl and D2).
 

Dietary administration of 1,5-naphthalenediamine produced an
 

increase in hepatocellular neoplasms in female mice, and it produced
 

a dose-related increase in thyroid neoplasms and compound-related
 

nonneoplastic thyroid lesions in both sexes. The compound-related
 

lesions are summarized below:
 

MALES FEMALES 

Con­ Low High Con­ Low High 

trol Dose Dose trol Dose Dose 

LIVER 

(Number of animals with 

tissues examined histopathologically) (39) (45) (43) (46) (49) (46) 

Hepatocellular Carcinoma 12 10 7 1 25 16 

Hepatocellular Adenoma 0 3 6 0 3 11 

THYROID 

(Number of animals with 

tissues examined histopathologically) (38) (46) (43) (44) (49) (45) 

Follicular-Cell Adenoma 

(Papillary or Follicular-Cell 

Adenoma, Papillary 

Cystadenoma)
 

Follicular-Cell Carcinoma
 

0

0

2 

8

1 

12 

16 

1

9 

2

2

2 

17 14 

1

4

2

6 

0

1

1 

1 

Follicular-Cell Hyperplasia
 

C-Cell Adenoma
 

C-Cell Carcinoma
 

0

0 

0

4 

2

0 

0

0 

42
 
In male mice, dietary administration of the compound did not
 

increase the incidence of hepatocellular neoplasms, whereas dosed
 

females showed a striking increase in hepatocellular carcinomas and
 

hepatocellular adenomas.
 

Grossly, hepatocellular neoplasms appeared as smooth, nodular,
 

rounded masses distorting the normal shape of the liver. Color
 

varied, many neoplasms appearing pale tan or dark red. Microscopically, hepatocellular carcinomas were expansive masses of hepatocytes
 

exhibiting loss of normal architectural pattern, the cells being
 

arranged in sheets or trabeculae instead of the normal lobules.
 

Nuclei were frequently uniform, although variable amounts of pleomorphism did occur. The cytoplasm was either basophilic or acidophilic,
 

sometimes varying from one region of the tumor to another, and was
 

frequently pale. Lesions classified as hepatocellular adenomas were
 

smaller, usually better differentiated, and were less pleomorphic
 

than the hepatocellular carcinomas.
 

The criteria for classification of thyroid neoplasms in mice
 

were the same as those used to classify thyroid neoplasms in rats.
 

The nonneoplastic thyroid lesions found in dosed mice were similar
 

to those in the rats but occurred in higher incidences. Hyperplasia
 

of follicular cells (focal, papillary or adenomatous) were found in
 

2/38 (5 percent) control, 12/46 (26 percent) low dose, and 9/43 (21
 

percent) high dose male mice. Abundant golden brown pigment was seen
 

in follicular epithelium, colloid, and macrophages. In the mice,
 

43
 
there were frequent foci of lymphocytes in the thyroid parenchyma and
 

occasional cystic areas filled with amorphous material containing
 

long clefts suggesting cholesterol crystals.
 

Three transitional-cell papillomas occurred in the bladder or
 

urethra of dosed mice (two high dose males and one high dose female),
 

but none occurred in controls.
 

Based upon the results of this pathologic examination, 1,5­

naphthalenediamine was carcinogenic to B6C3F1 mice, producing
 

hepatocellular neoplasms in females and thyroid neoplasms in both
 

sexes.
 

D. Statistical Analyses of Results
 

The results of the statistical analyses of tumor incidence in
 

mice are summarized in Tables 5 and 6. The analysis is included for
 

every type of tumor in either sex where at least two such tumors
 

were observed in at least one of the control or 1,5-naphthalenediamine-dosed groups and where such tumors were observed in at least 5
 

percent of the group.
 

For both male and female mice elevated incidences of thyroid tumors were observed in the dosed groups. In female mice the Cochran-


Artnitage test indicated a significant (P = 0.005) positive association
 

between dietary concentration and the incidence of C-cell carcinomas.
 

This was supported by a significant (P = 0.014) Fisher exact test for
 

the high dose group. For males the Cochran-Armitage test result
 

was also significant (P = 0.017), but the Fisher exact tests were
 

44
 
TABLE 5
 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS AT
 

SPECIFIC SITES IN MALE MICE TREATED WITH l.S-NAPHTHALENEDIAMINE*
 

TOPOGRAPHY : MORPHOLOGY
 

Lung: Alveolar/Bronchiolar Carcinoma
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Lung: Alveolar/Bronchiolar Adenoma or
 

Alveolar/Bronchiolar Carcinoma"
 

P Values0
 

Departure from Linear Trend
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Hematopoietic System: Malignant
 

Lymphoma"
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

CONTROL
 

2/39(0.05)
 

N.S.
 

109
 

4/39(0.10)
 

N.S.
 

P = 0.037
 

109
 

13/39(0.33)
 

P = 0.007(N)
 

LOW
 

DOSE
 

3/46(0.07)
 

N.S.
 

1.272
 

0.153
 

14.686
 

82
 

9/46(0.20)
 

N.S.
 

1.908
 

0.582
 

7.882
 

82
 

14/47(0.30)
 

N.S.
 

0.894
 

0.448
 

1.817
 

HIGH
 

DOSE
 

0/45(0.00)
 

N.S.
 

0.000
 

0.000
 

4.478
 

2/45(0.04)
 

N.S.
 

0.433
 

0.041
 

2.871
 

105
 

5/49(0.10)
 

P = 0.008(N)
 

0.306
 

0.094
 

0.829
 

Weeks to First Observed Tumor 100 82
 95
 
TOPOGRAPHY : MORPHOLOGY
 

Liver: Hepatocellular Carcinoma
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Liver: Hepatocellular Carcinoma or
 

Hepatocellular Adenoma^
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Thyroid: C-Cell Carcinoma
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

TABLE 5 (CONTINUED)
 

CONTROL
 

12/39(0.31)
 

N.S.
 

86
 

12/39(0.31)
 

N.S.
 

86
 

0/38(0.00)
 

P = 0.017
 

LOW
 

DOSE
 

10/45(0.22)
 

N.S.
 

0.722
 

0.318
 

1.620
 

88
 

13/45(0.29)
 

N.S.
 

0.939
 

0.453
 

1.981
 

88
 

0/46(0.00)
 

N.S.
 

HIGH
 

DOSE
 

7/43(0.16)
 

N.S.
 

0.529
 

0.198
 

1.306
 

105
 

13/43(0.30)
 

N.S.
 

0.983
 

0.473
 

2.071
 

105
 

4/43(0.09)
 

N.S.
 

Infinite
 

0.825
 

Infinite
 

105
 
TABLE 5 (CONCLUDED)
 

TOPOGRAPHY : MORPHOLOGY CONTROL 

LOW 

DOSE 

HIGH 

DOSE 

Thyroid: C-Cell Carcinoma or C-Cell 

Adenoma^ 

P Values0 

0/38(0.00) 

F = 0.044 

2/46(0.04) 

K:S. 

4/43(0.09) 

N.S. 

Relative Risk (Control)C 

Lower Limit 

Upper Limit 

Infinite 

0.246 

Infinite 

Infinite 

0.825 

Infinite 

Weeks to First Observed Tumor 105 105 

Thyroid: Papillary Adenoma, FollicularCell Adenoma, or Papillary Cystadenoma 

NOSb 0/38(0.00) 8/46(0.17) 16/43(0.37) 

P Values0 P < 0.001 P = 0.006 P < 0.001 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Infinite 

1.905 

Infinite 

Infinite 

4.523 

Infinite 

Weeks to First Observed Tuiror 105 98 

a

Treated groups received doses of 0.1 or 0.2 percent in feed.
 

Number of tumor-bearing animals/nuirber of animals examined at site (proportion).
 

£

The probability level for the Cochran-Armitage test is given beneath the incidence of tumors in
 

the control group when P < 0.05; otherwise, not significant (N.S.) is indicated. The probability
 

level for the Fisher exact test for the. comparison of a treated group with the control group is
 

given beneath the incidence of tumors in the treated group when P < 0.05; otherwise, not significant (N.S.) is indicated. For both Cochran-Armitage and Fisher exact tests a negative designation (N) indicates a lower incidence in the treated group(s) than in the control group.
 

6

The probability level of the test for departure from linear trend is given beneath the control
 

group when P < 0.05.
 
TABLE 6
 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS AT
 

SPECIFIC SITES IN FEMALE MICE TREATED WITH 1,5-NAPHTHALENEDIAMINE£
 

TOPOGRAPHY : MORPHOLOGY
 

Lung: Alveolar/Bronchiolar Carcinoma
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Lung: Alveolar/Bronchiolar Adenoma or
 

Alveolar/Bronchiolar Carcinoma"
 

P Values
 

Departure from Linear Trend
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Hematopoietic System: Leukemia or 

Malignant Lymphoma 

P Values0 

e 

Departure from Linear Trend
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

CONTROL
 

0/49(0.00)
 

N.S.
 

0/49(0.00)
 

N.S.
 

P = 0.005
 

13/49(0.27)
 

N.S.
 

P = 0.011
 

LOW
 

DOSE
 

1/48(0.02)
 

N.S.
 

Infinite
 

0.055
 

Infinite
 

89
 

10/48(0.21)
 

P = 0.001
 

Infinite
 

3.037
 

Infinite
 

89
 

19/50(0.38)
 

N.S.
 

1.432
 

0.760
 

2.781
 

HIGH
 

DOSE
 

3/46(0.07)
 

N.S.
 

Infinite
 

0.638
 

Infinite
 

91
 

5/46(0.11)
 

P = 0.024
 

Infinite
 

1.347
 

Infinite
 

91
 

5/46(0.11)
 

P = 0.045(N)
 

0.410
 

0.124
 

1.117
 

Weeks to First Observed Tumor 57 63
 105
 
TOPOGRAPHY : MORPHOLOGY
 

Liver: Hepatocellular Carcinoma
 

P Values0
 

Departure from Linear Trend
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Liver: Hepatocellular Adenoma or
 

Hepatocellular Carcinoma"
 

P Values0
 

g


Departure from Linear Trend
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Stomach: Squamous-Cell Papilloma
 

P Values0
 

g


Departure from Linear Trend
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

TABLE 6 (CONTINUED)
 

CONTROL
 

1/46(0.02)
 

P = 0.001
 

P < 0.001
 

109
 

1/46(0.02)
 

P < 0.001
 

P = 0.002
 

109
 

0/41(0.00)
 

N.S.
 

P = 0.017
 

LOW
 

DOSE
 

25/49(0.51)
 

P < 0.001
 

23.469
 

4.156
 

906.346
 

74
 

28/49(0.57)
 

P < 0.001
 

26.286
 

4.741
 

1030.801
 

74
 

3/47(0.06)
 

N.S.
 

Infinite
 

0.529
 

Infinite
 

105
 

HIGH
 

DOSE
 

16/46(0.35)
 

P < 0.001
 

16.000
 

2.683
 

646.516
 

99
 

27/46(0.59)
 

P < 0.001
 

27.000
 

4.874
 

1027.943
 

99
 

0/46(0.00)
 

N.S.
 
___ 

TOPOGRAPHY rMORPHOLOGY
 

Pituitary: Adenoma NOS, Chromophobe
 

Adenoma or Acidophil Adenoma
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Adrenal: Pheochromocytoma
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Thyroid: C-Cell Carcinoma
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

TABLE 6 (CONTINUED) 

CONTROL
 

3/34(0.09)
 

N.S.
 

109
 

3/46(0.07)
 

P = 0.040(N)
 

68
 

0/44(0.00)
 

P = 0.005
 

LOW
 

DOSE
 

4/35(0.11)
 

N.S.
 

1.295
 

0.238
 

8.188
 

105
 

0/44(0.00)
 

N.S.
 

0.000
 

0.000
 

1.731
 

1/49(0.02)
 

N.S.
 

Infinite
 

0.048
 

Infinite
 

105
 

HIGH
 

DOSE
 

1/30(0.03)
 

N.S.
 

0.378
 

0.007
 

4.424
 

106
 

0/44(0.00)
 

N.S.
 

0.000
 

0.000
 

1.731
 

6/45(0.13)
 

P = 0.014
 

Infinite
 

1.574
 

Infinite
 

105
 
TABLE 6 (CONCLUDED) 

LOW HIGH 

TOPOGRAPHY : MORPHOLOGY CONTROL DOSE DOSE 

Thyroid: C-Cell Adenoma or C-Cell 

Carcinoma^ 0/44(0.00) 2/49(0.04) 8/45(0.18) 

P Values P = 0.001 N.S. P = 0.003 __ _ 

Relative Risk (Control) Infinite Infinite 

Lower Limit 0.267 2.250 

Upper Limit Infinite Infinite 

Weeks to First Observed Tumor 105 41 

Thyroid: Papillary Adenoma, Follicular-Cell 

Adenoma, or Papillary Cystadenoma NOS^ 2/44(0.05) 17/49(0.35) 14/45(0.31) 

P Values0 P = 0.003 P < 0.001 P = 0.001 

Departure ficm Linear Trend P = 0.025 

Relative Risk (Control) 7.633 6.844 

Lower Limit 1.971 1.709 

Upper Limit 64.662 58.827 

Weeks to First Observed Tumor 80 105 91 

Treated groups received doses of 0.1 or 0.2 percent in feed.
 

Number of tumor-bearing animals/number of animals examined at site (proportion).
 

"The probability level for the Ccchran-Armitage test is given beneath the incidence of tumors in
 

the control group when P < 0.05; otherwise, rot significant (N.S.) is indicated. The probability
 

level for the Fisher exact test for the comparison of a treated group with the control group is
 

given beneath the incidence of tumors in the treated group when P < 0.05; otherxvise, net significant (N.S.) is indicated. For both Cochran-Armitage and Fisher exact tests a negative designation (N) indicates a lower incidence in the treated group(s) than in the control group.
 

The 95% confidence interval en the relative risk of the treated group to the control grcup.
 

2

The probability level of the test for departure from linear trend is given beneath the contrrl
 

group vhen P < C.05.
 
not. When incidences were combined so that the numerator represented
 

mice with either a papillary adenoma, a follicular-cell adenoma, or a
 

papillary cystadenoma of the thyroid, the Cochran-Armitage test indicated a significant positive association between dietary concentration
 

and tumor incidence for both males (P < 0.001) and females (P = 0.003).
 

These were supported by significant (P _<_ 0.006) Fisher exact test
 

results in each sex for comparisons of each dosed group to the control
 

group. Based on these results, the administration of 1,5-naphthalenediamine was associated with the incidence of thyroid neoplasms in both
 

male and female mice.
 

For females an increased incidence of hepatocellular carcinomas
 

was also observed among the dosed mice. The Cochran-Armitage test
 

indicated a significant (P = 0.001) positive association between dose
 

and incidence. This was supported by significant (P < 0.001) comparisons of both the high and low dose to the control group using
 

the Fisher exact test. Based on these results the administration of
 

1,5-naphthalenediamine was associated with the incidence of hepatocellular carcinomas in female mice.
 

For female mice, when the incidence of alveolar/bronchiolar
 

adenomas and alveolar/bronchiolar carcinomas were combined, an increased incidence in the dosed groups was noted. The Fisher exact
 

test was significant.for both the high (P = 0.024) and low (P = 0.001)
 

dose groups. The departure from linear trend was significant since
 

tumor incidence was increased more in the low dose than in the high
 

52
 
dose group. In historical control data compiled by this laboratory
 

for the NCI Carcinogenesis Testing Program, 17/275 (6 percent) of the
 

untreated female B6C3F1 mice had an alveolar/bronchiolar neoplasm.
 

Based upon these results the administration of 1,5-naphthalenediamine
 

was associated with the incidence of alveolar/bronchiolar neoplasms
 

in female mice.
 

For females the Fisher exact test comparing the incidence of
 

leukemia or malignant lymphoma in high dose mice with that in the
 

controls had a probability level in the negative direction of
 

P = 0.045, a marginal result which was not significant under the
 

Bonferroni criterion.
 

Also for females the Cochran-Armitage test showed a significant
 

(P = 0.040) negative association between dose and the incidence of
 

adrenal pheochromocytomas, but the Fisher exact tests were not significant.
 

In male mice the possibility of a negative association between
 

dose and the incidence of malignant lymphomas or leukemia was noted.
 

Based upon these statistical results the administration of 1,5­

naphthalenediamine was associated with the increased incidence of
 

thyroid neoplasms in male mice and of thyroid neoplasms, of hepatocellular carcinomas, and of alveolar/bronchiolar neoplasms in female
 

mice,
 

53
 
V. DISCUSSION
 

There were no significant positive associations between dietary
 

concentrations of 1,5-naphthalenediamine and mortality in either sex
 

of rats or mice. In all groups adequate numbers of animals survived
 

sufficiently long to be at risk from late-developing tumors.
 

Several uterine neoplasms occurred in dosed female rats at higher
 

incidences than in corresponding controls. There was a significant
 

positive association between dietary concentration of the compound
 

and the incidences of endometrial stromal polyps in female rats. In
 

addition, the high dose to control Fisher exact comparison was significant. Endometrial stromal sarcomas were observed in two low dose
 

and two high dose female rats, but not in controls. Uterine adenocarcinomas occurred at a higher incidence in the high dose female rat
 

group than in the control group, but the difference in tumor incidence
 

was not statistically significant.
 

The administration of 1,5-naphthalenediamine was associated with
 

an elevated incidence of clitoral gland neoplasms in female rats.
 

There was a significant positive association between the concentration
 

of the chemical added to the diet and the incidence of either adenomas
 

or carcinomas of the clitoral gland in female rats. The incidence of
 

either of these neoplasms in the high dose female rat group was
 

significant relative to the incidence in the control group.
 

Elevated incidences of thyroid neoplasms were observed among
 

dosed mice. For mice of both sexes there were significant positive
 

54
 
associations between dietary concentration of 1,5-naphthalenediamine
 

and the incidences of thyroid C-cell carcinomas. For the females the
 

high dose to control Fisher exact comparison supported the finding;
 

this was not true for males. When the mice were grouped so that the
 

numerator of the incidence represented those animals with a papillary
 

adenoma, a follicular-cell adenoma, or a papillary cystadenoma of
 

the thyroid, the Cochran-Armitage test was significantly positive for
 

both males and females and all the Fisher exact comparisons supported
 

the findings.
 

The incidence of hepatocellular carcinomas in female mice was
 

significantly associated with increased concentration of 1,5-naphthalenediamine. In addition, the high dose to control and the low dose
 

to control Fisher exact comparisons were significant. The incidence
 

of alveolar/bronchiolar adenomas was significant, relative to controls, in both the low dose and the high dose female mouse groups.
 

Under the conditions of this bioassay, 1,5-naphthalenediamine
 

was carcinogenic in female Fischer 344 rats, causing clitoral and
 

uterine neoplasms. 1,5-Naphthalenediamine was also carcinogenic for
 

B6C3F1 mice, producing thyroid neoplasms in males and neoplasms of the
 

thyroid, liver, and lung in females. Insufficient evidence was provided for the carcinogenicity of the compound in male Fischer 344 rats.
 

55
 
VI. BIBLIOGRAPHY
 

Anthony, H.M., and G.M. Thomas, "Tumors of the Urinary Bladder: An
 

Analysis of the Occupations of 1,030 Patients in Leeds, England."
 

Journal of the National Cancer Institute 45:879-895, 1970.
 

Armitage, P., Statistical Methods in Medical Research, Chapter 14.
 

J. Wiley & Sons, New York, 1971.
 

Berenblum, I., editor, Carcinogenicity Testing. International Union
 

Against Cancer, Technical Report Series, Vol. 2. International
 

Union Against Cancer, Geneva, 1969.
 

Chemical Abstracts Service, The Chemical Abstracts Service (CAS)
 

Ninth Collective Index, Volumes 76-85, 1972-1976. American
 

Chemical Society, Washington, D.C., 1977.
 

Cox, D.R., Analysis of Binary Data, Chapters 4 and 5. Methuen and
 

Co., Ltd., London, 1970.
 

Cox, D.R., "Regression Models and Life-Tables." Journal of the Royal
 

Statistical Society, Series "B" 34:187-220, 1972.
 

Freeman, H.G., G.F. Baxter, and G.T. Tiedeman, "Rapid Curing Resin
 

Compositions Comprising a Phenol-Aldehyde Condensation Polymer
 

Modified with an Aminonaphthalene." U.S. Patent 3,786,025
 

(Weyerhaeuser Co.) January 15, 1974; Chemical Abstracts 81,
 

78678u.
 

Fujiwara, T., K. Nagosawa, T. Tsumaki, S. Goto, and S. Fukuoka,
 

"Piperazine-Modified Aromatic Polyamide Fibers." Japan Kokai
 

74,116,322 (Asahi Chemical Industry Co., Ltd.) November 7,
 

1974; Chemical Abstracts 83, 12023m.
 

Gart, J.J., "The Comparison of Proportions: A Review of Significance
 

Tests, Confidence Limits, and Adjustments for Stratification."
 

International Statistical Institute Review 39:148-169, 1971.
 

Hall, C.M., "Pharmaceutical Compositions Containing 1,4,7,10-Tetrahydro-1,7-dioxoquino-8,7-h quinoline-3,9-dicarboxylic Acid
 

Derivatives or Salts." U.S. Patent 3,953,598 (Upjohn Co.)
 

April 27, 1976; Chemical Abstracts 85, 372484.
 

Hirai, Y., and R. Yamamoto, "Purification of 1,5-Naphthalenediisocyanate." Japan. Kokai 75 12,062 (Mitsui Toatsu Chemicals,
 

Inc.) February 7, 1975; Chemical Abstracts 82, 155968p.
 

56
 
International Agency for Research on Cancer, IARC Monographs on the
 

Evaluation of Carcinogenic Risk of Chemicals to Man: Some Aromatic Amines, Hydrazine and Related Substances, N-Nitroso Compounds, and Miscellaneous Alkylating Agents, Volume 4. IARC,
 

Lyon, France, 1974.
 

Kaplan, E.L., and P. Meier, "Nonparametric Estimation from Incomplete
 

Observations." Journal of the American Statistical Association
 

_53_:457-481, 1958.
 

Linhart, M.S., J.A. Cooper, R.L. Martin, N.P. Page, and J.A. Peters,
 

"Carcinogenesis Bioassay Data System." Computers and Biomedical
 

Research 7:230-248, 1974.
 

Miller, R.G., Simultaneous Statistical Inference. McGraw-Hill Book
 

Co., New York, 1966.
 

Pollock, J.R.A. and R. Stevens, editors, Dictionary of Organic
 

Compounds, 4th edition, Volume 4. Oxford University Press,
 

New York, 1965.
 

Sadtler Standard Spectra. Sadtler Research Laboratories, Philadelphia,
 

Pennsylvania. UV No. 7846.
 

Saffiotti, U., R. Montesano, A.R. Sellakumar, F. Cefis, and D.G.
 

Kaufman, "Respiratory Tract Carcinogenesis in Hamsters Induced
 

by Different Numbers of Administration of Benzo (a) Pyrene and
 

Ferric Oxide." Cancer Research 32:1073-1079, 1972.
 

Society of Dyers and Colourists. Colour Index, 2nd edition, Volume 3.
 

Yorkshire, England, 1956.
 

Stanford Research Institute, 1977 Directory of Chemical Producers,
 

U.S.A. Menlo Park, California, 1977.
 

Tarone, R.E., "Tests for Trend in Life-Table Analysis." Biometrika
 

62^:679-682, 1975.
 

Taube, C., "Trisazo Dye." Ger. Offen. 2,211,638 (Bayer A.-G.)
 

September 20, 1973; Chemical Abstracts 80, 4901u.
 

Wynder, E.L., J. Onderdonk, and N. Mantel, "An Epidemiological Investigation of Cancer of the Blader." Cancer 16:1388-1407, 1963.
 

57
 

Review of the Bioassay of 1,5-Naphthalenediamine*
 

for Carcinogenicity
 

by the Data Evaluation/Risk Assessment Subgroup
 

of the Clearinghouse on Environmental Carcinogens
 

June 29, 1978
 

The Clearinghouse on Environmental Carcinogens was
 

established in May, 1976, in compliance with DHEW Committee
 

Regulations and the Provisions of the Federal Advisory
 

Committee Act. The purpose of the Clearinghouse is to
 

advise the Director of the National Cancer Institute (NCI)
 

on its bioassay program to identify and to evaluate chemical
 

carcinogens in the environment to which humans may be exposed.
 

The members of the Clearinghouse have been drawn from
 

academia, industry, organized labor, public interest groups,
 

State health officials, and quasi-public health and research
 

organizations. Members have been selected on the basis of
 

their experience in carcinogenesis or related fields and,
 collectively, provide expertise in chemistry, biochemistry,
 

biostatistics, toxicology, pathology, and epidemiology.
 

Representatives of various Governmental agencies participate
 

as ad hoc members. The Data Evaluation/Risk Assessment
 

Subgroup of the Clearinghouse is charged with the responsibility of providing a peer review of reports prepared on
 

NCI-sponsored bioassays of chemicals studied for carcinogenicity. It is in this context that the below critique is given
 

on the bioassay of 1,5-Naphthalenediamine for carcinogenicity.
 

The reviewer agreed with the conclusion in the report
 

that 1,5-Naphthalenediamine was carcinogenic in treated
 

female rats and in both sexes of mice. He noted that the
 

study was conducted in a room in which other compounds were
 

under test. Based on the experimental findings, he concluded that 1,5-Naphthalenediamine may pose a carcinogenic
 

risk to humans. The reviewer moved that the report on the
 

bioassay of 1,5-Naphthalenediamine be accepted as written.
 

The motion was approved without objection.
 

Clearinghouse Members present:
 

Arnold L. Brown (Chairman), Mayo Clinic
 

Paul Nettesheim, National Institute of Environmental
 

Health Sciences
 

Verne Ray, Pfizer Medical Research Laboratory
 

Verald K. Rowe, Dow Chemical U.S.A.
 

Michael B. Shimkin, University of California at San Diego
 

Louise Strong, University of Texas Health Sciences Center
 

Subsequent to this review, changes may have been made
 

in the bioassay report either as a result of the review
 

or other reasons. Thus, certain comments and criticisms
 

reflected in the review may no longer be appropriate.
 

59
 

1>U . S . GOVERNMEN T PRINTIN G OFFICE : I 9 7 8-2 6 0'8 9 9 / 3 I 9 8
 

APPENDIX A
 

SUMMARY OF THE INCIDENCE OF NEOPLASMS
 

IN RATS TREATED WITH 1,5-NAPHTHALENEDIAMINE
 

TABLE Al 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN MALE RATS TREATED WITH 1,5-NAPHTHALENEDIAMINE 

CONTROL tONTR) LOW DOSE HIGH DOSE
 

01-0330 01-0260 01-0285
 

INIMALS INITIAHY IB STUDY 25 50 5ft
 INIMALS NECROPSIID 25 19 Ifl
 HHIBALS EXAMINED HISTOPATHOLOGICAILY** 25 »9 »9
 

INTEGUMENTARY SYSTEM
 

*SKIN (25) f»9) (50)
 SOnA MO OS CEtl PAPIIICHA 2 (8«) 1 (2V) 1 (2X)
 SEBACEOUS ADBNOCA1CINOHA 1 '.2%)
 

PIBROOS HISTIOCYTOBA 1 I2X)
 

*SOBCDT TISSUE	 (25) (U9) (50)
 PTBROMA	 1 ;««) 3 I6«) 2 ;»%)
 

RESPIRATORY fYSTEM
 

•1ARYHX	 35J ;»9) ;so)
 PJPIIICHA, SOS 1 (2*)
 

ITRACHHA fi») (16) (13)
 P»P!LLOMK, NOS 1 (6X)
 

ILONG (25) (19) (17)
 ADMOCAaCINOBA, NOS, HIIASTATIC 1 (2X)
 

ALTEOLAP/BtONCHIOLAR ADENON* 1 :2«) <t ;9%)
 

ALVTOLAR/BRONCHIOLAR CARCINOMA 2 3*)
 

C-CEir CAfiCINOHA, HETASTATIC ' ;2«)
 

S^BJCIODS ADENOCARCTNOBA, HETAST 1 12*)
 PHEOCHROHOCYTOHA, BPTASTATIC 1 ;»t)
 

HEHATOPOIETIC SYSTFB
 

•BOLTTP1E OPSANS	 j|Sj ;i»9) ;50)
 

LFOKEim.tiOS •< (2%)
 

DWDUrEREMTIATED LEOKERIA 10 ;20«) « ,'8X)
 

MYEIOM010CYTIC LEOKEBIA 1 :»X) 1 t2«)
 _ ^IBPHOgYTIC ^fOKEMI\	 „ „ ,2 ,;M!»
 

* NOMBER Of AHIMALS BITH TISSQE EXAMINED MICROSCOPIC*.lit
 

* NOMBfR Of ANINAIS NECRCPSTED
 

**EXCLUDES PARTIALLY AUTOLTZED ANIMALS
 

A-3
 
TABLE Al (CONTINUED) 

CONTROL(BITS) LCi DOSE HIGH DOSE 

01-0330 01-0280 01-0285 

tHAHDIBOLA" L. NODE ;U7)

 

r-CEU CAKCINOHA, BETASTATIC 1 (»«)

 SEBACEOOS ADENOCAHCIHOBA, BETJ SI 1 (2*)

 

NEOFILEBOHA, METASTATIC
 

*BEDIASTIN»I L.KODE
 

SEBACEOOS ADENOCAtCISOBA, BETAST 1 (2%)

 

IBESEHTIRir L. 10DE t»7)

 

1YBPHAHGIOBA
 

tLIVE" (119) 

tlHETFFERIHTHTED LEOKIBIA 1 (2*) 

tTHYMOS (35) (36) 

THYBOMA 1 (3*) 

BALIG.LYBPHOBA, LYUPHOCYTIC TYPE 

CTRCULAT08Y SYSTEM 

NONE 

DIGESTIVE SYSTEM 

tSUIVARY GLASD (25) (17) («6) 

ADENOCARCIHOMA, HOS 1 (2%) 

FIBROSAPCOBA 1 J2«) 

NEnRILEBOBA, MALI^NAIT 1 ;2«) 

*LIVE* (25) (49) (W9) 

NEOPIASTIC NODDLE 1 (US) 3 (6X) 2 (OX) 

HEPAIOCEILILAR CA1CINOMA 4 ;ex) 2 ;««) 

LYMPHANGIOBA 1 T2«) 

ISTOMACB tl*J C*') (47)

 

SQOAROQS CEIL PAPILL08A 1 (2*) 

OBTHARY SYSTEM 

»KIDNEY *0^J ^49J J4 8,J

 

1IPOHA 1 W*J

 

IORINAP1 El ADDER (25) (49) (»8) 

TRAH^IJIig.llAJ-CEII, nCSScI»OHi 1 1281 

I HOflBER OF AKIBA1S STTH TISSOI EXAMINED MICROSCOPICALLY 

* HOHBIP OF AKIBAIS RECROPSIED 

A-4 
TABLE A1 (CONTINUED) 

CONTROL (UNTR) LOW DOSE 

01-0330 01-0280 

BNDOCPINE SYSTEM
 

fPITOTTAPY (22) 

CARCINOMA,NOS 

ADENOMA, NOS 

CHROMOPBOBE ADENOMA 3 ;7X) 

ACIDOPHTI ADENOHA 

ACIDOPHIL CARCINOMIi 

BASOEHIL ADENOHA 

INTEFSTITIAL-CELL TOMOR, BETASTA 

tADRENAL (18) 

CP1TICAL ADENOHA 1 (2X) 

PHEOCBROHOCYTOBJ 

PBEOCBROMOCYTOBA, MALIGNANT 

NEOPOBLASTOBA 

4THYFOTD (211 (U7)
 

FOLLICOLAri-CELL ADENOMA 1 (5X)
 

C-CELL ADENOMA 2 ;txi
 C—CELL ^^RCTNOMA 2 ;iox) 3 I6X)
 

SEBACEODS JDENOCARCINOMA, BETAS! ' ;2X)
 PAPTLUSY CYSTADEHOCAHCINOMA.NOS
 

(2U)
 

AEENOBA, NOS
 

•PAHATBYROID (13)
 

tPANCPSATIC ISIETS (25) (H8)
 

ISLHT-CILL ADENOMA 1 ("X) 1 (2X)
 

ISLET-CELL CARCINOMA 1 ;2X)
 

RBPRODOCTIVE SYSTEM
 

•MAMMARY GLAND
 

ADENOCAFCINOMA, NOS
 

FIBROADXNOHA
 

*PREPUTIAI GLAND (U9)
 

CARCINOMA,NOS
 

ADENOMA, NOS
 

fTISTIS (25) (19) 

INTERSTITIAL-CELL TOHOR 21 (8UX) 4tt (90X) 

TOBOR. MHIGNA 

t BDHBEH OF ANIMALS HTTH TISSOE EXAMINED BICROSCOPICJILY
 

* ROBBER 01 ANIBAIS NECBCPSIED
 

A-5
 

HIGH EOSE
 

"1-0285
 

(Kit) 

1 (2%) 

7 -16X) 

2 ;5«) 

;2X)
 

|U7)


1 (2X)


5 ;nxj
 3 ;6X)
 

1 ;2X)
 

C28)
 

1 (IX)
 

(tc)


II (9%)


1
 ;2x>
 

(•50)
 

US (92X)
 
NEBVOOS SYSTEM 

IPSAIN


CAPCIHCBA, SOS, BITASTATIC


GlIiBA, HOS


lcmB*lLO!!


G1IOHA, HOS


SFECIAt SENSE npGMlS 

*E»B CAIHI

SQOAHOUS CE1I CIPCIFCRA


*ZIHBA1'S G1AND

SIBACtOUS ADENOCHHCIHOHA

HBSCOLOSKEIFTAL STSTEI 

10>IF 

EOCT CATIIIES
 

*EODT CAVITIES


BESOIHEITQBJ, NOS


MESOTHEUOHA, HALIGHAKT


*»BEOHIH»I CAVITT

OSTEOSABCOHA

All OTHI? STSTEHS 

TAIL 

I IDHBEP OF ANIHALS KITH TISSUE

* •DRBEP OP AHIBAIS NEC80CSIED 

TABLE Al (CONTINUED) 

CONTROL (OKTB) IC» DOSE HIGH DOSE 

01-0330 01-0280 01-0285 

 ;25) £WJ :«7)
 

1 (2*)
 

1 ;2U) 1 ;2X)
 

 (25)	 (19) (»7)
 

1 (2X)
 

 ;25) ytfy :so) 1 [t«»
 

 (25) [Vil (SO) 

1 (2«) 

 (25)	 («9J (50)
 

1	 (2X)
 

1 !2X)
 

 (25) (19> (50) 

1 '.2X) 

 EXABIIfED HICPOSCOPICAI.LT 

A-6 
TABLE Ai (CONCLUDED) 

CONTROL(OtlTR) LOW DOSE HIGH DOSE 

01-0330 01-0280 01-0285 

fNIMA L DISPOSITION SUNMR Y 

«.NIH»LS INITIALLY IN STDDY 2f Sft SO 

N»TORA L BE6TK9 5 1 8 

MORIBUN D SAC»IFIfE 3 1 5 

SCHEEOIFD SACRIFTi~E 

XCCIDEHTALIY KI1IED 

>NII<AI NJ5SISG 

9 INCIDDES MJTOLTZFD ^»IIH)^LS 

1CMPS SUM1JFY
 

TOTfL ANIMALS WI^H PPTMHRY TOMOPS* 21 «f «9
 

TOT«.L FPIMAPY TDCI^FS 35 tf 1,H$
 

TOT'L JNIMALS WI"H BENIGN TUMORS 21 *« 4%
 

BFNIGN TBMORS 2<) ST 81
 

TO"JL ANIMAL S WITH MJLIGNAN T TOHOES S 

TO^AL H»LIGSM)T TOMORS 5 

THTiL ANIMALS WITH SECONDARY 'ONORS* 2
 -
 TOT»! SECONDARY I'OI!< PS 2
 

TOTAL *NIH»LS WITH TUMORS ONCEP^MNBENIGN TR MMIGNfl^ 1 f t
 

TO^AI OlfEPTAIK TOfCFS J A S
 

TOTAL ANIMALS WITH TOMCRS DNCERTAINPJII«A''Y OR METASTATTC
 

TOTAL UNCiPTUN TBf'RS
 

* PF.IM'RY 1HMOPS: M-L TOHORS EXCEPT SECONDARY T'JHORS
 

t SECONDARY TUMORS: MST'STATir TnMORS OR THMORS INTHSTVE INTO tK ADJACENT 05GAH
 
TABLE A2 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN FEMALE RATS 

TREATED WITH 1,5-NAPHTHALENEDIAMINE 

CONTROL (U'HTR) LOW DOSE HIGH DOSE
 

02-0331 02-0280 02-0285
 

•NIMJLS INITIALLY IN STUDY 25 50 fO
 

JNIMALS NECBOgSIED 21 50 Si
 

ANIMALS EXAMINED HTSTOFHTHOLOGTCSLLY** 2<4 50 50
 

INTEGDM^NTABY SYSTEM
 

'SKIN (2«) (50) (50) 

SCOP MOOS CEII OPCINOt* 2 ;«*) 

fFSPIR>TQR Y SYSTEM 

ITRACHEA if-JJ ;16) ;10) 

PfPILICMt, , NOS 1 (10*) 

dLUNG l2») (50) (50) 

ALVFOLAP/BROtlCHIOLAR MINIMA 1 (2%) 1 (218) 

C-C5LL CARCINOMA , METASTWIC 1 '2X) 

ENDOMETRIA L ST'Omi SARC01A, MS" 1 '.2%) 

HEMATOPOIETTC SYSTEM
 

"inLTTPlE ORGANS ;2U) ;50) |*tt)
 MH.TG.IYMPHOMA, 1ISTT

CCYTIC TYPE 1 («*)
 

LEUKEMIA,NOS 1 (2%)
 ONDTFFERENTIATFD LEDKEHIA 6 ;12X) 1 ;2%)
 

MYEIOM010JYTIC LEUKEBIA 2 ;8%)
 

CI5CD1ATOSY SYSTEM
 

HONE
 

DIGESTIVE S1SIEM
 

«LIVER (21) (51) (»9)
 

OF ANIMAL S HITH TISSUE EXAMINED MICROSCOPICALLY 

* HOMBEP OF ANIBAL S MECROPSIEE 

** EXCLUDES PARTIALLY ADTOLYZED ANIMALS 

A-8 
TABLE A2 (CONTINUED) 

CrWT-nnOVT'!) LO W DOSE H3 BH fl l 

02-0330 02-0280 "2-0785 

B C""CTHC1J 1 (2%) 

II ST^OMS L SS<?CT1B, ISV 1 ;2X) 

(2<») (50) (U9 ) 

T .1 STtnmi S»PC11A, NV 1 ;2%) 

DFI'i1'>Y SYS1E1 

«KID»E Y J34). ;51) ;H9) 

LT OCI" 1 (2H) 1 (?*) 

« KTD"FY/PE;VI " (2U ) (50) (U9 ) 

T"^NSTTTO,<IL-"''LL P»PTLI011 1 (2*) 1 P*) 

KIJFTBJO Y EIADDE " j24) (IB) t«") 

"1S«NSrTICNft-'-''l!. "»PILIOM A 1 ("*) 

nr-oT«:E SYSTH* 

rs or T'JQMA t NO? 1 

«DEPC1», HOS 6 (29*1 1 ;2X) 1 

'c^»n«f "» ;iu*) tfi 

C»PCINO«!t

 1 ;2<) 

BfSOPHIL ADE'IO"? 1 ;2X) 

"PF T tt»''Y CYST'DE'inc? PCTflnwi ,MT;T i 'ju^ 

E"EW«l (W (50) ((19)
 

CnuTT-ft ipFNOIt ? ("X) 1 (2K)
 

CnsTI^AL CA^CTNC"" 1 '.2%\
 

FrLIT"OtAR-CElI C«F 

f-CELI *DaNOH« C-CEIt C»8

 

CYSHDE10MA, S03 

fKI^SL S HTTH TTSSHE EXAMINE D 1TCROSC1PICAILY 

A-9 
TABLE A2 (CONTINUED) 

COH^sm.. (HMT^ ) LOH DOSE HIGH DOSE 

C2-'J311 12-0280 02-0286 

»<M«icRBATI'- I?LETS ;22) t19) ' U 7^ 

•'SIE'-CEI.. '-fur-not^ 1 (•;<) 

FEI""lDnCTT»F SYSTEM 

• >!llH)M»Y Gl»N D {?«} (50) ( **^ )

ADEIC>"II, so-; 1 ;2») i !«} 

If "•"\ttJfTAI fppTirnw f 1

 '2*) 

FTP'sniDE^J"* 1 (17%) 5 ;10%) 13 ;•>«%) 

«rLlTnr>T. 5IiN") (21) (50) (10) fpFC"*JO'!*,'io$ 1 ;i%) 3 ;«x> 8 I 1 c ^^) 

ADE'IOIA, (OS ;io«) 

dn1"'1^!)? (2U) (US) (18) 

>EE'!0''»»C;Kr>C1 , toq 1 (lij) 1 (2H) 

ElfDnff"TTA! 'TPO1"! P^LY" 2 ,'8Si) 11 ;2<"l) 20 ,'12*) 

UMDoiPT' > T « i ^fo"'1 s?sco"" 1 ;ui) 2 ;ut) 1 ;u<) 

<f)TTfpuS/5N!TC1i rTPTU1 1 ! (21) (19) (18) *rfvo~*' 5

ciwo M s , MOC •> (uis) -> (f% ) 

fOV B P Y if^^ (1Q) (»") ijr finti^-cn.!. TIJI)C» 1 (2*) 

XEPV^tJ S SYSTEM 

ro1 

f-HPn«OFHObF CUTTVCC A , TNVAEIV F ' ' 1 (2X) 

«1TO«>, SOS 1 (2%) 1 ("*%) 

• H'BDEPI' H GLfW D ^|*I ;50) t?0) 

'EH" C»1<1 I2«) (50) (50>

 'QOIRIO0

 CEtl rA»CT < «OH» 1 ;«*) '

 

"ZYHPAt' S KlaSD (21) (50) (50) _ 

I VJT«I'""' OF 8 TISSOE * •ttlST"!! OP "NIHH1

? 

A-IO 
TABLE A2 (CONTINUED)
 

CONTO!. (U'lTP) f» D1?P HIGH DOS';
 oj-o-J^O ^2-0780 "2-0285
 

rnS-mOSKEL'TrtI SY?T51 

>ICNF 

BODY r*VTHF.5 

• EOE Y OVITIES (?«) , (50) 1*51 

"ESTTimiOBS, H«!IGJH»T 1 ;2%) 

*»Er"MrN« l CSVITY (2U) (50) jfel'J 

1 ;"%) 

(2t) (5')) ("*) 

"F" 1 ;2") 

'-.TH"P •5YEIFHS 

ATI. 

SOOii-^a " .III FH> T IICB« 1 

ST*01H S'PCOH, IE" 1 

KI«» T DT1":<-SrT?rn SIRiUPT 

"NI1JLS T<iTIT>II T T1' STUDY "><: ^0 "" 

n

f AJIllfLS HTTP ^ISSHE EXTTNE D rTCff'»SC1?I1:» III 

CF 
TABLE A2 (CONTINUED) 

LfH DOSE HIGH DOSE
 

12-0280 17-02P5
 

JTT-IJ pcT(., PY T!jHr-j>s* 17 ** »ft 

T-IL ETdipil Tl'I^R? 1U <ft 70 

TO"H <>)It!>Lf V-"V 1'TIGHJN T THMRPS 10 2t - 35 

•HJITRS 11 ff 33 

~r>T' L 4NIIHLS HI?H SrCCND^B Y TD1CFS* f J 

'^TPIT sF^orn'^ r TT"°SS t 1 

* ft , i«los«; It a 

C? il 

IJPS: *IT- "i)»op<; EXCEPT S3cosszrr T'Jioss TU>IOFS : «FTtSTfT T ~ ^nM^^ S 0°

 1nj'!'1I>S tnv«S''V? T.NTO AN JDJSCEK" 

A-12 
APPENDIX B
 

SUMMARY OF THE INCIDENCE OF NEOPLASMS
 

IN MICE TREATED WITH 1,5-NAPHTHALENEDIAMINE
 

TABLE Bl

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN MALE MICE 

TREATED WITH 1,5-NAPHTHALENEDIAMINE 

CONT^OI tt'N'"1?) !"W DOSE 

T • NIHAI S INITItli y K •'TODY SO 

SNTMUL S "ITSSINf; 2 

»1T«1IS NEC'CPSTED 

39 

SYSTE1
 

TISSUE («"') («?)
 

SYSTEM
 

2 (UX)
 

6 I11*)
 

»|SIT6»I»»T 1YHPHC1' , 'CS 11 1 (2«) 1 t'*J 

ftLIG.I,Y»PHO("'i, !Y«t>HOCY'TC TYPE 3 ;6%) S, HISTIOCYTT

C TfP*1 

(38 ) 

1 ,'3S) 

*1£SFN"'E''IC L. NODE (3f ) 

«?!TG.LY!«PHOK« , HISTIOCYTIC TY"E 1 I?*)

 

"ILIG'H'IT IYHPH1«^. «I TX ID TYPE 2

 

(13 ) (29) (U) 

CIPCOl» T 1KY SYSIEtt 

» NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY 

* NUMBER OF ANIMALS NECROPSIED 

"EXCLUDES PARTIALLY AUTOLYZED ANIMALS 

B-3 
TABLE Bl (CONTINUED) 

-01'r'=nL(Tlt|-';)

0*1-03^^

 I"K DOSE

 05~^28ei

 HIG H COSE 

Q5-02 Q 0 

T

V" (3° ) ' (U5 ) (Hi) 

HF.o«TiCi 1 LnT,AR

H"P» TOCFLLOHP

 fDENOlU

 rt^c-"tQ»\ 12 ;31*)

 3

 1n

 (7*)

 ;22%)

 * (10*) 

7 ;16*) 

SYSTEM 

'"-f^II ADENO» » 1 (2JS) 

ELJDDIE (37) {»5l (U1 ) 

MJ-.pmH3A

TO» NSITTO N t T_-"EIT P*PIL10 M *

 (-)5) |((-rj ( u o ) 

1 (2^) 

iPD^ p >J 1 T '*£) ^2) ^ U 2 ) 

1 (2*) 

(3«) (U6 ) (U3 ) 

1 (2%) 1 (2X) 

:~1If.-.-C''T.l ;3E'!Or^ r

"tjT IB-"?II cfc-Koi1" IL "DSKO"'

 71

 2

 ;i-;%)

 ;2%)

 ;UK) 

 1U

 1

 ;3'«) 

 ;2t> 

LI '-»nCTSO'"t « ;*>*) 

SYSTEM 

if f!JT(«iLF «T"H TISSOE I tY 

1? ^ST"*!" NEf"CESTE D 
TABLE Bl (CONTINUED) 

'•ONlrSni ('!»!"} IOW DOSS HIGH 

NON F 

C'VITIF * 

f*S> (if) (U9 ) 

M>, (IOS 

v.i O^HEB SY;IF«S 

ANII' I DISPOSITIO N S'JIfl'SY 

• ITHA' S •>MTT SIIY T> ) STUEY 1* ^0 5*1 

Hf-nst i DjifTH a

MOP TB'JN D 5HCfrF T O E

 »J

 4 * 

1

9 

 *** 

. 

SOHEEHI^ E =lic I'

T F -r

~E 

«CCT DEN'''LL Y KILLE D 

T'SMINU S^CFI^TOt %J 3*5 fa 

» N T I " »L CISSTKG "9 1 

HNT1M S «ITH TISSUE EX^W T '^E D HT C"1SCOPICU LY 

\0*M»Lt; NH^HOtS T ED 
TABLE Bl (CONCLUDED) 

1CK DOSE 

TS-028S 

HIGH DOSE 

15-0290 

-OTJL *'!T(1'-,,F HI^H PPTRl.SY TUC"PS*

ivirt* P^IMfY TO!"'"1S

.P SI'H BSTIGN Trifles? 

'I PCS'? 

t<* 

 '9 

41 

Sft 

21 

»f 

** 

12 

SECuVDiSY 

8ENTG M ".? PA 

TOTS! OVCSPTUN T'IMOFS 

! 1= «!FTtslr*"'rf 

i. U1SCEPTJTK "OMOH S 

!Y inroFS: t 

»."Y TUMOSS: TUK1I:,'5 T OSGSN 

8-6 
TABLE B2

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN FEMALE MICE 

TREATED WITH 1,5-NAPHTHALENEDIAMINE

 

T MITIULS INTTIAU Y k S"!1DY

I'lIM'!." M T S C I N G

NFC'OPTTED

•SKTK

•IIXED KFSiKCHYHH "DHPtl, flUTR'H

1 

p C'"CINOf» , BFTlST

'•inL^TPLB O'G'l'S

"HIIGMmT IYPFPCIH, NOS

«HTG.lT*PHOf<P , !YrPBiCYTTC TYPE

If t*T.lYMPHO«f. , HTSTJOCYTT TYP P

• QDNF 1»n°"« HEP»TOCFII.'n \" -f "CmOMA, MF-rasT

HEPfTncFiLnus '•sorniniix, HET«S1

i, 1TXFD ^Ypf

t!irDI>SII1«I I.KOD E

•:ONTF'H (UVT")

oe-T»T>

 5fl

 119

1)9

 £**>

»*1

 JS¥)

 ~1C (21?)

 1 ;2)f) 

 (UC )

1 ;?%) 

 (Utt)

0» WTTfl TISSUE EXM<INSD 

** EXCLUDES PARTIALLY AUTOLYZED ANIMALS
 

 L"W DOSS PIW 

•)6-'>285 if­

 50 "K 

1 

 50 15 

 tt4 U f 

 ;5i) :a*

1 

 (50) t«e 

1 

2 I"*) 

 ;^1) J6^ 

1 (2<) 

1 '.2«) 

(

u<; [43)

 1 (2<) 

1 (2*) 

1 J2*) 

 (US) |40 
TABLE B2 (CONTINUED) 

DOSE DC";" 

, ics (UU)

1 ('%) 

 (U5) 

HEP/TOC'IHn «° '•I'^IKCKd, 1fL-G1l«T lYPIPPOIt , NOS METJ.S T 2 («% ) 

««ITqm«» lYfDHOB* , M'KE D TYDE 

» LT V < F («€) (OR ) 

, NCS ('«) 

, « T X E C 

(»"*» 

» EY (US ) 

(«*) 

;U9 ) 

1 (•>«) 

*I'VF' 

(6%) 11 

' •"">CEILOL»P 

(18) 

1 (2%) 

(UK) 

piprticiH 

' C'Trifni's, 2 ;« 

•UK, 

»,H <!YSIE1 

,L.J;?I. i 
TABLE B2 (CONTINUED) 

CnvTP"! (DNTR ) LOW DOSE HTf!F DCS^ 

nf-rt33rt 15-0285 n<-OOQ Q 

SflDOPHT "DENON H 1 (1*) 

C»»N'I ((.6) (l|!l) (fl L) 

(UU ) (!*r<) («5 ) 

«DEHOi!

»<"H IrO! AS-CELI "DECOHf

fOLtlCDlAa-CELt CHK<-IN01i

 2

 2

 ;S%)

 fK)

 1

 7 (2«) ;11%)

 ­>

 1" 

1

 (OX ) 

 ;2 

f-CEL' 1DLNOMP 1 ;2X) 2 ;M 

PrF'lL»"Y CYSTfOENOI* , MO'1 9 ;18!5) U ;0%) 

''^^US l^ 1-) t^) (^3 ) 

- (2*) 1 (2f) 

"fST

(*") C*c) 1 (2*) i

6

-

1

) 

1

 ;2*> 

»ci;VOn<; SYSTEM 

SfCT'L SENS E ORGAN S 

>, HOS (U«S) (50)1 ;2*> C>P) 

n"5r:niosKlri,Efrti SYST^I 

N05i_ . 

« NUXBF1? OF fNIMJL S WITH TISSUE EXAMINE D 

* 1U1B"0 OF 
TABLE B2 (CONCLUDED) 

rONT""! (n>»TP) 1^« DOSE HIGH DOSE
 

06-0285 n6-02°0
 

(19) (16) 

« ' :SOTHf' TOnP, KOS 1 (2S) 

NON" 

Dt«TH » 

KriT'D 

T

!""ItIDE° M'r 

S'lHVFI 

21 37 

28 71 

23
 

T<-TM» I BENIG N 35
 

»T"H "I'lIGNAN -

IB 3d 

S1I1US "DMOPS* t 

TOTAL JHI1BLS SITU TrjBC"S UWFPT" 

BEH'";tI OS M^l^BVlUT 

TOTHL 'NT1AI.S SITH T«CCFS OKrECTf IH1BMOBS: «II "DHOBS EXCEPT SECONPAFY '
 

»Y "UfOSS: H'?ir»STATT': "OKOPS no TOfOHS ISTO AK 4EJACE1T 0"SAH
 

B-IO 
APPENDIX C
 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC
 

LESIONS IN RATS TREATED WITH 1,5-NAPHTHALENEDIAMINE
 

TABLE Cl

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN MALE RATS 

TREATED WITH 1,5-NAPHTHALENEDIAMINE

 

CO»TPOI (II1TR) LOW DOSF IT^F D1SE 

01-n3^ 01-0280 C1-02P C 

e Hi!TIAI f> T.Y IK STHDY ?« *iO

T«TFGII(!ENT«PY SYSTEM 

*SKTV

VE'JET^EIE ?OFET3»' P"DY

 (25) (U<< ) 1 J2H)  |f$| 

EPIDEHK'I INCTtisiTM CYST 1 ;2%) ( 

fB'JCESS,

•j^is

 KC?

 1 ;I»K) 

1 ;2X) 

TY^T, Nti« 1 ;2«) 

t;^(;pTP)i TQ^ Y SYSTEM 

r

N , f C'IT5/C'iPC»ITC * ' 1 (2%) 

01, <-f-i>Ot!TC 11 (22% ) 1 (2% ) 

"IF (25) (it'') ("7) 

, «OS 2 (11*) 

ILOSG (25) (J"*) (07) 

E"ORCHIi:CT'SIS u (Pit) 3 (6«) 

S"01CHOFK£nKON'rl':lE':ROTTZmG 1 («!() 

ABSCESS, SOS ' 1 '.2*) 

tBOUE 

I NOHFE" TF *ST1HS WITH TISSTE EX^TIllEO 

* IOKBP" or •ai'uts NECSOFSTE!:
 

**EXCLUDES PARTIALLY AUTOLYZED ANIMALS 
TABLE Cl (CONTINUED) 

L CD D1SE HTGP DOSE 

01-0280 "1-0285 

HfXOS T ri?OSIS 2 (IX) 

*ifKOT EU!:> D L. NODE (2&J (if) (47) 

.«, pi i**t CELI 2 ;e*) 

MY("IS |«"»| (35) (1*J 

<"YST, MOS 1 (3X) 

,,-,-,, Y SYTTFH 

1 (2X) 

25 (SIX) 1 

jjiPismnuT^ox, 

D'3PSTTTE SYSiFI " 

T P^EIVE 1 (2X) 1 (U%) 

DE'3E!»E')».'!ION, VO? 1 ;"*) 

1

i •«%> 1 ;2X) ' ,'"«) 3 ;exj 3 ;ex) 1 ;2X) ctOR-CEti. CH^VSE 1 ;?«) 

»tiGTEr'r»sis 1 ;ut) 

rT^BOSlS, HOS 1 <?X) 

jwrpif»<; fff} (U8) (US) 

FI^OSI', FOC»' 1 (2X) 

«-BOFHY, Fr><"H. 1 ;2X) 

H "J^J (I1) ("7) 

R, ACOT" 1 (2X) 

>L CE!l 1 ;2X) 

I (HIIBE" D F ANI"JH5 «I"H "ISSUE EXBMIIED >!ICP5SCOPIC'til 

« HI!K>"'H OF 'H'H*!* VE 
I

•cotov

Pf^STlTSd

tl^INUPY SYSTEM 

IKTDNEY

CYST, KOS

PYEIONF.PH3ITTS, FOCAL 

PYELONEPHRITIS , CHRONIC

NFPHBOS T S , NOS

HY"*6PLfSI», SPITHpLm

 

IKirNfY/CCPi'EX

lOITIIOCOLU" CYST

IKICNEY/PIIVI S

fALCDlUS , NOS 

fO°T»>' JY ElfDDE F

'•ILCULOS, NOS

INDCCRIN E SYSTEM 

lEITUITA'IY

HY'ISPI'SH, FOC^L 

»»D"HN> L

HYPFHD!»SI» , MOEDL' P 

•1DSEHHI COHTEX

HYPE8PI«SH, NOS 

HYPFRPIRSIt , "OCP1

•ADHEHM ftEDUIlf

HYPIRPUSI*, NOS

HYPEHPtSSIf, TOCJ.L 

CYST, NOS 

fOLt1C OL *8 CYS ^ NOS 

INFUNIHTION, CHPONIC 

HYPRPPI'SIil, C-CELI 

tPJPfHYROTD 

VDHPF" OF UIMS1S WITH TISStlE

-It »BTHHS HF 

TABLE Cl (CONTINUED) 

CONTROL (ONTR) ICW DOSE 

01-0331 01-0280 

;23, -.US, ' ' 5 (11*) 

;49) 

1 (2*) 

1 ;2X) 

21 IRtt*) U 2 ;86X) 

C>5) (US) 

1 (UX ) 

fast W9) 

ffs) (*9) 

2 (4%) 

J*|} 't'l 

{S<} (18) 

f»l (1(8) 

1 ;U3!) 

(2tt (2»)) (US) 1

1 (2«) 

(*8)

 (2*> a ;17X) 

(21) (tt-7) 

! -7 

(13) (2K) 

EXABISED (IICFOSCOPIC» LLY 

C-5 

HIGH DOS'! 

01-0781; 

;u6) 

;u8) 

1

 -.2*) 

15 ;31%) 2 ;&*) 

(U8) 

(U8 ) 

1 (2X) 

(US ) 

1 (2*) 

• a u) 

1(2X) 

(U8 ) 

1 (2«) 

(U8 ) 

3 (Pit) 

(U8S) 

2 (0%(i )

(07) 

« i 

(28) 

2.1. 
TABLE Cl (CONTINUED) 

CONTR 10» DOSE HIGH DOSH 

01-0280 01-0285 

IFfVpl-S^IC ISIET? (US ) 

BYPFR"I'SI» , NO'S (2*> 

SFPPOO'JCTTV SYSTEM 

r BISND .•50) 

ABSCISS, SOS 1 (2*) 

•«'1C«'T COC? (25)

1 ;i»X) 

(49) 

(25) (U9) 

TKFLAIM»TIOJI. ? (8%) 

3 

TKF T f im^xOK, 

• TijnFHY, iJO5 17 ; 

Y, NOS 

(25) (U9) 

17 ; 

, NOS 10 

1 12*) 

3 

STSTP1 

2 («*) 1 .(2*) 

STI1 (ttl) 

SFFCI" SF.H<=E 

•«g^T ,__ 

t BUBBS" OF J»T««LS HITH TISSIJF IX»!«I1ED "!J<~"0«;'~>PI<m,I,Y 

TF ^SIMUS HEC5CPSTED 
TABLE Cl (CONCLUDED) 

•ro»TROL(OHTH) LOW DOSE HIGH COSE 

"1-1330 01-0280 01-0285 

L SYSTE" 

NOUF 

Bt-DY f\VITTF.S 

"1EDOMIHU CAVT'Y

F^l

 ;25) Jl^)

 1 (2X)

 :51) 

? (tt%) 

A I L OTHF5 SYSTEMS 

SFTCIfL NOEFHOLOGY SU1F""t 

HO LESION FEPOBTED ft % 

AO"0/NE'-''OPSY/HO HISTO 1 

MJinLYSIS/NO ISC'iosy 1 

I NU1B2P OF AHIMJL S «TT H TISSBE ETHilHED MI'-'!OSCOPTCA LLY 

» NU1EIP IF »NT«»LS NE^FOPSTED 

fr-7 
TABLE C2

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN FEMALE RATS 

TREATED WITH 1,5-NAPHTHALENEDIAMINE

 

LOH DOSS HIG H EOSE 

02-" 330 02-0280 "2-02P5 

• UTNUL S INITT413.Y IN STtlDY 50 

«>l T MJ>r - NECEOPS'E O 50 

2U 50 50 

i"TtGUiiEN''»' >Y SYSTEM 

SY^TE H 

• I'fYN X (2«) (50) 1*0)

 

1 (2X> 

(2U) (50) 

(SO) 

2 (a*) 

I\, >IOS 

;n>ITJ, rH'PNTC NU'IN E 2 :« 

SYFT"1 

, »CS 

(2*) 1 

*SPLF"N (23) (50) (19) 

, NOS 

i pit) 

(H8) 

(2X) 

«MtHl)-:EOHc L. NOEE (19) (13) 

Of »MTHHS UI^H TISSUE m CI USD HICSOSTOPIOLLY 

* NH".Ef» OF MIMLS 11ECROPSIED 

**EXCLUDES PARTIALLY AUTOLYZED ANIMALS 
HYPIEPI'SIf, PISS1A CE1I 

'HI SYSTEM 

DEGEBF9«TI01, »CS 

OC^CTOH 

C^ICIFIC'TTON, FOOI 

WED!*' Tf
 

CALCTFIC'IION, HOS
 

C^GF^TIVE SYSTEM 

f-OHGESTTOM, CHR'-NTC 

CHOI^IGIPFIEPOSIS 

T'IFfBCT, FOCHt 

«FT")OPPHOSIS PIT^Y 

SA^TPHIIIC CYTP CH»NG E 

CI"\t-CBli CHANGE 

«PGIE"T> SIS 

HYPESFUSIB, 

, HOS 

fTPOPHY , F0f»l 

• STOHt'-H 

PTYPI» , HOS 

HY»EBPlf«!», BASUl CEIL 

• '•"inn 

SYS1EH 

» NnBSEH OF »NIH»1S »tTH TISSUE

• NOHBE* OF ^HIBUS HECPCPSIID 

TABLE C2 (CONTINUED) 

Log DOSE HIGH DOSH 

02-033T 12-0280 12-02P5 

2 (11*) 

I2U) 

(10*) 2 («*) 

(21) (SO)

 

1 (UK ) 

(21) 

(51) 

1 (2X) 

2 I"*) 

12 ; 

(SO) 

2 («) (2") 

(22) 

(2*) 

(50) 

2 !«) 3

 16 

(20) (tt7) (U6) 

3 (6%) 

1.J2SO __ 

 EXAMINE D 1IC"OSCOPICMIY 

C-9 
TABLE C2 (CONTINUED) 

i, NOS 

NFPHBOSIS , 10S 

NECBOSIS, MEDDLUm 

CM.riTIC»riOH, FO'TAL 

•KTDNEY/TBBOLE
 

OICIFICMIOH, SOS
 

EKDOr-" TME SYSfM 

HYPTPPl»£I« , NOS 

DUEN' I HEEUI H 

HYP'RPIASIS , HOS 

CYS-, >JOS 

IBCTS'IOK 

HYDFOMETFA 

EPIDBPMfl INCinSTOK CYST
 

5, NOS
 

B"PHY, NOS 

I Hn»B"T OF KTTH,TISSO» 

OF 

COSTS"!(UNTE)

02-0330

8 ;33%) 

1 ("») 

1 JSX) 

1 I1!*) 

ft* 

 1C» DCSE 

 02-0280 

1 (2X) 

7 

(50) 

;50)

1 (2X) 

(50) 

1 (2*) 

1 (2*) 

12 ;
 

(50)
 

1 (2X)
 

6
 

HIRH DOSE 

07-0285 

;U7) 

1 (2*) 

(18)
 

1 I2X)
 

3 ;6H)
 

"C-IO 
TABLE C2 (CONTINUED) 

. (UHfi) LOW DOSE HTGH CCSE 

02-0330 02-0280 02-0285 

TNE1»MH>1IOH, SOPPDBtTIVE 3 (13*) 

«, CHPOHIC

usi», CYSTIC

1, SQIMHODS 1 ;<(!{) 

1 ;2%) i ;e*) i ;?*) 

«OVi«Y/OVIDDCT

TKFLWKHTIOH, SOPPOFATIVE

ABSCESS, NCS

 (2*)

? (21«) (US) (18) 

2 ;<»«) 

i-YST, NOS

Tt'THMMATIOH, S CPD 0!>R'T'IVE

 2

 1

 (8%)

 ft*) 

8 (16*) 

TNFll"1MATIOtI, fUTE 1 J2X) 

'BSCE^j, SOS 1 ;2S) 

SEHVO'JS SYSTEM 

3, SCS ' 1 (2*) 

SEECTAI. SIHSE OR6»NS 

TION, NOS

antBT

 * 1

 1

 ;2X) 

 ;2«) 

EOEY CAVITIE S 

»SBDOHIH» I CAVITY (21) (50) (= 0) 

1H O^HSP SY£TE«S 

CS»»T"BOCCil POOCH 

CYST, >ICS 1 

SF»CI»l (lOBPHOIOGY S011«RY 

t NOME*" OF &HIHJLIS WITH TISSDE IXUHIHED MTCDOSCOPICALLY 

* HOSE?" OF «HT1»IS KB'-POFSIED 

C-ll 
APPENDIX D
 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC
 

LESIONS IN MICE TREATED WITH 1,5-NAPHTHALENEDIAMINE
 

TABLE Dl

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN MALE MICE 

TREATED WITH 1,5-NAPHTHALENEDIAMINE 

70t!THOI. (OUT") LOW DOSS H T G P DOSE1 

05-"331 15-0285 05-099(1 

im-uiiY in sinDY so *o ro »IISSING

 2 1 

RECPOPSTE D 39 If U9 

ISIIUIS IXJ1I1.ED HIS"OP4TBOIOGTr«HY*» 39 U7 U6 

I^TIGnHEH^JPY SYSTEM 

•SKTK ;39) ;»"») ;u<)) 

HPIDESHU Tji/'iosiT^ CYST 1 C<«) 

''"["SUE (39) (If) («9) 

ABSTSS, NO S 1 ;2«) 

, NCS •• (">») 

I, IYBPHCT E 1 p«) 

5TS 1 ('») 2 

"YTHPOFOIE^IS ' ;9U) 

^. NOPF (36) (U3) 

PLtSIf CUT. 1 (3%) 

«MESFNTE'Ir I. NODE (36) (t3) 

HYPPPPIISI'I IYMOHOID 1 

I lO'E"" OT »IIfi;i<! WTTH TISSUE EXAMINE D HTCPOSCOOICP T LY 

« N01FE" C? Mi TN»l c S"-':OF'!TEE 

"EXCLUDES PARTIALLY AUTOLYZED ANIMALS 

D-3 
TABLE Dl (CONTINUED) 

(OMTR) inW DOSE HIGH DOSE 

05-0285 P5-0290 

SYSISH 

IITVE" 

(2X) CLB»I>-Cr

It CHJNG F 

•SIS, F1C1L 

KpiHC'i'flC DUCT 

tiTL»T»TT O N , DOS 

(36) (U6) 

1 CX) 

(tO) 

«EOOEHI01 (37) (U7) (39) 

*MYLOTD"SIS 1 <3« ) 

IJFJUHD M 

M1YLOICOSIS 

(37, 

1 (31) 

(117) (39) 

1 (3«) 

IT1S01 (37) («7) (19) 

iHYLO'C^SIS ? (c

*) 

ITOI.ON 

pfPSS-TTCf, (37) (HO) 

2 (5X) 

(36) 

STSt^B 

» KTDVEY 

5, FOr^l 

2 (I*) 

FYETOHEFH8 T TIS , C^llIC 

FCT, HE1LED 

\CY10TOnSIS 

CJICTICH1'"'!, NOS 

(HE) 

(3%) 

(35) (15) 

«?i'iT0"t»i TISSUE (35) (U5) 

'BSCF5S, SOS 1 (2*1 

(39) (17) 

EXMIISED »t< OF »NIf!»L? 

0-4 
TABLE Dl (CONTINUED) 

LC M DOSE HTG P 

»TNH«Y El'ODE" 

CALCULUS, HOS O7

) (U5 ) 

(2%) 

(2%) 

(3P) C46) 

1 ('*) 

1 ;->*) 

' I1*) 

.-2*) HYOEROLfSIP, PJP'ILPPY 

HYP"FFI«SI», 

'8) 

1

 (f^l 

«pf RrpP 

-SI', NOE ^) 

1 (i») 

•««) 

DTH"»»T(-H, 

S SYS15M 

•147) 

1 

(ttO) 

SPECIAL SEN^E O'JGA'JS 

• HI)i!BEr OF 'N T «AL S WITH IC' 11Y * Ma«B»"> Of PNIMAI S •Trp 

0-5 
TABLE Dl (CONCLUDED) 

loi (I1VT1) L"W DOSE H'GH DOSE 

i-p" 05-028^ OS-C2°0 

'39) 'KQ) 

Mf i: 

BICDO--OPI'­ * '«t1CE'?

P 1" »NT«HS 

D-6
 
TABLE D2

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN FEMALE MICE 

TREATED WITH 1,5-NAPHTHALENEDIAMINE 

*"!. ('JVT" ) IC W DOS'! 

ST"CY 1 

tlEC^riPSIFD U 9 50 16 E It'll" 111*1) HT ST^P1THrnGir«TT.Y** U9 1(9 uf 

SYSTE M 

;snj 

f ucpTPSTflpy Sf 

(US ) 

CNTI , NOS 

2 

1u 

2 

*9r>NE rSRioii 

FTE'CSI": (2«)

 

HYPF.SPIHSIA, <«*)

 

(15) (it 8) 

u ;9J) 

•LYIPf HOCl OF ^HQ1! ' X (UU ) (-15) (ttf.) 

(Ul.) (itS) (•"•') 1 [2^) 

«101B'F t-YXPH NODE (1»U) (U5) (UO ) 

1 (2%) 

(UU ) .«) 111) 

« HOfB" OF »STX«I S W— H TISSUE E ", L1Y 

* 10XBFS OF UNIMfl S "F^BOPSIEC 

**EXCLHDES PARTIALLY AUTOLYZED AKIMALS 

D-7 
TABLE D2 (CONTINUED) 

CCWT^lCItlTB)

06-'133i

 IC» DOSE

 T6-C785

 HTQ H E^SE 

 06-C290 

tiyDlTCCHCI l ( 'VpDHrT D 1 (,,() 

• crip;. IY1"H NOE E (UK ) (<15) (10) 

HTIAT-iFI'aS-' S 1 (2*) 

•T.-,- nT JT O O Y SYSTF1 

T-iu-crj , «ic«! ' ' 1 (2\) 

CI^'-rfLL. rpvi'-T 1 (2X) 

*?-wrcT» 3 prC T,, T t FIT T S (38) (U7) («U ) 1 (2%) 

»-O(-CHY , "<rS * ;?f) 

HJMT'Mn i ("?) (H7 ) (U5 ) 

• 'L^UI (U2 ) (U7) (15) 

.^YLO-cnsiS ' (IX) 

*CYTOTDOSIS ' ' 3 ;6*> 1 ;?*) 

DYST'ST' , N05 1 ;2%) 

«TC"OSCOPIC!1 I1Y 

0-8 
TABLE D2 (CONTINUED) 

. (BM•"•«> ) ITU DCSS 

• FEPFU M 

(Utt) 

•P'FUIIU" 

HV"!;FPI8; 

O'J , M~I1T

^,10 ^

 F1CAL 1 P%) 

? 'U ^ ) 

HyorHD ' » ; 

s,

>,

 PtP T LI'B Y

 F1IlTf-'J!.tp-C-TI ? ;C1!) 

1

 '2%) ' ;u% ) 

('") (1^1 ('

r

) 

'"^"nlL 

(12%) 

(111.) (15) 

(UL ) (15) 1 (0*) 

tto ya p Y 

CYST, vos 

I"")

7 !'<)

('4 = )

 ^ (7% )

 (11) 

^ I"1!!) 

^r«oii^HtGir^CY' ; T U 9 X) 

J?nl";-^, rpr OK r c ? ") 

NE^^CPSIIC 

0-9 
TABLE D2 (CONCLUDED) 

L C H DOS" H T G H ETS E 

jp-0310 16-0265 1f-o?QO 

ftn r ULA q KEIf"~A l SYSTEI 

"-KEf-^L 1t)3rL* (HO ) (51) (U6 ) 

fpqC'^S , NOS 1 (2%) 

Er DY r* V T~TP c 

>-BD"« T tJ!l r»v T T Y

N pr- c -I = < pj ^ ;UQ ) ;si) •( (2% ) 

 ;ui) 

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