National Cancer Institute 

CARCINOGENESIS 

Technical Report Series 

No. 84 

1978 

BIOASSAY OF 

2,4-DIAMINOANISOLE SULFATE 

FOR POSSIBLE CARCINOGENICITY
 

CAS No. 615-05-4 

NCI-CG-TR-84 

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

Public Health Service 

National Institutes of Health 

BIDASSAY OF
 

2,4-DIAMINOANISOLE SULFATE
 

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) 78-1334
 

REPORT ON THE BIOASSAY OF 2,4-DIAMINOANISOLE SULFATE
 

FOR POSSIBLE CARCINOGEN1CITY
 

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
 

2,4-diaminoanisole sulfate 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 ca'pacity 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 2,4-diaminoanisole sulfate 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).
 

Histopathologic examinations were performed by Dr. D. W. Hayden
 

(3), Dr. A. S. Krishna Murthy (3) and Dr. A. Russfield (3) at the
 

Mason Research Institute, and the diagnoses included in this report
 

represent the interpretation of these pathologists. Histopathology
 

findings and reports were reviewed by Dr. R. L. Schueler (4).
 

Compilation of individual animal survival, pathology, and summary
 

tables was performed by EG&G Mason Research Institute (5); the statistical analysis was performed by Mr. W. W. Belew (6), and Dr. A. Chu (5),
 

using methods selected for the Bioassay Program by Dr. J. J. Gart (7).
 

This report was prepared at METREK, a Division of The MITRE Corporation (6) under the direction of the NCI. Those responsible for
 

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

(6), the task leader, Dr. M. R. Kornreich (6), the senior biologist,
 

Ms. P. Walker (6), the chemist, Dr. N. Zimmerman (6), and the technical editor, Ms. P. A. Miller (6). The final report was reviewed by
 

members of the participating organizations.
 

The statistical analysis was reviewed by members of the Mathematical Statistics and Applied Mathematics Section of the NCI: Dr.
 

J. J. Gart (7), Mr. J. Nam (7), Dr. H. M. Pettigrew (7), and Dr. R.
 

E. Tarone (7).
 

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), Dr. R.
 

A. Griesemer (1), Dr. H. A. Milman (1), Dr. T. W. Orme (1), Dr. R. A.
 

Squire (1,8), 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.	 Now with 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.	 Tracer Jitco, Inc., 1776 East Jefferson Street, Rockville,
 

Maryland.
 

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

Rockville, Maryland.
 

6.	 Th MITR e E Corporation, METREK Division, 1820 Dolley Madison
 

Boulevard, McLean, Virginia.
 

7.	 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.
 

8.	 No wit w h the Division of Comparative Medicine, Johns Hopkins
 

University, School of Medicine, Traylor Building, Baltimore,
 

Maryland.
 

iv
 
SUMMARY 

A bioassay of technical-grade 2,4-diaminoanisole sulfate for
 

possible carcinogenicity was conducted using Fischer 344 rats and
 

B6C3F1 mice. 2,4-Diaminoanisole sulfate was administered in the
 

feed, at either of two concentrations, to groups of 50 male and 50
 

female animals of each species. The time-weighted average dietary
 

concentrations used in the chronic bioassay were 0.12 percent for
 

the low dose rats and 0.5 percent for the high dose rats. The dietary concentrations used for low and high dose mice were 0.12 and
 

0.24 percent, respectively. After a 78-week period of chemical
 

administration, observation of the rats continued for an additional
 

29 weeks and observation of the mice continued for an additional 19
 

weeks. For each species, 49 or 50 animals of each sex were placed
 

on test as controls.
 

In both species, adequate numbers of animals in all groups survived long enough to be at risk from late-developing tumors.
 

In high dose male and female rat groups, the proportion of animals having one or more of the following malignant follicular-cell
 

thyroid tumors: papillary adenocarcinomas, follicular-cell carcinomas, papillary cystadenocarcinomas, or adenocarcinomas NOS, was
 

significantly greater than the proportion in corresponding control
 

groups. For high dose male rats the proportion of animals having
 

either a C-cell adenoma or a C-cell carcinoma was also significantly
 

increased.
 

The incidence of malignant tumors (squamous-cell carcinomas,
 

basal-cell carcinomas, or sebaceous adenocarcinomas) of the skin
 

and its associated glands were significantly increased among high
 

dose rats of both sexes.
 

Among high dose female mice, the combined incidence of thyroid
 

follicular-cell adenomas and carcinomas was significantly increased.
 

Among high dose male mice, the incidence of thyroid follicular-cell
 

adenomas was significantly increased, but no follicular-cell carcinomas were observed.
 

Under the conditions of this bioassay, technical-grade 2,4­

diaminoanisole sulfate was carcinogenic to both sexes of both species. In Fischer 344 rats dietary administration of the chemical
 

induced increased incidences of malignant tumors of the skin and
 

its associated glands and malignant thyroid tumors in each sex.
 

In B6C3F1 mice, dietary administration of 2,4-diaminoanisole sulfate induced thyroid tumors in each sex.
 

v 

TABLE OF CONTENTS
 

I. INTRODUCTION
 

II. MATERIALS AND METHODS
 

A. Chemicals
 

B. Dietary Preparation
 

C. Animals
 

D. Animal Maintenance
 

E. Selection of Initial Concentrations
 

F. Experimental Design
 

G. Clinical and Histopathologic Examinations
 

H. Data Recording and Statistical Analyses
 

III. CHRONIC TESTING RESULTS: RATS
 

A. Body Weights and Clinical Observations
 

B. Survival
 

C. Pathology
 

D. Statistical Analyses of Results
 

IV. CHRONIC TESTING RESULTS: MICE
 

A. Body Weights and Clinical Observations
 

B. Survival
 

C. Pathology
 

D. Statistical Analyses of Results
 

V. DISCUSSION
 

VI. BIBLIOGRAPHY
 

APPENDIX A SUMMARY OF THE INCIDENCE OF NEOPLASMS IN
 

RATS TREATED WITH 2,4-DIAMINOANISOLE SULFATE
 

APPENDIX B SUMMARY OF THE INCIDENCE OF NEOPLASMS IN
 

MICE TREATED WITH 2,4-DIAMINOANISOLE SULFATE
 

APPENDIX C SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC
 

LESIONS IN RATS TREATED WITH 2,4-DIAMINOANISOLE SULFATE
 

APPENDIX D SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC
 

LESIONS IN MICE TREATED WITH 2,4-DIAMINOANISOLE SULFATE
 

Page
 

1
 

5
 

5
 

5
 

6
 

6
 

10
 

11
 

14
 

16
 

21
 

21
 

21
 

24
 

32
 

50
 

50
 

50
 

53
 

55
 

64
 

67
 

A-l
 

B-l
 

C-l
 

n-i
 

vii
 
LIST OF ILLUSTRATIONS
 

Figure Number Page 

1 GROWTH CURVES FOR 2,4-DIAMINOANISOLE SULFATE CHRONIC STUDY RATS 

22 

2 SURVIVAL COMPARISONS OF 2,4-DIAMINOANISOLE 

SULFATE CHRONIC STUDY RATS 

23 

3 GROWTH CURVES FOR 2,4-DIAMINOANISOLE SULFATE CHRONIC STUDY MICE 

51 

4 SURVIVAL COMPARISONS OF 2,4-DIAMINOANISOLE 

SULFATE CHRONIC STUDY MICE 

52 

LIST OF TABLES 

Table Number Page 

1 DESIGN SUMMARY FOR FISCHER 344 RATS—2,4­

DIAMINOANISOLE SULFATE FEEDING EXPERIMENT 

12 

2 DESIGN SUMMARY FOR B6C3F1 MICE—2,4-DIAMINOANISOLE SULFATE FEEDING EXPERIMENT 

13 

3 EPITHELIAL TUMORS OF THE INTEGUMENTARY SYSTEM FOUND IN FISCHER 344 RATS 

26 

4 ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS 

AT SPECIFIC SITES IN MALE RATS TREATED WITH 

2,4-DIAMINOANISOLE SULFATE 

33 

5 ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS 

AT SPECIFIC SITES IN FEMALE RATS TREATED 

WITH 2,4-DIAMINOANISOLE SULFATE 

40 

6 ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS 

AT SPECIFIC SITES IN MALE MICE TREATED WITH 

2,4-DIAMINOANISOLE SULFATE 

56 

7 ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS 

AT SPECIFIC SITES IN FEMALE MICE TREATED 

WITH 2,4-DIAMINOANISOLE SULFATE 

59 

viii
 
IX
 

I. INTRODUCTION
 

2,4-Diaminoanisole sulfate (NCI No. C01989), a component of many
 

hair dyes, was selected for bioassay by the National Cancer Institute
 

because of the increased incidence of bladder cancer observed among
 

workers in the dye manufacturing industry (Wynder et al., 1963;
 

Anthony and Thomas, 1970). Occupational exposure to several classes
 

of chemicals, including aromatic amino-compounds, is believed to
 

contribute to this increased cancer risk (Wynder et al., 1963), and
 

2,4-diaminoanisole sulfate is especially suspect because it is structurally analogous to the known carcinogen 2,4-diaminotoluene sulfate
 

(ito et al., 1969). The widespread exposure to 2,4-diaminoanisole
 

sulfate among the general population and the possibility of an increased cancer risk among hairdressers (Anthony and Thomas, 1970)
 

were additional factors in its selection for testing.
 

The Chemical Abstracts Service (CAS) Ninth Collective Index (1977)
 

*
 

name for this compound is 4-methoxy-l,3-benzenediamine sulfate. On
 

hair dye labels this compound is commonly called 4-methoxy-m-phenylenediamine sulfate or 4-MMPD. It is also called 2,4-diamino-l-methoxybenzene sulfate; and 2,4-DAA sulfate.
 

2,4-Diaminoanisole sulfate is one of the most frequently used
 

couplers in permanent hair dye formulations (Corbett and Menkart,
 

1973). Imines, produced within the hair shaft by the oxidation of
 

_
 

The CAS registry number is 615-05-4.
 

1
 
a primary intermediate, such as p-phenylenediamine, react with 2,4­

diaminoanisole sulfate to produce blue or purple-blue colors (Corbett,
 

1971; Corbett and Menkart, 1973). The maximum concentration of the
 

compound in hair dye preparations is about 1.5 percent; however, in
 

actual use, the dye base is diluted with an equal volume of hydrogen
 

peroxide solution (Menkart, 1977).
 

2,4-Diaminoanisole sulfate can also be used as an oxidation base
 

to dye furs (Society of Dyers and Colourists, 1971a; as cited in Urso,
 

1977) and as an intermediate for the production of textile dye, C.I.
 

(Colour Index) Basic Brown 2 (Society of Dyers and Colourists, 1971b;
 

as cited in Urso, 1977).
 

Specific production statistics for 2,4-diaminoanisole sulfate are
 

not available; however, the listing of the compound in the 1977 Directory of Chemical Producers, U.S.A. (Stanford Research Institute,
 

1977) implies that 2,4-diaminoanisole sulfate is produced in commercial quantities (greater than 1000 pounds or $1000 in value annually).
 

The estimated annual usage in the United States is 30,000 pounds
 

(Menkart, 1977).
 

Exposure to 2,4-diaminoanisole sulfate may occur at chemical and
 

dye production facilities and among hairdressers using dyes which
 

contain the compound. Dermal contact occurs for all those whose hair
 

is colored with dyes containing 2,4-diaminoanisole sulfate. About 40
 

percent of U.S. women are regular users of hair dyes, and approximately
 

three out of every four dollars spent on hair coloring is spent on the
 

2 
permanent type of dye (Corbett and Menkart, 1973). Additionally,
 

exposure may result from unreacted portions of 2,4-diaminoanisole
 

sulfate reaching rivers and streams via domestic wastewater.
 

2,4-Diaminoanisole sulfate can be absorbed through the skin.
 

It has been recovered after dermal application in the urine of Rhesus
 

monkeys and humans (Maibach, 1977).
 

No data concerning the effects of acute or chronic exposure to
 

2,4-diaminoanisole sulfate in humans are available. No evidence of
 

carcinogenicity was found after up to 18 months of topical applications of hair dye formulations containing 2,4-diaminoanisole sulfate
 

to 200 random-bred Swiss Webster mice of both sexes (Burnett et al.,
 

1975). Seven topical applications of preparations containing 2,4­

diaminoanisole sulfate to 60 female Charles River CD rats during
 

gestation produced no teratogenic effects (Burnett et al., 1976).
 

2,4-Diaminoanisole sulfate has been found to induce frame shift
 

reversions from a histidine-requiring mutant back to prototype in
 

Salmonella typhimurium strain TA 1538 (Ames et al., 1975). Of nine
 

hair dye components found to be mutagenic by Ames et al. (1975),
 

2,4-diaminoanisole sulfate was the most active in reverting TA 1538.
 

Mutagenicity was also indicated by marked difference of inhibition
 

for growth (DIG) in the DNA repair test in Escherichia coli strains
 

B/r WP2 (trp~) and WP100 (trp~, uvrA~, recA~) (Nishioka, 1976). In
 

a recessive lethal assay utilizing Drosophila melanogaster, 2,4­

diaminoanisole sulfate exhibited weak mutagenic activity (Blijleven,
 

3 
1977). Mammalian mutagenicity studies have been performed both in
 

vivo and in vitro. In a forward mutational assay system which
 

utilized the thymidine kinase heterozygous locus of mouse lymphoma
 

cells, 2,4-diaminoanisole sulfate gave a weak response, causing a
 

dose-related increase in mutation frequency and in the ratio of induced to spontaneous mutations (Palmer et al., 1977). The compound
 

was not mutagenic to germ cells in a dominant lethal study of Charles
 

River CD rats following intraperitoneal administration of 20 mg/kg
 

three times weekly for 8 weeks to 20 males (Burnett et al., 1977).
 

4 
II. MATERIALS AND METHODS 

A. Chemicals
 

A single batch of technical-grade 2,4-diaminoanisole sulfate, in
 

powder form, was purchased from American Hoechst and chemical analysis
 

was performed by Mason Research Institute, Worcester, Massachusetts.
 

The range of the experimentally determined melting point (135° to
 

138°C) was relatively narrow. No literature value was reported for
 

this compound. Thin-layer chromatography resulted in a major spot
 

and a single additional spot. Infrared analysis was not inconsistent
 

with the structure of the compound.
 

Throughout this report the term 2,4-diaminoanisole sulfate is used
 

to represent this technical-grade material.
 

B.	 Dietary Preparation
 

The basal laboratory diet for both treated and control animals
 

(R)
 was Wayne Lab-Blox (Allied Mills, Inc., Chicago, Illinois). 2,4­

Diaminoanisole sulfate was administered to the treated animals as a
 

component of the diet.
 

Quantities of the chemical were weighed out and hand-blended in
 

an aluminum bowl with an aliquot of the ground feed. Once visual
 

homogeneity was attained, the mixture was placed in a 6 kg capacity
 

Patterson-Kelley twin-shell stainless steel V-blender along with the
 

remainder of the meal. The blender was sealed and operated for 20
 

minutes. The mixtures were placed in double plastic bags, and stored
 

in the dark at 4°C. The mixture was prepared once weekly.
 

5 
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. Mice and high dose treated and control rats were supplied by
 

Charles River Breeding Laboratories, Inc., Wilmington, Massachusetts.
 

Low dose treated and control rats were supplied by Laboratory Supply
 

Company, Inc., Indianapolis, Indiana. Treated rats and high dose mice
 

were received in separate shipments from their controls. Low dose
 

treated and control mice arrived on the same day.
 

Upon arrival, two males and two females per species per shipment
 

were sacrificed and examined for parasites and other signs of disease.
 

Rats and mice to be used in the bioassay were quarantined by species
 

for 2 weeks prior to initiation of the test. Animals were assigned
 

to groups and distributed among cages so that average body weight per
 

cage was approximately equal for a given species and sex.
 

D. Animal Maintenance
 

All animals were housed by species in rooms having a temperature
 

range of 20° to 28°C. These rooms were not humidity controlled. In­

(R) (R)
 coming 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-hourdaily cycle.
 

6
 
Rats were housed five per cage by sex. During quarantine and
 

for the first 13 months of study, rats were kept in galvanized-steel
 

wire-mesh cages suspended above newspapers. Newspapers under cages
 

were replaced daily, and cages and racks washed weekly. For the remainder of the study, all rats were housed in suspended solid-bottom
 

polycarbonate cages equipped with disposable nonwoven fiber filter
 

sheets. Clean bedding and cages were provided twice weekly. Corn­

®


cob bedding (SAN-I-CEL , Paxton Processing Company, Paxton, Illinois)
 

was used for the entire period of polycarbonate caging for low dose
 

®
 rats and their controls. SAN-I-CEL was also used for high dose rats
 

and their controls during the first 8 months of polycarbonate caging.
 

Hardwood chips (Aspen bedding, American Excelsior Company, Baltimore,
 

Maryland) were used for these groups for the final 3 months of study.
 

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 cages. During quarantine and periods of chemical administration, cages were fitted with
 

perforated stainless steel lids. During the observation period which
 

followed, stainless steel wire bar lids were used. Both types of
 

lids were from Lab Products, Inc., Garfield, New Jersey. Nonwoven
 

fiber filter bonnets were used over cage lids. Low dose mice and
 

their controls were housed ten per cage for the first 18 months of
 

study and five per cage thereafter. The number of high dose rats per
 

cage was reduced to five after 12 months, and the number of high dose
 

7 
control rats per cage was reduced to five after 11 months. Clean
 

cages, lids, and bedding were provided three times per week when cage
 

populations were ten and twice per week when cage populations were
 

• ®
 reduced to five. Ab-sorb-dri hardwood chips (Wilner Wood Products
 

Company, Norway, Maine) were used as bedding for the first month of
 

study for high dose mice and their controls, and for the first 8
 

®
 months of study for low dose mice and their controls. SAN-I-CEL
 

was used for the next 12 months for all mice. Corncob bedding (Bed­

(R)
 o-Cobs , The Andersons Cob Division, Maumee, Ohio) was used for the
 

remainder of the study for low dose mice and their controls, and for
 

the next 8 months for high dose mice and their controls. For the
 

remainder of the study, high dose mice and their controls were provided with Aspen bedding. Reusable filter bonnets and pipe racks
 

were sanitized every 2 weeks throughout the study.
 

Water was available from 250 ml 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.
 

During the 78-week period of chemical administration all dosed
 

animals were supplied with Wayne Lab-Blox meal containing the
 

appropriate concentrations of 2,4-diaminoanisole sulfate. Control
 

animals had untreated meal available. Meal was supplied throughout
 

the study to all mice and to low dose rats and their controls in
 

Alpine® aluminum feed cups (Curtin Matheson Scientific, Inc., Woburn,
 

8
 
Massachusetts) containing stainless steel baffles. High dose rats
 

and their controls were fed from Alpine feed cups for the first 11
 

months of study and from stainless steel gangstyle feed hoppers
 

(Scientific Cages, Inc., Bryan, Texas) thereafter. During the untreated observation period, mice were fed pelleted Wayne Lab-Blox
 

from a wire bar hopper incorporated into the cage lid, and rats were
 

fed pelleted Wayne Lab-Blox on the cage floor. Food hoppers were
 

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

daily in Alpine feed cups.
 

High dose, low dose, and low dose control rats in the 2,4-diaminoanisole sulfate study were housed in a room with other rats receiv-


JU
 

ing diets containing 5-nitro-o-toluidine (99-55-8); hydrazobenzene
 

(530-50-7); 2-aminoanthraquinone (117-79-3); 3-amino-9-ethylcarbazole
 

hydrochloride; 6-nitrobenzimidazole (94-52-0); 1-nitronaphthalene
 

(86-57-7); and APC (8003-03-0). High dose control rats were in a
 

room with other rats receiving diets containing amitrole (61-82-5);
 

2-methyl-l-nitroanthraquinone (129-15-7); and 3-nitro-p-acetophenetide
 

(1777-84-0).
 

High dose mice were housed in a room with other mice receiving
 

diets containing 5-nitro-o-anisidine (99-59-2); 2,5-toluenediamine
 

sulfate (6369-59-1); 5-nitro-o-toluidine (99-55-8); hydrazobenzene
 

(530-50-7); 3-amino-9-ethylcarbazole hydrochloride; 6-nitrobenzimidazole (94-52-0); and 1-nitronaphthalene (86-57-7). Low dose mice and
 _
 

CAS registry numbers are given in parentheses.
 

9
 
all control mice were housed in a room where other mice were receiving diets containing the following compounds: amitrole (61-82-5);
 

N,N-dimethyl-p-nitrosoaniline (138-89-6); 5-nitro-o-anisidine (99-59­

2); 2,5-toluenediamine sulfate (6369-59-1); 2,4-dinitrotoluene (121­

14-2); 2-aminoanthraquinone (117-79-3); 3-amino-4-ethoxyacetanilide
 

(17026-81-2); 3-amino-9-ethylcarbazole hydrochloride; l-amino-2­

methylanthraquinone (82-28-0); 4-nitroanthranilic acid (619-17-0);
 

1-nitronaphthalene (86-57-7); 5-nitroacenaphthene (602-87-9); 3-nitrop-acetophenetide (1777-84-0); and APC (8003-03-0).
 

E. Selection of Initial Concentrations
 

In order to establish the maximum tolerated concentrations of
 

2,4-diaminoanisole sulfate for administration to treated 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. 2,4­

Diaminoanisole sulfate was incorporated into the laboratory diet and
 

supplied ad libitum to five of the six rat groups and five of the six
 

mouse groups in concentrations of 0.075, 0.125, 0.209, 0.348, and
 

0.580 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 4 weeks, followed by a 2-week observation period during which animals were fed the basal diet. Survivors were sacrificed at the end of the test, and gross necropsies
 

were performed.
 

10
 
The initial high concentrations chosen for administration during
 

the chronic test were those causing no deaths, no gross compound-


related abnormalities, and no mean body weight depression in excess
 

of 15 percent relative to controls during the 6-week subchronic test.
 

No gross abnormalities were noted in rats. One male rat died at
 

each of 3 dosages: 0.075, 0.125, and 0.58 percent. Only the latter
 

death was judged compound-related. The initial high concentration
 

chosen for rats in the chronic study was 0.125 percent, but at a later
 

time a group of rats was placed on test at a higher concentration.
 

No gross abnormalities were observed in mice. One male mouse
 

died at a concentration of 0.58 percent. The initial high concentration used for the mouse chronic study was 0.24 percent.
 

F. Experimental Design
 

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

treated and untreated observation periods, and time-weighted average
 

concentrations) are summarized in Tables 1 and 2.
 

The dietary concentrations of 2,4-diaminoanisole sulfate initially
 

administered to rats in the chronic study were 0.125 and 0.06 percent.
 

After 10 weeks, the 0.125 percent concentration was reduced to 0.12
 

percent in order to standardize all doses at two decimal places and
 

facilitate dose formulation. The rat group initially receiving a
 

concentration of 0.06 percent was sacrificed after 10 months and no
 

histopathologic examinations were performed because the dose level
 

11
 
TABLE 1
 

DESIGN SUMMARY FOR FISCHER 344 RATS
 

2,4-DIAMINOANISOLE SULFATE FEEDING EXPERIMENT
 

INITIAL 

GROUP 

SIZE 

2,4-DIAMINOANISOLE SULFATE

CONCENTRATION 

(PERCENT) 

 OBSERVATION PERIOD

TREATED UNTREATED

(WEEKS) (WEEKS)

 TIME-WEIGHTED 

 AVERAGE 

CONCENTRATION3 

MALE 

LOW DOSE CONTROL 50 0 0 107 0 

HIGH DOSE CONTROL 49 0 0 109 0 

LOW DOSE 50 0.125 

0.12 

0 

10

68 

29 

 0.12 

HIGH DOSE 50 0.5 

0 

78

29 

 0.5 

FEMALE 

LOW DOSE CONTROL 50 0 0 108 0 

HIGH DOSE CONTROL 50 0 0 110 0 

LOW DOSE 50 0.125 

0.12 

0 

10

68 

29 

 0.12 

HIGH DOSE 50 0.5 

0 

78

29 

 0.5 

12
 
TABLE 2
 

DESIGN SUMMARY FOR B6C3F1 MICE
 

2,4-DIAMINOANISOLE SULFATE FEEDING EXPERIMENT
 

INITIAL 

GROUP 

SIZE 

ANISOLE SULFATE 

CONCENTRATION 

(PERCENT) 

OBSERVATION PERIOD 

TREATED UNTREATED 

(WEEKS) (WEEKS) 

MALE 

LOW DOSE CONTROL 50 0 0 96 

HIGH DOSE CONTROL 50 0 0 96 

LOW DOSE 50 0.12 

0 

78 

19 

HIGH DOSE 50 0.24 

0 

78 

18 

FEMALE 

LOW DOSE CONTROL 50 0 0 97 

HIGH DOSE CONTROL 50 0 0 96 

LOW DOSE 50 0.12 

0 

78 

19 

HIGH DOSE 50 0.24 

0 

78 

18 

13
 
was considered, on the basis of weight depression, to be too low. A
 

new treated group, receiving 0.5 percent, was started approximately
 

11 months after the initiation of the chronic study. Throughout
 

this report the rat groups receiving an initial concentration of 0.5
 

percent are referred to as the high dose groups and the rat groups
 

receiving an initial concentration of 0.125 percent are referred to
 

as the low dose groups. The treated rats were supplied with dosed
 

feed for 78 weeks, followed by a 26-week observation period.
 

The initial dietary concentrations of 2,4-diatninoanisole sulfate administered to mice in the chronic study were 0.24 and 0.12
 

percent. Throughout this report those mouse groups receiving a concentration of 0.24 percent are referred to as the high dose groups
 

and those mouse groups receiving 0.12 percent are referred to as the
 

low dose groups. These treated mice were supplied with dosed feed
 

for 78 weeks, followed by a 13-week observation period.
 

All rats and mice were approximately 6 weeks old at the time
 

that they were placed on test. The rats that became the low dose
 

group were placed on test one month after their controls. The new
 

high dose rats were placed on test one month before the high dose
 

controls. The mice that became the low dose group were placed on
 

test at the same time as their controls. The new high dose mice
 

were placed on test one month before the high dose control mice.
 

G. Clinical and Histopathologic Examinations
 

Animals were weighed immediately prior to initiation of the
 

experiment. From the first day, all animals were inspected twice
 

14
 
daily for mortality. Body weights were recorded twice weekly for
 

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

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, or gross lesions taken from sacrificed animals and, whenever
 

possible, from animals found dead.
 

Slides were prepared from the following tissues: skin, subcutaneous tissue, 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, testis,
 

prostate, brain, ear, uterus, mammary gland, ovary and Zymbal's
 

gland.
 

Tissues for which slides were prepared were preserved in 10 percent buffered formalin, embedded in paraffin, sectioned, and stained
 

with hematoxylin and eosin prior to microscopic examination. An
 

15
 
occasional section was subjected to special staining techniques for
 

more definitive diagnosis.
 

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 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
 

16
 
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
 

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
 

17
 
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
 

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
 

18
 
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. Cox1s methods of comparing
 

these curves were used for two groups; Tarone1s 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
 

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
 

c
 

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
 

19
 
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 limit 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,
 

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.
 

20
 
III. CHRONIC TESTING RESULTS: RATS
 

A. Body Weights and Clinical Observations
 

Mean body weights of high dose male and female rats were depressed relative to their controls throughout the study. No consistent depression in mean body weight was apparent for low dose male or
 

female rats (Figure 1).
 

Among high dose rats, one male had a cutaneous lesion below the
 

ear and two females had black, crusted vaginal lesions. The color of
 

the coats of male and female high dose rats changed to a light brown.
 

Palpable subcutaneous masses were observed in ten high dose control
 

male rats and one high dose control female rat. Among high dose control females, white discoloration of the eye was observed in one rat,
 

exopthalmia in a second rat, and acute alopecia in a third rat. One
 

low dose control male had a crusted cutaneous lesion on the dorsolateral surface. No other clinical abnormalities were observed in rats
 

of any group.
 

B. Survival
 

The estimated probabilities of survival for male and female rats
 

in the control and 2,4-diaminoanisole sulfate-dosed groups are shown
 

in Figure 2.
 

For male rats the Cox tests for positive association between increased dosage and accelerated mortality were not significant. Five
 

rats from each dosed and control group were sacrificed in week 78 in
 

addition to the 10 low dose controls sacrificed in week 29. There
 

21
 
TIME ON TEST (WEEKS) 

FIGURE 1 

GROWTH CURVES FOR 2,4-DIAMINOANISOLE SULFATE CHRONIC STUDY RATS 

22 
TIME ON TEST (WEEKS) 

FIGURE 2 

SURVIVAL COMPARISONS OF 2,4-DIAMINOANISOLE SULFATE CHRONIC STUDY RATS 

23 
were adequate numbers of male rats at risk from late-developing
 

tumors with 54 percent (27/50) of the high dose, 60 percent (30/50)
 

of the low dose, 61 percent (30/49) of the high dose controls, and
 

54 percent (27/50) of the low dose controls alive until the termination of the study.
 

For female rats the Cox test showed a significant (P < 0.001)
 

association between increased dosage and accelerated mortality in comparing the high dose to the high dose control. Five rats from each
 

dosed and control group were sacrificed in week 78 as well as 10 low
 

dose controls which were sacrificed in week 29. Survival was good
 

in all groups until about week 80. With 44 percent (22/50) of the
 

high dose, 58 percent (29/50) of the low dose, 74 percent (37/50) of
 

the high dose control, and 46 percent (23/50) of the low dose control
 

animals surviving until the termination of the experiment, adequate
 

numbers of female rats 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).
 

A variety of neoplasms were found in both the control and treated
 

groups (Appendix A). The incidences of most of these lesions could
 

not be related to feeding with 2,4-diaminoanisole sulfate. However,
 

there was an increased incidence of neoplasms of the integumentary
 

system and of the thyroid gland in the treated rats.
 

24
 
Tumors of the integumentary system are summarized in Table 3.
 

Epithelial neoplasms of the skin, preputial/clitoral gland, and
 

Zymbal's gland/ear canal were increased in treated groups, particularly in males.
 

Squamous-cell carcinomas appeared grossly as hard, sometimes
 

irregular masses which were often ulcerated and poorly demarcated
 

from surrounding tissue. They were composed of more or less well-


differentiated squamous cells which grew in whorls or strands into
 

adjacent structures. Mitoses were often abundant. Epithelial pearl
 

formation, hyalinization of cytoplasm, and the presence of intercellular bridges were used as criteria of squamous differentiation.
 

Basal-cell carcinomas appeared grossly as firm, rounded epithelial masses which were usually of small to moderate size. They were
 

composed of small, dark undifferentiated basal cells originating from
 

the epidermis and growing down into the dermis in masses exhibiting
 

polarization of the outside cell layer or occasionally forming a lace-


like pattern.
 

Sebaceous adenomas and adenocarcinomas also appeared grossly as
 

firm masses, their color and consistency varying greatly with the
 

degree of secondary ulceration and squamous differentiation. Microscopically, they consisted of a variable mixture of basal cells and
 

rounded pale cells containing small clear vacuoles. When the tumor
 

was well-differentiated with considerable histologic resemblance to
 

normal sebaceous glands, it was classified as an adenoma. When
 

25
 
TABLE 3
 

EPITHELIAL TUMORS OF THE INTEGUMENTARY SYSTEM FOUND IN
 

26 

Tissue/Lesion
 

SKIN (including dermis , subcutis,
 

and tail)
 

Number of Animals with Tissues
 

Examined Histopathologically
 

Squamous-Cell Papilloma
 

Squamous-Cell Carcinoma
 

Basal-Cell Carcinoma
 

Sebaceous Adenoma
 

Sebaceous Adenocarcinoma
 

PREPUTIAL/CLITORAL GLAND
 

Number of Animals with Tissues
 

Examined Histopathologically
 

Adenoma NOS , Cystadenoma,
 

or Papillary Adenoma
 

Carcinoma NOS
 

Squamous-Cell Papilloma
 

Squamous-Cell Carcinoma
 

ZYMBAL'S GLAND/EAR CANAL
 

Number of Animals with Tissues
 

Examined Histopathologically
 

Sebaceous Adenocarcinoma
 

Squamous-Cell Carcinoma
 

FISCHER 344 RATS
 

MALES
 

Low High
 

Dose Dose Low


Control Control Dose


(46)	 (48) (48)
 

0 0 1
 

0 0 0
 

0 0 1
 

0 0 0
 

0 0 1
 

(46) (48) (48)
 

0 0 2
 

0 0 0
 

0 0 0
 

0 0 0
 

(46)	 (48) (48)
 

0 0 1
 

0 0 0
 

 High
 

 Dose
 

(49)
 

0
 

5
 

2
 

1
 

3
 

(49)
 

5
 

4
 

0
 

0
 

(49)
 

4
 

2
 

Low


Dose


Control


(49)
 

0
 

0
 

0
 

0
 

0
 

(49)
 

0
 

0
 

0
 

0
 

(49)
 

0
 

0
 

FEMALES
 

 High
 

 Dose


 Control


(50)
 

0
 

0
 

1
 

0
 

0
 

(50)
 

2
 

0
 

1
 

0
 

(50)
 

0
 

0
 

 Low


 Dose


(49)
 

0
 

0
 

0
 

0
 

0
 

(49)
 

4
 

0
 

0
 

1
 

(49)
 

0
 

0
 

High
 

 Dose
 

(49)
 

0
 

0
 

1
 

0
 

0
 

(49)
 

6
 

2
 

0
 

0
 

(49)
 

7
 

0
 
peripheral cell masses were poorly differentiated and tended to
 

invade into adjacent tissues, they were classified as sebaceous
 

adenocarcinomas. Particularly in the sebaceous tumors which arose in
 

the ear canal, there was occasionally sufficient superficial squamous
 

differentiation so as to suggest a squamous carcinoma.
 

Tumors of the preputial and clitoral glands were round, firm,
 

fluctuant subcutaneous masses on the ventral surface in the genital
 

area. When cut, they appeared as cysts which were usually filled
 

with green pasty material. These tumors were characterized microscopically by the presence of at least a few large cells having
 

abundant cytoplasm packed with coarse, brightly acidophilic granules.
 

Some of these tumors exhibited enough superficial squamous differentiation to be classified as squamous-cell neoplasms. Others, with
 

minimal squamous change, were classified as adenomas if they consisted of masses of large granulated cells with large, round, vesicular nuclei surrounding a central cavity and having a well-defined,
 

intact outer border. If the outer border was irregular and consistent with invasion of surrounding tissue, the tumors were diagnosed
 

as carcinomas. It was difficult to distinguish between varieties
 

of these tumors since there was a continuous spectrum from adenoma to
 

adenocarcinoma to squamous-cell carcinoma.
 

27
 
The incidence of neoplasms observed in the thyroid are tabulated
 

Below: 

Low High 

Dose Dose Low High 

MALES Control Control Dose Dose 

Thyroid 

(Number of Animals with Tissues 

Examined Histopathologically) (45) (48) (47) (49) 

Adenoma NO S 1 0 0 0 

Adenocarcinoma NOS 2 0 1 14 

Papillary Adenocarcinoma 0 0 0 1 

Follicular-Cell Adenoma 0 0 0 1 

Follicular-Cell Carcinoma 0 0 0 1 

Papillary Cystadenoma NO S 0 0 0 2 

Papillary Cystadenocarcinoma NO S 0 O i l 

C-Cell Adenoma 1 0 4 8 

C-Cell Carcinoma 0 1 0 3 

Low High 

Dose Dose Low High 

FEMALES Control Control Dose Dose 

Thyroid 

(Number of Animals with Tissues 

Examined Histopathologically) (47) (45) (46) (49) 

Adenoma NO S 1 0 1 0 

Adenocarcinoma NO S 2 0 0 4 

Follicular-Cell Adenoma 0 0 0 2 

Follicular-Cell Carcinoma 0 1 0 3 

Papillary Cystadenoma NO S 0 0 1 0 

Papillary Cystadenocarcinoma NO S 0 0 1 3 

C-Cell Adenoma 1 1 2 4 

C-Cell Carcinoma 0 1 0 3 

The thyroids of the treated rats, especially those receiving the 

high dose of the compound, tended to be enlarged and dark red to black 

in color. Microscopically, golden brown pigment granules were found 

*
 Follicular-cell tumors were reported as adenoma NOS, follicular-cell
 

adenoma, papillary cystadenoma NOS, adenocarcinoma NOS, papillary
 

adenocarcinoma, papillary cystadenocarcinoma NOS, and follicular-cell
 

carcinoma.
 

28
 
within follicular cells, within macrophages, and lying free in the
 

colloid. This hyperpigmentation was observed in 47/49 high dose males
 

and 49/49 high dose females. A few thyroids exhibited focal lymphocytic infiltrate. In part, thyroid enlargement was due to follicular
 

cysts (9/49 high dose males and 12/49 high dose females) or hyperplasia (either follicular or C-cell), and in part to the presence of
 

tumors. These tumors were of two varieties, follicular-cell and
 

C-cell origin. Feeding rats 2,4-diaminoanisole sulfate tended to
 

increase the incidence of follicular tumors more than those of C-cell
 

origin.
 

Follicular-cell tumors were usually rounded, unilateral thyroid
 

masses having a semitranslucent red-tan or red-brown cut surface.
 

Histologically these tumors exhibited varying degrees of differentiation and were reported as adenoma NOS, follicular-cell adenoma,
 

papillary cystadenoma, adenocarcinoma NOS, papillary adenocarcinoma,
 

papillary cystadenocarcinoma, and follicular-cell carcinoma. Adenomas were well-defined collections of follicles clearly differentiated
 

from the normal thyroid tissue by increased basophilia, variation in
 

follicle size, and increased cellularity. Papillary cystic patterns
 

were sometimes present. There was no rigid histological differentiation between follicular adenomas and carcinomas. However, the latter
 

tended to be larger, to have a higher nuclear/cytoplasmic ratio, and
 

to exhibit more nuclear pleomorphism. Often, carcinomas exhibited
 

capsular or extracapsular invasion.
 

29
 
C-cell tumors were usually unilateral, round thyroid masses but
 

on section were somewhat more opaque and tended to have yellowish
 

streaks. Microscopically, C-cell adenomas were round, well-


circumscribed collections of cells having uniform ovoid nuclei and a
 

moderate amount of indistinct pale acidophilic cytoplasm. Sometimes
 

small cystic areas lined by flat cells and filled with colloid-like
 

material were noted in these tumors. C-cell carcinomas tended to
 

be larger, and both cells and nuclei occasionally assumed a spindle
 

shape. Peripheral cells invaded the thyroid capsule and often grew
 

beyond it.
 

The response of the mammary gland appeared equivocal since there
 

was wide variation in the numbers of mammary tumors found in the low
 

dose control females as compared with the high dose control females
 

as shown in the following table:
 

Low High 

Dose Dose Low High 

Control Control Dose Dose 

Mammary Gland, Mammary Duct, and 

Lactiferous Duct 

Number of Animals with Tissues 

Examined Histopathologically (49) (50) (49) (49) 

Adenocarcinoma NOS 1 0 2 2 

Papillary Cystadenoma NOS 0 0 1 0 

Papillary Cystadenocarcinoma NOS 1 0 2 0 

Fibroadenoma 4 19 16 3 

Adenoma NOS 1 0 0 0 

Mammary tumors were present as subcutaneous masses on the ventral
 

surface ranging from firm to soft and fluctuant. When cut, they
 

30
 
sometimes exuded milky fluid, often contained within grossly visible
 

cysts. The most frequently observed mammary neoplasm was a fibroadenoma composed of well-differentiated glands embedded in a copious
 

fibrous connective tissue stroma. In some cases, the glands were
 

compressed by the stroma; in others, they showed intracellular secretion vacuoles and were dilated by fluid. Often, these secretory
 

fibroadenomas were associated with large galactoceles. A few tumors
 

composed of masses of well-differentiated mammary glands having
 

little associated stroma were classified as adenomas. Papillary
 

cystic variants of adenomas were sometimes observed. Adenocarcinomas
 

contained a high proportion of glands in comparison to stroma, and
 

the glands tended to be poorly differentiated, sometimes breaking
 

down into isolated cells which invaded adjacent tissue. The nuclei
 

of the malignant tumors tended to have heavy chromatin borders, vesicular architecture, and prominent nucleoli. Mitoses were sometimes
 

noted. Some tumor cells appeared to be lactating. Others formed
 

papillary cystic patterns.
 

Both treated and control rats exhibited a spectrum of nonneoplastic degenerative and inflammatory lesions which could not be related
 

to compound administration.
 

Results of this histopathologic examination indicate that under
 

the conditions of this study, 2,4-diaminoanisole sulfate was carcinogenic to Fischer 344 rats because it increased the incidence of
 

31
 
integumentary neoplasms, particularly in males, and because it
 

increased the incidence of thyroid neoplasms in male and female
 

rats.
 

D. Statistical Analyses of Results
 

The results of the statistical analyses of tumor incidence in
 

rats are summarized in Tables 4 and 5. 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 2,4-diaminoanisole sulfate-


dosed groups and where such tumors were observed in at least 5 percent
 

of the group. Because a number of low dose control rats of both sexes
 

were sacrificed in week 29, all Fisher exact tests involving these
 

controls have been based exclusively on rats surviving at least 30
 

weeks. Due to the differences in starting times between the low dose
 

and the high dose experiments, no Cochran-Armitage tests have been
 

performed.
 

Large numbers of thyroid tumors were noted in treated rats, with
 

significant incidences of carcinomas in both male and female rats.
 

For males the Fisher exact test showed a significant (P = 0.001) increase in adenocarcinomas NOS in the high dose group compared to the
 

high dose control. For females when incidences were combined so that
 

the numerator represented rats with either an adenocarcinoma NOS, a
 

follicular-cell carcinoma, or a papillary cystadenocarcinoma NOS, the
 

Fisher exact test indicated a significantly (P = 0.006) greater
 

incidence in the high dose than in the high dose control group. For
 

32
 
33 

TABLE 4
 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS AT
 

SPECIFIC SITES IN MALE RATS TREATED WITH 2,4-DIAMINOANISOLE SULFATE*
 

TOPOGRAPHY: MORPHOLOGY 

LOW DOSE

CONTROL*

 HIGH DOSE 

 CONTROL 

LOW 

DOSE* 

Thyroid: C-Cell Adenoma or C-Cell 

Carcinoma^ 1/35(0.03) 1/48(0.02) 4/47(0.08) 

P Values0 N.S. 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

2.979 

0.314 

143.426 

Weeks to First Observed Tumor 107 109 78 

Thyroid: Adenocarcinoma NOS 2/35(0.06) 0/48(0.00) 1/47(0.02) 

P Values0 N.S. 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

0.372 

0.007 

6.887 

Weeks to First Observed Tumor 107 107 

Thyroid: Adenoma NOS, Follicular-Cell 

Adenoma, or Papillary Cystadenoma 

NOSb 1/35(0.03) 0/48(0.00) 0/47(0.00) 

P Values0 N.S. 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

0.000 

0.000 

13.884 

Weeks to First Observed Tumor 107 

HIGH
 

DOSE
 

10/49(0.20)
 

P = 0.004
 

9.796
 

1.484
 

4.551
 

72
 

14/49(0.29)
 

P = 0.001
 

Infinite
 

4.279
 

Infinite
 

74
 

3/49(0.06)
 

N.S.
 

Infinite
 

0.590
 

Infinite
 

72
 

*Includes only those rats surviving at least 30 weeks,
 
TABLE 4 (CONTINUED)
 

34 

TOPOGRAPHY: MORPHOLOGY
 

Thyroid: Adenocarcinoma NOS, Papillary
 

Adenocarcinoma, Follicular-Cell
 

Carcinoma or Papillary
 

Cystadenocarcinoma NOS"
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Pituitary: Adenoma NOS or Chromophobe
 

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
 

LOW DOSE
 

CONTROL*
 

2/35(0.06)
 

107
 

12/32(0.38)
 

101
 

6/35(0.17)
 

107
 

HIGH DOSE
 

CONTROL
 

0/48(0.00)
 

9/38(0.24)
 

85
 

8/47(0.17)
 

107
 

LOW
 

DOSE*
 

2/47(0.04)
 

N.S.
 

0.745
 

0.057
 

9.877
 

107
 

12/44(0.27)
 

N.S.
 

0.727
 

0.342
 

1.491
 

78
 

5/46(0.11)
 

N.S.
 

0.634
 

0.159
 

2.163
 

107
 

HIGH
 

DOSE
 

17/49(0.35)
 

P = 0.001
 

Infinite
 

5.306
 

Infinite
 

74
 

8/40(0.20)
 

N.S.
 

0.844
 

0.318
 

2.206
 

74
 

9/47(0.19)
 

N.S.
 

1.125
 

0.422
 

3.061
 

74
 

*Includes only those rats surviving at least 30 weeks.
 
35 

TABLE 4 (CONTINUED) 

LOW DOSE HIGH DOSE
 

TOPOGRAPHY '.MORPHOLOGY CONTROL* CONTROL
 

Testis: Interstitial-Cell Tumor 33/35(0.94) 42/47(0.89)
 

P Values0
 —_


Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor 78 78
 

Preputial Gland: Carcinoma NOS 0/36(0.00) 0/48(0.00)
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Preputial Gland: Adenoma NOS, Papillary
 

Adenoma or Cystadenoma NOS^ 0/36(0.00) 0/48(0.00)
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

*Includes only those rats surviving at least 30 weeks, 

LOW
 

DOSE*
 

47/48(0.98)
 

N.S.
 

0.963
 

0.815
 

1.307
 

78
 

0/48(0.00)
 

2/48(0.04)
 

N.S.
 

Infinite
 

0.224
 

Infinite
 

100
 

HIGH
 

DOSE
 

39/49(0.80)
 

N.S.
 

0.891
 

0.763
 

1.082
 

71
 

4/49(0.08)
 

N.S.
 

Infinite
 

0.909
 

Infinite
 

107
 

5/49(0.10)
 

P = 0.030
 

Infinite
 

1.237
 

Infinite
 

98
 
36 

TABLE 4 (CONTINUED)
 

TOPOGRAPHY: MORPHOLOGY

LOW DOSE

 CONTROL*

 HIGH DOSE 

 CONTROL 

Preputial Gland: Carcinoma NOS, 

Adenoma NOS, Papillary Adenoma, or 

Cystadenoma NOSb

P Values0 

 0/36(0.00) 0/48(0.00) 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

Zymbal's Gland or Ear Canal: 

Sebaceous Adenocarcinomab 0/36(0.00) 0/48(0.00) 

P Values0 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

Zymbal's Gland or Ear Canal or Skin of 

Ear: Squamous-Cell Carcinoma or 

Sebaceous Adenocarcinomab 0/36(0.00) 0/48(0.00) 

P Values0 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

*Includes only those rats surviving at least 30 weeks.
 

LOW
 

DOSE*
 

2/48(0.04)
 

N.S.
 

Infinite
 

0.224
 

Infinite
 

100
 

1/48(0.02)
 

N.S.
 

Infinite
 

0.040
 

Infinite
 

79
 

1/48(0.02)
 

N.S.
 

Infinite
 

0.040
 

Infinite
 

79
 

HIGH
 

DOSE
 

8/49(0.16)
 

P = 0.003
 

Infinite
 

2.288
 

Infinite
 

98
 

4/49(0.08)
 

N.S.
 

Infinite
 

0.909
 

Infinite
 

74
 

8/49(0.16)
 

P = 0.003
 

Infinite
 

2.241
 

Infinite
 

78
 
37 

TABLE 4 (CONTINUED) 

LOW DOSE HIGH DOSE LOW HIGH 

TOPOGRAPHY: MORPHOLOGY CONTROL* CONTROL DOSE* DOSE 

Hematopoietic System: Leukemia or 

Malignant Lymphoma 2/36(0.06) 6/48(0.13) 2/48(0.04) 1/49(0.02) 

P Values0 N.S. N.S. ___ Relative Risk (Control) —_ 0.750 0.163 

Lower Limit 0.057 0.004 

Upper Limit 9.957 1.274 

Weeks to First Observed Tumor 79 93 91 107 

Body Cavities: Mesothelioma NOS or 

Malignant Mesothelioma 0/36(0.00) 2/48(0.04) 4/48(0.08) 3/49(0.06) 

P Values0 N.S. N.S. ___ 

Relative Risk (Control) _— Infinite 1.469 

Lower Limit 0.703 0.176 

Upper Limit Infinite 16.960 

Weeks to First Observed Tumor 106 92 97 

Lung: Alveolar /Bronchiolar Adenoma, 

or Alveolar /Bronchiolar Carcinoma 0/36(0.00) 1/48(0.02) 3/47(0.06) 3/49(0.06) 

P Values0 N.S. N.S. 

Relative Risk (Control) Infinite 2.939 

Lower Limit 0.465 0.246 

Upper Limit Infinite 151.056 

Weeks to First Observed Tumor 109 107 98 

*Includes only those rats surviving at least 30 weeks. 
38

 

TABLE 4 (CONTINUED) 

TOPOGRAPHY: MORPHOLOGY

LOW DOSE

 CONTROL*

 HIGH DOSE 

 CONTROL 

Skin: Squamous-Cell Carcinoma (Excluding Skin of Ear)b

P Values0 

 0/36(0.00) 0/48(0.00) 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

Skin: Squamous-Cell Carcinoma, BasalCell Carcinoma, or Sebaceous Adenocarcinoma (Excluding Skin of Ear)b

 0/36(0.00) 0/48(0.00) 

P Values0 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

Skin or Subcutaneous Tissue: Fibromab 0/36(0.00) 3/48(0.06) 

P Values0 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 95 

*Includes only those rats surviving at least 30 weeks.
 

LOW
 

DOSE*
 

0/48(0.00)
 

N.S.
 

2/48(0.04)
 

N.S.
 

Infinite
 

0.224
 

Infinite
 

107
 

5/48(0.10)
 

N.S.
 

Infinite
 

0.956
 

Infinite
 

100
 

HIGH
 

DOSE
 

3/49(0.06)
 

N.S.
 

Infinite
 

0.590
 

Infinite
 

71
 

7/49(0.14)
 

P = 0.007
 

Infinite
 

1.904
 

Infinite
 

72
 

2/49(0.04)
 

N.S.
 

0.653
 

0.057
 

5.450
 

107
 
39

 

TABLE 4 (CONCLUDED)
 

LOW DOSE HIGH DOSE LOW HIGH 

TOPOGRAPHY :MORPHOLOGY CONTROL* CONTROL DOSE* DOSE 

Skin or Subcutaneous Tissue: 

Fibrosarcoma^ 0/36(0.00) 1/48(0.02) 1/48(0.02) 3/49(0.06) 

P Values0 N.S. N.S. 

Relative Risk (Control) Infinite 2.939 

Lower Limit 0.040 0.246 

Upper Limit Infinite 151.056 

Weeks to First Observed Tumor 102 107 43 

Pancreatic Islets: Islet-Cell Adenoma 2/32(0.06) 0/46(0.00) 5/45(0.11) 0/48(0.00) 

P Values0 N.S. N.S. 

Relative Risk (Control) 1.778 

Lower Limit 0.315 

Upper Limit 17.842 

Weeks to First Observed Tumor 107 78 

*Includes only those rats surviving at least 30 weeks.
 o
 

Treated groups received time-weighted average doses of 0.12 or 0.5 percent in feed.
 

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

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. A negative designation (N) indicates a lower incidence
 

in the treated group than in the control group.
 

The 95% confidence interval on the relative risk of the treated group to the control group.
 
TABLE 5
 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS AT
 

SPECIFIC SITES IN FEMALE RATS TREATED WITH 2,4-DIAMINOANISOLE SULFATE£
 

TOPOGRAPHY : MORPHOLOGY
 

Thyroid: C-Cell Adenoma or C-Cell
 

Carcinoma
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Thyroid: Adenocarcinoma NOS
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Thyroid: Papillary Cystadenoma


Papillary Cystadenocarcinoma


P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

 NOS or
 

 NOS°
 

LOW DOSE HIGH DOSE LOW 

CONTROL* CONTROL DOSE* 

1/38(0.03) 2/45(0.04) 2/46(0.04) 

N.S. 

1.652 

0.090 

95.360 

107 110 107 

2/38(0.05) 0/45(0.00) 0/46(0.00) 

N.S. 

0.000 

0.000 

2.784 

107 

0/38(0.00) 0/45(0.00) 2/46(0.04) 

N.S. ___ Infinite 

0.246 

Infinite 

107 

40 

*Includes only those rats surviving at least 30 weeks.
 

HIGH
 

DOSE
 

7/49(0.14)
 

N.S.
 

3.214
 

0.654
 

30.445
 

87
 

4/49(0.08)
 

N.S.
 

Infinite
 

0.854
 

Infinite
 

78
 

3/49(0.06)
 

N.S.
 

Infinite
 

0.554
 

Infinite
 

78
 
TABLE 5 (CONTINUED)
 

41

 

LOW DOSE
 

TOPOGRAPHY: MORPHOLOGY CONTROL*
 

Thyroid: Adenocarcinoma NOS,
 

Follicular-Cell Carcinoma, or
 

Papillary Cystadenocarcinoma NOS 2/38(0.05)
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor 107
 

Pituitary: Carcinoma NOS or
 

Adenocarcinoma NOS" 2/37(0.05)
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor 107
 

Pituitary: Carcinoma NOS, Adenocarcinoma NOS, Adenoma NOS, or
 

Chromophobe Adenoma

•i0 20/37(0.54)
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor 76
 

HIGH DOSE LOW HIGH 

CONTROL DOSE* DOSE 

1/45(0.02) 1/46(0.02) 10/49(0.20) 

N.S. P = 0.006 

0.413 9.184 

0.007 1.395 

7.641 388.645 

109 107 78 

0/40(0.00) 3/47(0.06) 0/38(0.00) 

N.S. N.S. 

1.181 

0.143 

13.582 

92 

17/40(0.43) 20/47(0.43) 16/38(0.42) 

N.S. N.S. 

0.787 0.991 

0.489 0.555 

1.298 1.756 

78 78 78 

^Includes only those rats surviving at least 30 weeks.
 
42 

TABLE 5 (CONTINUED) 

TOPOGRAPHY :MORPHOLOGY 

LOW DOSE 

CONTROL* 

HIGH DOSE 

CONTROL 

Adrenal: Cortical Adenoma 

P Values0 

0/37(0.00) 1/49(0.02) 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 110 

Adrenal: Pheochromocytoma 

P Values0 

2/37(0.05) 3/49(0.06) 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 108 109 

Mammary Gland and Ducts:

P Values0 

 Fibroadenoma 4/39(0.10) 19/50(0.38) 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 101 107 

*Includes only those rats surviving at least 30 weeks. 

LOW
 

DOSE*
 

0/47(0.00)
 

N.S.
 

3/47(0.06)
 

N.S.
 

1.181
 

0.136
 

12.842
 

100
 

16/49(0.35)
 

P = 0.011
 

3.184
 

1.139
 

12.083
 

92
 

HIGH
 

DOSE
 

6/49(0.12)
 

N.S.
 

6.000
 

0.769
 

269.766
 

107
 

7/49(0.14)
 

N.S.
 

2.333
 

0.570
 

13.275
 

106
 

4/49(0.08)
 

P = O.OOl(N)
 

0.215
 

0.058
 

0.589
 

107
 
43 

TABLE 5 (CONTINUED)
 

TOPOGRAPHY :MORPHOLOGY 

LOW DOSE 

CONTROL* 

HIGH DOSE 

CONTROL 

Mammary: Ad eno carcinoma NOS or

Papillary Cystadenocarcinoma NOS 

, 

2/39(0.05) 0/50(0.00) 

P Values0 —_ 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 100 

Uterus or Uterus/Endometrium: Adenocarcinoma NOS^ 4/38(0.11) 1/50(0.02)
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor 95 109
 

Uterus: Endometrial Stromal Polyp 10/38(0.26) 10/50(0.20)
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor 78 78
 

*Includes only those rats surviving at least 30 weeks,
 

LOW
 

DOSE*
 

4/49(0.08)
 

N.S.
 

1.592
 

0.243
 

16.921
 

92
 

1/46(0.02)
 

N.S.
 

0.207
 

0.004
 

1.978
 

92
 

18/46(0.39)
 

N.S.
 

1.487
 

0.749
 

3.155
 

80
 

HIGH
 

DOSE
 

2/49(0.04)
 

N.S.
 

Infinite
 

0.302
 

Infinite
 

95
 

3/46(0.07)
 

N.S.
 

3.261
 

0.273
 

167.332
 

86
 

3/46(0.07)
 

P = 0.050(N)
 

0.326
 

0.061
 

1.173
 

65
 
44 

TABLE 5 (CONTINUED)
 

TOPOGRAPHY: MORPHOLOGY

LOW DOSE 

 CONTROL* 

HIGH DOSE 

CONTROL 

Uterus or Uterus/Endometrium:

cinoma NOS; or Uterus/Endome

Carcinoma, Papillary Carcinom

Adenocarcinomab

 Adenoc

trium: 

a, or 

ar­

 4/38(0.11) 1/50(0.02) 

P Values0 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 95 109 

Clitoral Gland: Adenoma

Cystadenoma NOSb

 NOS or 

 0/39(0.00) 2/50(0.04) 

P Values0 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 104 

Clitoral Gland: Adenoma NOS, 

Cystadenoma NOS or SquamousPapillomabCell 

 0/39(0.00) 3/50(0.06) 

P Values0 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 104 

*Includes only those rats surviving at least 30 weeks.
 

LOW
 

DOSE*
 

5/46(0.11)
 

N.S.
 

1.033
 

0.299
 

6.185
 

92
 

4/49(0.08)
 

N.S.
 

Infinite
 

0.743
 

Infinite
 

80
 

4/49(0.08)
 

N.S.
 

Infinite
 

0.743
 

Infinite
 

80
 

HIGH
 

DOSE
 

6/46(0.13)
 

P = 0.044
 

6.522
 

0.836
 

292.687
 

86
 

6/49(0.12)
 

N.S.
 

3.061
 

0.581
 

29.826
 

95
 

6/49(0.12)
 

N.S.
 

2.041
 

0.464
 

11.991
 

95
 
45 

TABLE 5 (CONTINUED)
 

LOW DOSE HIGH DOSE
 

TOPOGRAPHY -.MORPHOLOGY CONTROL* CONTROL
 

Clitoral Gland: Adenoma NOS, Cystadenoma NOS, Squamous-Cell Papilloma,
 

Squamous-Cell Carcinoma, or
 

Carcinoma NOSb 0/39(0.00) 3/50(0.06)
 

P Values
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor 104
 

Zymbal's Gland: Sebaceous Adenocarcinomab

 0/39(0.00) 0/50(0.00)
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Hematopoietic System: Leukemia or
 

Malignant Lymphomab 4/39(0.10) 5/50(0.10)
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor 101 104
 

*Includes only those rats surviving at least 30 weeks.
 

LOW
 

DOSE*
 

5/49(0.10)
 

P = 0.049
 

Infinite
 

1.012
 

Infinite
 

80
 

0/49(0.00)
 

1/49(0.02)
 

N.S.
 

0.199
 

0.004
 

1.911
 

107
 

HIGH
 

DOSE
 

8/49(0.16)
 

N.S.
 

2.721
 

0.699
 

15.105
 

83
 

7/49(0.14)
 

P = 0.006
 

Infinite
 

1.979
 

Infinite
 

86
 

0/49(0.00)
 

P = 0.030(N)
 

0.000
 

0.000
 

0.809
 
TABLE 5 (CONCLUDED)
 

Treated groups received time-weighted average doses of 0.12 or 0.5 percent in feed.
 

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

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. A negative designation (N) indicates a lower incidence
 

in the treated group than in the control group.
 

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

46 
males the combined incidence of C-cell adenomas and C-cell carcinomas
 

of the thyroid was significant (P = 0.004) when comparing the high
 

dose to the high dose control group. These results indicate that the
 

administration of 2,4-diaminoanisole sulfate was associated with the
 

incidence of thyroid neoplasms in both male and female rats.
 

For males and females a number of sebaceous adenocarcinomas of
 

the Zymbal's gland were noted in dosed rats. For females the Fisher
 

exact test showed that the incidence was significantly (P = 0.006)
 

greater in the high dose group than in the high dose control group.
 

For males, when incidences were combined so that the numerator represented rats with a sebaceous adenocarcinoma or a squamous-cell carcinoma of either the Zymbal's gland, the ear canal, or the skin of the
 

ear, the Fisher exact test comparing high dose to high dose control
 

was significant (P = 0.003). In historical data on untreated Fischer
 

344 rats compiled by this laboratory for the NCI Carcinogenesis Testing Program, 2/334 (0.01 percent) males and 0/336 females had one of
 

these types of tumor. Based upon these results the administration of
 

2,4-diaminoanisole sulfate was associated with an increased incidence
 

of Zymbal's gland/ear canal/skin of ear neoplasms in male and female
 

rats.
 

In males, when incidences were combined so that the numerator
 

represented rats with either a squamous-cell carcinoma, a basal-cell
 

carcinoma, or a sebaceous adenocarcinoma of the skin (excluding skin
 

of the ear), the incidence was significantly (P = 0.007) greater in
 

47
 
the high dose group than in the high dose control. When preputial
 

tumor incidences in male rats were combined to include carcinomas NOS,
 

adenomas NOS, papillary adenomas, or cystadenomas NOS, the comparison
 

of high dose to high dose control was significant (P = 0.003). Based
 

upon these results the administration of 2,4-diaminoanisole sulfate
 

was associated with an increased incidence of skin tumors and of neoplasms of the preputial gland in male rats.
 

In the female rats the incidence of fibroadenomas of the mammary
 

gland and ducts was significantly (P = 0.011) higher in the low dose
 

group than in the low dose control. The comparison of high dose to
 

high dose control, however, showed a significant (P = 0.001) negative
 

association. Compared to the historical control data compiled by
 

Mason Research Institute for the NCI Carcinogenesis Testing Program
 

(105/585 [18 percent] untreated female Fischer 344 rats developed a
 

spontaneous mammary fibroadenoma), the observed incidence was somewhat
 

lower (4/39 or 10 percent) than expected in the low dose control and
 

higher (19/50 or 38 percent) than expected in the high dose control.
 

In female rats when the incidences of uterine adenocarcinomas
 

NOS or of uterine/endometrial carcinomas, papillary carcinomas, or
 

adenocarcinomas were pooled, the comparison of high dose to control
 

had a probability level of P = 0.044—a marginal result which was not
 

significant under the Bonferroni criterion. Similarly, results for
 

endometrial stromal polyps, malignant lymphomas or leukemias, and
 

48
 
clitoral neoplasms in the females and for fibromas of the integumentary system in the males were not significant under the Bonferroni
 

inequality.
 

In summary, there were statistically significant incidences of
 

neoplasms of the thyroid, of the Zymbal's gland, of the skin, and of
 

the preputial gland in male rats and increased incidences of neoplasms
 

of the thyroid and of the Zymbal's gland in female rats.
 

49
 
IV. CHRONIC TESTING RESULTS: MICE
 

A. Body Weights and Clinical Observations
 

No marked mean body weight depression was observed in any group
 

of mice treated with 2,4-diaminoanisole sulfate (Figure 3).
 

No clinical abnormalities were observed in treated or control
 

mice during the bioassay.
 

B. Survival
 

The estimated probabilities of survival for male and female mice
 

in the control and 2,4-diaminoanisole sulfate-dosed groups are shown
 

in Figure 4.
 

For male mice the Cox tests for positive association between increased dosage and accelerated mortality were not significant. Five
 

mice were sacrificed from the high dose and from each control group
 

in week 78; five additional high dose control mice were sacrificed in
 

week 49. With 82 percent (41/50) of the high dose, 92 percent (46/
 

50) of the low dose, 78 percent (39/50) of the high dose control, and
 

84 percent (42/50) of the low dose control mice alive at the termination of the study, there were adequate numbers of male mice at risk
 

from late-developing tumors.
 

For female mice the Cox tests did not show a significant positive
 

association between increased dosage and accelerated mortality. As
 

with the males, five mice were sacrificed from the high dose and from
 

each control group in week 78 with five additional high dose control
 

mice having been sacrificed in week 49. There were adequate numbers
 

50 
TIME ON TEST (WEEKS) 

FIGURES 

GROWTH CURVES FOR 2,4-DIAMINOANISOLE SULFATE CHRONIC STUDY MICE 

51 
TIME ON TEST (WEEKS) 

FIGURE 4 

SURVIVAL COMPARISONS OF 2,4-DIAMINOANISOLE SULFATE CHRONIC STUDY MICE 

52 
of female mice at risk from late-developing tumors with 78 percent
 

(39/50) of the high dose, 76 percent (38/50) of the low dose, 76
 

percent (38/50) of the high dose control, and 74 percent (37/50) of
 

the low dose control mice surviving until the end of the study.
 

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).
 

With the exception of thyroid and hematopoietic neoplasms, the
 

tumors listed in Appendix B appeared with approximately equal frequency in treated and control mice or appeared in insignificant numbers.
 

There was an increase in follicular-cell hyperplasia and
 

follicular-cell neoplasia in treated mice as shown in the following
 

table: 

Low

Dose

Control

 High 

 Dose

 Control

 Low

 Dose

 High 

 Dose 

MALES 

Thyroid 

(Number of Animals with Tissues 

Examined Histopathologically)

Adenocarcinoma NO S

Follicular-Cell Adenoma

Hyperplasia NO S

Hyperplasia, Follicular-Cell

 (47)

 0

 1

 0

 1

 (45)

 0 

0

 0 

0

 (46)

1 

0

1 

0

 (45) 

0 

 11 

0 

 12 

FEMALES 

Thyroid 

(Number of Animals with Tissues 

Examined Histopathologically) (43) (44) (42) (45) 

53
 
Follicular-Cell Adenoma
 

Follicular-Cell Carcinoma
 

Hyperplasia NOS
 

Hyperplasia, Follicular-cell
 

Hyperplasia, Papillary
 

Hyperplasia, Adenomatous
 

Adenomas were nodular lesions composed of neoplastic follicles
 

lined with cuboidal or columnar epithelium. These adenomas were
 

usually better circumscribed than those lesions classified as focal
 

hyperplasia. In the high dose mice, occasional follicular cells in
 

adenomas contained a golden brown pigment which also appeared in the
 

colloid.
 

Follicular-cell carcinomas were much larger, often involving an
 

entire thyroid lobe and extending through the capsule into surrounding
 

tissues. They were composed of columnar follicular cells exhibiting
 

some degree of nuclear pleomorphism. In some areas, the carcinomas
 

were well-differentiated, forming follicular structures and in other
 

areas the tumor cells were more poorly differentiated.
 

The thyroids of the treated mice usually contained enlarged
 

follicles filled with dense colloid and lined by flat epithelium with
 

pyknotic nuclei. Follicular-cell hyperplasia was defined as the
 

occurrence of a focal area in which follicular epithelium suddenly
 

became cuboidal to columnar, the nucleus enlarged and oval in shape,
 

and the colloid either became vacuolated or diminished in quantity.
 

Papillary and adenomatous variants of hyperplasia were seen, and
 

6 

2 

0

1

0

0 

0

0

1

3 

7

0 

0

0 

0

0 

2

1 

0

0

0

0

0

0 

54
 
often, especially in the male mice, it was difficult to distinguish
 

a sharp boundary between hyperplasia and adenoma formation.
 

In female mice only there was an increased incidence (5/48 low
 

dose control, 2/50 high dose control, 14/45 low dose, 9/50 high dose)
 

of malignant lymphoma (NOS, undifferentiated, lymphocytic, histiocytic and/or mixed type) which occurred in multiple organs, spleen,
 

mesenteric lymph node, or liver.
 

A variety of other nonneoplastic lesions which were observed
 

both in control and treated mice could not be related to compound
 

administration.
 

This histopathologic examination indicates that under the conditions of this study, dietary administration of 2,4-diaminoanisole
 

sulfate increased the incidence of both thyroid follicular-cell
 

hyperplasia and thyroid follicular-cell neoplasms in B6C3F1 mice of
 

both sexes and possibly malignant lymphoma in female mice.
 

D. Statistical Analyses of Results
 

The results of the statistical analyses of tumor incidence in
 

mice are summarized in Tables 6 and 7. 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 2,4-diaminoanisole sulfate-


dosed groups and where such tumors were observed in at least 5 percent
 

of the group. Because a number of high dose control mice of both
 

sexes were sacrificed in week 49, all Fisher exact tests involving
 

these controls have been based exclusively on mice surviving at least
 

55
 
56 

TABLE 6
 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS AT
 

SPECIFIC SITES IN MALE MICE TREATED WITH 2,4-DIAMINOANISOLE SULFATE£
 

TOPOGRAPHY : MORPHOLOGY 

LOW DOSE 

CONTROL 

HIGH DOSE 

CONTROL* 

LOW 

DOSE 

HIGH 

DOSE* 

Hematopoietic:

P Values0 

 Malignant Lymphoma 4/48(0.08) 5/44(0.11) 6/49(0.12) 

N.S. 

4/49(0.08) 

N.S. 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

1.469 

0.373 

6.670 

0.718 

0.152 

3.131 

Weeks to First Observed Tumor 96 96 80 85 

Lung: Alveolar/Bronchiolar Carcinoma 

P Values0 

6/48(0.13) 5/44(0.11) 2/48(0.04) 

N.S. 

6/48(0.13) 

N.S. 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

0.333 

0.034 

1.752 

1.100 

0.302 

4.252 

Weeks to First Observed Tumor 96 96 97 69 

Lung: Alveolar/Bronchiolar Adenoma or 

Alveolar/Bronchiolar Carcinoma^3 

P Values0 

6/48(0.13) 10/44(0.23) 3/48(0.06) 

N.S. 

12/48(0.25) 

N.S. 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

0.500 

0.085 

2.194 

1.100 

0.487 

2.556 

Weeks to First Observed Tumor 96 96 97 69 

^Includes only those mice surviving at least 50 weeks
 
TABLE 6 (CONTINUED)
 

LOW DOSE HIGH DOSE
 

TOPOGRAPHY -.MORPHOLOGY CONTROL CONTROL*
 

Liver: Hepatocellular Carcinoma 7/48(0.15) 6/43(0.14)
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor 78 78
 

Liver : Hepatocellular Adenoma or
 

Hepatocellular Carcinoma'11 7/48(0.15) 8/43(0.19)
 

P Values0
 

LOW
 

DOSE
 

14/49(0.29)
 

N.S.
 

1.959
 

0.814
 

5.224
 

97
 

14/49(0.29)
 

N.S.
 

HIGH
 

DOSE*
 

11/49(0.22)
 

N.S.
 

1.609
 

0.600
 

4.869
 

12/49(0.24)
 

5

N.S.
 

7 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 78 78 

Thyroid: Follicular-Cell Adenoma 1/47(0.02) 0/40(0.00) 

P Values0 

Relative Risk (Control) _ — 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 96 

*Includes only those mice surviving at least 50 weeks.
 

1.959
 

0.817
 

5.224
 

97
 

0/46(0.00)
 

N.S.
 

0.000
 

0.000
 

19.034
 

1.316
 

0.550
 

3.368
 

78
 

11/45(0.24)
 

P < 0.001
 

Infinite
 

2.976
 

Infinite
 

78
 

78 
TABLE 6 (CONCLUDED)
 

Treated groups received time-weighted average doses of 0.12 or 0.24 percent in feed.
 

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

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. A negative designation (N) indicates a lower incidence
 

in the treated group than in the control group.
 

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

58 
TABLE 7
 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS AT
 

SPECIFIC SITES IN FEMALE MICE TREATED WITH 2,4-DIAMINOANISOLE SULFATE£
 

TOPOGRAPHY: MORPHOLOGY 

LOW DOSE 

CONTROL 

Lung: Alveolar /Bronchiolar Carcinoma 

P Values0 

1/46(0.02) 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 96 

Lung: Alveolar /Bronchiolar Adenoma or 

Alveolar/Bronchiolar Carcinoma^ 

P Values0 

4/46(0.09) 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 96 

Hematopoietic:

P Values0 

 Malignant Lymphoma 5/48(0.10) 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 96 

HIGH DOSE
 

CONTROL*
 

1/45(0.02)
 

78
 

3/45(0.07)
 

78
 

2/45(0.04)
 

96
 

LOW
 

DOSE
 

3/43(0.07)
 

N.S.
 

3.209
 

0.270
 

164.399
 

69
 

5/43(0.12)
 

N.S.
 

1.337
 

0.308
 

6.314
 

69
 

14/45(0.31)
 

P = 0.013
 

2.987
 

1.118
 

9.703
 

94
 

HIGH
 

DOSE*
 

0/50(0.00)
 

N.S.
 

0.000
 

0.000
 

16.794
 

4/50(0.08)
 

N.S.
 

1.200
 

0.215
 

7.805
 

78
 

9/50(0.18)
 

P = 0.038
 

4.050
 

0.899
 

36.946
 

78
 

59

 

*Includes only those mice surviving at least 50 weeks.
 
60

 

TABLE 7 (CONTINUED) 

TOPOGRAPHY : MORPHOLOGY 

LOW DOSE 

CONTROL 

HIGH DOSE 

CONTROL* 

LOW 

DOSE 

HIGH 

DOSE* 

Pituitary:

c 

P Values 

 Adenoma 2/42(0.05) 1/38(0.03) 5/38(0.13) 

N.S. 

5/34(0.15) 

N.S. 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 97 96 

2.961 

0.571 

29.586 

97 

5.588 

0.670 

255.444 

82 

Thyroid: Follicular-Cell Adenoma 

P Values0 

0/43(0.00) 0/41(0.00) 0/42(0.00) 6/45(0.13) 

P ­ 0.017 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

Infinite 

1.468 

Infinite 

78 

Thyroid: Follicular-Cell Adenoma or 

Follicular-Cell Carcinoma13 

P Values0 

0/43(0.00) 0/41(0.00) 0/42(0.00) 8/45(0.18) 

P = 0.004 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

Infinite 

2.096 

Infinite 

78 

*Includes only those mice surviving at least 50 weeks. 
TABLE 7 (CONCLUDED)
 

Treated groups received time-weighted average doses of 0.12 or 0.24 percent in feed.
 

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

°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. A negative designation (N) indicates a lower incidence
 

in the treated group than in the control group.
 

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

61 
50 weeks. Due to the differences in starting times between the high
 

dose and the low dose experiments, no Cochran-Armitage tests have
 

been performed.
 

A high incidence of follicular-cell thyroid tumors were noted
 

in both male and female treated mice. For the males the Fisher
 

exact test indicated a significantly (P < 0.001) greater incidence
 

of follicular-cell adenomas in the high dose than in the high dose
 

control group; no follicular-cell carcinomas were observed. For the
 

females when incidences were combined so that the numerator represented mice with either a follicular-cell adenoma or a follicularcell carcinoma of the thyroid, the Fisher exact test again indicated
 

a significantly (P = 0.004) greater incidence in the high dose than
 

in the high dose control. Historical records collected by Mason Research Institute for untreated B6C3F1 mice in the NCI Carcinogenesis
 

Testing Program indicated 1/350 (0.28 percent) male and 0/350 female
 

mice with either a follicular-cell adenoma or a follicular-cell carcinoma of the thyroid.
 

A number of malignant lymphomas were also observed in the female
 

mice. The Fisher exact test indicated a significantly elevated
 

incidence in the low dose group (P = 0.015) compared to the low dose
 

control group. The comparison of high dose to high dose control had
 

a probability level of P = 0.038, a marginal result which was not
 

significant under the Bonferroni criterion. In the historical data
 

62
 
43/350 (12 percent) of the untreated females had leukemia or a malignant lymphoma. However, other groups of control female mice at this
 

laboratory have had incidences of malignant lymphomas of 10/49 (20
 

percent) and 11/50 (22 percent).
 

Based upon these results the administration of 2,4-diaminoanisole
 

sulfate was associated with the incidence of follicular-cell thyroid
 

tumors in both male and female mice and may have been associated with
 

the incidence of malignant lymphomas in female mice.
 

63
 
V. DISCUSSION
 

In this bioassay, adequate numbers of rats and mice in all groups
 

survived long enough to be at risk from late-developing tumors. Adverse clinical signs and depression of mean group body weight were
 

observed in high dose rats. No growth retardation or other adverse
 

clinical signs were observed among low dose rats or mice dosed at
 

either level.
 

Follicular-cell thyroid tumors found in rats included adenomas
 

and adenocarcinomas NOS; papillary adenocarcinomas; follicular-cell
 

adenomas and carcinomas; and papillary cystadenomas and adenocarcinomas. For purposes of statistical analysis, incidences of malignant
 

follicular-cell tumors of the thyroid were combined; C-cell neoplasms
 

were analyzed separately. For high dose groups of both sexes, the
 

proportion of rats having malignant follicular-cell tumors of the
 

thyroid was significantly greater than in control rats. For the high
 

dose male rats, the combined incidence of C-cell adenomas and C-cell
 

carcinomas was significantly greater than the incidence in the corresponding control group.
 

An increased incidence of epithelial tumors of the integumentary
 

system was observed among treated rats. These tumors were found in
 

the skin, preputial gland, clitoral gland, Zymbal's gland, and ear
 

canal. The incidence of malignant skin tumors (squamous-cell carcinoma, basal-cell carcinoma, or sebaceous adenocarcinoma) was significantly increased in the high dose male group but not in other groups
 

64
 
of treated rats. Malignant tumors (sebaceous adenocarcinomas or
 

squamous-cell carcinomas) of the ear (Zymbal's gland, ear canal or
 

skin of the ear) were significantly increased among high dose male
 

and female rats compared to their respective controls. Among high
 

dose male rats a significantly increased incidence of preputial gland
 

tumors was observed including carcinomas NOS, cystadenomas, papillary
 

adenomas, and adenomas NOS.
 

Among male and female mice both thyroid follicular-cell hyperplasia and thyroid follicular-cell neoplasia were related to administration of 2,4-diaminoanisole sulfate. Among high dose male mice,
 

the incidence of follicular-cell adenomas was significantly increased,
 

but no follicular-cell carcinomas were observed. Among high dose
 

female mice, the combined incidence of follicular-cell adenoma and
 

follicular-cell carcinoma was significantly increased. The eight
 

female mice with either a follicular-cell adenoma or follicular-cell
 

carcinoma or both included only two animals in which follicular-cell
 

carcinoma was observed.
 

The incidence of malignant lymphomas was increased among female
 

mice at both dose levels, but statistical significance was established
 

only for the low dose group. The incidence of malignant lymphomas for
 

the low dose controls was 5/48 (10 percent); however, other groups of
 

untreated control female mice at this laboratory have had incidences
 

of this lesion of 10/49 (20 percent) and 11/50 (22 percent). Thus,
 

although the incidence of malignant lymphomas was increased among both
 

65
 
dose levels of female mice when compared to their respective controls,
 

the relationship of development of this lesion to the administration
 

of the test chemical was not clearly evident. This uncertainty is
 

based on the variability of the incidence of these tumors in untreated
 

control female mice and the lack of a significant dose-related effect
 

at this site in this study.
 

Under the conditions of this bioassay, technical-grade 2,4­

diaminoanisole sulfate was carcinogenic to both sexes of both species.
 

In Fischer 344 rats dietary administration of the chemical induced
 

increased incidences of malignant tumors of the skin and its associated glands and malignant thyroid tumors in each sex. In B6C3F1 mice,
 

dietary administration of 2,4-diaminoanisole sulfate induced thyroid
 

tumors in each sex.
 

66
 
VI. BIBLIOGRAPHY
 

Ames, B.N., H.O. Kammer, and E. Yamasaki, "Hair Dyes are Mutagenic:
 

Identification of a Variety of Mutagenic Ingredients." Proceedings of the National Academy of Science, U.S.A. 72(6):2423-2427,
 

1975.
 

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.
 

Blijleven, W.G.H., "Mutagenicity of Four Hair Dyes in Drosophila
 

Melanogaster." Mutation Research 48:181-186, 1977.
 

Burnett, C., E.I. Goldenthal, S.B. Harris, F.X. Wazeter, J. Strausburg,
 

R. Kapp, and R. Voelker, "Teratology and Percutaneous Toxicity
 

Studies on Hair Dyes." Journal of Toxicology and Environmental
 

Health 1:1027-1040, 19761
 

Burnett, C., B. Lanman, R. Giovacchini, G. Wolcott, and R. Scala,
 

"Long-term Toxicity Studies on Oxidation Hair Dyes." Food and
 

Cosmetics Toxicology 13:353-357, 1975.
 

Burnett, C., R. Loehr, and J. Corbett, "Dominant Lethal Mutagenicity
 

Study on Hair Dyes." Journal of Toxicology and Environmental
 

Health 2:657-662, 1977.
 

Chemical Abstracts Service. The Chemical Abstracts Service (CAS)
 

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

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

Corbett, J.F., "Hair Dyes." The Chemistry of Synthetic Dyes, Vol. 5.
 

Academic Press, Inc., New York, 1971.
 

Corbett, J.F., and J. Menkart, "Hair Coloring." CUTIS 12:190-197,
 

1973.
 

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

Co., Ltd., London, 1970.
 

67
 
Cox, D.R., "Regression Models and Life-Tables." Journal of the Royal
 

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

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.
 

Ito, N., Y. Hiasa, Y. Konishi, and M. Marugami, "The Development of
 

Carcinoma in Liver of Rats Treated with m-Toluylenediamine and
 

the Synergistic and Antagonistic Effects with Other Chemicals."
 

Cancer Research 29:1137-1145, 1969.
 

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

Observations." Journal of the American Statistical Association
 

_5^: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.
 

Maibach, H., "Percutaneous Penetration of Hair Dyes." Paper submitted
 

to the U.S. Food and Drug Administration, Washington, D.C.,
 

August 1977.
 

Menkart, J., Senior Vice President-Technology, Clairol, Stamford,
 

Connecticut. Personal communication, June 8, 1977.
 

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

Co., New York, 1966.
 

Nishioka, H., "Detection of Carcinogenicity of Color Cosmetics in
 

Bacterial Test Systems." Mutation Research 38:345, 1976.
 

Palmer, K.A., A. Denunzio, and S. Green, "The Mutagenic Assay of Some
 

Hair Dye Components, Using the Thymidine Kinase Locus of L5178Y
 

Mouse Lymphoma Cells." Journal of Environmental Pathology and
 

Toxicology 1:87-91, 1977.
 

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, Color Index, 3rd edition, Volume 3,
 

Yorkshire, England, 1971a.
 

Society of Dyers and Colourists, Color Index, 3rd edition, Volume 4,
 Yorkshire, England, 1971b.
 

68
 
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.
 

Urso, S., Research Analyst, Chemical-Environmental Program, Chemical
 

Industries Center, Stanford Research Institute, Menlo Park,
 

California. Personal communication, June 22, 1977.
 

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

69
 

APPENDIX A

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS 

IN RATS TREATED WITH 2,4-DIAMINOANISOLE SULFATE 

TABLE Al

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN MALE RATS TREATED 

WITH 2,4-DIAMINOANISOLE SULFATE 

LOW DOSE HIGH DOSE CONTROL (UNTR) CONTROL (UNTR) LCW DOSE 

01-0037 01-01 18 01-0048 

ANIMALS INITIALLY IN STUDY 5 0 35 0 5 0 

ANIMALS NECROPSIED 4 6 48 4 8 

ANIMALS EXAMINED HISTOPATHOLOGICALLY** 4 6 4 8 4 8 

INTEGUMENTABY SYSTEM 

•SKIN (46) (48) (4 8) 

SQUAMOUS CELL CARCINOMA 

BASAL-CELL CARCINOMA 1 (2%) 

SEBACEOUS ADENOMA 

SEBACEOUS ADENOCARCINOMA 1 (2%) FIBROMA 3 (6%) 

FIBROSARCOMA 1 (2%) 

*SUBCUT TISSUE (46) (48) (48) 

SARCOMA, NCS 1 (2%) FIBROMA 3 (6%) 2 (4%) 

FIBROSARCOMA 1 (2%) 

RESPIRATORY SYSTEM 

•TRACHEA (45) (48) (47) 

ADENOCARCINOMA, NOS# METASTATIC 1 (2%) 

#LUNG (46) (48) (47) 

ADENOCARCINOMA, NOS, METASTATIC 1 (2%) ALVEOLAR/BRONCHIOLAR ADENOMA 3 (6%) ALVEOLAR/BKCNCHIOLAR CARCINOMA 1 (2X) SEBACEOUS ADENOCARCINOMA, METAST 

PHEOCHROHCCYTOMA, METASTATIC 1 (2%) FIBROSARCOMA 1 (2%) 

HEMATOPOIETIC SYSTEM 

•MULTIPLE ORGANS (46) (48) (48) 

MALIGNANT LYMPHOMA, NOS 1 J2%L 

# NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY • NUMBER OF ANIMALS NECROPSIED ••EXCLUDES

 PARTIALLY AUTOLYZED ANIMALS 

9 50 ANIMALS WERE INITIALLY IN THE STUDY, BUT ONE ANIMAL WAS FOUND TO BE A FEMALE IN

HIGH DOSE 

01-0109 

50 

49 

4 9 

(49) 

5 (10%) 

2 (4%) 

1 (2%) 

3 (6%) 

1 (2%) 

1 (2%) 

(49) 

1 (2%) 

2 (4%) 

(49) 

(49) 

2 (4%) 

1 (2%) 

1 (2%) 

(49) 

A MALE GROUP. 

A-3 
TABLE Al (CONTINUED)
 

LOW DOSE 

CONTROL (UNTR) 

01-0037 

HIGH DOSE CONTROL(UNTR) 

01-0118 

LEUKEMIA,NOS 

UNDIFFERENTIATED LEUKEMIA 

MYELONONOCYTIC LEUKEMIA 

MONOCYTIC LEUKEMIA 

1

1

 (2%) 

 (2%) 

1

4

 (2%) 

 (8%) 

•SKIfJ 

MALIG,LYMPHOMA, HISTIOCYTIC TYPE 

(46) (48) 

«LYMPH NODE 

ADENOCARCINOMA, NOS, METASTATIC 

(38) 

1 (3%) 

(44) 

«THYMUS 

FIBROSARCOMA 

(38) (23) 

CIRCULATORY SYSTEM 

«HEART (46) (48) 

FIBROSARCOMA 

DIGESTIVE SYSTEM 

«SALIVARY GLAND (38) (47) 

ADENOCARCINOMA, NOS 1 (2%) SARCOMA, NCS 1 (2%) 

«LIVhR (46) (48) 

NhOPLASTIC NODULE 

HLPATOCELLULAR CARCINOMA 1 (2*) 

«PANCREAS (42) (46) 

ACINAR-CELL ADENOMA 

•DUODENUM (43) (46) 

FIBROSARCOMA 

#ILEUM (43) (46) 

SARCOMA, NOS 1 (2%) 

«COLON (43) (46) 

ADENOCARCINOMA, NOS 

URINAaY SYSTEM 

«KIDNEY/PELVIS (46) (48) 

TRANSITIONAL-CELL PAPILLOMA 

# NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY 

* NUflfaER OF ANIMALS NECROPSIED 

LOW DOSE 

01-0048 

(48) 

1 (2%) 

(42) 

(37) 

1 (3%) 

(47) 

1 (2%) 

(45) 

1 (2%) 

(48) 

2 (4%) 

(45) 

(«5) 

1 (2%) 

(45) 

(4 3) 

1 (2%) 

(48) 

HIGH DOSE 

01-0109 

1 (2%) 

(49) 

(47) 

(21) 

(49) 

(47) 

(48) 

1 (2X) 

1 (2%) 

(48) 

1 (2%) 

(47) 

(47) 

(40) 

(49) 

1 (2%) 

A-4 
TABLE A1 (CONTINUED) 

LOW DOSE CONTROL (UNTR) 

01-0037 

HIGH DOSE CONTROL(UNTS) 

01-01 18 

LOW DOSE 

C1-0048 

HIGH DOSE 

01-0109 

ENDOCEINE SYSTEM 

«PITUITARY 

ADENOMA, NCS 

CHROMOPHOBE ADENOMA 

(41) 

2 (5%) 

10 (24%) 

(38) 

9 (24%) 

(44) 

12 (27%) 

(40) 

8 (20%) 

#ADRr.NAL 

ADENOCARCINOMA, NOS, M£TASTATIC 

PHEOCHROMOCYTOMA 

PHEOCHROMOCYTOMA, MALIGNANT 

(43) 

1 (2%) 

6 (14%) 

(47) 

7 (15%) 

1 (2%) 

(46) 

5 (11%) 

(47) 

9 (19%) 

tTHYHOID 

ADENOMA, NOS 

ADENOCARCINOMA, NOS 

PAPILLARY ADENOCARCINOMA 

FOLLICULAR-CELL ADENOMA 

FOLLICULAR-CELL CARCINOMA 

C-CELL ADENCMA 

C-CELL CARCINOMA 

PAPILLARY CYSTADENOMA, NOS 

PAPILLARY CYSTADENOCARCINOMA,NOS 

(45) 

1 (2%) 

2 (4%) 

1 (2%) 

(48) 

1 (2%) 

(47) 

1 

4 

1 

(2%) 

(9%) 

(2%) 

(49) 

14 

1 

1 

1 

8 

3 

2 

1 

(29%) 

(2%) 

(2%) 

(2%) 

(16%) 

(6%) 

(4%) 

(2%) 

#PARATHYROID 

AJENOMA, NCS 

(32) (28) 

1 (4%) 

(30) (24) 

•PANCREATIC ISLETS 

ISLET-CEIL ADENOMA 

(42) 

2 (5%) 

(46) (45) 

5 (11%) 

(48) 

REPRODUCTIVE SYSTEM 

•MAMMARY GLANE 

FIBROADENOMA 

(46) (48) (4 8) 

2 (4%) 

(49) 

•PREPUTIAL GLAND 

CARCINOMA,NOS 

ADENOMA, NCS 

PAPILLARY ADENOMA 

CYSTADENCMA, NOS 

(46) (48) (4 8) 

2 (4%) 

(49) 

4 

4 

1 

(8%) 

(8%) 

(2%) 

«PROSTATE 

ADENOMA, NOS 

PARAGANGLIOMA, NOS 

(45) 

1 J2%1 

(44) (47) 

1 (2%) 

(47) 

# NUMoER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY 

* NUHBER OF ANIMALS NECROPSIED 

A-5
 
«TESTIS

 

INTERSTITIAL-CELL TUMOR 

NERVOUS SYSTEM 

#BRAIN 

GLIOMA, NOS

 

ASTROCYTOMA 

SPECIAL SENSE CRGANS 

•EAR 

SQUAMOUS CELL CARCINOMA 

*EAR CANAL 

SQUAMOUS CELL CARCINOMA 

SEBACEOUS ADENOCARCINOMA 

•ZYMoAL'S GLAND 

SfcBACEOUS ADENOCARCINOMA 

MUSCULOSKELETAL SYSTEM 

NONfc 

BODY CAVITIES 

•BODY CAVITIES 

MESOTHELIOMA, NOS 

MuSOTrfELICrA, MALIGNANT 

•MEDIASTINUM 

FIBROSARCOMA 

ALL OTHER SYSTEMS 

TAIL 

SQUAMOUS CELL PAPILLOMA 

# NUMBER OF ANIMALS WITH TISSUE

* NUMriER OF ANIMALS NECROPSIID 

TABLE Al (CONTINUED) 

LOW DOSE CONTROL (UNTR) 

01-0037 

(45) 

33 (73%) 

(44) 

1 (2%)

 

(46) 

(46) 

(46) 

(46) 

(46) 

HIGH DOSE CONTROL (UNTR) 

01-0 1 18 

(47) 

42 (89*) 

(48) 

1 (2%) 

(48) 

(48) 

(48) 

(48) 

2 (4%) 

(48) 

 EXAMINED MICROSCOPICALLY 

LCW DOSE 

01-0048 

(48) 

47 (98*) 

(46) 

1 (2%) 

(48) 

(48) 

(4 8) 

1 (2%) 

(48) 

4 (8%) 

(48) 

1 (2%) 

1 

HIGH DOSE 

01-0109 

(49) 

39 (80%) 

(49) 

(49) 

1 (2%) 

(49) 

1 (2%) 

1 (2%) 

(49) 

3 (6%) 

(49) 

2 (4%) 

1 (2%) 

(49) 

A-6 
TABLE Al (CONCLUDED)
 

LOW DOSE HIGH DOSE CONTROL (UNTR) CONTROL(UNTR) LOW DOSE HIGH DOSE 

01-0037 01-01 18 01-0048 01-0109 

ANIMAL DISPOSI1ION SUMMARY 

ANIMALS INITIALLY IN STUDY 50 50 50 50

 

NATURAL DEATHS 6 6 8 7

 

MORIBUND SACRIFICE 2 8 7 11 

SCHEDULED SACRIFICE 15 5 c 5 

ACCIDENTALLY KILLED 

TERMINAL SACRIFICE 27 30 3 0 27

 

ANIMAL MISSING 

ANIMAL DELETED(WRONG SEX) 1

 

3 INCLUDES AOTOLYZED ANIMALS 

TUMOR SUMMARY 

TOTAL ANIMALS WITH PRIMARY TUMORS* 3 4 44 4 8 47

 

TOTAL PRIMARY TUMORS 61 8 0 109 1 2 9 

TOTAL ANIMALS WITH BENIGN TUMORS 33 4 3 4 8 45

 

TOTAL BENIGN TUMORS 55 6 2 87 7 9

 

TOTAL ANIMALS WITH MALIGNANT TUMORS 5 17 12 3 4

 

TOTAL MALIGNANT TUMORS 5 18 18 47

 

TOTAL ANIMALS WITH SECONDARY TUMORS# 1 1 1

 

TOTAL SECONDARY TUMORS 4 1 1

 

TOTAL ANIMALS WITH TUMORS UNCERTAINBENIGN OR MALIGNANT 1 4 3

 TOTAL UNCERTAIN TUMORS 1 4 3

 

TOTAL ANIMALS WITH TUMORS UNCERTAIN-

PRIdARY OB METASTATIC

 

TOTAL UNCERTAIN TUMORS 

* PRIHARY TUMORS: ALL TUMORS EXCEPT SECONDARY TUMORS
 

# SECONDARY TUMORS: METASTATIC TUMORS OR TUMORS INVASIVE INTO AN ADJACENT ORGAN
 

A-7
 
TABLE A2

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN FEMALE RATS TREATED WITH 

2 4-DIAMINOANISOLE SULFATE 

LOW DOSE HIGH DOSE CONTROL(UNTR) CONTROL (UNTR) 

02-0037 02-01 18 

ANIMALS INITIALLY IN STUDY 50 50 

ANIMALS NECRCFSIED 4 9 50 

ANIMALS EXAMINED HISTOPATHOLOGICALLY* * 49 5 0 

INTEGUMENTARY SYSTEM 

• SKIN (49) (50) 

BASAL-CELL CARCINOMA 1 (2%) F1BROSARCCMA 

•SUBCUT TISSUE (49) (50) FIBROMA 1 (2%) FIBROSARCOMA 1 (2*) 

RESPIRATORY SYSTEM 

#LUNG (49) (50) 

SQUAMOUS CELL CARCINOMA, METASTA 1 (2%) ADENOCARCINOMA, NOS, METASTATIC 1 (2%) HEPATOCELLULAR CARCINOMA, METAST 1 (2%) ALVEOLAR/BRONCHIOLAR ADEN3MA 1 (2%) 1 (2%) C-CELL CARCINOMA, METASTATIC 

HEMATOPOIETIC SYSTEM 

•MULIIPLE ORGANS (49) (50) 

MALIGNANT IYMPHOMA, NOS 

MALIG.LYMPHOMA, LYMPHOCYTIC TYPE 2 (4%) UNDIFFERENTIATED LEUKEMIA 1 (2%) MYELOMONOCYTIC LEUKEMIA 3 (6%) MONOCYTIC LEUKEMIA 2 (4%) 

#SPLEEN (49) ("8) 

HUMANGIOSARCOM A 

UNDIFFERENTIATED LEUKEMIA 1 (2%) 

«RENAL LYMPH NODE (4 1) (47) 

ADENOCARCINOMA, NOS, METASTATIC 1 (2%) 

CIRCULATORY SYSTEM 

NONc 

# NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY 

* NUMBER OF ANIMALS NECROPSIED 

**EXCLUDES PARTIALLY AUTOLYZED ANIMALS 

LOW DOSE 

02-0048 

50 

49 

4 9 

(49) 

(4 9) 

2 (4%) 

(48) 

(49) 

1 (2%) 

(4 3) 

HIGH DOSE 

02-0109 

50 

49 

4 9 

(49) 

1 (2%) 

1 (2%) 

(49) 

1 (2%) 

1 (2%) 

(49) 

1 (2X) 

1 (2%) 

(49) 

(49) 

1 (2X) 

(41 ) 

A-8 
TABLE A2 (CONTINUED)
 

LOW DOSE HIGH DOSE CONTROL (UNTR) CONTROL(UNTR) 

02-0037 02-0118 

DIGESTIVE SYSTEM 

•LIVER (49) (50)

 

ADENOCARCINOMA, NOS, METASTATIC 1 (2%)

 

NEOPLASTIC NODULE 

HEPATOCELLULAR CARCINOMA 2 (4%)

 

#STOrtACH (48) (48)

 

SQUAMOUS CELL PAPILLOMA 

•ILEUM (47) (48)

 

LEIOMYOSARCOMA 1 (2%)

 

URINAHY SYSTEM 

•KIDNEY (49) (50)

 TUBULAR-CELL ADENOMA

 

ENDOCEINE SYSTEM 

•PITUITARY (43) (40)

 

CARCINOMA,NOS 

ADENOMA, NOS 3 (7%) 17 (43%)

 

ADENOCARCINOMA, NOS 2 (5%)

 

CHROMOPHOBE ADENOMA 15 (35%)

 

•ADRhNAL (46) (49)

 

CORTICAL ACENOMA 1 (2%)

 

PHEOCHRCMCCYTOMA 2 (4%) 3 (6%)

 

•ADRENAL MEDULLA (46) (<*9)

 

GANGLIONEUROMA 1 (2%)

 

•THYROID (47) (45)

 

ADENOMA, NOS 1 (2%)

 

ADENOCARCINOMA, NOS 2 (4%)

 

FOLLICULAR-CELL ADENOMA 

FOLLICULAR-CELL CARCINOMA 1 (2%)

 

C-CELL ADEKOMA 1 (2%) 1 (2%)

 

C-CELL CARCINOMA 1 (2%)

 

PAPILLARY CYSTADENOMA* NOS 

• NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMbER OF ANIMALS NECROPSIED
 

LOW DOSE 

02-0048 

(48) 

(47) 

(46) 

(48) 

(47) 

3 (6%) 

18 (38%) 

(47) 

3 (6%) 

(47) 

(4 6) 

1 (2%) 

2 (4%) 

1 11%) 

HIGH DOSE 

02-0109 

(49) 

1 (2%) 

(<*9) 

1 (2%) 

(47) 

(49) 

1 (2%) 

(38) 

16 (42%) 

(49) 

6 (12%) 

7 (14%) 

(49) 

(49) 

* (8%) 

2 (4%) 

3 (6%) 

4 (8%) 

3 (6%) 

A-9
 
TABLE A2 (CONTINUED) 

LOW DOSE CONTROL (UNTR) 

02-0037 

"cWo^UNTR) 

02-0118 

LOW DOSE 

02-0048 

HIGH DOSE 

02-0109 

PAPILLARY CYSTADENOCARCINOMA,NOS 1 (2%) 3 (6%) 

«PANCREATIC ISLETS 

ISLET-CELL ADENOMA 

ISLET-CELL CARCINOMA 

(46) (48) 

2 (4%) 

(46) 

1 (2%) 

(47) 

REPRODUCTIVE SYSTEM 

•MAMMARY GLANE 

AUENOMA, NOS 

ADENOCARCIWOMA, NOS 

PAPILLARY CYSTADENOMA, NOS 

PAPILLARY CYSTADENOCARCINOMA,NOS 

FIBROADENOMA 

(49) 

1

1

1

4

 (2%) 

 (2%) 

 (2%) 

 (8%) 

(50) 

19 (38%) 

(<*9) 

2 (4%) 

1 (2%) 

2 (4%) 

16 (33%) 

(49) 

2

3

 (4%) 

 (6%) 

•MAMrtARY DUCT 

FIBROADENOMA 

(49) (50) (49) (49) 

1 (2%) 

•LACTIFEROUS DUCT 

FIBROADENCMA 

(49) (50) (49) 

1 (2%) 

(49) 

•CLirORAL GLAND 

CARCINOMA,NOS 

SQUAMOUS CELL PAPILLOMA 

SQUAMOUS CELL CARCINOMA 

ADENOMA, NCS 

CYSTADENCMA, NOS 

(49) (50) 

1 (2%) 

2 (4%) 

(49) 

1 (2%) 

1 (2%) 

3 (6%) 

(49) 

2

6

 (4%) 

 (12%) 

IUTERUS 

ADENOCARCINOMA, NOS 

ENDOMETRIAL STROMAL

ENDOM3TRIAL STROMAL

 POLYP 

 SARCOMA 

C+8) 

4 (8%) 

10 (21%) 

(50) 

1 (2%) 

10 (20%) 

1 (2%) 

(46) 

1

18

 (2%) 

 (39%) 

(46) 

2

3

 (4%) 

 (7%) 

«UTERUS/ENDCMETRIUM 

CARCINOMA,NOS 

PAPILLARY CARCINOMA 

ADENOCARCINOMA, NOS 

SARCOMA, NCS 

(48) (50) (46) 

4

2

 (9%) 

 (4%) 

(46) 

2

1

1

 (4%) 

 (2%) 

 (2%) 

#OVAt(Y 

PAPILLARY CYSTADENOCARCINOMA,NOS 

GKANULOSA-CELL TUMOR 

(47) (49) 

1 (2%) 

(46) 

1

1

 (2%) 

 (2%) 

(47) 

NERVOUS SYSTEM 

NONE 

# NUMBER

* NUMBER

 OF

 OF

 ANIMALS

 ANIMALS

 WITH TISSUE

 NECROPSIED 

 EXAMINED MICROSCOPICALLY 

Α-IO
 
TABLE A2 (CONTINUED)
 

LOW DOSE CONTROL(UNTR) 

02-0037 02-01 18 

SPECIAL SENSE ORGANS 

•EAR	 CANAL (49) (50)
 

FIBROMA 1 (2%)
 

•ZYMBAL'S	 GLAND (49) (50)
 

St-BACEOUS ADENOCARCINOMA
 

MUSCULOSKELETAL SYSTEM 

NONE 

BODY CAVITIES 

•BODY	 CAVITIES (49) (50)
 

MESOTHELIOMA, MALIGNANT 1 (2%)
 

ALL OTHER SYSTEMS 

SIT E UNKNOWN 

SQUAMOUS CELL CARCINOMA 1 

ANIMAL DISPOSITION SUMMARY 

ANIMALS INITIALLY IN STUDY 50 50 

NATURAL DEATHS 5 5 MORIBUND SACRIFICE 7 3 

SCHEDULED SACRIFICE 15 5 ACCIDENTALLY KILLED 

TERMINAL SACRIFICE 2 3 37 

ANIMAL MISSING 

# NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

• NUMBER OF ANIMALS NECROPSIED
 

LCW DOSE 

02-0048 

(4 9) 

(4 9) 

(49) 

50 

6 

10 

5 

29 

HIGH DOSE 

02-0109 

(49) 

(49) 

7 (14%) 

(49) 

50 

9
 

14
 

5
 

22 

Α-II
 
TABLE A2 (CONCLUDED)
 

LOW DOSE CONTROL(UNTR) 

02-0037 

HIGH DOSE CONTROL(UNTR) 

02-01 18 

LCW DOSE 

02-0048 

HIGH DOSE 

02-0109 

TUMOR SUMMAR* 

TOTAL ANIMALS WITH PRIMARY

TuTAL PRIMARY TUMORS 

 TUMORS* 3 2 

56 

38 

7 3 

4 4 

87 

37 

86 

TOTAL ANIMALS WITH BENIGN

TOTAL BENIGN TUMORS 

 TUMORS 2 7 

39 

35 

59 

4 0 

67 

29 

51 

TOTAL ANIMALS WITH MALIGNANT

TOTAL MALIGNANT TUMORS 

 TUMORS 1 5 

17 

12 

13 

15 

19 

2 2 

34 

TOTAL AWIMALS WITH SECONDARY

TOTAL SECONDARY TUMORS 

 TUMORS* 2 

4 

1 

1 

2 

2 

TOTAL ANIMALS WITH TUMORS

BENIGN OR MALIGNANT 

TOTAL UNCERTAIN TUMORS 

 UNCERTAIN1 

1 

1 

1 

1 

1 

TOTAL ANIMALS WITH TUMORS

PRIHARY OR METASTATIC 

TOTAL UNCERTAIN TUMORS 

 UNCERTAIN­

• PRIMARY TUMORS: ALL TUMORS EXCEPT SECONDARY TUMORS 

# SECONDARY TUMORS: METASTATIC TUMORS OR TUMORS INVASIVE INTO AN ADJACENT ORGAN 

A-I2
 
APPENDIX B 

SUMMARY OF THE INCIDENCE OF NEOPLASMS 

IN MICE TREATED WITH 2,4-DIAMINOANISOLE SULFATE 

TABLE B1 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN MALE MICE 

TREATED WITH 2,4-DIAMINOANISOLE SULFATE 

LOW DOSE 

CONTROL (UMTR) 

05-0037 

HIGH DOSE 

CONTROL (UNTR) 

05-0118 

LOW DOSE 

05-0048 HIGH DOSE 05-0108 

ANIMALS INITIALLY IN STUDY 

ANIMALS MISSING 

ANIMALS NECPOPSIED 

ANIMALS EXAMINED hiSTOFATHOLOGICALLΥ** 

50 

48 

48 

50 

1 

49 

49 

50 

49 

49 

50 

49 

49 

INTEGUMENTARY SYSTEM 

NONE 

RESFIEATOHY SYSTEM 

• LUNG 

HEPATOCELLULAF CARCINOMA, METAST 

ALVEOLAP/BRONCHIOLAR ADEKOMA 

ALVEOLAB/BRONCHIOLAR CAFCINOMA 

(48) 

6 (13%) 

(49) 

1 (2%) 

5 (10%) 

5 (10%) 

(48) 

1 (2%) 

2 (4%) 

(48) 

2 (4%) 

7 (15%) 

6 (13%) 

HEMATOPOIETIC SYSTEM 

•MULTIPLE ORGANS 

MALIGNANT LYMPHOMA, NOS 

MALIG.LYMPHOMA, HISTIOCYTIC TYPE 

MALIGNANT LYMPHOMA, MIXED TYPE 

(*»8) 

2 (4%) 

2 (4%) 

(49) 

3 (6%) 

(49) 

2 (H%) 

(49) 

1 (2%) 

2 (4%) 

#SPLEEN 

HEMANGIOMA 

3EMANGIOSARCOMA 

UALIG.LYMPHOMA, LYMPHOCYTIC

MALIG.LYMPHOMA, HISTIOCYTIC

 TYPE 

 TYPE 

(47) 

1 (255) 

C*9) 

1

1

 (2%) 

 (2%) 

(48) 

1 (2%) 

1 (2%) 

(48) 

2 (4%) 

•LYMPH NODE 

MALIG.LYMPHOMA, HISTIOCYTIC TYPE 

(44) (<*2) 

1 (2%) 

(45) (45) 

«MESENTERIC L. NODE 

tfALIGNANT LYMPHOMA, MIXED TYPE 

(44) ("2) (45) (45) 

1 (2%) 

tSMALL INTESTINE 

MALIG.LYMPHOMA, HISTIOCYTIC TYPE 

(48) (49) (47) 

1 (2%) 

(47) 

tPEYERS PATCH 

MALIG.LYMPHOMA, HISTIOCYTIC TYPE 

(48) (49) (47) 

2 (4%) . 

(47) 

# NUMBER OF ANIMALS WITH TISSU E EXAMINED

* NUMBER OF ANIMALS NECROPSIED 

**EXCLUDES PARTIALLY AUTOLYZED ANIMALS 

 MICROSCOPICALLY 

B-3
 
TABLE Bl (CONTINUED) 

LOW DOSE HIGH DOSE
 

CONTROL(UNTR) CONTROL (UNTR)
 

05-0037 05-0118
 

CIRCULATORY SYSTEM
 

NONE
 

DIGESTIVE SYSTEM
 

#LIVER 

HEPATOCELLULAR ADENOMA 

HEPATOCELLULAR CAf-CINOMA 

HEMANGIOMA 

KEMANGIOSARCOMA, METASTATIC 

HEMANGIOSARCOM*, UNC PRIM OP MET 

(48) 

7

1

 (15*) 

(2%) 

(48) 

2

6

1

 (4%) 

 (13%) 

 (2%) 

«STOMACH 

SQUAMOUS CELL CAFCINOXA 

(47) 

1 (2%) 

(48) 

•SMALL INTESTINE 

ADENOCAFXINOKA, NOS 

(48) (49) 

URINABY SYSTEM
 

•KIDNEY	 (47) (49)
 

TUBULAR-CELL ADINCKA
 

ENCOCRINE SYSTEM
 

•PITUITARY	 (42) (40)
 

ADENOMA, NOS
 

•ADRENAL	 (45) (44)
 

CORTICAL ADENOMA
 

PHEOChROMOCYTOMA 1 (2%)
 

•THYROID	 (47) (45)
 

ADENOCARCINOMA, NOS
 

FOLLICULAR-CELL ADENOMA 1 [2%)
 

•PARATHYROID	 (29) (24)
 

ADENOMA, NOS
 

•PANCREATIC	 ISLETS (48) (47)
 

I-SLET-CELL ADENOMA
 

• NUKBUR OF ANIMALS WITh TISSUE EXAMINED MICROSCOPICALLY
 

* NUMEER OF ANIMALS NECROP3IED
 

LOW DOSE
 

05-0048
 

(49)
 

14	 (29%)
 

(46)
 

(47)
 

1 (2%)
 

(48)
 

1 (2%)
 

(40)
 

(47)
 

1 (2%)
 

1 (2%)
 

(t6)
 

1 (2%)
 

(26)
 

(44)
 

1 (2%)
 

HIGH DOSE
 

05-0108
 

(49)
 

1 (2%)
 

11 (22%)
 

1 (2%)
 

(48)
 

(47)
 

(49)
 

(43)
 

1 (2%)
 

(40)
 

1 (3%)
 

(45)
 

11 (2H%)
 

(22)
 

1 (5%)
 

(47)
 

B-4
 
TABLE Bl (CONTINUED) 

REPRODUCTIVE sYSTF 1
 

#TESTIS
 

INTERS^ITIAL-C^L L TUMOR
 

NERVOUS SYSTEM
 

NONE
 

SPECIAL SENSE ORGANS
 

•HARDERIAN	 GLAND
 

ADENOFA, hOS
 

MUSCULOSKELETAL SYSTEM
 

NONE
 

BOEY CAVITIES
 

NONE
 

ALL C1HER SYSTEMS
 

NONE
 

ANIMAL DISPOSITION SIMMARY
 

ANIMALS INITIALLY IN STUDY
 NATURAL DEATH®
 

MORIBUND SACRIFICE
 

SCHEDULED SACRIFICE
 

ACCIDENTALLY KILLED
 TERMINAL SACRIFICE
 

ANIMAL MISSING
 

d INCLUDES AUTOLYZhD ANIMALS
 

LOW DOSE
 CONTROL(UNTR)
 05-0037
 

(47)
 

(48)
 

50
 

3
 

5
 

42
 

HIGH DOSE
 CONTROL(UNTR\
 05-0118
 

(48)
 

i^9)
 

50
 

10
 

39
 

1
 

LOW DOSE
 05-0048
 

(46)
 

(49)
 

50
 

4
 

46
 

HIGH DOSE
 05-0108
 

(48)
 

1 (2%)
 

(49)
 

1 (2%)
 

50
 

4
 

5
 

41
 

# NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMALS NLCROPSILD
 

B-5
 
TABLE B1 (CONCLUDED) 

LOW DOSE CONTROL(UKTP) 05-0037 HIGH DOSE CONTROL (ONTR) 05-0118 LOW DOSE 05-0048 HIGH DOSE 05-0108 

TUMOR SUMMAPY 

TOTAL ANIMALS WITH PFIfAPY TUMORS* TOTAL PRIMARY ^UMOFS 

17 

21 22 

26 24 30 33 47 

TOTAL ANIuALS WTT

J BIMGN TOTAL BFNIGN TinCFS TUMORS 2 

3 8 8 5 5 22 24 

TOTAL ANIMALS ΗΙΊ MALIGNANT

TOTAL MALIGNAN" TUfOPS 

 TUMORS 15 

18 

15 

17 

21 

25 

21 

23 

TOTAL ANIMALS Wl^H SECONDARY TOTAL SECONDARY ^UMOPS TUMORS# 1 

1 3 3 

TOTAL ANIMALS Wlr

 rf TUMOBS UNCERTAINBENIGN OR MALIGNANT TOTAL UNCERTAIN TUhORS 

TOTAL fNIMALS WITH T0KOP5 UNCERTAIN­ PRIMARY OR METASTATIC TOTAL UNCERTAIN TUKORS 1 

1 

* PRIMARY TUTORS: ALL TU^OPS FXCEPT SECONDARY TUMORS 

# SECONDAPY TUMORS: KfcTAbTATIC TUMORS OR TUMORS INVASIVE INTO AN ADJACENT ORGAN 

B-6
 
TABLE B2

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN FEMALE MICE TREATED 

WITH 2,4-DIAMINOANISOLE SULFATE 

LOW DOSE HIGH DOSE
 CONTROL (UNTP) CONTROL(UNTR) LOW DOSE HIGH DOSE
 06-0037 06-0118 06-0048 06-0108
 

ANIKALS INITIALLY IN STUDY 50 50 50 50
 ANIMALS MISSING 1
 

ANIMALS NLCFOPSIED 48 50 45 50
 ANIMALS EXAMINED HISTGPATHOLOGICALLY** 47	 so
 44 50
 

INTEGUMENTAFY SYSTEM
 

*SUBCUT TISSUE (48) (50) (50)
 *IBRCSABCOM* 1 (2%)
 LEIOMYOSARCOMA 1 (2%)
 

RESPIRATORY SYSTEM
 

«LUNG (46) (50) (43) (50)
 

CARCINOMA, NOS, METASTATIC 1 (2%)
 

ALVEOLAR/uRONCHIOLAF ADENOMA 3 <7T) 2 (4%) 2 (5%) 4 (8%)
 

ALVEOLAR/BRONCFIOLAF CAFCINOMA 1 (2%) 1 (2%) 3 (7%)
 

HEMATOPOIETIC SYSIIM
 

•MULTIPLE ORGANS	 (48) (50) (45) (50)
 

MALIGNANT LYMPHOMA, NOS 1 (2%) 2 (4%) 1 (2%) 1 {2%)
 

MALIG.LYMPHOMA, UNDIFFEP-TYPE 2 (4%) 3 (6%)
 

MALIG.LYMPHOMA, LYMPHOCYTIC TYPE 1 (2%)
 

MALIG.LYMPHOMA, HISTIOCYTIC TYPE 2 (4%) 4 (9%) 1 (2%)
 

MALIGNANT LYMPHOMA, MIXED TYPE 1 (2%)
 

«SPLEEN	 (46) (49) (44) (50)
 

HEMANGIOSAECOMA 1 {27)
 MALIGNANT LYMPHOMA, NOS 3 (7%)
 

HALIG.LYMPHOMA, HISTIOCYTIC TYPE 1 (22) 2 (5%)
 

•MESENTERIC	 L. NODL (39) (44) (38) (43)
 

MALIG.LYMPHOMA, HISTIOCYTIC TYPE 2 (5%)
 

MALIGNANT LYMPHOMA, MIXED TYPE 1 (2*)
 

•LIVER	 (47) (50) (42) (50)
 

MALIGNANT LYMPHOMA, MIXED TYPE 1 12%)
 

# NUMEER OF ANIMALS WITh TISSUE EXAMINED MICROSCOPICALLY 

* NUMBER OF ANIMALS NECROPSIED 

••EXCLUDES PARTIALLY AUTOLYZED ANIMALS 

B-7 
•THYMUS
 

LYMPHANGIOMA
 

MALIGNANT LYMPHOMA, NOS
 

CIBCULATOBY SYSTEM
 

NONE
 

DIGESTIVE SYSTEM
 

#LIVER
 

HEPATOCELLULAR CARCINOMA
 

FIBROSARCOMA
 

•STOMACH
 

SQUAMOUS CELL PAPILLOMA
 

•COLON
 

LEIOMYOSARCOMA
 

URINABY SYSTEM
 

NONE
 

ENDCCBINE SYSTEM
 

•PITUITARY
 

CARCINOMA,NOS
 

ADENOMA, NOS
 

CHROMOPHOBE ADENCKA
 

•ADRENAL
 

CORTICAL ADENOhA
 

PHEOCHPCKCCYTOFA
 

•THYROID
 

FOLLICULAR-CFLL ADINOMA
 

FO1LICULAR-CELL CAFCINOMA
 

REEBCEUCTIVE SYSTEM
 

*MAMMARY GLAND
 

CARCINOMA.NOS
 

• NUMEER OF ANIMALS KITH TISSUE

• NUKBLR OF ANIMALS NECROFSIFD 

TABLE B2 (CONTINUED) 

LOW DOSE HIGH DOSE
 

CONTROL(UNTR) CONTROL (UNTR)
 

06-0037 06-0118
 

(31) (21)
 

1	 (3%)
 

(47)	 (50)
 

1 (2*) 1 (2%)
 

1 (2%)
 

(44)	 (49)
 

1 (22)
 

(40)	 (38)
 

1 (3%)
 

(42)	 (4 2)
 

1 (2%)
 

2 (5%) 1 (2%)
 

2 (5%)
 

(45) (48)
 

1 (2*)
 

(43) (44)
 

(48) (50)
 

 EXAMINED MICROSCOPICALLY 

•
 

LOW DOSE
 

06-0048
 

(36)
 

(42)
 

1 (2%)
 

(40)
 

(37)
 

(38)
 

5 (13%)
 

(41)
 

(42)
 

(45)
 

1 (2%)
 

HIGH DOSE
 

06-0108
 

(32)
 

1 (3%)
 

(50)
 

2 (4%)
 

(49)
 

(41)
 

(34)
 

5 (15%)
 

(49)
 

2 (4%)
 

(45)
 

6 (13%)
 

2 (4%)
 

(50)
 

B-8 
TABLE B2 (CONTINUED) 

ADENOCARCINOMA, NOS
 

iUTERUS
 

ADENOCARCINOMA, NOS
 LEIOMYOSARCOMA
 

ENDOrtTRIAL STPOMiL POLYP
 

#OVARY/OVIDUCT
 

PAPILLARY ADENOMA
 

#OVARY
 

GRANULOSA-CELL TUMOR
 

TUBULAR ADuNOMA
 

NERVCUS SYSTEM
 

NONE
 

SPECIAL SENS* ORGANS
 

•HAFDERIAN GLAND
 

PAPILLARY ADhNOPA
 

MUSCULOSKELETAL SYSTEM
 

NONE
 

BCEY CAVITIES
 

•BODY CAVITIES
 

MESOTHELIOMA, MALIGNANT
 

ALL CTHEE SYSTEMS
 

NDNF
 

LOW DOSE
 

CONTROL (UNTR)
 

06-0037
 

(45)
 

1 (2%)
 

3 (7%)
 

(45)
 

(45)
 

1 (2%)
 

(48)
 

(<*8)
 

HIGH DOSE
 

CCNTROL(UNTR)
 

06-0118
 

(47)
 

(47)
 

1 (2X)
 

(48)
 

(50)
 

1 (2%)
 

(50)
 

LOW DOSE
 

06-0048
 

1 (2%)
 

1 (2X)
 

(41)
 

(40)
 

1 (3%)
 

(45)
 

(*5)
 

1 (2%)
 

HIGH DOSE
 

06-0108
 

(46)
 

1 (2%)
 

(46)
 

(43)
 

1 (2%)
 

(50)
 

(50)
 

# NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMALS NLXRCPS1ED
 

B-9 
TABLE B2(CONCLUDED)
 

LOW DOSE HIGH DOSE
 CONTROL (UNTR) CONTROL (UNTR) LOW DOSE HIGH DOSE
 06-0037 06-0 118 06-0043 06-0108
 

ANIFAL CISFCSITICN SUMMARY
 

ANIMALS INITIALLY IN STJD\ 50 50 50 50
 NATURAL DEATHS 6 2 1 1 3
 

MORIBUND SACRIFICE 2 2
 

SCHEDULED SACRIFICE 5 10 5
 

ACCIDENTALLY KILLED 1
 

TERMINAL SACRIFICE 37 38 8 39
 

ANIKA1 MISSING 1
 

a> INCLUDES AUTOLYZZD ANIMALS
 

TUMCE SUMMARY
 

TOTAL ANIMALS WITn PPIMARY TUMORS* 20 10 25 30
 

TOTAL PRIMARY TUMOFS 24 11 30 34
 

TOTAL ANIMALS WITH BENIGN TUMORS 11 7 7 17
 

TOTAL BENIGN TUMOFS 11 7 7 19
 

TOTAL ANIMALS WITH MALIGNANT TUMORS 11 4 21 14
 

TCTAL MALIGNANT TUMORS 13 4 22 14
 

TOTAL ANIMALS fcllH SECONDARY TUMOPS# 1
 

TCTAL SECONDAPY TUTORS 1
 

TOTAL ANIMALS WITH TUMORS UNCEFTAIN­ BEMGN OR MALIGNANT 1 1
 TOTAL UNCERTAIN TUhORS 1 1
 

TOTAL ANIMALS WITH TUMORS UNCERTAIN­ PRIMARY OR METASTATIC
 TOTAL UNCERTAIN TUMORS
 

• PRIMARY TUMORS: ALL 1UMORS TXCEPT SECONDARY TUMOPS
 

* SECONDARY TUMORS: METASTATIC TUMORS OR TUMORS INVASIVE INTO AN ADJACENT ORGAN
 

B-IO
 
APPENDIX C

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC 

LESIONS IN RATS TREATED WITH 2,4-DIAMINOANISOLE SULFATE 

TABLE CI
 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN MALE RATS
 

TREATED WITH 2,4-DIAMINOANISOLE SULFATE
 

LOW DOSE 

CONTROL(UNTR) 

01-0037 

HIGH DOSE 

CONTROL (UNTR) 01-0118 LOW DOSE 01-0048 HIGH DOSE 01-0109 

ANIMALS

ANIMALS

AMMAL S

 INITIALLY IN STUDY 

 NECBOPSIED 

 EXAMINED HISTOPATHOLOGICALLY * 

50 

46 

* 46 

350 

48 

48 

50 

48 

48 

50 

49 

49 

INTEGUMENTARY SYSTEM
 

•SKIN	 (46) (48) (48) (49)
 

CYST, NOS 1 (2*)
 

EFTDEP1AL INCLJSION CYST 2 (4%)
 

•SUPCUT TISSUE	 (46) (US) (48) (49)
 

ABSCESS, NOS 1 (2*)
 

MFTAFLASIA, ObSEOUS 1 (2%)
 

RESFIFATO&Y SYSTEM
 

#TRACHEA (45) (48) (47) (49)
 

INFLAMMATION, NOS 9 (20%) 2 (4%)
 

ABSCESS, NOS 1 (2%)
 INFLAMMATION, CHPONIC 10 (22%)
 

#LUNG/EP0NCHU5 (46) (48) (47) (49)
 

BRONCHIECTASIS 1 (2%) 1 (2%)
 INFLAMMATION, NOS 7 (15%)
 

INFLAMMATION, CHPOM C 8 (17%)
 

#BRCNCHIAL MUCOUS GLA (46) (48) (47) (49)
 

AESCESS, NOS 1 (2%)
 

NECROSIS, NOS 1 (2%)
 HYPERPLASIA, ADENOMATOUS 1 (2?)
 

#LUNG/EPONCHIOLE (46) (48) (47) (49)
 INFLAMMATION, NOS 1 (2%)
 

INFLAMMATION, ?OCAL 1 (2%)
 

#LUNG (46) (43) (47) (49)
 

ATELECTASIS 1 (2%)
 

CONGESTION, NU3 1 (2%) 1 12%)
 

# NUMEER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY 

* NUMBER OF ANIMALS NECROPSIED 

**EXCLUDES	 PARTIALLY AUTOLYZED ANIMALS 

 5 0 ANIMALS WERE INITIALLY IN THE STUDY, PUT ONE ANIKAL WAS FOUND TO BE A FEMALE IN A MALE GROUP. 
TABLE CI (CONTINUED)
 

LOW DOSE HIGH DOSE 

CONTROL(UNTR) CONTROL (UNTR) 

01-0037 01-0118 

EDEMA, NOS 

INFLAMMATION, NOS 

1 

1 (25!) 

(2%) BRONCHOPNEUMONIA, FOCAL 

INFLAMMATION, FOCAL 3 (7%) INFLAMMATION, INTERSTITIAL 1 (2%) 4 (8%) 

INFLAMMATION, SUPPURATIVE 1 (2%) 

BRONCHOPNEUMONIA SUPPURATIVE INFLAMMATION, NECROTIZING 1 (2%) 

PNEUMONIA, CHRONIC riUPINE 1 (2%) 1 (2%) 

INFLAMMATION, CHRONIC 1 (2%) 

PERIVASCULITIS 5 (11*) HYPERPLASIA, EPITHELIAL 1 (2%) HYPERPLASIA, ADENOMATOUS 

HEMATOPOIETIC SYSTEM
 

#2ONE MARROW (44) (4 7)
 

ERYTHROPQirSIS
 

#SPLSEN (46) (48)
 

THROMBOSIS, NOS 1 (27)
 FIBROSIS 1 (2%) 1 (2%)
 INFARCT, HEALED 1 (2?)
 FIGMENTATION, NOS
 

HEMOSIDEROSIS 1 (2%)
 RETICULOCYTOSIS 1 (2%)
 

HYPERPLASIA, HEMATOPOIETIC 9 (19%)
 HYPERPLASIA, ERYTHFOID 12 (26%) 10 (21%)
 HYPERPLASIA, 3ETICULUM CZLL 8 (17%)
 HYPERPLASIA, LYMPHOID
 

HEMATOPOIESIS
 

#SPL£NIC CAPSULE («46) (48)
 

INFLAMMATION, FOCAL
 

•SPLENIC	 RED PULP (46) (48)
 

INFLAMMATION PROLIFERATIVE
 

#LYHPH NODE (38) (44)
 

HEMOFRHAGE 1 (2%)
 

INFLAMMATION, NOS 1 (3%)
 HYPERPLASIA, NOS 1 (3%)
 PLASKACYTOSIS 1 (2%)
 HYFEFPLASIA, RETICULUM CELL 3 18%)
 

# NUMBER OF ANIMALS WITH TISSU£ EXAMINED MICROSCOPICALLY
 

* NUMBER 0? ANIMALS NECROPSIED
 

LOH DOSE
 01-0048
 

1 (2%)
 

1 (2%)
 

(45)
 

1 (2%)
 

(48)
 

1 (2%)
 

1 (2%)
 

(48)
 

1 (2%)
 

(48)
 

(42)
 

HIGH DOSE
 01-0109
 

1 (2%)
 

1 (2%)
 

1 (2*)
 

2 (4%)
 

(45)
 

(48)
 

1 (2%)
 

(48)
 

(48)
 

1 (2%)
 

(47)
 

C-4
 
TABLE CI (CONTINUED) 

LOW DOSE 

CONTROL(UNTR) 

01-0037 

HIGH DOSE 

CONTROL (UNTR) 

01-0118 

LOW DOSE 

01-0048 

HIGH DOSE 

01-0109 

HYPERPLASIA, LYMPHOID 3 (7%) 

«MEDIASTINAL L.NODE 

PLASMACYTOSIS 

(38) 

1 (3%) 

(4 4) (42) (47) 

1 (2%) 

«MESENTERIC L. NODE 

PLASMACYTOSIS 

(38) (44) (42) (47) 

1 (2%) 

CIRCULATORY SYSTEM 

*LYMPhATIC VESSELS 

INFLAMMATION, NOS 

(46) 

1 (2%) 

(48) (48) (49) 

tHEART 

trERIART^lTIS 

(46) (48) (47) (49) 

1 (2%) 

#i1YOCARDIUM 

INFLAMMATION, NOS 

INFLAMMATION, INTEFSTITIAL 

INFLAMM\TION, CHRONIC FOCAL 

FIBROSIS 

CEGENEFATION, NOS 

FIGMFNTATION, NOS 

(46) 

1 (2%) 

22 (4h«) 

3 (7 ) 

7 (15%) 

(48) 

23 (48%) 

12 (25X) 

(47) 

7 (15%) 

3 (6%) 

(49) 

29 (59%) 

1 (2%) 

#ENDOCAFDIU1 

INFLAMMATION,

INFLAMMATION,

 FOCAL 

 ACUTE/CHRONIC 

(46) (48) (47) (49) 

1 (2%) 

1 '2%) 

•AORTA 

INFLAMMATION, CHRONIC FOCAL 

(46) 

1 (2%) 

(48) (48) (49) 

•PULMONARY ARTERY 

INFLAMMATION PROLIFERATIVE 

HYPEFTPOPHY, NOS 

(46) 

 (2') 

(48) (48) (49) 

1 (2%) 

DIGESTIVE SYSTEM 

•SALIVARY GLAND 

INFLAMMATION, CHRONIC FOCAL 

(38) (47) (45) (47) 

1 (2%) 

•SUBMAXILLARY GLAND 

PLASMACYTOSIS 

(38) (47) (45) (47) 

1 (2%) 

•LIVER 

FIBROSIS SEPTAL LIVER 

(46) (48) 

2 (4XL 

(48) (48) 

# NUMBER OF ANIMALS

* NUMEfiR OF ANIflALS

 WITH TISSUE EXAMINED

 NECROPSIED 

 MICRCSCOPICALLY 

C-5
 
TABLE CI (CONTINUED)
 

LOW DOSE HIGH DOSE CON1FOL(UNTfc) 01-0037 CONTROL (UNTR) 01-0 118 

NECROSIS, FOCAL 

NECROSIS, COAGULATIVE 3 (1%) 

1 (2%) 

2 <*•*) 

METAMORPHOSIS FATTY 1 {21) HYPERTROPHY, NOS HYPERPLASIA, NODULAR 

HYPEFPLASIA, NOS HYPERPLASIA, FOCAL 23 (50%) 15 (31%) 

HYPERPLASIA, DIFFUSE 

ANGIICTAoIS 1 (2%) 

*LIVER/CENTRILOfaULAR (46) (48) 

DEGENERATION, NOS 

NECROSIS, NOS 1 (2%) 

«LJVER/PERIPORTAL (46) (48) 

FIBROSIS 1 (2%) 

#LIVEP/KEPATOCYTES (46) (48) 

HYPERPLASIA, NODULAR 

HYPEFPLASIA, NOS HYPERPLASIA, FOCAL 

•EILE DUCT (46) (48) 

INFLAMMATION, NOS 6 (13%) 3 (6%) 

HYPERPLASIA, NOS 32 (70%) 43 (90%) 

HYPERPLASIA, FOCAL 1 (2%) 

#PANCREAS (42) (46) 

INFLAMMATION, NOS 10 (24%) 17 (37%) PERIARTERITIS 

HYPEFPLASIA, INTRADUCTAL 1 (23) 

#PANCPEATIC ACINUS (42) (4 6) 

ATROPHY, NOS 4 (10%) ATROPHY, FOCAL 

HYPERPLASIA, FOCAL 1 (2%) 

#ESOPHAGUS (46) (45) 

DYSPLASIA, NOS 1 (2%) 

«STOMACH (45) (4 8) 

EPIDERMAL INCLUSION CYST 1 (2*) ' INFLAMMATION, NOS 1 (2%) ULCER, NOS 2 (4%) 

ULCER,. FOCAL
 

# NUMBER OF ANIMALS KITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMALS NECROPSIED
 

LOW DOSE
 01-0048
 

4 (8%)
 

1 (2%)
 2 (4%)
 

1 (2%)
 5 (10%)
 

(48)
 

1 (2%)
 

(<*8)
 

(48)
 

1 (2%)
 

12 (25%)
 

(48)
 

(45)
 

(45)
 

2 (4%)
 

(47)
 

(45)
 

.1. {2%i
 

HIGH DOSE
 01-0109
 

1 (2%)
 

(48)
 

(48)
 

(48)
 

2 (4%)
 11 (23%)
 

(49)
 

(48)
 

1 (2%)
 

(48)
 

6 (13%)
 

1 (2%)
 

(47)
 

(47)
 

1 (2%)
 

C-6
 
TABLE CI (CONTINUED)
 

LOW DOSE CONTROL(UNTR) 01-0037 HIGH DOSE CONTROL (UNTR) 01-0118 

HYPERPLASIA, NOS 

HYPERPLASIA, BASAL CELL 

HYPERKERATOSIS 

ACANTHOSIS 

6

1

1

 (13%) 

(2%) 

 (2%) 

1 (2%) 

2 (4%) 

2 (4%) 

#SMALL INTESTINE 

INFLAMMATION, ACUTE/CHRONIC 

(43) (46) 

#S.INTESTINE/MUC"OSA 

PIGMENTATION, NOS 

(43) (46) 

#PEYERS PATCH 

HYPERPLASIA, NOS 

(43) 

7 (16%) 

(46) 

12 (26%) 

#DUCDENAL MUCOSA 

PIGMENTATION, NOS 

(43) (46) 

#ILEUM 

INFLAMMATION, NOS 

(43) (46) 

2 (4%) 

#COLON 

ULCER, FOCAL 

NEMATODIASIS 

PARASITISM 

(43) 

3 (7?) 

(46) 

3 (7%) 

UFINARY SYSTEM
 

•KIDNEY	 (46) (48)
 

GLOMERULONEPHRITIS, NOS 33 (12%) 47 (98%)
 

INFLAMMATION, INTEBSTITIAL 1 (2%)
 FIBROSIS, DIFFUSE 6 (13%)
 

NEPHROSIS, NOS
 

HYPEPPIGMENTATION
 

#KI2NEY/TUEULE (46) (48)
 

PIGMENTATION, MOS
 

#KIDNEY/PELVIS (46) (48)
 

CALCIFICATION, FOCAL
 

#UBINARY BLADDER (42) (43)
 

INFLAMMATION, NOS 1 (2%)
 

HYPERPLASIA^ EPITHELIAL 3 (7*1 1 12%1
 

# NUMEER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

• NUMEER OF ANIMALS NECROPSIED
 

LOW DOSE
 01-0048
 

(45)
 

(45)
 

(45)
 

(45)
 

(45)
 

(43)
 

1 (2%)
 

(48)
 

45 (94%)
 

(48)
 

(48)
 

(45)
 

HIGH DOSE
 01-0109
 

(47)
 

1 (2%)
 

(47)
 

2 (4%)
 

(47)
 

(47)
 

30 (64%)
 

(47)
 

(40)
 

(49)
 

1 (2%)
 

47 (96%)
 

1 (2%)
 

(49)
 

2 (4%)
 

(49)
 

1 (2%)
 

(48)
 

C-7
 
TABLE Cl (CONTINUED) 

LOW DOSE 

CONTROL (UNTP) 01-0037 

HIGH DOSE 

CONTROL(UNTR) 01-0118 LOW DOSE 01-0048 HIGH DOSE 01-0109 

ENDOCRINE SYSTEM 

«PITUITARY 

HYPERPLASIA, NOS 

HYPERPLASIA, FOCAL 

HYPERPLASIA, CHFCMOPHOBE-CELL 

(41) 

3 (7T) 

2 (5%) 

(38) 

1 (3%) 2 (5%) 

(44) 

7 (16%) 

(40) 

«ADRENAL CORTEX 

HYPERTROPHY, FOCAL HYPERPLASIA, NODULAR 

HYPERPLASIA, NOS 

HYPERPLASIA, FOCAL 

(43) 

1 (2%) 

1 (2t) 

(47) (46) 

4 (9%) 

(47) 

2 (4%) 

•ADRENAL MEDULLA 

NECROSIS, NOS 

CALCIFICATION, NOS HYPERPLASIA, NODULAR 

HYPOPLASIA, NOS 

HYPERPLASIA, FOCAL 

(43) 

1 (2%) 

1 (2*) 

1 (2*) 6 (14%) 

(47) 

1 (2%) 

4 (9%) 

(46) 

1 (2%) 

1 (2%) 

1 (2%) 

(47) 

3 (6%) 

1 (2%) 

#THYBOID 

FOLLICULAR CYST, NCS 

LYMPHOCYTIC INFLAMMATORY INFILTR 

INFLAMMATION, ACUTF/CHPONIC 

PIGMFNTATION, NOS 

HYPEPPIGKENTATION 

HYPERPLASTIC NODULE 

HYPERPLASIA, CYSTIC 

HYPERPLASIA, ADENOMATOUS 

HYPERPLASIA, C-CELL 

HYPERPLASIA, FOLLICULAR-CELL 

(45) 

1 (2%) 1 (27) 

(48) 

3 (6%) 

(47) (49) 

9 (18%) 

1 {2%) 

1 (2%) 

2 (4%) 

47 (96%) 

1 (2%) 

2 (4%) 

3 (6%) 

1 (2%) 

•PARATHYROID 

HYPERPLASIA, NOS 

(32) (28) 

1 (4%) 

(30) (24) 

1 (4%) 

«PANCREATIC ISLETS 

HYPERPLASIA, NOS 

(42) 

2 (S*) 

(46) 

1 (2%) 

(45) (48) 

2 (4%) 

RIFECEUCTIVr SYSTEM 

•MAMMARY GLAND 

GALACTOC£LE 

HYPERPLASIA, NOS 

(46) 

5 (11%) 

(48) 

2 (4%) 

4 (Q%1 

(48) 

1 (2%) 1 (2%1 

(49) 

# NUMEEP OF ANIMALS «ITH TISSUE EXAMINED MICROSCOPICALLY 

* NU1EER OF ANIMALS NECPOPSIED 

C-8
 
TABLE CI (CONTINUED) 

LOW DOSE 

CONTROL(UNTF) 01-0037 

HIGH DOSE 

CONTROL (UNTR) 01-0118 

LOW DOSE 

01-0048 HIGH DOSE 01-0109 

•EBEPUTIAL GLAND 

CYSTIC DUCTS 

-JLCER, NOS 

ABSCESS, NOS 

HYPEBPLASIA, NOS 

HYPEPPLASIA, EPITHELIAL 

(46) 

1 (2*) 

1 (2*) 

(48) (48) (49) 

1 (2%) 

1 (2%) 

1 (2%) 

•PROSTATE 

INFLAMMATION, NOS 

INFLAMMATION, FOCAL INFLAMMATION, \CUTF 

INFLAMMATION ACUTE AND CHRONIC 

INFLAMMATION, ACUTE/CHRONIC DEGENERATION, NOS 

ATROPHY, NOS 

HYPERPLASIA, FOCAL 

HYPERPLASIA, PAPILLARY 

HYPERPLASIA, ADENOMATOUS 

METAPLASIA, SQUAMOUS 

(45) 

21 (47%) 

3 (7%) 

5 (11%) 

2 (4*) 

5 (11%) 

(44) 

17 (39%) 

(47) 

1 (2%) 

5 (11%) 3 (6%) 

1 (2%) 

1 (2%) 

(47) 

11 (23%) 

1 (2%) 

1 (2%) 1 (2%) 

*SEMINAL VESICLE 

ATPOPHY, NOS 

HYPERPLASIA, EPITHELIAL 

(46) (4 8) (48) 

35 (73%) 

(49) 

11 (22%) 

1 (2%) 

•COAGULATING GLAND 

ATROPHY, NOS 

(46) (4 8) (*8) 

3 (6%) 

(49) 

#TESTIS 

MINERALIZATION ATROPHY, NOS 

ASPERMATOGENESIS 

HYPERPLASIA, INTERSTITIAL CBLL 

(45) 

2 (4%) 

1 (2%) 19 (42%) 

(47) 

1 (2%) 6 (13%) 

3 (6%) 

(48) 

3 (6%) 

1 (2%) 

(49) 

4 (8%) 

3 (6%) 

#TES1IS/TUBULE 

DEGENERATION, NOS 

(45) 

6 (13%) 

(47) (48) 

3 (6%) 

(49) 

19 (39%) 

NERVOUS SYSTEM 

NONE 

SPECIAL SENSE ORGANS 

•EYE 

CATARACT 

(46) (48) (48) (**9) 

1 (2%) 

# NUMBFR OF ANIMALS WITH TISSUE EXAMINED

• NUMEER OF ANIMALS NECROPSIED 

 MICROSCOPICALLY 

C-9
 
TABLE CI (CONCLUDED)
 

LOW DOSE HIGH DOSE
 CONTROL(UNTR) CONTROL(UNTR)
 

01-0037 01-0118
 

•EYE/RETINA	 (46) (48)
 

DEGENERATION, NOS
 

•EYE/LACBIMAL Gi,AKD	 (46) (48)
 

INFLAMMATION, NOS
 

MUSCULOSKELETAL SYSTEM
 

•CA3TILAGE,NOS	 (46) (48)
 

CYST, NUS 1 (27)
 

BOCY CAVITIIS
 

•ABDOMINAL CAVITY	 (46) (48)
 

NECROSIb, FAT
 

•PERITONEUM	 (46) (48)
 

INFLAMMMION, CHRONIC FOCAL
 

•PLEURA	 (46) (48)
 

INFLAMMATION, FI2PIN0US
 

•EPICARDIUM	 (46) (48)
 

INFLAMMATION, NOS
 

INFLAMMATION, CHRONIC
 

•MESENTERY	 (46) (48)
 

PERIARTERITIS
 

ALL CTHEP SYSTEMS
 

OMENTUM
 

NECROSIS, FAT 2
 

SPECIAL MCREHOLOGY SUMMARY
 

AUTO/NECROPSY/HISTC PSRF 1
 

AUT01YSIS/NO NECROPSY 4 1
 

# NUMEER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

• NUHEER OF ANIMALS NECROPSIED
 

LOW DOSE
 01-0048
 

(48)
 

(48)
 

1 (2%)
 

(48)
 

(48)
 

1 (2%)
 

(48)
 

1 (2%)
 

(48)
 

1 (2%)
 

(48)
 

1 (2%)
 1 (2%)
 

(48)
 

2 (4%)
 

2
 

HIGH DOSE
 01-0109
 

(49)
 

1 (2%)
 

(49)
 

(49)
 

(49)
 

(49)
 

(49)
 

(49)
 

(49)
 

C-10
 

1 
TABLE C2

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN FEMALE RATS 

TREATED WITH 2,4-DIAMINOANISOLE SULFATE 

LOW DOSE 

CONTROL (UNTR) 

02-0037 

HIGH DOSE 

CONTROL (UNTR) 

02-0118 

ANIMALS INITIALLY IN STUDY 50 

ANIMALS NECRCPSIED 49 

ANIMALS EXAMINED HISTOPATHOLOGICALLY ** 49 

50 

50 

50 

INTEGUMENTARY SYSIEM 

• SKIN (49) (50)
 INFLAMMATION, NOS 1 (2%)
 GRANULOMA, FOFSIGN BODY
 

•SUBCUT TISSUE	 (49) (50)
 

MINERALIZATION 1 (2%)
 AESCESS, NOS 1 (2%)
 

RZSEIEATOBY SYSTEM
 

tTRACHEA (48) (4 9)
 

INFLAMMATION, NOS 9 (19%)
 

INFLAMMATION, CHRONIC 10 (21%)
 

FCLYP, INFLAMMATORY 1 (2%)
 

«LUNG/EBONCHUS (49) (50)
 

BRONCHIECTASIS 1 (2%)
 

INFLAMMATION, NOS 1 (2%) 3 (6%)
 INFLAMMATION, CHFONIC 9 (18%)
 

HYPERPLASIA, FOCAL
 

#IUNG/ERONCHIOLE (49) (50)
 

INFLAMMATION, NOS 1 <2S)
 

#LUNG (49) (50)
 

INFLAMMATION, NOS 1 (2%)
 INFLAMMATION, FOCAL 7 (14%)
 

INFLAMMATION, INTERSTITIAL 2 (4%) 6 (12%)
 

INFLAMMATION, SUPPURATIVE
 

BRONCHOPNEUMONIA SUPPURATIVE
 

BRONCHOPNEUMONIA NECROTIZING
 

ABSCESS^. NOS
 

# NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY 

• NUMEEE OF ANIMALS NECEOPSILD 

**EXCLUDES PARTIALLY AUTOLYZED ANIMALS 

LOW DOSE
 02-0048
 

50
 

49
 

49
 

(49)
 

(49)
 

(48)
 

(48)
 

1 (2%)
 

(48)
 

(48)
 

1 (2%)
 

HIGH DOSE
 02-0109
 

50
 

49
 

49
 

(49)
 

1 (2%)
 

(49)
 

(48)
 

(49)
 

2 (4%)
 

(49)
 

(49)
 

1 (2%)
 6 (12%)
 

2 (4%)
 

C-ll 
TABLE C2 (CONTINUED) 

ERCNCHOPNEUMONIA, CHRONIC
 

PERIVASCULITIS
 

CYTCMEGALY
 

HYPOPLASIA, EPITHFLIAi.
 

FYPEFPLASIA, ADuNCfATOUS
 

HEMATCFOJETTC bYSTEf*
 

#ECTE PARRCfc
 

OSTEOSCLEROSIS
 MYELOSCLEROSIS
 

#SPJ EEN
 

HYPEPPIG^ENTATION
 

FEMOSIDEFOSIS
 

HYPERPLASIA, NOS
 

HYPFPPLAbIA, HEMATOPOIETIC
 

HYPERPLASIA, EFYThl-OID
 

HYPERPLASIA, PLASMA CELL
 

HYPERPLASIA, RETICULUM CELL
 HEMATOPOIESIS
 

ERYTHFOPCIESIS
 

«SPLENIC CAPSULE
 

HEMORRHAGIC CYsT
 

FIBROSIS, FOCAL
 

#LYMPH NCDE
 

INFLAMMATION, fcOS
 

HYPERPLASIA, NOS
 

FLASPACYTOSIS
 

HYPERPLASIA, PLASMA CELL
 

HYPERPLASIA, LYMPHOID
 

#AETCMINAL LYMPh NODE
 

PIASMACYTOSIS
 

CIRCULATORY SYSTEM
 

«HEART
 PERIARTERITIS
 

PERIVASCULITIS
 

tMYCCARDIUM
 

TNFLAMMATION- NOS
 

LOW DOSE
 

CONTROL (UNTR)
 

02-0037
 

6 (12%)
 

(48)
 

(49)
 

1 (2%)
 

3 (67)
 17 (35%)
 

1 (2*)
 11 (22%)
 

(49)
 

(41)
 

3 (79?)
 

2 (5%)
 

3 0*)


1 (2%)
 

(41)
 

(49)
 

(49)
 

1 I2f)
 

HIGH DOSE
 

CONTROL (UNTR)
 

02-0118
 

1 (2%)
 

(46)
 

1 (2%)
 

(4 8)
 

12 (25%)
 

25 (52%)
 19 (40%)
 

(4 8)
 

1 (2%)
 

(47)
 

1 (2%)
 

4 (9%)
 

(47)
 

(50)
 

(50)
 

1 (2%)
 

LOW DOSE
 

02-0048
 

1 (2%)
 

(43)
 

(48)
 

2 (4%)
 

2 (4%)
 

10 (21%)
 

1 (2%)
 

(<*8)
 

(43)
 

(43)
 

1 (2%)
 

(48)
 

1 (2%)
 

(48)
 

HIGH DOSE
 

02-0109
 

1 (2%)
 

1 (2%)
 

(45)
 

1 (2%)
 

(49)
 

2 (4%)
 

1 (2%)
 

(49)
 

1 (2%)
 

(41)
 

(41)
 

(48)
 

2 (4%)
 

(48)
 

# NUMBER OF ANIMALS WiTu

 TISSUE EXAMINED MICFOSCOPICALLY
 

* NUMEER OF HIJIMALS NECROPSILD
 

C-I2
 
TABLE C2 (CONTINUED)
 

LOW DOSE CONTROL (UNTR) 02-0037 

HIGH DOSE CONTROL(UNTK) 02-0118 LOW DOSE 02-0048 HIGH DOSE 02-0109 

INFLAMMATION, INTERSTITIAL 

INFLAMMATION, ACUTE/CHRGNIC 

FIBROSIS 

DEGENERATION, NOS 

24 

5 

(49%) 

(10%) 

23 

15 

(46%) 

(30%) 

1 (2%) 1 

11 

(2%) 

(23%) 

«ENDOCARDIUM 

INFLAMMATION, hOS 

INFLAMMATION, SUPPURATIVE 

INFLAMMATION, ACUTE/ChRONIC 

INFLAMMATION PROLIFERATIVE 

(49) (50) 

1 (2%) 

(48) (48) 

1 

1 

1 

(2%) 

(2%) 

(2%) 

*PORTAL VEIN 

THROMBUS, MUHAL 

(49) 

1 (2*) 

(50) C*9) (49) 

DIGESTIVE SYSTEM 

•LIVER 

FIBROSIS 

PERIVASCULITIS NECROSIS, FOCAL 

NECROSIS, COAGULATIVE 

rETAMORPHOSIS FATTY HYPERPLASIA, NODULAR 

HYPERPLASTIC NODULF 

HYPERPLASIA, FOCAL 

HYPERPLASIA, DIFFUSE 

ANGIECTASIS 

HYPERPLASIA, ERYTHPOID 

HEMATOPOIESIS 

(49) 

1 

1 

4 

2 

1 

1 

22 

1 

(2*) 

(2%) 

(8%) 

(4$) 

(2 S> (2%) 

(45%) 

(2%) 

(50) 

2 

6 

38 

1 

2 

(*%) 

02%) 

(76%) 

(2%) 

<«*%) 

(48) 

1 

1 

4 

2 

(2%) 

(2%) 

(8%) 

(4%) 

(49) 

1 

1 

2 

(2%) 

(2%) 

(4%) 

tLIVER/CENTRILOBULAR 

DEGENERATION, NOS NECROSIS, NOS 

(49) (50) (48) 

1 (2%) 

(49) 

2 

4 (4%) (8%) 

•LIVER/PERIPORTAL 

INFLAMMATION, ACUTF/CHRONIC 

(49) (5 0) (48) (<*9) 

1 (2%) 

#LIVER/HEPATOCYTES 

HYPERPLASIA, NOS 

HYPERPLASIA, FOCAL 

HYPERPLASIA, DIFFUSE 

(49) (50) (48) 

1 

31 (2%) (65%) 

(49) 

1 

28 

2 

i2%) 

'57%) 

'4%) 

•BILE DUCT 

INFLAMMATION. NOS 

(49) 

5 (10%) 

(50) 

1 (2%) 

(49) (49) 

# NUMEER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY 

• NUMEER OF ANIMALS NECROPSIED 

C-I3
 
TABLE C2 (CONTINUED)
 

LOW DOSE 

CONTROL (UN~T) 

02-0037 

HIGH DOSE 

CONTROL (UNTR) 

02-0118 

LOW DOSE 

02-0048 

HIGH DOSE 

02-0109 

INFLAMMATION, CHFONIC 

hYFEFPLASIA, KOS 

HYPERPLASIA, FOCAL 

27 (55%) 32

1

 (64%) 

 (2%) 

1 (2%) 

•TANCREAS 

INFLAMMATION, NOS 

(46) 

7 (1S%) 

(43) 

6 (13%) 

(46) 

1 (2%) 

(47) 

•PANCREATIC DUCT 

HYPERPLASIA, NGS 

(46) 

1 (2?) 

(48) (46) (47) 

•PANCREATIC ACINUS 

ATPOPHY, NOS 

HYPEFPLASIA, DIFFUSE 

(46) 

2 (4%) 

C+8) (46) 

1 (2%) 

(47) 

6 (13%) 

1 (2%) 

#STCMACh 

INFLAMMATION, NCS 

ULCER, NCS 

INFLAMMATION, FCCAL 

FROSICN 

HYPERPLASIA, nPITFFLIAL 

HYPEFPLASIA, BASAL CELL 

ACANTHOSIS 

(48) 

2 (4?) 

2 (47) 

1 (2£) 

(4 8) 

1 (2%) 

2 (4%) 

(47) 

1 (2%) 

1 (2%) 

(49) 

1 (2%) 

#GASTPIC MUCOSA 

HYPEFPLASIA, NOS 

(^8) 

1 {21) 

(48) (47) (49) 

•FEYEPS PATCh 

HYPERPLASIA, NOS 

(47) 

6 (13%) 

(4 8) 

15 (31%) 

(46) (47) 

•EUOCENAL MUCOSA 

PIGMENTATION, NCS 

(47) (48) (46) (47) 

24 (51%) 

•COLON 

NEMATODIASIS 

TARASITTSM 

(43) 

3 (7%) 

(46) 

2 (4%) 

(45) (40) 

URINARY SYSTEM 

•KIDNEY 

HYDRONEPHROSIS 

GLOMERULONEPHRITIS, NOS 

INFLAMMATION, INTERSTITIAL 

GLOMERULONEPHRITIS, MEMBRANOUS 

INFLAMMATION. CHRONIC 

(49) 

1 (2£) 

33 (67%) 

1 (2S) 

1 (2?) 

1 (2\\ 

(50) 

43 (86%) 

(48) 

31 (65%) 

1 (2%) 

(49) 

# NUMBER OF ANIMALS WITH TISSUE

• NUMBER OF ANIMALS NECROPSIED 

 EXAMINED MICROSCOPICALLY 

C-I4
 
TABLE C2 (CONTINUED)
 

FIBROSIS, DIFFUSE
 

NEPHFCSIS, NOS
 

IKIDNEY/CORTEX
 

CYST, NOS
 

#KIDNEY/GLO!1ERULUS
 

NEPHROSIS, NOS
 

•KIZNEY/TUBULE
 

PIGMENTATION, NOS
 

•UFINAPY BLADDER
 

INFLAMMATION, t!OS
 

ENECCRINI SYSTEM
 

«PITUITARY
 

PERIVASCULITIS
 

HYPERPLASIA, NOS
 

HYPERPLASIA, FOC\L
 HYPERPLASIA, CHRCMCPHO3E-CELL
 

•ADRENAL
 

METAMORPHOSIS FATTY
 

«ADRENAL COSTEX
 

NODULE
 

HYPERPLASIA, NODULAE
 HYPERPLASIA, NOS
 HYPERPLASIA, FOCAL
 

•ADRENAL MEDULLA
 

HYPERPLASIA, NODULAR
 

HYPEPPLASIA, NOS
 

HYPERPLASIA, FOCAL
 

tTHYBOID
 

CYSTIC FOLLICLES
 

FOLLICULAR CYST, NOS
 

LYMPHOCYTIC INFLAMMATORY INFILTR
 

HYPEPPIGdZNTATION
 

HYPERPLASIA, C-CELL
 

HYPESPLASIA, FOLLICULAF-CSLL
 

LOW DOSE
 CONTROL(UNTR)
 

02-0037
 

(49)
 

(49)
 

(49)
 

(41)
 

1 (2%)
 

(43)
 

2 (5»)
 

1 (27)
 

(46)
 

(46)
 

1 [2%)
 

7 (15S)
 

4 (9?)
 

(47)
 

1 (27)
 

HIGH DOSE
 CONTROL (UNTR)
 

02-0118
 

1 (2%)
 

(50)
 

(50)
 

(50)
 

(46)
 

(40)
 

1 (3%)
 

3 (8%)
 

(49)
 

1 (2%)
 

(49)
 

(49)
 

3 (6%)
 

3 (6%)
 

(4 5)
 

1 (2%)
 

1 (2%)
 

LOW DOSE
 

02-0048
 

(48)
 

1 (2%)
 

(48)
 

1 (2*)
 

(48)
 

(43)
 

(47)
 

(47)
 

(47)
 

1 (2%)
 

1 (2%)
 

(47)
 

(46)
 

HIGH DOSE
 02-0109
 

23 (47%)
 

(49)
 

(49)
 

(49)
 

1 (2%)
 

(46)
 

(38)
 

(49)
 

(49)
 

8 (16%)
 

(49)
 

*• (8X)
 

(49)
 

12 (24%)
 

1 (2%)
 

49 (100%)
 15 (31%)
 

# NUMSER OF ANIMALS WITH TISSUE EXAMINED MICFOSCOPICALLY
 

* NUfiEER OF ANIMALS KECROPSIED
 

C-15
 
TABLE C2 (CONTINUED)
 

LOW DOSE 

CONTROL(UNTP) 

02-0037 

HIGHI DOSE 

CONTPOL(UNTR) 

02-0118 

LOW DOSE 

02-0048 

HIGH DOSE 

02-0109 

tPANCPEATIC ISLETS 

hYPfPPLASIA, NOS 

(46) 

1 (2%) 

(48) (46) (47) 

REEFCEUCTIVE SYSTEM 

•MAhMARY GLAND 

CALACTCCELE 

hYPERPLiSIA,

FYPERPLA£IA,

HYPERPLASIA,

 MOS 

 PAPILLARY 

 ADENOMATOUS 

(49) 

5 

17 

1 

(10%) 

(35%) 

(2%) 

(50) 

16 

8 

(32%) 

(16%) 

(49) 

8 

1 

(16%) 

(2%) 

(49) 

•MAFMAFY DUCT 

FIBROSIS 

(49) (5 0) (49) 

1 (2%) 

(49) 

•CLITOFIS 

HYPERPLASIA, EPITHELIAL 

(49) (50) (49) (49) 

1 (2%) 

•CLITORAL GL1KD 

hYPEFiPL^SIA, NOS 

(49) (50) (49) (49) 

2 (4%) 

iUTERUS 

HYDROMETRA 

INFLAMMATION, SUPPUFATIVE 

TYOrETRi 

INFLAMMATION, ACUTE 

ABSCESS, NOS 

hYPEFFLPSIA, ADENOMATOUS 

tCLYT, INFLAMMATORY 

(46) 

3 

1 

2 

5 

(6*) 

{2%) 

(4*) 

(10%) 

(50) 

1 (2%) 

(46) 

4 

2 

(9%) 

(<*%) 

4 

2 

1 

(9%) 

(4%) 

(2%) 

#UT£RUS/ENDOMETRIUM 

INFLAMMATION, I OS 

INFLAMMATION, FOCAL 

INFLAMMATION, SUPPMFATIVE 

INFLAMMATION, iCUTE 

INFLAMMATION, ACUTc/CHFONIC 

hYPEFPLASIA, NCS 

hYFEFPL»SIA, EPITHELIAL 

tYPEIPLASIA, CYSTIC 

hYPEFFLASIA, ADENOMATOUS 

(*»8) 14 

1 

2 

1 

2 

1 

(29%) 

(2*) 

(4%) 

(2«) 

(47) 

(2%) 

(50) 

22 

6 

1 

(44%) 

(12%) 

(2%) 

(46) 

1 

9 

4 

2 

1 

22 

(2%) 

(20%) 

(9%) 

(<*%) 

(2%) 

(48%) 

(46) 

1 

1 

4 

1 

5 

(2%) 

(2%) 

(9%) 

(2%) 

(11%) 

#OVARY/OV1DUCT 

INFLAMMATION,

INFLirMATION.

 NOS 

 SUPPURATIVE 

(48) 

1 (2%) 

(50) 

10 

2 

(20%) 

(4%) 

(46) (46) 

1 

2 

(2%) 

I***) 

# NUtlEEP OF ANIMALS WITH TISSUc EXAMINED MICPCSCOPICALLY 

* NUMEER OF ANIMALS NECROPSIED 

C-16
 
LOW DOSE
 

CONTROL (UNTR)
 

02-0037
 

(47)
 

4 (97)
 

1 (2%)
 

1 (2%)
 

(U9)
 

(49)
 

(49)
 

(U9)
 

1
 

HIGH DOSE
 

CONTROL (UNTR)
 

02-0118
 

(4 9)
 

8 (16%)
 

(50)
 

 (2%)
 

(50)
 

1 (2%)
 

(50)
 

1 (2%)
 

(50)
 

1
 

LOW DOSE
 

02-0048
 

(46)
 

2 (4%)
 

1 (2%)
 

(49)
 

1 (2%)
 

1 (2%)
 

1 (2%)
 

(49)
 

1 (2%)
 

(49)
 

(49)
 

1 (2%)
 

1
 

1
 

HIGH DOSE
 02-0109
 

1 {2%)
 

(47)
 

(49)
 

1 i2%)
 

(49)
 

1 (2%)
 

<*9)
 

0*9)
 

1
 

INFLAMMATION, CHRONIC
 

#OVARY
 

CYST, NOS
 

INFLAMMATION, ΡCUTE/CHFONIC
 

INFLAMMATION, FOCAL GRANULOMATOU
 

HYPERPLASIA, INTERSTITIAL CELL
 

NERVOUS SYSTIM
 

NONE
 

SFECIAL SuNSE ORGANS
 

•ΕΥΓ
 

SYNECHIA, ANTEPIOF
 SYNECHIA, POSTERIOR
 

CATAFACT
 

*EYE/RETINA
 

DEGENERATION, NOS
 

ATROPHY, NOS
 

*HARDERIAN GLAND
 

HYPERPLASIA, NOS
 

MUSCULOSKELETAL SYSTFM
 

•SKflETAL	 MUSCLE
 

METAPLASIA, OSSEOUS
 

BOIY CAVITIES
 

NONE
 

All CTHEP SYSTEMS
 

OMENTUM
 

NECROSIS^ FAT
 

SPECIAL MORPHOLOGY SUMMARY
 

NO LESION REPORTED
 AUTOLYSIS/NO MXFOPSY
 

TABLE C2 (CONCLUDED)
 

t NUMEER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 • NUMEER OF ANIMALS NECEOPSIED
 

C-17
 

APPENDIX D

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC 

LESIONS IN MICE TREATED WITH 2,4-DIAMINOANISOLE SULFATE 

TABLE Dl
 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN MALE MICE
 

TREATED WITH 2,4-DIAMINOANISOLE SULFATE
 

ANIMALS INITIALLY IN STUDY
 

ANIIULS KISSING
 

ANIMALS NiSCEvOPSIED
 

ANIMU S EXAMINED HISTOPATHOLOGICALLY
 

INTEGUMENTARY SYSTEM
 

•SKIN
 

HEMORRHAGE
 

INFLAMMATION, NOS
 

INFLAMMATION, FOCAL
 

INFLAMMATION, NECROTIZING
 

FIBROSIS
 

ALOPECIA
 

•SUFCUT	 TISSUE
 

ABSCESS, NOS
 

NECROSIS, NOS
 

RESPIRATORY SYSTEM
 

•LUNG/ERONCHUS
 

INFLAMMATION, NUS
 

INFLAMMATION, FOCAL
 

#LUNG/ERONCHIOLE
 

INFLAMMATION, FOCAL
 

«LUNG
 

INFLAMMATION, NOS
 

INFLAMMATION, INTEFSTITIAL
 

HYPERPLASIA, EPITHELIAL
 

HYPERPLASIA, ALVEOLAR EPITHELIUM
 

#LUNG/ALV£OLI
 

INFLAMMATION, FOCAL
 

FIBROSIS, FOCAL
 

HEMATOPOIETIC SYSTEM
 

«BONE MARROW
 

HYPERPLASIA. NOS
 

• NUMBER OF ANIMALS WITH TISSU E

• NUMBER OF ANIMALS NECROPSIED 

**EXCLUDES PARTIALLY AUTOLYZED ANIMALS 

LOW DOSE
 

CONTROL (UNTR)
 

05-0037
 

50
 

U8
 

** U8
 

(48)
 

1 (2%)
 

1 (22)
 

(48)
 

1 (2%)
 

(48)
 

1 (2S)
 

1 (2%)
 

(48)
 

(48)
 

1 (2%)
 

14 (29%)
 

2 (4%)
 

(48)
 

2 (4%)
 

1 (2%)
 

(47)
 

HIGH DOSE
 

CONTROL (UNTR)
 

05-0118
 

50
 

1
 

49
 

49
 

(49)
 

1 (2%)
 

3 (6%)
 

1 (2%)
 

(49)
 

(49)
 

1 (2X)
 

(49)
 

1 (2%)
 

(49)
 

10 (20%)
 

(49)
 

(48)
 

 EXAMINED MICPCSCOPICALLΥ 

LOW DOSE
 

05-0048
 

50
 

49
 

49
 

(49)
 

(49)
 

1 (2%)
 

(48)
 

(48)
 

(48)
 

3 (6%)
 

(48)
 

(46)
 

HIGH DOSE
 

05-0108
 

50
 

49
 

49
 

(49)
 

1 (2%)
 

(49)
 

3 (6%)
 

(48)
 

(48)
 

(48)
 

2 (4%)
 

(48)
 

(48)
 

1 '2%L .
 

D-3
 
TABLE Dl (CONTINUED)
 

LOW DOSE CONTROL (UNTR) 

05-0037 

HIGH DOSE CONTROL (UNTR) 

05-0118 

HYPERPLASIA, DIFFUSE 

HYPERPLASIA, HEMATOPOIETIC 

#SPLEEN 

INFLAMMATION, NOS 

FIBROSIS, FOCAL ATROPHY, NOS 

HYPERPLASIA, NOS ANGIECTASIS 

(47) 

1 

2 

(2%) 

(4%) 

(49) 

6 (12%) 

RETICULOCYTOSX5 

HYPERPLASIA, HEMATOPOIETIC 

HYPERPLASIA, ERYTHEOID 

HYPERPLASIA, LYMPHOID 

HEMATOPOIESIS 

2 

2 

2 

(4%) 

(4%) 

(4*) 

1 

5 

1 

(2%) 

(10%) 

(2%) 

GRANULOPOIESIS 

#LYMPH NODE 

HEMORRHAGIC CYST INFLAMMATION, NOS 

DEGENERATION, CYSTIC 

HYPERPLASIA, NOS 

RETICULOCYTOSIS 

PLASKACYTOSIS 

(44) 

1 

13 

1 

2 

(2%) 

(30%) 

(2%) 

(5%) 

(42) 

10 

1 

2 

(24%) 

(2%) 

(5%) 

HYPERPLASIA, HEMATOPOIETIC 

HYPERPLASIA, LYMPHOID 

MYELOID METAPLASIA 

1 

2 

2 

(2%) 

(5%) 

(5%) 

3 (7%) 

•MEDIASTINAL L.NODE 

NECROSIS, NOS 

PLASMACYTOSIS 

(44) 

1 (2%) 

(4 2) 

tABDOMINAL LYMPH NODE 

HYPERPLASIA, RETICULUM CFLL 

(44) (4 2) 

•PANCREATIC L.NODE 

INFLAMMATION, HOS 

(44) 

1 (2%) 

(4 2) 

•MESENTERIC L- NODE 

HEMORRHAGE 

INFLAMMATION, NOS INFLAMMATION, ACUTE/CHRONIC 

INFLAMMATION, CHPONIC 

AMYLOIDOSIS 

(44) 

1 

9 (2%) 

(20%) 

(42) 

HYPERPLASIA, NOS PLASMACYTOSIS
 

• NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMALS NECROFSIED
 

LOW DOSE
 05-0048
 

(48)
 

1
 

1
 

1
 

1
 

1
 

1
 

(45)
 

(45)
 

(45)
 

1
 

(45)
 

(45)
 

2
 

9
 

1
 

1
 

1
 

(2%)
 

(2%)
 

(2%)
 

(2%)
 

(2%)
 

(2%)
 

(2%)
 

(4%)
 

(20%)
 

(2%)
 

(2%)
 

(2%)
 

HIGH IOSE
 05-0*08
 

15 (31%)
 

31 165%)
 

(48)
 

2 (4%)
 

2 (4%)
 

1 (2%)
 

(45)
 

1 (2%)
 

(45)
 

1 (2%)
 

(45)
 

(45)
 

(45)
 

1 (2%)
 

JLI22]_.
 

D-4
 
TABLE Dl (CONTINUED)
 

LOW DOSE HIGH DOSE 

CONTROL (UNTR) CONTROL (UNTR) 05-0037 05-0118 

HYPERPLASIA, RETICULUM CELL 

HYPERPLASIA, LYMPHOID 

•THYMUS (34) (28) 

NECROSIS, NOS 1 (3*) 

#THYMIC MEDULLA (34) (28) 

HYPERPLASIA, LYMPHOID 

CIRCULATORY SYSTEM
 

#HEABT (48) (49)
 

MINERALIZATION 1 (2%)
 

PERIARTERITIS
 

ARTERIOSCLEROSIS, NOS
 

#HEART/V£NTRICLE (48) (49)
 

MELANIN 2 (4%)
 

•MYOCARDIUM	 (48) (49)
 

INFLAMMATION, INTERSTITIAL 2 (4*)
 

INFLAMMATION PROLIFERATIVE
 

FIBROSIS 5 (10%)
 

DEGENERATION, NOS
 

•BLOOD VESSEL	 (48) (4 9)
 

INFLAMMATION, NOS 2 (4%)
 

•PULMONARY	 ARTERY (48) (49)
 

MINERALIZATION 2 (4%)
 

•SPLENIC ARTERY	 (48) (49)
 

THROMBOSIS, NOS
 

DIGESTIVE SYSTEM
 

#SALIVAFY GLAND (47) (48)
 

INFLAMMATION, NOS 2 (4?)
 INFLAMMATION, ACUTE/CHRONIC
 

PERIVASCULAR CUFFING 1 (2%)
 

«LIVER (48) (48)
 

INFLAMMAriONx_ACUTE^CHROKIC
 

# NUMEER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

• NUMEER OF ANIMALS NECROPSIED
 

LOW DOSE
 05-0048
 

1 (2%)
 

1 (2%)
 

(35)
 

(35)
 

1 (3%)
 

(48)
 

1 (2%)
 

(48)
 

(48)
 

1 (2%)
 

(49)
 

(49)
 

(49)
 

(47)
 

14 (30%)
 

(49)
 

HIGH DOSE
 05-0108
 

1 (2%)
 

(33)
 

(33)
 

(47)
 

3 (6%)
 

(47)
 

(47)
 

1 (2%)
 

(49)
 

(49)
 

(49)
 

1 (2%)
 

(47»
 

(49)
 

1 (2%)
 

D-5
 
TABLE Dl (CONTINUED)
 

LOW DOSE HIGH DOSE CONTROL (UNTR) CONTROL (UNTR) 

05T 0037 05-0118 

NECROSIS, FOCAL 13 (27%) 9 (19%) 

AMYLOIDOSIS 

METAMCPPHOSIS ?ΑΓΤΥ 3 (6*) 

riYPEFPLASIA, NODULAR 2 (Hf) 

riYPEFPLASTIC NODULE 1 (2%) 

HYPEFPLAbIA, FOCAL 1 (2%) HYPEFPLPSIA, DIFFUSE 

ANGIECTaoIS 1 (2%) MYELOID METAPLASIA 1 (2%) 

#LIVEP/riEPATOCYTc3 (48) (48) 

OEGENFPATION, NOS 1 (27) 

HYPERTROPHY, DIFFUSE 

• GALLBLALD-fcR (48) (49) 

INFLAMMATION, FOCAL 1 (2%) 

*EILE DUCT (48) (49) 

INFLAMMATION, CHPOM C 

•PANCREAS (48) (47) 

INFLAMMATION, NOS 7 (1*%) 1 (2%) 

INFLAMMATION, FOCAL 1 (2%) 

INFLAMMAIION ACTIVL CHRONIC 

DfcGENEPAHON, CYSTIC 1 (2%) METAMORPHOSIS FATTY 1 (2%) 

#PANCRE?TIC DUCT (48) (47) 

CALCIFICATION, NOS 

HYPERPLASIA, NOS 1 (2%) 

#PANCPEATIC ACINUS (48) (47) 

HYPERTROPHY, FOCAL 1 (2*) HYPERPLASIA, FOCAL 1 (2%) 

•STOMACH (47) (4 8) 

INFLAMMATION, NOS 13 (2fi%) 

ULCER, NOS 1 (2%) INFLAMMATION, FOCAL 1 (2%) 2 (4%) 

INFLAMMATION, INTEFSTITIAL 1 (2%) 

INFLAMMATION, NECROTIZING 1 (2%) INFLAMMATION, ACUTE FOCiL 

INFLAMMATION, ACUTE/CHROMC 

hYPEFPLASIA, NOS 1 (2%) HYPERPLASIA­ FOCAL 1 i2%) 1 (2%) 

# NUKEER OF ANIMALS KITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMEEE OF ANIMALS NECROPSIED
 

LOW DOSE
 

05-0048
 

1 (2%)
 

2 (4%)
 

1 (2%)
 

1 (2%)
 

(49)
 

(49)
 

(49)
 

1 (2%)
 

(44)
 

1 (2%)
 

(44)
 

(44)
 

(46)
 

1 (2%)
 2 (4%)
 

HIGH DOSE
 05-0108
 

1 (2%)
 

1 (2%)
 

(49)
 

1 (2%)
 

(49)
 

(49)
 

(47)
 

(47)
 

1 (2%)
 

(47)
 

1 (2%)
 

(48)
 

1 (2%)
 

D-6
 
TABLE Dl (CONTINUED)
 

LOW DOSE 

CONTROL (UNTR) 

05-0037 

HIGH DOSE 

CONTROL (UNTR) 

05-0118 

LOW DOSE 

05-0048 

HIGH DOSE 

05-0108 

HYPERKERATOSIS 

ACANTHOSIS 

3

3

 (6%) 

 (6*) 

1

1

 (2%) 

(2%) 

#GASTRIC MUCOSA 

HYPERPLASIA, FOCAL 

(47) 

1 (2f) 

(48) (46) (48) 

#SMALL INltSTINS 

INFLAMMATION, ACUTL/CHRCNIC 

hYPEFPLASIA, ADENOMATOUS 

HYPERPLASIA, LYPPhCID 

(48) (49) (47) 

1 (2%) 

(47) 

1 (2%) 

1 (2%) 

#PEYERS PATCH 

HYPERPLASIA,

FYPEPFLASIA,

 NUS 

 LYMPHOID 

(48) 

2 (U?) 

(49) 

7 (14%) 

(47) 

1 (2%) 

(47) 

2 (4%) 

#ILEUM 

HEMORRHAGE 

INFLAMMATION, NOS 

AMYLOIDOSIS 

ATROPHY, NOS 

HYPERPLASIA, NOS 

(48) 

1 (27) 

2 (4%) 

(^9) (47) 

1

1

1

 12%) 

 (2%) 

 (2%) 

(47) 

#COLON 

PARASITISM 

(45) 

1 (22) 

(43) 

3 (7%) 

(46) (39) 

OFINAEY SYSTEM 

•KIDNEY 

GLOMERULONEPHRITIS, NOS 

INFLAMMATION, NOS 

INFLAMMATION, IKTEFSTITIAL 

INFLAMMATION, SUPPURATIVE 

PERIVASCULITIS 

GLOMERULCSCLEFOSIS, NOS 

(47) 

6 (13%) 

1 (2%) 

23 (49%) 

(49) 

2 (4%) 

16 (33%) 

(48) 

1 (2%) 

15 (31%) 

1 (2%) 

1 (2%) 

1 (2%) 

(49) 

1 (2%) 

1 (2%) 

tKIDNEY/GLOMERULUS 

AMYLOIDOSIS 

(47) (49) (48) 

1 (2%) 

(49) 

#KIDNEY/TUbULE 

NECROSIS, FOCAL 

(47) 

1 (2?) 

(49) (48) (49) 

#KICN£Y/PELVIS 

INFLAMMATION,

INFLAMMATION,

 NOS 

 ACUTL _ _ 

(47) (49) (48) 

1 (2%) 

1(2% ) 

(49) 

# NUMEER OF ANIMALS WITH TISSUE EXAMINED MICEGSCOPICALLΥ 

• NUMEER OF ANIMALS NECROPSIED 

D-7
 
TABLE Dl (CONTINUED)
 

LOW DOSE CONTROL (UNTR) 05-0037 HIGH DOSE CONTROL (UNTR) 05-0118 LOH DOSE 05-0048 HIGH DOSE 05-0108 

INFLAMMATION, ACUTF/CHRONIC 11 (23%) 

•URINARY BLADDER 

INFLAMMATION, NOS HYPERPLASIA, EPIThtLIAL 

(48) 

4 (8%) 9 (19%) 

(4 8) 

4 (8%) 

(48) (47) 

ENCCCRINE SYSTEM 

•PITUITARY 

HYPERPLASIA, NOS 

HYPERPLASIA, FOCAL 

(<*2) 

3 P%) 

3 (7%) 

(40) (40) (43) 

•ADRENAL 

AMYLOIDOSIS 

HYPERPLASIA, NOS 

(45) 

3 (7%) 

(47) 

1 (2%) 

(40) 

#ADRENAL/CAPSULE 

HYPERPLASIA, NOS 

(«•5) (44) 

3 (7%) 

(47) (40) 

•ADRENAL CORTEX 

NODULE 

HYPERTROPHY, FOCAL 

HYPERPLASIA, NOS 

<*5) 

1 (2%) 

1 (2*) 1 (2*) 

(44) C»7) 

1 (2%) 

1 (2%) 

(40) 

•ADRENAL MEDULLA 

DEGENERATION, NOS 

(45) 

1 (2%) 

(44) (47) (40) 

•THYROID 

LYMPHOCYTIC INFLAMFATOR\ INFILTR HYPERPLASIA, NOS 

HYPERPLASIA, PAPILIARY 

HYPERPLASIA, FOLLICULAR-CELL 

(47) 

1 (2J) 

1 (2%) 

1 (2%) 

(4 5) (46) 

1 (2%) 

(45) 

12 (27%) 

•THYROID FOLLICLE 

DEGENERATION, NOS 

(47) (4 5) (U6) (45) 

44 (98%) 

•PARATHYROID 

HYPERPLASIA, NOS 

HYPERPLASIA, FOCAL 

(29) (24) (26) (22) 

1 (5%) 1 (5%) 

•PANCREATIC ISLETS 

HYPERPLASIA, NOS 

(48) 

2 (4%) 

(4 7) (44) (47) 

REPBOEUCTIVI SYSTEM 

•PREPUTIAL GLAND 

ABSCESS . NOS 

(48) 

2 (U%) 

(49) 

1 12%) 

(49) (*9) 

• NUMEER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY 

* NUMEER OF ANIMALS NECROPSILD 

D-8
 
TABLE Dl (CONTINUED) 

•PBCSTATE
 

INFLAMMATION, FOCAL
 

INFLAMMATION, ACUTE
 

HYPERPLASIA, PAPILLARY
 

#TESIIS/TUBULE
 

DEGENERATION, NOS
 

CALCIFICATION, FOCAL
 

•EPIDIDYMIS
 

INFLAMMATION, NOS
 

NERVOUS SYSTEM
 

tERAIN/KENINGES
 

INFLAMMATION, ACUTE/CHRONIC
 

«BRAIN
 

INFLAMMAIION, ACUTE/CHRONIC
 

•CEREBRAL CORTEX
 

MINERALIZATION
 

SPECIAL SENSE ORGANS
 

NONE
 

MUSCULOSKELETAL SYSTEM
 

•SKELETAL	 MUSCLE
 

THROMBOSIS, NOS
 

INFLAMMATION, ACUTK FOCAL
 

BOEY CAVITIES
 

•ABDOMINAL CAVITY
 

NECROSIS, FAT
 

•PLEURA
 INFLAMMATION PROLIFERAT E
 

LOW DOSE
 

CONTROL(UNTR)
 

05-0037
 

(48)
 

(47)
 

4 (9%)
 

(48)
 

(48)
 

(48)
 

(48)
 

3 (6%)
 

(48)
 

(48)
 

(48)
 

HIGH DOSE
 

CONTROL (UNTR)
 

05-0118
 

(49)
 

(48)
 

(49)
 

1 (2%)
 

(49)
 

(49)
 

(49)
 

(4 9)
 

(49)
 

(49)
 

LOW DOSE
 

05-0048
 

(48)
 

1 (2%)
 

1 (2%)
 

1 (2%)
 

(46)
 

7 (15%)
 

(49)
 

(48)
 

("8)
 

(48)
 

(49)
 

(49)
 

(49)
 

HIGH DOSE
 

05-0108
 

(43)
 

(48)
 

1 (2%)
 

(49)
 

(47)
 

1 (2%)
 

(47)
 

1 (2%)
 

(47)
 

(49)
 

1 (2%)
 1 (2%)
 

(49)
 

1 (2X)
 

(49)
 

1_I2%1 .
 

# NUKEER OF ANIMALS WITL TISSUE EXAMINED MICROSCOPICALLY
 

• NUMEER OF ANIMALS KLCROPSILD
 

D-9
 
TABLE Dl (CONCLUDED)
 

LOW DOSE HIGH DOSE
 

CONTROL (UNTR) CONTROL (UNTR)
 

05-0037 05-0118
 

•KESENTEBY	 (48) (49)
 

FERIAFTERITIS
 

NECROSIS, FOCAL
 

ALL CTHEB JYSTEMS
 

ADIPOSE TISSUE
 

INFLAMMATION, HLlTil 1
 

OMENTUM
 

NECROSIS, FAT 1
 

SPECIAL MCFTHOLOGY SUMMAnY
 

NC LESION REPOPTIT 5
 

ANIMAL KISSING/NO NECROPSY 1
 

AUTOLYSIS/NO NSCPOPSY 2
 

# NUMEER C¥ ANIMALS WITH IISSI'E EXAMINED MICROSCOPICALLY
 

• NUMEER OF ANIMALS KECPOPSTED
 

LOW DOSE
 

05-0048
 

(49)
 

5
 

1
 

HIGH DOSE
 

05-0108
 

(49)
 

1 (2X)
 1 (2%)
 

1
 

1
 

D-IO
 
TABLE D2

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN FEMALE MICE 

TREATED WITH 2,4-DIAMINOANISOLE SULFATE 

LOW DOSE HIGH DOSE 

CONTROL (UNTR) 

06-0037 

CONTROL (UNTR) 

06-0118 

ANIMALS INITIALLY I

AKIMALS KISSING 

ANIMALS NECECFSIED 

ANIMALS EXAMINED

N STUDY 

riISTOPATHOLOGICALLY 

50 

48 

50 

50 

50 

INTEGUMENTARY SYSTuM
 

*SK1N (48) (50)
 

FFIDERMAL INCLUSION CYST
 

•SUFCUT	 TISSUE (48) (50)
 

MINERALIZATION 1 (2%)
 

ABSCESS, NOS 1 (2%)
 

FIBROSIS 1 {2%)
 

BESFIFATORY SYSTEM
 

#LUNG/ERONCHUS (46) (50)
 

INFLAMMATION, iOCAL 1 (27) 1 (2%)
 

#LUNG/ERONCHIOLE (46) (50)
 

HYPERPLASIA, NOS 1 (2%)
 

#LUNG (46) (50)
 

INFLAMMATION, INTERSTITIAL 10 (22%) 14 (28%)
 

HYPERPLASIA, EPITHELIAL 3 (7%)
 

HYPERPLASIA, ALVEOLAP EPITHELIUM
 

HEMATOPOIETIC SYSTEM
 

#BCKE MARROW (45) (49)
 

HYPOPLASIA, NOS
 

HYPERPLASIA, DIFFUSE
 

MYELOFIBROSIS 1 (2%)
 

MYELOSCLEROSIS
 

HYPERPLASIA, HEMATOPOIETIC
 

tSPLEEN (46) (49)
 

ATROPHY^ NOS
 

# NUMEER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY 

* NUMBER OF ANIMALS NECSOPSIED 

**EXCLUDES PARTIALLY AUTOLYZED ANIMALS 

LOW DOSE 

06-0048 

50 

1 

45 

44 

HIGH DOSE 

06-0108 

50 

50 

50 

(45) 

(45) 

(50) 

1 {2%) 

(50) 

(43) 

(43) 

(43) 

1 (2%) 

1 (2%) 

(50) 

(50) 

(50) 

(41) 

1 (2%) 

(44) 

1 12%) _ 

(*8) 

1 (2%) 

2 (4%) 

29 (60%) 

11 (23%) 

38 (79%) 

(50) 

1(2% ) 

D-ll
 
TABLE D2 (CONTINUED) 

LOW DOSE 

CONTROL(UNTR) 

06-0037 

HIGH DOSE 

CONTROL (UNTR) 

06-0118 

hYPEFPLASIA, NOS 

ANGIECTASIS 

HYPERPLASIA, ΗΓΚΑΤΟΡΟΙΕΤΙΟ 

HYPERPLASIA, EFYTHPOID 

HYPERPLASIA, LYMPHOID 

HEMATOPOIESIS 

fYELOPCIESIS 

16 (35%) 

6 (13%) 

10 (22%) 

1 (2%) 

1 (2%) 

9 (18%) 

6 (12%) 

2 (4%) 

•HEMCLYMPH NODES 

INFLAMMATION, NOS 

HYPERPLASIA, NOS 

(46) (49) 

2 (4%) 

1 (2%) 

•LYMPH NOEE 

CYST, NOS 

INFLAMMATION, NGS 

HYPERPLASIA, NOS 

fcETICULOCYTOSIS 

HYPERPLASIA, HEMATOPOIETIC 

HYPERPLASIA, LYMPHOID 

MYELOID METAPLASIA 

(39) 

1 0*) 15 (38%) 

1 (37) 

1 (3%) 

2 (5%) 

1 (3%) 

(44) 

9 (20%) 

3 (7%) 

1 (2%) 

1 (2%) 

4 (9%) 

«MEDIASTINAL L.NODE 

HYPERPLASIA, LYMPHOID 

(39) (44) 

•MESENTERIC L. NODE 

INFLAMMATION, ACUTE/CHRONIC 

HYPERPLASIA, RETICULUM CELL 

(39) (44) 

•THYMUS 

INFLAMMATION, NOS 

GRANULOMA, NOS 

HYPERPLASIA, LYMPHOID 

(3D (21) 

•THYMIC MEDULLA 

HYPERPLASIA, NOS 

HYPERPLASIA, LYMPHOID 

(31) (21) 

CIPCULATCRY SYSTEM
 

•HEART	 (46) (50)
 

PERIARTERITIS
 

•HEART/VENTSICLE	 (46) (50)
 

MELANIN 4 (9%)
 

• NUMbER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMEER OF ANIMALS NECROPSIED
 

LOW DOSE
 

06-0048
 

3 (7%)
 

(-4 4)
 

(38)
 

(38)
 

1 (3%)
 

(38)
 

2 (5%)
 

2 (5%)
 

(36)
 

1 (3%)
 

(36)
 

2 (6%)
 

(**3)
 

0*3)
 

HIGH DOSE
 

06-0108
 

1 (2%)
 

2 (4%)
 

(50)
 

(43)
 

(43)
 

(43)
 

(32)
 

1 (3%)
 

1 (3%)
 

1 (3%)
 

(32)
 

1 (3%)
 

(50)
 

2 (U%)
 

(50)
 

D-I2
 
TABLE D2 (CONTINUED)
 

• NUMEER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMEER OF ANIMALS NECROPSIED
 

D-I3
 
TABLE D2 (CONTINUED)
 

LOW DOSE HIGH DOSE 

CONTROL(UNTR) CONTROL (UNTR) 

06-0037 06-0118 

HYPERPLASIA, NOS 1 (2%) HYPERPLASIA, EPITHELIAL 1 (2%) HYPERPLASIA, ADENOMATOUS 1 (2%) HYPERKERATOSIS 

ACANTHOSIS 

1 

1 (2T)

(2%) 2 (4%) 

#GASTRIC MUCOSA (44) (49) 

HYPERPLASIA, FOCAL 1 (2%) 

•PEYERS PAICH (44) (48) 

HYPERPLASIA, NOS 1 (2*) 7 (15%) 

URINARY SYSTEM
 

•KIDNEY	 (46) (50)
 

HYDRONEPHROSIS
 

GLOMERULONEPHRITIS, NOS 14 (30%) 4 (8%)
 

GLOMERULONEPHRITIS, FOCAL 1 (2%)
 

INFLAMMATION, INTERSTITIAL 16 (35%) 12 (24%)
 

PERIARTERITIS
 

GLOMERULOSCLEROSIS, NOS
 

#KIDNEY/TUEULE (46) (50)
 

MINERALIZATION 1 (2%)
 

•KIDNEY/PELVIS	 (46) (50)
 

INFLAMMATION, ACUTT/CHRONIC
 

«URINARY BLADDER (46) (4 8)
 

INFLAMMATION, NOS 4 (9%)
 HYPERPLASIA, iPITHFLIAL 10 (22%) 1 (2%)
 

ENEOCRINE SYSTEM
 

•PITUITARY	 (42) (4 2)
 

HYPERPLASIA, FOCAL 6 (14%)
 

#ADRENAL/CAPSULE (45) (48)
 

HYPERPLASIA, NOS 5 (10%)
 

•ADRENAL CORTEX	 (45) (48)
 

NODULE 3 (7%) 1 (2%)
 

NECPOSISX NOS
 

• NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMALS NECROPSILD
 

LOW DOSE
 06-0048
 

1 (3%)
 1 (3%)
 

(40)
 

(39)
 

(44)
 

1 (2%)
 

2 (5%)
 

(44)
 

(44)
 

10 (23%)
 

(41)
 

1 (2%)
 

(38)
 

1 (3%)
 

(41)
 

(41)
 

2 (5%)
 

HIGH DOSE
 06-0108
 

(49)
 

(49)
 

(50)
 

4 (8%)
 

2 (4%)
 2 (4%)
 

(50)
 

(50)
 

(48)
 

(34)
 

(49)
 

(49)
 

1-12*1 .
 

D-14
 
TABLE D2 (CONTINUED) 

LOW DOSE 

CONTROL(UNTR) 

06-0037 

HIGH DOSE 

CONTROL (UNTP) 

06-0118 

LOW DOSE 

06-0048 

HIGH DOSE 

06-0108 

FYPEFPLAbIA,

HYPERPLASIA,

HYPEFFLASIA,

 NODULAR 

 NCS 

 DIFFUSE 

1 (2%) 

1 

2 

1 

(2%) 

<5X) 

(2%) 

#THYROID 

FOLLICULAR CYST, NOS 

INFLAMMATION, NOS 

INFLAMMATION, FOCAL 

PEEIAPTEHITIS 

HYPERPLASIA, NOS 

HYPEFPLASIA, PAPILLAFY 

hYPEPPLASIA, ADENOMATOUS 

HYPERPLASIA, FOLLICULAP-CELL 

(43) 

1 

1 

{2%) 

(2%) 

(44) 

1 

2 

1 

(2%) 

(5%) 

(2%) 

(42) 

1 

7 

3 

(2%) 

(17%) 

(7%) 

(45) 

1 (2%) 

1 (2%) 

#THYEOID FCLLICLZ 

LiGEKERATION, >'OS 

hYPEFTRCtHi, NUS 

(43) (44) (42) 

1 (2%) 

(45) 

44 (98%) 

REEECEUCTIVJ : SYSTEf 

•MAMMARY GLAND 

GALACTOCELE 

HYPERPLASIA, NOS 

(48) 

1 

4 

(2%) 

(8X) 

(50) 

1 {2%) 

(45) (50) 

#UTERUS 

FYDROMETRA 

FYOMETRA 

ABSCESS, NOS 

INFLAMMATION, CHFONIC 

FIBROSIS 

METAPLASIA, SQUAMOUS 

(45) 

1 

3 

1 

(2%) 

(2%) 

(47) 

13 (28%) 

(41) 

2 

1 

2 

1 

2 

(5%) 

(2%) 

(5%) 

(2%) 

(5%) 

(46) 

9 (20%) 

•UTEBUS/ENDGMETRIUM 

HEMORRHAGIC CYST 

INFLAMMATION, NOS 

INFLAMMATION, SUFPUPATIVF 

INFLAMMATION, ACUTE 

ABSCESS, NOS 

INFLAMMATION, ACUTE/ChHONIC 

HYPEPPLASIA, NOS 

HYPERPLASIA, PAPILLARY 

HYPERPLASIA, CYSTIC 

HYPERPLASIA, ADENOMATOUS 

HYPERPLASIA, STROM?! __ 

(45) 

10 

4 

4 

18 

1 

(22%) 

(9*) 

(9%) 

(40%) 

(2%) 

(47) 

8 

8 

6 

(17%) 

(17%) 

(13%) 

(41) 

2 

2 

1 

1 

1 

29 

(5%) 

(5%) 

(2%) 

(2%) 

(2%) 

(71%) 

(46) 

1 (2%) 

2 (4%) 

1 (2%) 

21 (46%) 

2_IMI 

# NUhBER CF ANIMALS

• NUHEER OF ANIMALS

 WITH TISSUE EXAMINED

 NECROPSIED 

 MICROSCOPICALLY 

D-15
 
LOW DOSE 

CONTROL(UNTR) 

06-0037 

HIGH DOSE 

CONTROL (UNTR) 

06-0118 

LOW DOSE 

06-0048 HIGH DOSE 06-0108 

tlKTAPLASIA, SQUiMGUS 1 (2%) 

#CVARY/OVIDUCT 

INFLAMMATION, hOS 

INFLAMMATION, SUPPURATIVE 

AESCESS, NOS 

TNFLAMMATION, CflPOMC 

(45) 

5 (11%) 

(47) 

4 (9%) 

1 (2%) 

(41) 

3 (7%) 

1 (2%) 

(46) 

1 (2%) 

tOVARY 

CYST, NCS 

INFLAMMATION, NOS 

LYMPHOCYTIC IN^LAMr ΑΤΟ*3

 Υ INFILTP 

INFLAMMATION, SUPPURATIVE 

AESCESS, NOS INFLAMMATION, CFRONIC 

SCLEFOSIS 

PERIARTERITIS DEGENERATION, CYSTIC 

NECROSIS, COArULPTIVE 

ATROPHY, NOS 

(45) 

3 (7Jt) 

4 (9%) 

1 (2*) 

10 (22%) 

4 (9X) 

1 (2%) 

(48) 

10 (21%) 

4 (8%) 

1 (2%) 

3 (6%) 

(40) 

6 (15%) 

1 (3%) 

2 (5%) 

6 (15%) 

1 (3%) 

(*3) 

2 (5%) 

1 (2%) 

1 (2%) 

NEBVCUS SYSTEM 

«ERAIN/MENINGES 

INFLAMMATION, ACUTF/CHRONIC 

PERIARTERITIS PERIVASCULITIS 

(44) (48) (41) (49) 

2 (4%) 

1 (2%) 

1 (2%) 

SPECIAL SENSE ORGANS 

NONE 

MUSCUICSKEIETAL SYSTiP 

•BONE 

RESORPTION 

(48) 

3 (6f) 

(50) («5) (50) 

BOEY CAVITIES 

•ABDOMINAL CAVITY 

STFATITIS 

(48) (50) (45) 

1 (2%) 

(50) 

# NUMEER OF ANIMALb WITH TISSUE EXAMINED

• NUMEEP OF ANIMALS NECROPSIED 

 MICROSCOPICALLY 

D-16
 
TABLE D2 (CONCLUDED)
 

LOW DOSE HIGH DOSE
 

CONTROL (UKTR) CONTROL (UNTR)
 

06-0037 06-0118
 

NECROSIS, FAT
 

ALL OTHER SYSTEMS
 

OMFNTUM
 

NECROSIS, FAT
 

SPECIAL MORPHOLOGY SUMMARY
 

NO LESION REFOPTED
 

ANIMAL MISSING/NO NECROPSY
 

AUTO/NECROPSY/HISTO PEFF
 

AUTO/NECFOPSY/NO HISTO
 

AUTOLYSIS/NO NECROPSY
 

# NUMBER OF ANIMALS WITΗ TISSUE EXAMINED


* NUMBER OF ANIMALS NFCROPSILL
 

1
 

1 3
 

2 1
 

1
 

2
 

 MICROSCOPICALLY
 

LOW DOSE HIGH DOSE 

06-0048 06-0108 

2 (U*) 

1 

1 

1 

4 

D-17
 


DHEW Publication No. (NIH) 78-1334