National Cancer Institute 

CARCINOGENESIS 

Technical Report Series 

No. 54 

1978 

BIOASSAY OF 

2,4-DINITROTOLUENE 

FOR POSSIBLE CARCINOGENICITY 

CAS No. 121-14-2 

NCI-CG-TR-54 

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

Public Health Service 

National Institutes of Health 

BIOASSAY OF
 

2 ,4-DINITROTOLUENE
 

FOR POSSIBLE CARCINOGENICITY
 

Carcinogenesis Testing Program
 

Division of Cancer Cause and Prevention
 

National Cancer Institute
 

National Institutes of Health
 

Bethesda, Maryland
 

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

Public Health Service
 

National Institutes of Health
 

DHEW Publication No. (NIH) 78-1360
 

REPORT ON THE BIOASSAY OF 2,4-DINITROTOLUENE
 

FOR POSSIBLE CARCINOGENICITY
 

CARCINOGENESIS TESTING PROGRAM
 

DIVISION OF CANCER CAUSE AND PREVENTION
 

NATIONAL CANCER INSTITUTE, NATIONAL INSTITUTES OF HEALTH
 

CONTRIBUTORS: This report presents the results of the bioassay of
 

2,4-dinitrotoluene conducted for the Carcinogenesis Testing Program,
 

Division of Cancer Cause and Prevention, National Cancer Institute
 

(NCI), National Institutes of Health, Bethesda, Maryland. This bioassay was conducted by Mason Research Institute, Worcester, Massachusetts, initially under direct contract to the NCI and currently under
 

a subcontract to Tracer Jitco, Inc., prime contractor for the NCI
 

Carcinogenesis Bioassay Program.
 

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

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

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

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

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

Histopathologic examinations were performed by Dr. R. W. Fleischman (3), Dr. D. W. Hayden (3), Dr. A. S. Krishna Murthy (3), Dr. A.
 

Russfield (3), and Dr. D. S. Wyand (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 (6).
 

Compilation of individual animal survival, pathology, and
 

summary tables was performed by EG&G Mason Research Institute (7);
 

the statistical analysis was performed by Mr. W. W. Belew (5) and
 

Dr. A. Chu (7), using methods selected for the Bioassay Program by
 

Dr. J. J. Gart (8).
 

This report was prepared at METREK, a Division of The MITRE
 

Corporation (5) under the direction of the NCI. Those responsible
 

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

Thomas (5), the task leader, Dr. M. R. Kornreich (5), the senior
 

biologist, Ms. P. Walker (5), and the technical editor, Ms. P. A.
 

Miller (5). The final report was reviewed by members of the participating organizations.
 

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

Dr. J. J. Gart (8), Mr. J. Nam (8), Dr. H. M. Pettigrew (8), and
 

Dr. R. E. Tarone (8).
 

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 (1,9), and Dr. J. M. Ward (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.	 Midwest Research Institute, 425 Volker Boulevard, Kansas City,
 

Missouri.
 

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

Boulevard, McLean, Virginia.
 

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

Maryland.
 

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

Rockville, Maryland.
 

8.	 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.
 

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

University, School of Medicine, Traylor Building, Baltimore,
 

Maryland.
 
SUMMARY
 

A bioassay of practical-grade 2,4-dinitrotoluene for possible
 

carcinogenicity was conducted using Fischer 344 rats and B6C3F1 mice.
 

2,4-Dinitrotoluene was administered in the feed, at either of two
 

concentrations, to groups of 50 male and 50 female animals of each
 

species. For male and female rats, the high and low time-weighted
 

average dietary concentrations of 2,4-dinitrotoluene were 0.02 and
 

0.008 percent, respectively. For male and female mice, the high and
 

low time-weighted average concentrations were 0.04 and 0.008 percent,
 

respectively. After a 78-week period of compound administration,
 

observation of the rats continued for an additional 26 weeks and
 

observation of the mice continued for 13 additional weeks.
 

For the chronic rat bioassay, 25 rats of each sex were placed on
 

test as high dose controls, and 50 rats of each sex served as the low
 

dose controls. For the mice, 50 males and 50 females were placed on
 

test as controls for each of the high dose and low dose groups.
 

In both species the survival in all groups was adequate for
 

statistical analysis of late-appearing tumors.
 

In the male rats, a significantly increased incidence of fibroma
 

of the skin and subcutaneous tissue occurred in both the high and the
 

low dose groups when compared to their respective controls. A statistically significant incidence of fibroadenoma of the mammary gland
 

occurred in the high dose female rats.
 

Among the mice a variety of tumors was observed but none were
 

considered to be associated with the dietary administration of 2,4­

dinitrotoluene.
 

Under the conditions of this bioassay dietary administration
 

of 2,4-dinitrotoluene to Fischer 344 rats induced benign tumors
 

(i.e., fibroma of the skin and subcutaneous tissue in males and
 

fibroadenoma of the mammary gland in females). No evidence was
 

provided for the carcinogenicity of the compound in B6C3F1 mice of
 

either sex.
 

TABLE OF CONTENTS
 

Page
 

1. INTRODUCTION 1
 

II. MATERIALS AND METHODS 3
 

A. Chemicals 3
 

B. Dietary Preparation 3
 

C. Animals *
 

I). Animal Maintenance /*
 

E. Selection of Initial Concentrations 9­

F. Experimental Design 9
 

G. Clinical and Histopathologic Examinations 12
 

H. Data Recording and Statistical Analyses 14
 

III. CHRONIC TESTING RESULTS: RATS 19
 

A. Body Weights and Clinical Observations 19
 

B. Survival 21
 

C. Pathology 21
 

D. Statistical Analysis of Results 24
 

IV. CHRONIC TESTING RESULTS: MICE 34
 

A. Body Weights and Clinical Observations 34
 

B. Survival 34
 

C. Pathology 37
 

D. Statistical Analyses of Results 37
 

V. DISCUSSION 43
 

VI. BIBLIOGRAPHY 45
 

APPENDIX A SUMMARY OF THE INCIDENCE OF NEOPLASMS IN RATS
 

TREATED WITH 2,4-DINITROTOLUENE A-l
 

APPENDIX B SUMMARY OF THE INCIDENCE OF NEOPLASMS IN MICE
 

TREATED WITH 2,4-DINITROTOLUENE B-l
 

APPENDIX C SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC
 

LESIONS IN RATS TREATED WITH 2,4-DINITROTOLUENE C-l
 

APPENDIX D SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC
 

LESIONS IN MICE TREATED WITH 2,4-DINITROTOLUENE D-l
 

vii
 
LIST OF ILLUSTRATIONS
 

Figure Number Page 

1 'GROWTH CURVES FOR 2,4-DINITROTOLUENE 

CHRONIC STUDY RATS 20 

SURVIVAL COMPARISONS OF 2,4-DINITROTOLUENE CHRONIC STUDY RATS 22 

GROWTH CURVES FOR 2,4-DINITROTOLUENE 

CHRONIC STUDY MICE 35 

SURVIVAL COMPARISONS OF 2,4-DINITROTOLUENE CHRONIC STUDY MICE 36 

LIST OF TABLES 

Table Number Page 

DESIGN SUMMARY FOR FISCHER 344 RATS— 

2,4-DINITROTOLUENE FEEDING EXPERIMENT 10 

DESIGN SUMMARY FOR B6C3F1 MICE—2 ,4-

DINITROTOLUENE FEEDING EXPERIMENT 11 

ANALYSES OF THE INCIDENCE OF PRIMARY 

TUMORS AT SPECIFIC SITES IN MALE RATS 

TREATED WITH 2,4-DINITROTOLUENE 25 

ANALYSES OF THE INCIDENCE OF PRIMARY 

TUMORS AT SPECIFIC SITES IN FEMALE RATS 

TREATED WITH 2,4-DINITROTOLUENE 29 

ANALYSES OF THE INCIDENCE OF PRIMARY 

TUMORS AT SPECIFIC SITES IN MALE MICE 

TREATED WITH 2,4-DINITROTOLUENE 38 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS 

AT SPECIFIC SITES IN FEMALE MICE TREATED 

WITH 2,4-DINITROTOLUENE 40 

viii
 
LIST OF TABLES (Concluded) 

Table Number Page 

Al SUMMARY OF THE INCIDENCE OF NEOPLASMS 

IN MALE RATS TREATED WITH 2,4-DINITROTOLUENE A^-3 

A2 SUMMARY OF THE INCIDENCE OF NEOPLASMS 

IN FEMALE RATS TREATED WITH 2,4-DINITROTOLUENE A1

"8 

Bl SUMMARY OF THE INCIDENCE OF NEOPLASMS IN 

MALE MICE TREATED WITH 2,4-DINITROTOLUENE B ~ 3 

B2 SUMMARY OF THE INCIDENCE OF NEOPLASMS IN 

FEMALE MICE TREATED WITH 2,4-DINITROTOLUENE B-6 

Cl SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC 

LESIONS IN MALE RATS TREATED WITH 2,4-

DINITROTOLUENE C-3 

C2 SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC 

LESIONS IN FEMALE RATS TREATED WITH 2,4-

DINITROTOLUENE C^12 

Dl SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC 

LESIONS IN MALE MICE TREATED WITH 2,4-

DINITROTOLUENE D-3 

D2 SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC 

LESIONS IN FEMALE MICE TREATED WITH 2,4-

DINITROTOLUENE D-8 

IX
 

I. INTRODUCTION
 

2,4-Dinitrotoluene (NCI No. C01945), a precursor in the synthesis
 

of azo dyes, was selected for bioassay by the National Cancer Institute along with other dye intermediates in an attempt to elucidate
 

those chemicals which may be responsible for the increased incidence
 

of bladder cancer observed among workers in the dye manufacturing
 

industry (Wynder et al., 1963; Anthony and Thomas, 1970). Aromatic
 

nitro compounds are one of several classes of chemicals thought to
 

contribute to the increased cancer risk in this industry (Wynder et
 

al., 1963). The structural relationship of 2,4-dinitrotoluene to the
 

known carcinogen 2,4-diaminotoluene was also a factor in its selection
 

for testing (Weisburger, 1976; Hiasa, 1970).
 

The Chemical Abstracts Service (CAS) Ninth Collective Index
 

*
 (1977) name for this compound is l-methyl-2,4-dinitrobenzene. It is
 

also called 2,4-dinitrotoluol and 2,4-DNT.
 

Precise production figures for 2,4-dinitrotoluene are not available; however, the U.S. International Trade Commission (1977) reported
 

a combined production of 272,610,000 pounds for the 2,4- and 2,6-dinitrotoluene isomers in 1975.
 

Aside from its use by the dye manufacturing industry, 2,4-dinitrotoluene is used by the munitions industry as a modifier for smokeless
 

powders and, to a limited extent, as a gelatinizing and waterproofing
 

*
 

The CAS registry number is 121-14-2.
 
agent in military and commercial explosive compositions (Institute of
 

Makers of Explosives, 1977). 2,4-Dinitrotoluene experiences widespread application as a chemical intermediate for the production of
 

toluene diisocyanate (TDI) which, in turn, is consumed in the production of flexible polyurethane foams; however, most TDI producers use
 

toluene as the starting material, generating 2,4-dinitrotoluene as a
 

captive intermediate in the process (Urso, 1977).
 

The risk of exposure to 2,4-dinitrotoluene is greatest for
 

workers in the dye and explosives industries and at chemical plants
 

producing TDI. The general population may also experience exposure
 

as a result of discharge of 2,4-dinitrotoluene into rivers and
 

streams from munitions plants (Simon et al., 1977).
 

2,4-Dinitrotoluene can cause anemia, methemoglobinemia, cyanosis,
 

and liver damage (Sax, 1975). It is rapidly absorbed through the
 

intact skin; however, toxic levels may also be reached by inhalation
 

or ingestion (Manufacturing Chemists Association, 1966). Studies by
 

Simon et al. (1977), suggest that 2,4-dinitrotoluene is not a potent
 

mutagen in the rat, as determined by dominant lethal mutation experiment s.
 
II. MATERIALS AND METHODS
 

A. Chemicals
 

Practical-grade 2,4-dinitrotoluene was purchased from J. T.
 

Baker Chemical Company. Analysis by the manufacturer suggested a
 

purity greater than 95 percent. The observed melting point (67° to
 

70°C) suggested a compound of relative purity due to the narrow range
 

and its close proximity to the literature value (71°C). Thin-layer
 

chromatography visualized with ultraviolet light indicated at least
 

one impurity. Ultraviolet analysis showed a peak at 250 nm which is
 

consistent with the value reported in the literature (252 nm).
 

Throughout this report the term 2,4-dinitrotoluene is used to
 

represent this practical-grade material.
 

B. Dietary Preparation
 

The basal laboratory diet for both treated and control animals
 

®
 

was Wayne Lab-Blox (Allied Mills, Inc.). 2,4-Dinitrotoluene was
 

weighed under an exhaust hood and ground in a mortar and pestle with
 

®
 

an aliquot of ground Wayne Lab-Blox meal. Once visual homogeneity
 

was attained, the mixture and the remainder of the feed to be administered to the treated animals were placed into a 6 kg capacity
 

Patterson-Kelly twin shell stainless steel V-blender. After blending
 

for 20 minutes, the mixture was sealed in double plastic bags and
 

stored in the dark at 4°C. Mixtures were used for only 1 week.
 
C. Animals
 

Two animal species, rats and mice, were used in the carcinogenicity bioassay. Fischer 344 rats and both C57BL/6CR and B6C3F1 mice
 

were obtained through contracts of the Division of Cancer Treatment,
 

National Cancer Institute. High and low dose rats and mice, and high
 

dose control rats were supplied by Charles River Breeding Laboratories, Wilmington, Massachusetts. Low dose control animals were
 

supplied by ARS/Sprague-Dawley, Madison, Wisconsin. The low dose
 

groups were received in shipments separate from their respective
 

controls. Upon arrival, a sample of animals was examined for parasites and other signs of disease. The rats and mice to be tested
 

were quarantined for 2 weeks prior to initiation of the bioassay.
 

Animals were assigned to groups and distributed among cages so that
 

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

and species.
 

D.	 Animal Maintenance
 

All animals were housed by species in rooms having a temperature
 

range of 23° to 34°C and a range in relative humidity of 10 to 85
 

®
 percent. Incoming air was filtered through Tri-Dek 15/40 denier
 

Dacroir filters providing six changes of room air per hour. Fluorescent lighting was provided on a 12-hour-daily cycle.
 

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

for the first 11 months of study, high dose rats and their controls
 
were maintained in galvanized- and stainless-steel wire-mesh cages
 

suspended above newspapers. Low dose and low dose control rats were
 

kept in these cages for the first 13 months of study. Newspapers
 

under the cages were replaced daily and cages and racks washed weekly.
 

For the remainder of the study, all rats were housed in suspended
 

polycarbonate cages equipped with disposable nonwoven fiber filter
 

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

dose rats and their controls were provided with hardwood chip bedding
 

®
 (Ab-sorb-dri , Wilner Wood Products Co.) for the first 9 months that
 

®
 they were housed in polycarbonate cages. SAN-I-CEL corncob bedding
 

(Paxton Processing Co.) was used for these animals for the next 12
 

months. High dose rats and their controls were provided with SAN-I­

®
 CEL for the first 12 months that they were housed in polycarbonate
 

cages. For the remainder of the study, Bed-o'Cobs (The Anderson's
 

Cob Division) was provided in all treated and control rat cages.
 

Stainless steel cage racks (Fenco Cage Products) 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 following chemical administration, stainless steel wire bar lids were
 

substituted. Both types of lids were supplied by Lab Products, Inc.
 

Nonwoven fiber filter bonnets were used over cage lids. All mice
 

were housed 10 per cage for the first part of the study. Cage
 
populations for high dose, high dose control, low dose, and low dose
 

control mice were reduced to five per cage after 12 months, 14 months,
 

19 months, and 19 months, respectively. Cages, lids, filters, and
 

bedding were provided three times per week when cage populations were
 

ten and twice per week when cage populations were reduced to five.
 

Bedding was of the same brands as those used for rats. Ab-sorb-dri
 

was used for 2 months (high dose), 4 months (high dose controls), and
 

9 months (low dose and low dose controls) prior to SAN-I-CEL being
 

used for 12 months. A second type of corncob bedding (Bed-o'Cobs )
 

was then used for the remainder of the bioassay. 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. Food and water were available ad libitum.
 

Pelleted Wayne Lab-Blox was supplied to all animals during the
 

quarantine and final observation periods. During the period of chemical administration, all treated animals received dosed Wayne Lab-


Blox . Control animals received untreated meal. Alpine aluminum
 

feed cups (Curtin Matheson Scientific, Inc.) containing stainless
 

steel baffles were used to distribute powdered feed to low dose rats,
 

their controls, and all mice during the entire study, and to high dose
 

rats and their controls for the first 13 months of chemical administration. High dose rats and their controls were fed from stainless
 
steel gangstyle feed hoppers (Scientific Cages, Inc.) during the last
 

5 months of chemical administration. During the observation period,
 

rats were fed pellets on the cage floor and mice were fed pellets
 

from a wire bar hopper incorporated into the cage lid. Food hoppers
 

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

daily in Alpine feed cups.
 

Low dose, low dose control, and high dose rats were housed in a
 

*
 

room in which other rats were receiving diets treated with acetylaminofluorene (53-96-3); dulcin (150-69-6) and L-arginine glutamate
 

(4320-30-3); sodium nitrite (7632-00-0); L-arginine glutamate (4320­

30-3); N-butyl urea (592-31-4); N,N-dimethyl-p-nitrosoaniline (138­

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

acid (619-17-0); 1,5-naphthalenediamine (2243-62-1); N-(l-naphthyl)
 

ethylenediamine dihydrochloride (1465-25-4); 2-chloro-p-phenylenediamine sulfate (61702-44-1); aniline hydrochloride (142-04-1); and
 

p-anisidine hydrochloride (20265-97-8). High dose control rats
 

shared a room with other rats receiving diets treated with 5-nitro-otoluidine (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); 2,4-diaminoanisole
 

sulfate (615-05-4); and APC (8003-03-0).
 

*
 

CAS registry numbers are given in parentheses,
 
Low dose, low dose control, and high dose mice were housed in a
 

room in which other mice were receiving diets treated with: 2,5­

toluenediamine sulfate (6369-59-1); 2-aminoanthraquinone (117-79-3);
 

3-amino-4-ethoxyacetanilide (17026-81-2); 3-amino-9-ethylcarbazole
 

hydrochloride; l-amino-2-methylanthraquinone (82-28-0); 5-nitro-oanisidine (99-59-2); 1-nitronaphthalene (86-57-7); 4-nitroanthranilic
 

acid (619-17-0); 5-nitroacenaphthene (602-87-9); 2,4-diaminoanisole
 

sulfate (615-05-4); 3-nitro-p-acetophenetide (1777-84-0); and N,Ndimethyl-p-nitrosoaniline (138-89-6). High dose control mice shared
 

a room with other mice receiving diets treated with 2-methyl-l-nitroanthraquinone (129-15-7); p-cresidine (120-71-8); fenaminosulf
 

(140-56-7); 4-chloro-m-phenylenediamine (5131-60-2); and cinnamyl
 

anthranilate (87-29-6).
 

E. Selection of Initial Concentrations
 

Six-week subchronic toxicity studies were conducted with Fischer
 

344 rats and C57BL/6CR mice in order to determine the high concentrations for administration during the chronic bioassay. Animals of
 

each species were distributed among four groups, each consisting of
 

five males and five females. 2,4-Dinitrotoluene was administered in
 

the feed for 4 weeks. Three animal groups of each species received
 

dietary concentrations of 0.00375, 0.0075, and 0.015 percent. A
 

fourth group of each species served as a control, receiving only the
 

basal laboratory diet. No deaths occurred at any dose level tested.
 
All animals were sacrificed at the end of the test and gross necropsies were performed.
 

A dosage inducing no mortality and resulting in a depression in
 

mean group body weight of approximately 15 percent relative to controls was to be selected as the initial high dose. When weight gain
 

criteria were not applicable, mortality data alone were utilized.
 

The initial high dose selected for administration to rats and
 

mice in the chronic study was 0.008 percent. However, for the reasons indicated below, the initial high doses utilized for rats and
 

mice in the chronic study were 0.02 and 0.04 percent, respectively.
 

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 the time-weighted
 

average concentrations) are summarized in Tables 1 and 2.
 

The treated and control rats were all approximately 6 weeks old
 

at the time they were placed on test. The initial dietary concentrations of 2,4-dinitrotoluene administered to rats were 0.0075 and
 

0.00375 percent. After week 19, the higher dose was changed from
 

0.0075 to 0.008 percent to facilitate dose formulation. The rat
 

group receiving 0.00375 percent was sacrificed after 51 weeks and no
 

histopathologic examinations were performed because the dose levels
 

being used in the chronic bioassay were considered, on the basis of
 

weight depression, to have been too low. A new group, receiving
 
TABLE 1
 

DESIGN SUMMARY FOR FISCHER 344 RATS
 

2,4-DINITROTOLUENE FEEDING EXPERIMENT
 

INITIAL 2,4-DINITRO- OBSERVATION PERIOD TIME-WEIGHTED
 

GROUP TOLUENE TREATED UNTREATED AVERAGE
 

SIZE (PERCENT) (WEEKS) (WEEKS) CONCENTRATION3
 

MALE
 

LOW DOSE
 

CONTROL 50 0 0 104 0
 

HIGH DOSE
 

CONTROL 25 0 0 104 0
 

LOW DOSE 50	 0.0075 19 0.008
 

0.008 59
 

0 26
 

HIGH DOSE 50	 0.02 78 0.02
 

0 26
 

FEMALE
 

LOW DOSE
 

CONTROL 50 0 0 104 0
 

HIGH DOSE
 

CONTROL 25 0 0 104 0
 

LOW DOSE 50	 0.0075 19 0.008
 

0.008 59
 

0 26
 

HIGH DOSE 50 0.02 78 0.02
 

26
 

a	 _	 —
 2 (concentration x weeks received)
 Timp— UPI crVifpH avpra OP crtnr pnfra fi m
 receiving chemical)
 

10
 
TABLE 2
 

DESIGN SUMMARY FOR B6C3F1 MICE
 

2,4-DINITROTOLUENE FEEDING EXPERIMENT
 

MALE 

LOW DOSE CONTROL 

INITIAL 

GROUP 

SIZE 

50 

2,4-DINITROTOLUENE 

(PERCENT) 

0 

OBSERVATION PERIOD 

TREATED 

(WEEKS) 

0 

UNTREATED 

(WEEKS) 

91 

HIGH DOSE CONTROL 50 0 0 91 

LOW DOSE 50 0.008 

0 

78 

13 

HIGH DOSE 50 0.04 

0 

78 

13 

FEMALE 

LOW DOSE CONTROL 50 0 0 91 

HIGH DOSE CONTROL 50 0 0 91 

LOW DOSE 50 0.008 

0 

78 

13 

HIGH DOSE 50 0.04 

0 

78 

13 

11
 
0.02 percent, was started with a new control. Throughout this report
 

the groups initially receiving a concentration of 0.0075 percent and
 

their controls are referred to as the low dose and low dose control
 

groups, respectively, while tiie groups receiving 0.02 percent and
 

their controls are referred to as the high dose and high dose control
 

groups, respectively. These treated rats were supplied with dosed
 

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

The treated and control mice were all approximately 6 weeks old
 

at the time they were placed on test. The initial dietary concentrations of 2,4-dinitrotoluene administered to mice were 0.008 and
 

0.00375 percent. The mouse groups receiving 0.00375 percent were
 

sacrificed after 29 weeks because the groups receiving 0.008 percent
 

did not demonstrate desired weight depression. A new group, receiving
 

0.04 percent, was started with a new control group. Throughout this
 

report the groups initially receiving a concentration of 0.008 percent and their controls are referred to as the low dose and low dose
 

control groups, respectively, while the groups receiving 0.04 percent
 

and their controls are referred to as the high dose and high dose
 

control groups, respectively. Treated mice were supplied with
 

dosed feed for a total of 78 weeks, followed by a 13-week observation
 

period.
 

G. Clinical and Histopathologic Examinations
 

Animals were weighed immediately prior to initiation of the
 

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

12
 
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 and bile
 

duct (mice), pancreas, esophagus, stomach, small intestine, large
 

intestine, kidney, urinary bladder, pituitary, adrenal, thyroid,
 

parathyroid, test is, prostate, brain, ear, uterus, mammary gland, and
 

ovary.
 

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
 

13
 
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
 

14
 
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) for testing two groups for
 

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

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
 

15
 
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.
 

A time-adjusted analysis was applied when numerous early deaths
 

resulted from causes that were not associated with the formation of
 

tumors. In this analysis, deaths that occurred before the first tumor was observed were excluded by basing the statistical tests on
 

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

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

tumor was found, comparisons were based exclusively on animals that
 

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

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

Cochran-Armitage tests, etc.) were followed.
 

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

incidence of tumors. Curves of the proportions surviving without an
 

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

during which animals died naturally or were sacrificed was entered as
 

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

16
 
these curves were used for two groups; Tarone's extension to testing
 

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

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

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

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
 

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
 

17
 
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.
 

18
 
III. CHRONIC TESTING RESULTS: RATS
 

A. Body Weights and Clinical Observations
 

In males, mean body weight depression was evident in the high
 

dose group, when compared to the high dose controls, as early as week
 

16 (Figure 1). At the end of the bioassay these high dose males
 

weighed approximately 25 percent less than their controls. Very
 

slight mean body weight depression was recorded for the low dose
 

males when compared to their controls. The same general weight depression patterns were observed in females. The exceptions were: the
 

unexplained large weight gain and subsequent loss in the low dose
 

females as compared to their controls during weeks 36 to 68; the
 

negative and positive peaks observed in high dose males and females,
 

respectively, in week 10; and a mean group body weight in the high
 

dose females approximately 18 percent less than their controls at the
 

end of the bioassay.
 

Clinical observations recorded for rats were primarily limited
 

to palpable subcutaneous masses (one low dose female, one low dose
 

control female, and three high dose males) and ulcerative inguinal
 

lesions (one high dose male, one low dose control male, and one low
 

dose control female). The only other clinical signs reported were
 

pale discoloration of the eye in one high dose male and an abscess
 

on the ventral surface in one high dose female.
 

19
 
750- •750 

MALE RATS ——— LOW CONTROL 

—'—•"-' HIGH CONTROL 

600 — - 600 LOW DOSE 

____ _ HIGH DOSE 

o 

450­ -450 T 

(3 

8 — 300 

CD 

150 — -150 

T~ 

15 30 45 60 75 90 120 

TIME ON TEST (WEEKS) 

750- -750 

FEMALE RATS _—_——_ LOW CONTROL 

—'•—'—" HIGH CONTROL 

600 — - 600 

LOW DOSE 

____ _ HIGH DOSE 

I- 450- -450 

I 

— LU 

g 

> 

g 300- -300 

m 

150 — -150 

\ I 

15 30 45 60 75 90 105 120 

TIME ON TEST (WEEKS) 

FIGURE 1 

GROWTH CURVES FOR 2,4-DINITROTOLUENE CHRONIC STUDY RATS 

20 
B. Survival
 

The estimated probabilities of survival for male and female
 

rats in the control and 2,4-dinitrotoluene-treated groups are shown
 

in Figure 2. Because the experiments for the low dose and high dose
 

rats were conducted at different times, each was assigned its own set
 

of controls.
 

For male rats neither the high dose nor the low dose group
 

experienced a significantly different survival rate from its corresponding control group. Despite the sacrifice of five males from each
 

group in week 78, survival was relatively good: 58 percent (29/50)
 

of the high dose, 58 percent (29/50) of the low dose, 52 percent
 

(13/25) of the high dose control, and 64 percent (32/50) of the low
 

dose control group survived until the end of the study.
 

For female rats neither the high dose nor the low dose group
 

had a significantly different survival rate from its corresponding
 

control group. Five females were sacrificed from each group in week
 

78. Survival, however, was relatively good as 52 percent (26/50) of
 

the high dose, 48 percent (12/25) of the high dose control, 62 percent (31/50) of the low dose, and 62 percent (31/50) of the low dose
 

control survived until the end of the study.
 

C. Pathology
 

Histopathologic findings on neoplasms in rats are tabulated in
 

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

tabulated in Appendix C (Tables Cl and C2).
 

21
 
PROBABILITY OF S

PROBABILITY OF SURVIVAL 

en 

3J 

O 

O 

s­ 1

•a 

z 

en 

O 

-n 

N 

o 

633 

Ig m 

m 

m c 

m en 

m 

o 

33 

O 

Z 

o 

CO 

a 

en 

~T 

o 
There appeared to be an increase in the occurrence of integumentary tumors in low and high dose males. The predominant tumor
 

type was the fibroma (skin/subcutaneous tissue 7/49 low dose, 13/49
 

high dose), with the sporadic occurrence of squamous-cell papillomas
 

(1/49 low dose), basal-cell carcinomas (1/49 low dose), fibrosarcomas
 

(1/49 low dose; 2/49 high dose), and lipomas (3/49 high dose). None
 

of these integumentary tumor types were observed in control males.
 

Among females, a slight increase in fibromas of the skin was noted
 

among the high dose group. The fibromas were circumscribed, well-


differentiated masses composed of mature fibroblasts enmeshed in
 

bundles and whorls of collagen.
 

In assessment of other organ systems, there was a high incidence
 

of fibroadenomas of the mammary gland in high dose females. The
 

histologic appearance of these tumors was basically similar to those
 

described by Hallowes and Young (1973). However, certain histologic
 

variations were seen in these fibroadenomas which were not noted by
 

Hallowes and Young. Marked variation in the epithelial/stromal ratio
 

of the tumors was noted and many contained large dilated ducts of
 

galactoceles which contained secretion. Within a focal area of a
 

fibroadenoma in one low dose female, transformation to an intraductal
 

carcinoma was noted. In the affected area, cells were more hyperchromatic, piled upon each other, and there were numerous mitotic figures.
 

In one high dose female with multiple mammary tumors, four were fibroadenomas and one was an adenocarcinoma. Lobular hyperplasia of mammary
 

23
 
tissue adjacent to the fibroadenomas frequently occurred and, in some
 

areas, there was marked basophilia of the cytoplasm and hyperchromicity of nuclei in the hyperplastic lobules. In general, the remaining
 

organ systems showed a similar variety and incidence of neoplasms in
 

the chemically treated and control groups, and these were considered
 

to be part of the general background level of neoplasms in Fischer
 

344 rats.
 

Certain unusual neoplasms occurred in a low incidence in some
 

treated groups and not in controls. These included one hemangiosarcoma in the subcutis, one hemangiosarcoma of the urinary bladder,
 

and one adenocarcinoma of the prostate gland in the high dose males.
 

However, these are not considered to be related to chemical administration.
 

The incidence and variety of nonneoplastic, degenerative, proliferative, and inflammatory lesions were similar in control and
 

chemically treated rats.
 

Based on the increase of fibromas in male rats and fibroadenomas
 

in high dose female rats observed in this histopathologic examination,
 

2,4-dinitrotoluene appeared to induce benign tumors in the Fischer
 

344 rat.
 

D. Statistical Analyses of Results
 

The results of the statistical analyses of tumor incidence in
 

rats are summarized in Tables 3 and 4. The analysis for every type
 

24
 
TABLE 3 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS AT 

SPECIFIC SITES IN MALE RATS TREATED WITH 2,4-DINITROTOLUENE£ 

LOW DOSE HIGH D.OSE LOW 

TOPOGRAPHY: MORPHOLOGY CONTROL CONTROL DOSE 

Liver: Hepatocellular Carcinoma 0/45(0.00) 0/25(0.00) 3/49(0.06) 

P Values0 N.S. 

Relative Risk (Control) Infinite 

Lower Limit 0.554 

Upper Limit Infinite 

Weeks to First Observed Tumor 96 

SJ 

Ul 

Subcutaneous Tissue or Skin: Fibroma 0/46(0.00) 0/25(0.00) 7/49(0.14) 

P Values0 P ­ 0.008 

Relative Risk (Control) Infinite 

Lower Limit 1.827 

Upper Limit Infinite 

Weeks to First Observed Tumor 96 

Subcutaneous Tissue: Lipoma 0/46(0.00) 0/25(0.00) 0/49(0.00) 

P Values0 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

HIGH 

DOSE 

3/48(0.06) 

N.S. 

Infinite 

0.321 

Infinite 

108 

13/49(0.27) 

P = 0.003 

Infinite 

2.106 

Infinite 

85 

3/49(0.06) 

N.S. 

Infinite 

N.S. 

Infinite 

108 
TABLE 3 (Continued) 

TOPOGRAPHY : MORPHOLOGY 

LOW DOSE 

CONTROL 

HIGH DOSE 

CONTROL 

Pancreatic Islets: Islet-Cell Adenoma 

or Islet-Cell Carcinoma'3 

P Values0 

1/45(0.02) 2/25(0.08) 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 105 109 

Hematopoietic System:

P Values0 

 Leukemia 3/46(0.07) 4/25(0.16) 

N.S. 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

—_ 

Weeks to First Observed Tumor 105 85 

Pituitary:

Adenoma 

P Values0 

 Adenoma NOS or Basophil 

9/44(0.20) 3/21(0.14) 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 105 78 

LOW
 

DOSE
 

3/45(0.07)
 

N.S.
 

3.000
 

0.252
 

153.830
 

102
 

4/49(0.08)
 

N.S.
 

1.252
 

0.224
 

8.138
 

78
 

5/44(0.11)
 

N.S.
 

0.556
 

0.159
 

1.689
 

78
 

HIGH
 

DOSE
 

3/48(0.06)
 

N.S.
 

0.781
 

0.097
 

8.952
 

78
 

3/49(0.06)
 

N.S.
 

0.383
 

0.061
 

2.111
 

64
 

0/35(0.00)
 

P = 0.048(N)
 

0.000
 

0.000
 

0.979
 
TOPOGRAPHY : MORPHOLOGY 

Adrenal: Pheochromocytoma 

P Values0 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

Thyroid: C-Cell Adenoma or C-Cell Ni Carcinoma"3 

—I 

P Values0 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

Testis: Interstitial-Cell Tumor 

P Values 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

TABLE 3 (Continued) 

LOW DOSE 

CONTROL 

6/45(0.13) 

96 

3/42(0.06) 

105 

44/45(0.98) 

HIGH DOSE 

CONTROL 

2/25(0.08) 

109 

0/23(0.00) 

19/24(0.79) 

LOW 

DOSE 

3/46(0.07) 

N.S. 

0.489 

0.084 

2.140 

106 

3/41(0.07) 

N.S. 

1.537 

0.186 

17.606 

96 

43/46(0.93) 

N.S. 

0.956 

0.914 

1.058 

HIGH 

DOSE 

3/45(0.07) 

N.S. 

0.833 

0.104 

9.528 

108 

5/47(0.11) 

N.S. 

Infinite 

0.637 

Infinite 

108 

46/49(0.94) 

N.S. 

1.186 

0.963 

1.411 

Weeks to First Observed Tumor 77 78 78 72 
TABLE 3 (Concluded)
 

Treated groups received time-weighted average concentrations of 0.008 or 0.020 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 of the relative risk of the treated group to the control group.
 

CO
 
TABLE 4

 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS AT 

SPECIFIC SITES

TOPOGRAPHY : MORPHOLOGY 

Liver: Hepatocellular Carcinoma 

P Values0 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

NS 

Subcutaneous Tissue: Fibroma 

P Values0 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

Mammary Gland: Fibroadenoma 

P Values0 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

 IN FEMALE RATS TREATED WITH 2,4-DINITROTOLUENE£ 

LOW DOSE HIGH DOSE LOW HIGH 

CONTROL CONTROL DOSE DOSE 

1/47(0.02) 0/23(0.00) 0/49(0.00) 1/50(0.02) 

N.S . N.S. 

0.000 Infinite 

0.000 0.025 

17.891 Infinite 

107 109 

0/48(0.00) 0/23(0.00) 0/49(0.00) 3/50(0.06) 

—_ N.S. ___ Infinite 

0.285 

Infinite 

100 — — 

9/48(0.19) 4/23(0.17) 12/49(0.24) 23/50(0.46) 

N.S. P = 0.016 ___ ___ 1.306 2.645 

0.559 1.062 

3.183 9.435 

92 109 83 69 
TABLE 4 (Continued)
 

TOPOGRAPHY : MORPHOLOGY
 

Hematopoietic System: Leukemia
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Pituitary: Adenoma NOS or
 

Chromophobe Adenoma"-1
 

UJ
 

o 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
 

2/48(0.04)
 

105
 

19/46(0.41)
 

71
 

2/47(0.04)
 

105
 

HIGH DOSE
 

CONTROL
 

2/23(0.09)
 

106
 

8/21(0.38)
 

109
 

2/23(0.09)
 

109
 

LOW
 

DOSE
 

2/49(0.04)
 

N.S.
 

0.980
 

0.074
 

13.043
 

107
 

22/45(0.49)
 

N.S.
 

1.184
 

0.718
 

1.955
 

78
 

2/49(0.04)
 

N.S.
 

0.959
 

0.072
 

12.769
 

78
 

HIGH
 

DOSE
 

4/50(0.08)
 

N.S.
 

0.920
 

0.145
 

9.724
 

93
 

14/40(0.35)
 

N.S.
 

0.919
 

0.446
 

2.154
 

78
 

0/50(0.00)
 

N.S.
 

0.000
 

0.000
 

3.177
 
TABLE 4 (Concluded) 

TOPOGRAPHY : MORPHOLOGY 

LOW DOSE 

CONTROL 

HIGH DOSE 

CONTROL 

LOW 

DOSE 

HIGH 

DOSE 

Thyroid: C-Cell Adenoma or

Carcinoma 

P Values0 

 C-Cell 

2/45(0.04) 3/21(0.14) 2/45(0.04) 

N.S. 

6/48(0,13) 

N.S. 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

1.000 

0.075 

13.270 

0.875 

0.213 

5.047 

Weeks to First Observed Tumor 105 109 94 100 

u> Uterus: Endome trial Stromal Polyp 

P Values0 

15/46(0.33) 6/23(0.26) 14/47(0.30) 

N.S. 

11/49(0.22) 

N.S. 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

0.914 

0.464 

1.789 

0.861 

0.343 

2.539 

Weeks to First Observed Tumor 78 86 91 78 

Treated groups received time-weighted average concentrations of 0.008 or 0.020 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 of the relative risk of the treated group to the control group.
 
of tumor that was observed in more than 5 percent of any of the 2,4­

dinitrotoluene-dosed groups of either sex is included. The Cochran-


Armitage test was not used in these analyses since the low dose and
 

the low dose control were started at a different time from the high
 

dose and the high dose control.
 

For male rats the Fisher exact test showed that the high dose
 

group had a significantly (P = 0.003) higher incidence of fibromas of
 

the subcutaneous tissue and skin than the high dose control. For the
 

comparison of low dose to low dose control the Fisher exact test was
 

also significant (P = 0.008). In the historical data compiled by
 

this laboratory for the NCI Bioassay Program, 23/584 (3 percent) of
 

the untreated male Fischer 344 rats had fibromas of the subcutaneous
 

tissue or skin.
 

Based on these results the statistical conclusion is that the
 

administration of 2,4-dinitrotoluene to male Fischer 344 rats was
 

associated with the increased incidence of fibromas of the subcutaneous tissue or skin.
 

In females the Fisher exact test indicated a significant (P =
 

0.016) increase in fibroadenomas of the mammary gland in the high
 

dose compared to the high dose control. The historical data indicated
 

115/585 (20 percent) untreated female Fischer 344 rats had a riammary
 

fibroadenoma.
 

Based upon these results the statistical conclusion is that the
 

administration of 2,4-dinitrotoluene to the high dose female Fischer
 

32
 
344 rats was associated with the increased incidence of fibroadenomas
 

of the mammary gland.
 

The possibility of a negative association between administration
 

and incidence was noted for pituitary adenomas in male rats.
 

33
 
IV. CHRONIC TESTING RESULTS: MICE
 

A. Body Weights and Clinical Observations
 

When compared with their respective controls, high and low dose
 

mice of both sexes exhibited mean body weight depression by week 30
 

(Figure 3). Approximate weight gain, expressed as a percentage of
 

the weight gained by their respective control groups at the end of
 

the bioassay, was 91 percent for low dose males, 82 percent for high
 

dose males, 89 percent for low dose females, and 76 percent for high
 

dose females.
 

No clinical observations were reported for any treated or control mice of either sex.
 

B. Survival
 

The estimated probabilities of survival for male and female
 

mice in the control and 2,4-dinitrotoluene-treated groups are shown
 

in Figure 4. Because the low and high dose groups were tested at
 

different times, each was assigned its own control group.
 

The statistical tests did not indicate a significant positive
 

relationship between dosage and mortality for either sex. In male
 

mice survival was high despite the sacrifice of five high dose and
 

five high dose control males in week 78 and of five of the low dose
 

control males in week 79. Seventy-eight percent of the high dose,
 

74 percent of the high dose control, 90 percent of the low dose,
 

and 82 percent of the low dose control males survived until the end
 

of the test.
 

34
 
-50 

.--'•• — 40 

/ S \£ 

*' ---"- "^ 

-30 

-20 

LOW CONTROL 

HIGH CONTROL 

-10 LOW DOSE 

HIGH DOSE 

l^ I ~ I ' I 

15 60 75 90 105 120 

TIME ON TEST (WEEKS) 

LOW CONTROL 

—'• HIGH CONTROL 

LOW DOSE 

HIGH DOSE 

T 15 45 60 75 105 120 

TIME ON TEST (WEEKS) 

FIGURE 3 

GROWTH CURVES FOR 2,4-DINITROTOLUENE CHRONIC STUDY MICE 

35 
PROBABILITY OF SURVIVAL PROBABILITY O

33 

O 

o s- X 

(/>

O 

en 

O 

O 

ig m m 

en 

m 

m 

O 

o 

I 

0 

o 
In females five of the high dose treated and five of the high
 

dose control mice were sacrificed in week 78, as well as five low
 

dose controls in week 79. Seventy-two percent of the high dose, 70
 

percent of the high dose control, 84 percent of the low dose, and 78
 

percent of the low dose control survived until the end of the study.
 

Thus in both sexes survival was adequate for meaningful statistical analyses.
 

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).
 

There appeared to be no increase in the incidence of neoplasms
 

in the treated mice compared with their corresponding control groups.
 

With few exceptions, the same variety of neoplasms occurred in the
 

chemically treated and control groups. This spectrum of neoplasms
 

was similar to that expected in untreated B6C3F1 mice.
 

The incidence and variety of nonneoplastic, degenerative,
 

proliferative, and inflammatory lesions was similar in the control
 

and chemically treated mice.
 

This histopathologic examination of B6C3F1 mice treated with
 

2,4-dinitrotoluene provided no evidence of carcinogenicity.
 

D. Statistical Analyses of Results
 

The results of the statistical analyses of tumor incidence in
 

mice are summarized in Tables 5 and 6. The analysis for every type
 

37
 
___ 

TABLE 5 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS AT 

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

LOW DOSE HIGH DOSE LOW 

TOPOGRAPHY MORPHOLOGY CONTROL CONTROL DOSE 

Liver: Hepatocellular Carcinoma 12/46(0.26) 10/45(0.22) 6/47(0.13) 

P Values N.S. 

Relative Risk (Control) 0.489 

Lower Limit 0.165 

Upper Limit 1.281 

Weeks to First Observed Tumor 93 93 92 LO 

00 

Hematopoietic System: Malignant Lymphoma 2/46(0.04) 2/46(0.04) 1/48(0.02) 

P Values0 N.S. 

Relative Risk (Control) —_ 0.479 

Lower Limit 0.008 

Upper Limit 8.888 

Weeks to First Observed Tumor 93 97 93 

Lung: Alveolar/Bronchiolar Carcinoma 2/46(0.04) 4/45(0.09) 3/48(0.06) 

P Values0 N.S. 

Relative Risk (Control) 1.437 

Lower Limit 0.173 

Upper Limit 16.575 

Weeks to First Observed Tumor 93 97 93 

HIGH 

DOSE 

9/48(0.19) 

N.S. 

0.844 

0.335 

2.095 

59 

3/49(0.06) 

N.S. 

1.408 

0.169 

16.250 

78 

1/48(0.02) 

N.S. 

0.234 

0.005 

2.254 

93 
TABLE 5 (Concluded) 

LOW DOSE HIGH DOSE LOW HIGH 

TOPOGRAPHY : MORPHOLOGY CONTROL CONTROL DOSE DOSE 

Lung: Alveolar/Bronchiolar Adenoma or 

Alveolar/Bronchiolar Carcinomab 7/46(0.15) 11/45(0.24) 3/48(0.06) 2/48(0.04) 

P Values0 N.S. P = 0.005(N) 

Relative Risk (Control) 0.411 0.170 

Lower Limit 0.072 0.019 

Upper Limit 1.679 0.725 

Weeks to First Observed Tumor 79 78 93 93 

10
 

vO Hematopoietic System: Hemangiosarcoma 

or Hemangiomab 0/46(0.00) 0/46(0.00) 3/48(0.06) 0/49(0.00) 

P Values0 N.S. 

Relative Risk (Control) Infinite

 

Lower Limit 0.578

 

Upper Limit Infinite

 

Weeks to First Observed Tumor 93 —« —— 

Treated groups received time-weighted average concentrations of 0.008 or 0.040 percent in feed.
 

D

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 of the relative risk of the treated group to the control group.
 
___ 

___ 

ANALYSES OF THE

SPECIFIC SITES IN FEMALE

TOPOGRAPHY : MORPHOLOGY 

Liver: Hepatocellular Carcinoma 

P Values0 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

Hematopoietic System: Malignant Lymphoma 

P Values0 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

Lung: Alveolar/Bronchiolar Adenoma 

P Values 

Relative Risk (Control) 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

TABLE 6 

 INCIDENCE OF

 MICE TREATED

LOW DOSE 

CONTROL 

4/46(0.09) 

93 

5/46(0.11) 

79 

0/46(0.00) 

— _ 

 PRIMARY TUMORS AT 

 WITH 2,4-DINITROTOLUENE* 

HIGH DOSE 

CONTROL 

4/45(0.09) 

78 

11/46(0.24) 

—_ 

94 

1/45(0.02) 

97 

LOW 

DOSE 

1/46(0.02) 

N.S. 

0.250 

0.005 

2.401 

94 

4/46(0.09) 

N.S. 

0.800 

0.169 

3.480 

94 

3/46(0.07) 

N.S. 

Infinite 

0.603 

Infinite 

84 

HIGH 

DOSE 

1/50(0.02) 

N.S. 

0.225 

0.005 

2.167 

93 

7/50(0.14) 

N.S. 

0.586 

0.211 

1.509 

68 

0/48(0.00) 

N.S. 

0.000 

0.000 

17.480 
___ 

TABLE 6 (Concluded)
 

LOW DOSE HIGH DOSE LOW HIGH 

TOPOGRAPHY rMORPHOLOGY CONTROL CONTROL DOSE DOSE 

Stomach: Squamous-Cell Papilloma 0/45(0.00) 3/42(0.07) 0/44(0.00) 0/41(0.00) 

P Values0 «__ N.S. 

Relative Risk (Control) 0.000 

Lower Limit 0.000 

Upper Limit 1.692 

Weeks to First Observed Tumor 97 

Pituitary: Adenoma NOS or Chromophobe
 

Adenoma" 4/37(0.11) 6/37(0.16) 3/38(0.08) 0/34(0.00)
 

P Values N.S. P = 0.017(N)
 

Relative Risk (Control) 0.730 0.000
 

Lower Limit 0.114 0.000
 

Upper Limit 4.022 0.670
 

Weeks to First Observed Tumor 93 97 94
 

Treated groups received time-weighted average concentrations of 0.008 or 0.040 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 of the relative risk of the treated group to the control group.
 
of tumor that was observed in more than 5 percent of any of the
 

2,4-dinitrotoluene-dosed groups of either sex is included.
 

There were no tumors in either sex having a statistically significant positive association between chemical administration and
 

incidence. As such, there was no convincing evidence of carcinogenicity in B6C3F1 mice at the dose levels used in this experiment.
 

To provide additional insight into the possible carcinogenicity
 

of this compound, 95 percent confidence intervals on the relative
 

risk have been estimated and entered in the tables based upon the
 

observed tumor incidence rates. In many of the intervals shown in
 

Tables 5 and 6, the value one is included; this indicates the absence
 

of statistically significant results. It should also be noted that
 

many of the confidence intervals have an upper limit greater than
 

one, indicating the theoretical possibility of tumor induction in
 

mice by 2,4-dinitrotoluene that could not be established under the
 

conditions of this test.
 

The possibility of a negative association between administration
 

and incidence was observed for pituitary adenomas in female mice and
 

for alveolar/bronchiolar neoplasms in male mice.
 

42
 
V. DISCUSSION
 

Under the conditions of this bioassay, dietary administration of
 

2,4-dinitrotoluene was associated with increased incidences of fibromas of the subcutaneous tissue and skin in male rats and fibroadenomas
 

of the mammary gland in female rats, but there were no increased incidences of tumors in treated mice of either sex when compared to controls. No significant 'association was demonstrated between chemical
 

administration and mortality in either species.
 

In male rats integumentary tumors (i.e., fibromas) were the only
 

neoplasms observed at statistically significant incidences. The incidences of skin and subcutaneous tissue fibromas were 0/46, 0/25, 7/49
 

(14 percent), and 13/49 (27 percent) in low dose control, high dose
 

control, low dose, and high dose males, respectively. The only group
 

among the female rats exhibiting these tumors was the high dose (3/50
 

or 6 percent). Statistical analyses of the incidences of these subcutaneous fibromas indicated a significant positive increase in incidence for the high dose males compared to the high dose control males.
 

There were certain unusual neoplasms (i.e., hemangiosarcoma in
 

the subcutis, hemangiosarcoma of the urinary bladder, and prostate
 

gland adenocarcinoma) that occurred at low incidences in high dose
 

but not low dose or control male rats. These tumors were not considered to be related to chemical administration.
 

In female rats the only neoplasm observed at a significantly
 

increased incidence was fibroadenoma of the mammary gland. This
 

43
 
tumor occurred at incidences of 9/48 (19 percent), 4/23 (17 percent),
 

12/49 (24 percent), and 23/50 (46 percent) in the low dose control,
 

high dose control, low dose, and high dose groups, respectively. The
 

comparison of the high dose group to its control group indicated a
 

statistically significant increase in incidence in the dosed females.
 

There were no neoplasms occurring at statistically significant
 

incidences in mice of either sex.
 

Under the conditions of this bioassay dietary administration of
 

2,4-dinitrotoluene to Fischer 344 rats induced benign tumors (i.e.,
 

skin and subcutaneous tissue fibromas in male Fischer 344 rats and
 

mammary fibroadenomas in female Fischer 344 rats). No evidence was
 

provided for the carcinogenicity of the compound in B6C3F1 mice of
 

either sex.
 

44
 
VI. BIBLIOGRAPHY
 

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

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

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

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

J. Wiley & Sons, New York, 1971.
 

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

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

Union Against Cancer, Geneva, 1969.
 

Chemical Abstracts Service, The Chemical Abstracts Service (CAS)
 

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

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

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

Co., Ltd., London, 1970.
 

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

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

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.
 

Hallowes, R.C., and S. Young, "Tumours of the Mammary Gland." Pathology of Tumours in Laboratory Animals, Volume 1. V.S. Turusov,
 

editor. International Agency for Research on Cancer, Lyon,
 

France, pp. 31-74, 1973.
 

Hiasa, Y.J., "m-Toluenediamine Carcinogenesis in Rat Liver." Journal
 

of the Nors Medical Association 21:1-19, 1970.
 

Institute of Makers of Explosives, New York, New York. Personal
 

communication, May 16, 1977.
 

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

Observations." Journal of the American Statistical Association
 

53_: 457-481, 1958.
 

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

"Carcinogenesis Bioassay Data System." Computers and Biomedical
 

Research 7:230-248, 1974.
 

45
 
Manufacturing Chemists Association, "Properties and Essential Information for Safe Handling and Use of Dinitrotoluenes ." Chemical
 

Safety Data Sheet SD-93, 1966.
 

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

Co., New York, 1966.
 

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.
 

Sax, N.I., Dangerous Properties of Industrial Materials. Van Nostrand
 

Reinhold Company, New York, 1975.
 

Simon, G.S., R.G. Tardiff, and J.F. Borzelleca, "Possible Mutagenic
 

Effects of 2,4-Dinitrotoluene: A Dominant Lethal Study in the
 

Rat." Abstracts of Papers. Society of Toxicology Sixteenth
 

Annual Meeting. Toronto, Canada, March 27-30, 1977.
 

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

, 1975.
 

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

Industries Center, Stanford Research Institute, Menlo Park,
 

California. Personal communication, May 11, 1977.
 

U.S.	 International Trade Commission, Synthetic Organic Chemicals.
 

United States Production and Sales, 1975. USITC Publication
 

804, Washington, D.C., 1977.
 

Weisburger, E.K., Chief, Carcinogen Metabolism and Toxicology Branch,
 

National Cancer Institute, Bethesda, Maryland. Personal communication, 1976.
 

Wynder, E.L., J. Onderdonk, and N. Mantel, "An Epidemiological
 

Investigation of Cancer of the Bladder." Cancer 16:1388-1407,
 

1963.
 

46
 
APPENDIX A
 

SUMMARY OF THE INCIDENCE OF NEOPLASMS
 

IN RATS TREATED WITH 2,4-DINITROTOLUENE
 

TABLE A1

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN MALE RATS TREATED WITH 2,4-DINITROTOLUENE 

LOW DOSE HIGH DOSE CONTROL (UNTS ) COSTFCL (DNTE) L OU DOSE HIG H DOSE 

01-0030 01-0084 01-OC25 01-0087 

JltilKAL S INITIALL Y I N STUC Y 50 25 50 50 

ANIMAL S NECROPSIE D 46 25 49 49 

ANIMAL S EXAMINE D HISTOPATHOLOGICAL LY ** 45 25 49 49 

INTEGUMENTAR Y SYSTEM
 

*SKIN (46) (25) (49) (49) 

SQUAMOU S CEL L PAPILLOM A 1 (2% ) 

SQOAMCD S CEL L CAFCINOM A 

BASAL-CEL L CBRCINOM A 1 (2*) 

FIEROH A 2 <4X ) 

*SUBCUT TISSUE (46 ) (25) (49) (49 ) FIBROM A 5 (10X) 13 (27% ) 

FIBROSARCOMA 1 <2X) 2 (H%) 

LIPOM A 3 (6*) ANGIOSAPCCM B 1 (2%) 

HEEPIRATOP Y SJSTEfl 

IIPACHEA ("5) (11) (44) (49)
 

SQOAF10US CELL CAPCINOdA 1 (2%)
 

ILONG (45) (25) (48) (49)
 

ALVEOLAH/BtcSCHICLAP ADENOMA 1 (2%) 2 <8X)
 

ALVEOLAR/ERCNCHIOLAF CARCINC»IA 1 <4S)
 PHEOCHHOBOCYTOMJ , H^TASTATIC 1 <4X)
 FIBROSARC01A, METSSTATIC 1 (2*)
 

HEMATOPOIETIC SYSTI"
 

*«UITIPLE ORGANS (46 ) (25) (49) MALIGNAN T IYMFHCMB , NOS 

LEUKEMIA,NOS 

DNEIFFERENTIBTED IEUKEMI A 

MYELOMONOCYTIC IEPKEMIA 1 (2%) 3 (6%) 

LYMPHOCYTIC LEUKEHIA 2 (4« ) 

G£ANULOCYTJC_LJUKEJ5JA 

t NUMBE R O F ANIMAL S KIT H TISSUE EXAMINE D MICROSCOPICALL Y 

* NUMBE R C E ANIMAL S NECEOPSIE D 

"EXCLUDES PARTIALLY AUTOLYZED ANIMALS 

A-3 
•SPLEEN
 

IEICMYCMA
 

MYELOMONOCYTIC LEUKEMIA
 

CIRCULATORY SYSTEM
 

NONE
 

DIGESTIVE SYSTEM
 

tSALIVARY GLAND
 

FIBFOSARCCSA, HETASTATIC
 

»IIVER
 

HEPATCCFIIULAR CAFCINOMA
 

•STOMACH
 

SQUAMCIIS CSLL FAPILLOMA
 

EASAL-CELL CARCINOM A
 

HUUODENUM
 

ACENOCAPCINC'A, NCS
 

KILEUM
 

CYSTACENCCAECINCMA, NCS
 

URINAR Y SYSTEM
 

(URINARY BLACDEF
 

TBANSITICNAI-CFLL PAPILLOMA
 

HEMANGICSAPCOI1A
 

ENDOCRINE SYSTEM
 

HPITUIIAR Y
 

CARCINCMA.NCS
 

ADENOKA, NCS
 

EASOPHIL ADENOMA
 

tACFENAL
 

CORTICAL ADESC^A
 

I HU1BE B CI ANIMAL S WIT H TISSUE

* MUflBE E O F ANIHAL S KECHCESIE D 

TABLE Al (CONTINUED) 

LOW DOSE ' HIGH DOSE 

CCNTEOL(UNTE ) COH1BCL(CSTR)

01-0030 01-0081

(U5 ) (25) 

(13) (21)
 

(15) (25)
 

(15) (21)
 

1 (1*)
 

1 (IX)
 

("5) (21)
 

(15) (21)
 

(11) (23)
 

(11) (21)
 

9 (20%) 1 (5X)
 

2 (10X)
 

(15)	 (25)
 1 J2%)
 

 EXAMINE D KICROSCOPICAII Y 

A-4 

 LOU DOSE 

 01-0025 

(18)
 

1 (2*)
 

(13)
 

(19)
 

3 (6X)
 

(15)
 

(15)
 

(15)
 

1 (2X)
 

(11)


1 (2X)
 

(11)


2 (5X)
 

5 (11X)
 

(16)
 

HIGH DOSE
 

01-0087
 

(19) 

(18)
 

1 (2*)
 

(18)
 

3 (6X)
 

(19)
 

(19)
 

1 <2X)
 

(19)
 

(19)
 

1 (2K)
 

(35)
 

(15)
 

„
 
TABLE Al (CONTINUED)
 

LOW DOSE HIGH DOSE CCNTROL(USTH) CCNTEOL (CN1R) 

01-0030 01-008H 

EHECCHFOMOOTOr A 6 (13%) 2 (8X) 

OHEOCHROKOCYTOMP , MALIGNAN T 2 (8*) 

IAERENA L MEDIJ11 A ( U 5) (25) 

GANGLICNEUPCM A 

tlHYRCI D (12) (23) 

EOLIICULAR-CEL 1 CARCINOM A 

C-C'.ll ADF.SCM A 2 (5%) 

C-CFLL CARCINOr A 1 (2*) 

UtABATHYECl r (-12) (15) ADEtCfA , NC S 

KIANCREAIIC ISLF1S (45) (25) 

ISIET-CE1L ADEt-CHP . 1 (211) 2 (6*) ISLET-CEIL CABCINC1A 

REPRODUCTIV E SYSTBK 

*HA1MAR Y GLAN D (U6) (25) 

ADENOC6RCIHOI1S , NO S 

FIEFOAEENOM A 1 (<4X) 

*IPEEUTIA 1 GIAN E (46 ) (25) 

CABCINOMA,NC S 1 <UX) ADENOHA , NOS 1 (2%) 1 (t«) 

KFFCSTAT E (US ) (23) ADENOCARCIJJOKA , SC S 

I1ESTIS (U5 ) (21) INTERSTITIAL-CFLL TUMCR Ht (98S) 19 (79%) 

HEBVOU S SYSTEM 

«ERAIS (U5) (25) GLIOMA , NO S 

SEECIA L SENS ^ OPGAf S 

'HARDEBIA N GLAN D (t6) (25) _ ADENOMA^ NOS _ _ „ ^ _ 

» NU1BER OF AMI1ALS «ITH TISSUE yXABINE D MICROSCOPICALLY
 

* NUCBEP CE ANI1ALS NECFOPSIEP
 

A-5
 

LOB DOSE 

01-0025 

3 <7X ) 

(U6 ) 

1 (2*) 

(11) 

3 (7*) 

(27) 

(U5) 

3 <7X > 

(U9) 

1 (2X) 

(«9) 

(15) 

(16) 

13 (93X) 

(15) 

1 (2X) 

(19) 

1 (2*) 

HIGH DOSE 

01-0087 

3 (7X) 

(15) 

(17) 

2 (IX) 1 (2X) 

t (9X) 

(30) 

1 (3X) 

(18) 

2 (IX) 

1 (2X) 

(19) 

1 (2X) 

(19) 

1 (2X) 

(18) 

1 (2X) 

(19)

46 (91X) 

(19) 

(19) 
TABLE Al (CONTINUED)
 

LOW DOSE HIGH DOSE CCNTEO L (IJNTB) CONTBCL ( INTR )

01-0030 01-0081

*EA3 CANA L (16) (25)

SQUAKOD S CELL CARCINOM A 1 (US) 

MUSCULOSKELETJ L SYSTE M 

KCtiE 

ECD Y CAVITIE S 

»ECE Y CAVITIE S <U6 ) (25)

1ESCTBELICM , 90S 3 (7%)

MESOTHELIOMA , PAL1GNAN T 

*HEDIASTINU M («6 ) (25)

SLVEO L AE/EBONCKICIA E CA , METAST A 1 (14%) 

*FLEOB A ((46) (25)

<SLVEOLftR/B?ONCKICIAF . CA , METSST A 1 (14*) 

A L L OTHEP SYSTEMS 

NCN E 

ANIMA L CISPOSITIO N SUHP1AE Y 

ANIMAL S INITIALL Y I N STDD Y 50 25

NATUBA I DEATH S 7 3

 

MORIBUN D SACRIFIC E 6 «

SCHEDULE D SACPIEICE 18 5

ACCIDENTA1L Y KI1IE D 

IEEHINA L SACRIFIC E 19 13

ANIMA L MUSING 

a INCLUDE S AUTCLYZE D ANIMAL S 

I NUMBER OF ANIMALS HUH TISSUE EXAMINED MICSO SCOPICALLY
 

* NUMBER CB ANIMAL S NICEOFSIED
 

 LOW DOSE 

 01-0025 

 (U9 ) 

 (U9 ) 

1 (2*) 

 ((49) 

 (M9 ) 

 50 

6 

 10 

5 

 29 

HIG H DOSE 

01-0087 

(149) 

(149) 

1 (2% ) 

1 (2X) 

(149) 

(19) 

50 

5 

11 

5 

29 

A-6
 
TABLE Al (CONCLUDED)
 

LOU DOSE HIGH DOSE 

CONTBOl(OHIB) COHTBCL(CHTB) LOW DOSE HIGH DOSE 

01-0030 01-0081 01-0025 01-0087 

TUM0 5 SUMMAR Y 

TCTA L ANIMAL S HI1H PRIMAR Y TUMORS * u u 22 U7 48 

TOTAL PRIMAR Y 10SCBS 75 11 81 92 

TOTA L ANIMAL S BIT H BENIG N TU«CP S 14 20 «5 U6 

TCTAL BENIG N tl'MOSS 65 31 70 69 

TOTAL ANIMAL S WITH MALIGNAN T TUKCF S 7 9 12 18 

TOTAL MALIGNAK T TUMORS 7 10 13 22

 

TCTAL ANIMALS WITH SJCONDAEY TUKCKS* 2
 

TOTAL SECONDARY TU1CBS 3
 

TOTAL ANIMALS KITH TUMORS OSCBRTAINEIMGN CB KALIGNfN-r 3 1
 

TOTAL tlNCEBTAIS TUN05S 3 1
 

TOTAL ANIMALS WITH TUMORS UNCISTAIKIFIMAFY OR MF1AS7ATIC
 

TOTAL UNCERTAIN TUMORS
 

* PRIMARY TUMORS: ALL TUMORS EXCEPT SECCNDABY TUHOBS
 

I SICONCAFY TUMOFJ: PIJTASTATIC TUKCBS OR TUMCBS INVASIVE INTO AH ADJACENT ORGAN
 

A-7
 
TABLE A2

 

SUMMARY OF THE INCIUbNCE OF NEOPLASMS IN FKMALE RATS 

TREATED WITH 2,4-DINITROTOI.UF,NF

 

LOW DOSE HIGH DOSE
 CONTROL(UNTB) CONTROL (UNTR) LOU DOSE HIGH DOSE
 

02-0030 02-008U 02-0025 02-0087
 

ANIMALS INITIALLY IN STUDY 50 25 50 50
 ANIMALS NECPOP£IEC 48 23 «9 50
 ANIMAL S EXAMINED HISTOFATHOLOGICALL Y ** HI 23 H9 50
 

INTEGONENTAPY SYSTEC
 

»SKIN (U8) (23) (K9) (50)
 SEBACEOUS ADENCCAECINOMA 1 (U«)
 

FIEEOMA 1 (2*)
 FIEROSARCOMA 1 (2%)
 FIEROADENOMA 1 (2%)
 

*SU8CUT TISSUE (18) (23) (49) (50)
 SQUAMOUS CELL CAFCINOMA 1 (2X)
 FIEFOMA 3 <6X)
 

RESPIRATORY SYSTEF
 

• LUNG (47) (23) (49) (50)
 SQUAMCUS CEII CAPCINCMA 1 (2%)
 AIVEOLAP/BPCNCHIOLAP ADENOMA 1 (i»X) 1 (2X)
 ALVEOLAR/ERCNCKIOLA? CARCINOMA 1 (2%)
 

HEMATOPOIETIC SYSTEM
 

*HUITIPLE CPGANS (48) (23) (49) (50)
 

MALIGNANT LY!«PFOIi8, NOS 1 (2%)
 LEUKEMIA, NOS 1 (2X)
 UNEIFFEEENTISTID LEUKEMIA 2 (9%)
 

1YELOMONOCYTIC LEUKEMIA 1 (2%) 1 (2X) 4 (8X)
 

*SUBCUT TISSUE (48) (23) (49) (50)
 

HAIIG.LYBPHCMA, HISTIOCYTIC TYPE 1 (2X)
 

ISFIEEN («7) (23) (49) (50)
 

"YF.LCKCNCCYTIC LEUKEMIA 1 (2X)
 

CIRCULATORY SYSTEM
 

NONE
 

# NUMBE R O F ANIMAL S WIT H TISSUE EXACINE E BICRCSCOPICftLLY 

* KUMBE P Cf ANIMAL S NECBOPSIE D 

**EXCLUDES PARTLALLY AUTOLYZED ANIMALS 

A-8 
TABLE A2 (CONTINUED) 

LOW DOSE CONTRO L (UNTS) 

02-0030 

HIGH DOSE CONTROL (UNIR) 

02-0081 

LOil DOSE 

02-0025 

HIGH DOSE 

02-0087 

CIGESTIV E SYSTI M 

ILIVER 

NEOPLASTIC SODULE 

HEPATOCEILULAR CASCINOBA 

(17) 

1 (2X) 

(23 ) 

2 (9X) 

(19) (50) 

1

1

 (2X) 

 <2X) 

ISTOBACH 

3QUAMCUS CELL EAPIH01A 

(16) (23) (19) 

1 (2X) 

(18) 

URINAR Y SYSTEM 

NONE 

IliECCBIN E SYSTE M 

tEIIUITARY 

CASCIKCMA.NO S 

ADENOMA, NOS 

CHROHOPHOEE ADENOB A 

(16) 

1 (2X) 

19 (11X) 

(21) 

1 <5X) 

7 (33X) 

(15) 

2 (IX) 

22 (19%) 

(10) 

1 (3X) 

1« (35X) 

IUDSENA L 

CORTICAL ADENCK A 

PHEOCHSOBCCYTCBA 

PHEOCHROBOCYTOHA , MALIGNAN T 

(17) 

1

2

 (2%) 

 (IX) 

(23) 

2 (9X) 

1 (KX ) 

(19) 

2 (OX) 

(50) 

1 (2*) 

fTHYRCI D 

ACEHOBA , NCS 

POLLICULAR-CFLL CARCINOMA 

C-CELL ADENOM A 

C-CELL CARCINOM A 

(45) 

2 (IX) 

(21) 

1 (5X) 

2 (10X) 

1 (5X) 

(15) 

1

2

 (2X) 

 (IX) 

(18) 

2 (IX) 

« <8X) 

IEANCBEATIC ISLITS 

ISLHT-CELL iDESCM A 

(16) 

2 (1%) 

(22) (17) (18) 

BEERODUCTIV E SYSTEM 

*«A«MABY GLAN E 

ADENOCARCIKCBA , NOS 

PAPILLAR Y CYSTADEtiCMA, NOS 

PAPILLARY CYSTADENOCARCINO(1A,lvCS 

INTRADUCTA L CARCINOM A 

JILEIillllISS £«CT_c;£RCiji2^ 

(18) 

2 (IX) 

(23) 

2 (9X) 

1 J1X1 

(19) 

1 (2X) 

1 <2X ) 

, .._ LL _ L , 

(50) 

1 (2X) 

I NUMBER OF ASIWALS HITH TISSUE EXAMINED

* KU1BER Of ANIBALS 8ECFOESIED 

 MICROSCOPICALLY 

A-9
 
TABLE A2 (CONTINUED) 

LOW DOSE HIGH DOSE CCNTECI(aSTE) CONTBOL (CSTE) LOW DOSE HIGH DOSE 

02-0030 02-008t 02-0025 02-0087 

"IE50ABENCMA 9 (19%) « (17X) 12 (24X) 23 (16%) 

»CIITCRAL GLAN D 

SQUAMOU S CELL FAPIILOB A 

(«8) (23) (19) (50) 

1 <2X ) 

'VAGIN A (18) (23) (i»9) (50) FIEEOSABCOM A 

LYMPHANGIOSAECOf« A 

1

1

 (2*) 

 (2*) 

IUTEBOS (46) (23) (U 7) (49) ESDOMETRIA L STFCdAL fOLYP 15 (33*) 6 (26%) 1 M (30X) 11 (22%) 

lUTEBUS/INCOIlITBIU H 

CAECISC?iA,!10S 

("6)

1 <2X) 

 (23) (H7 ) (19) 

SEEVCU S SYSTE M 

NONE 

SEICIAL SENS E OEGAN S 

*HA3 CANA 1 

FIBHOSAECCfA 

(«8)

1 <2X) 

 (23) <M9 ) (50) 

MUSCUIOSKEIETA I SYSTEM 

NON E 

ECD Y CAVITIE S 

NON E 

A LL OTHER SYSTEM S 

SITE UNKNOW N 

* KUdBE B Cf ANIMAL S SI1H TISSUE IJJMINE D MICSOSCOPICALIY 

* KOflBE B CF ANIMAL S KECECFSIE D 

A-IO 
TABLE A2 (CONCLUDED) 

LOW DOSE HIGH DOSE CCNTBCL (INTE) COSIEOL (CN1E) LOB DOSE HIGH DOSE 02-0030 02-0084 02-0025 02-0087 

ANIMA L DISPOSITION SUKPAP Y 

ANIKAI S INITIALL Y I N STUD Y 50 25 50 50

 NATURA L DEATHS! 6 3 4 7

 MORIBUN D SACRIFIC E 8 5 10 12

 SCHECU1EC SACRIFIC E 17 5 5 5

 ACCIDENTALL Y KILLE D

 

TERMINA L SACRIFIC E 19 12 31 26

 

*NIBA L HISSIN G 

a INCLUDE S AUTCLYZE D ANIMAL S 

1UPCF SUHPAH Y 

TOTA L ANIBALf i KIT H EET«AP Y TUMOBS * 38 19 41 40 

TQTft I PRI"«P Y Ttl»CF S 66 31 62 69 

TCIA L AHIMAL S KIT H tFMIG N TUMOR S 36 18 40 39 

TOTA L EEtiIG N '''IJ^C1? 5U 23 54 56 

TOTAL ANIK8L S WIT H MAIIGNAN T TUPC5 S 9 8 8 11 

TOTA L HALIGNAK T 1UMCR E 12 9 8 12 

TOTA L AKIBAL S JIT H STC010ASY TI1BCFS * 

TOTAL SECONDAR Y TOMC5 S 

TOTA L ANIMAL S HIT F THMOK S UNCEPTAIli -

EE>iIG N CF MALIGNAK 1 2 

TOTAL UNCFBTAI N IIIICFS 2 

TOTA L ANIMAI S WIT H T"NORS '!SCE?TBIlifBIMAF Y C F METAEIAII C

 

TOTAL 'INCEFTAI S 1UTP5 

* PBIHAR Y TljrOPS : AI L TUMOR S EXCEP T SECCNDAn Y TUMOR S 

I EECONCAF Y TIIfOFE : HETASTJTI C TUP!CBS OP, TUBOR S IdVASIV E INTO A N ADJACEN T OEGA N 

A-ll 

APPENDIX B
 

SUMMARY OF THE INCIDENCE OF NEOPLASMS
 

IN MICE TREATED WITH 2,4-DINITROTOLUENE
 

TABLE Bl

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN MALE MICE TREATED WITH 2,4-DINITROTOLUENE 

LOW DOSE HIGH DOSE
 CONTROL (UHTR) COSTROL (ONTR) LOU DOSE HIGH DOSE
 

05-0030 05-0077 05-0025 05-0088
 

AflFALS IllITIAILY IN STUCY 50 50 50 50
 ANIMALS NECRCPSIED 46 48 49
 .« "*6
 

ANIMALS EXAMINED HISTOPATHOIOGICA IIY ''* 46 45 48 19
 

ISTEGUMENTABY SYS1IM
 

*SKIN (46) (46) (48)
 (19)
 HEHANGIOSAFCOMA 1 (2X)
 

RESPIRATORY SYSTEM
 

tlUNG (46) (45) (48) (48)
 HEPATCCEL1ULAB CABCINOMA, METPET 1 (2%) 1 (2K)
 

ALVEOLAR/BBONCHICIAS ADENOMA 5 (11*) 7 (16*) 1 (2*)
 

ALVEOLAR/ERONCFIOIAB CARCINOMA 2 (4*) 1 (9*) 3 (6*) 1 (2*)
 

HEMATOPOIE1IC SYSTEM
 

•MULTIPLE ORGANS (46) (16) (48) (49)
 MALIGNANT LYMPhOMA, NOS 1 (2*) 1 (2X)
 MALIG.LYMPHOMA, HISTIOCYTIC TYEI 1 (2%) 1 (2*) 1 (2*)
 

*SPLEE!i (46) (45) (47) (11)
 HEMANGIOMA 1 (2<)
 HEMANGIOSABCCMP 1 (2X)
 MALIG.LYMPHOMA, HISTIOCYTIC TYEI 1 (2*)
 

JMANDIBOLAR L. NCDE (34) (35) (40) (34)
 

MALIG.IYSPHOMA, HISTIOCYTIC TYPE 1 (3%)
 

IEEYEBS PATCH (46) (43) (45) (45)
 MALIGNANT lYtFHCf*, NOS 1 (2*)
 

CIEC'JLATOPY SYSTEM 

NON E 

* NUHESR OP ANIMAL S KITH TISSUE EXAMSE D MICBOSCOPICALLY 

* SUCBEE Of ANIMAL S SICROPSIED 

**EXCLUDES PARTIALLY AUTOLYEED ANIMALS 

B-3 
DIGESTIVE SYSTEM
 

HIVES
 

HEPATCCELIULAB CAECINOMA
 

HEMANGIOMA
 

ISTOMACH
 

SCUAMOOS CEII FAEILLCMA
 

U5INA8Y SYSTEM
 

NONE
 

EHDCCHINE SYS1IP!
 

*fITUITAEY
 

AUENOCA, NCS
 

HTHYBCID
 

ADENOMA, NCS
 

ADBNOCAPCTNOMA, NCS
 

BEPEODUCTIVE SYSTEB
 

ITESTIS
 

SEMINCMA/DYSGEPMIBCMA
 

N5EVOIJS SYSTEM
 

NONE
 

SEICIA1 SENSE ORGANS
 

*HAHDEBIAN GLAND
 

PAPILLARY CYSTADENOKA, NOS
 

*IAF CANAL
 

SQOAMCUS CELL CARCINOMA
 

t!USCIJLOSKELETAL SYSTEB
 

NCNE
 

I liUMBE B CB ANIMAL S SI1H TISSU E

* MI1BEE CE ANIMAL S fECECFSIE D 

TABLE Bl (CONTINUED) 

LOW DOSE HIGH DOSS
 CCNTBOL (UNTB) CCNTBOL (CUTE)


05-0030 05-0077


(U6) (<t5)


12 (26%) 10 (22X)


(45)	 (42)


1 <2X)
 

(39)	 (36)


1 (3%)
 

<4M) (KO)


2 (5%)
 

(16)	 (US)


1 (2«)
 

(46)	 (46)


1 (2X)
 

(46)	 (46)


1 <2X)
 

 EJJRINE D MICPOSCOPICAL1 Y 

B-4 

 LOB DOSE HIGH DOSE
 

 05-0025 05-0088
 

 (K7) (18)
 

6 (13X) 9 (19X)
 

1 (2X)
 

 (K5) (45)
 

 (33) (U2)
 

 (42)	 (41)
 

1 (2X)
 

 (47) (47)
 

 (148) (»9)
 

 (K8) (49)
 
TABLE Bl (CONCLUDED)
 

LOW DOSE CCliTFCLlUNTB)

05-0030

 HIGH DOSE CONTBOL ( CNTR)

 05-0077

 LOU DOSE

 05-0025

 HIGH DOSE 

 05-0088 

EOEY CAVITIES 

NONE 

All CTHE R SYSTEM S 

NON E 

ANIMA L CISECSITIC N SUMMAR Y 

ANIMAI S INITIALL Y I N STUD Y 

"IATUESL DE8TH S 

MCHIBUK D SACBIFICJ 

SCHEDULE D SAC51FICE 

ACCIDENTALL Y KIIIE D 

TERMINA L SACBIFICE 

ANIMA L MISSIM G 

50

1 

5 

3 

U1

 50

7 

1 

5

 37

 50

5 

 U5

 50 

6 

5 

 39 

S INCLUDE S AUTCIYZE D ANIMAL S 

TDtC F SUMMAF Y 

TOTAL AJilBAl S WIT H EFIHAS Y

TOTA L PFIBAE K TU10PS 

 TUBOFS * 19

26

 21

 25

 13

 15

 13 

 11 

TCTAL AKIMAL S VillH BENIG N

TOTA L BFNIG N TUHCF S 

 TDMOE S 6 

7 

8 

8 

3 

3 

1 

1 

TOTAL ANIMAL S HIT H MALIGNAN T

TCTAL BALIGNASl TCHOES 

 TUPCB S 16

19

 15

 17

 10

 12

 13 

 13 

TOTAL ANIMALS WITH SECCHDAPY TDMC5S*

TOTAL SECCNDASY TOMOES 1 1 1 1 

TOTAL ANIMALS WITH TUMOBS UBCEBTAIN­ EESIGN CB MALIGN8N1 

TCTAL UNCE5TAIH TUMORS 

TOTAL ANIMALS WITH TUMORS UNCEBTAIN­ EBIBABY CB flEIASTA1IC 

TOTAL UNCEFTAIN TUHOES 

* PRIMARY TUMORS: SLL TUMORS EXCIET SECCNCARY TUMOBS t SECCNCABY TUMOPS: KE1ASTATIC TUMOFS OE TUMORS INVASIVE INTO AN ADJACENT ORGAN 

B-5
 
TABLE B2

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN FEMALE MICE TREATED WITH 2,4-DINITROTOLUENE 

ANIMAL S INITIAIIY IK STUDY
 

ANIMALS MISSING
 

ANIMALS NECBOFSIED
 

mitiULS EXAMINED HISTCFATHOLCGICALL Y:
 

INTEGUMENTARY SYSTEM
 

*SKIN
 

?IEROSARCO"A
 

RESPIRATORY SYSTEM
 

ILUSG
 

CARCINOMA, NOS, "ETASTATIC
 

ALVEOLAR/B50NCHIOIAF ADENOMA
 

ALVEOLAH/ERONCEIOLAF. CARCINOMA
 

HEMATOPOIETIC SYSTEM
 

*«OITIPLE CRGANS
 

MALIGNAN T LYMPHOMA , NOS
 

MALIG.LYMPROMA, UN tl EE ER-TYPE
 

MALIG.LYMPHOMA, HISTIOCYTIC TYPE
 

LYMPHOCYTIC IEUKEMIA
 

*SFtEEN
 

HEHANGIOSAECOMA
 

MALIGNANT 1YMPHO»A, NCS
 

MALIG.LYNPEOMA, HISTIOCYTIC TYEE
 

•LIVES
 

MALIG.LYFFHCMA, HISTIOCY1IC TYPE
 

KUPFFEB-Cfill SAFCCMA
 

IEEYEBS PATCH
 

MALIG.LYMPHCMA, HISTIOCYTIC TYPE
 

CIFC'lLATOFY SYSTE M 

NON E 

LOW DOSE
 CONTROL (ONTR)
 

06-0030
 

50
 

l»7
 

*» 47
 

(47)
 

(46)
 

1 <2X)
 

1 (2X)
 

(47)
 

2 (4*)
 

2 (4X)
 

(45)
 

1 (2%)
 

(46)
 

1 (2X)
 

(45)
 

HIGH DOSE
 CONTROL (DSTR)
 

06-0077
 

50
 

46
 

46
 

(46)
 

2 (4X)
 

(45)
 

1 (2X)
 

(46)
 

3 (7X)
 

1 (2X)
 

6 (13X)
 

1 (2X)
 

(13),
 

(45)
 

(43)
 

1 <2X)
 

LOU DOSE
 

06-0025
 

50
 

3
 

46
 

46
 

(46)
 

(46)
 

3 (7X)
 

1 (2X)
 

(46)
 

2 (4X)
 

1 (2X)
 

(44)
 

1 <2X)
 

(46)
 

1 (2X)
 

(42)
 

HIGH DOSE
 

06-0088
 

50
 

50
 

50
 

(50)
 

(48)
 

(50)
 

4 (8X)
 

2 (IX)
 

(48)
 

1 (2X)
 

(50)
 

(44)
 

* NUMBE R O F ANIMAL S WIT H TISSU E EXBMKE D MICROSCOPICALL Y 

* KUW8E F Cf ANIMAL S NICFOPSIED 

**EXCLUDES PARTIALLY AUTOLYZED ANIMALS 

B-6 
TABLE B2 (CONTINUED)
 

LOW DOSE HIGH DOSE coaTpoL(oin:H) CONTBCL(tliTB)

06-0030 06-0077

DIGESTIVE SYSTEM 

f IIVE E (U6 ) (15)

CARCINCMA , DOS, MEIASTAIIC 1 (2%)

 HEPATOCELIULAB CABCINOMA K (9*) 1 <9X)

ISTQHAC H (15) (112)

SQUAMCUS CELL FAPILLCMA 3 <7X)

 

DB1NAB Y SYSTEM 

NONE 

ItiCOCBIN E SYSTE M 

IEI1UITAB Y (3-7) (37)

ACENOMA , HOS 3 (8X) 6 (16S)

CHEOMOFHCBE ADENOM A 1 (3») 

«AEBENA L (41*) (13)

CORTICAL ACENCC A 1 (2*)

 

• 1HYBOIC (11) (30)

PAPILIABY CABCINCfl A 

IEA1CBEA1IC ISLETS (39) (11)

ISLET-CEIL ADEKCH A 1 (2«) 

REFBCDUCTIV E SYSTE M 

*BAMBAS Y G1AN C ( U 7) (16)

ADENOCARCINCMA , HO S 1 (2*)

 

FIEBOSDBNOMA 1 (2%)

 

• DTEBU S (13) (13)

ADENOCABCIHONA , DCS 

i-NDOBETBIA L STROflAl EOLY P 1 (2«) INDOMETBIA 1 STROHA L SARCOM A

 

IUTEBUS/EKDCMETPIU H (13) (13)

CABCINCnA.NC S 1 (2*)

 

ICVABY/OVIDUC 1 (<t3 ) (13)

JAPTL1AH Y AEEtiCB A - -J-J2S1 

f NUMBEB CI ANIMALS WITH TISSUE EXAMINED MICEOSCOPICALIY
 

» tHJIBEB CI ANIMALS NECBOESIED
 

B-7
 

 LOW DOSE 

 06-0025 

 (16) 

1 (2%) 

 (11) 

 (38) 

3 (8*) 

 (12) 

 (10) 

 (11) 

 (16) 

 (11) 

 (11) 

 (11) 

HIGH DOSE 

06-0088 

(50) 

1 (2*) 

(11) 

(31) 

(15) 

(12) 

1 <2») 

(11) 

(50) 

(13) 

1 (2*) 

1 (2*) 

(13) 

(13) 

— _ — _ — 
TABLE B2 (CONTINUED) 

LOW. DOSE HIGH DOSE 

CCNTBOL (UHTR ) CONTBCL(CNTR ) LOU DOSE HIGH DOSE 

06-0030 06-0077 06-0025 06-0088 

INTFAtDCTAL 5AEIIIOMA 1 (2%) 

*CVARY (44) (41) (44) (37) 

IUTICMA 1 (2X) 

N1PVCUS SYSTEB 

NONE 

SEICIAL SEKSE OBGANS 

»HAaDESIAN GLAND (1*7) (46) (46) (50) 

PAPILLARY CYSTADENOMA, NOS 2 (4X) 

NOSC'ILCSKEIETAL SYSTEH 

NONE 

BCDY CAVITIES 

*AEECMINA L CAVITY (47) (46) (46) (50) 

HEMANGIOSAFCOMA 1 (2%) 

All CTREE SYSTEMS 

NONE 

ANIMA L IISECSITICN SUMMAS Y 

BNIHAIS INITISIIY IK STUDY 50 50 50 

NATUEAI DEATHS 5 

MOEIBUND SACBIFICI 1 

SCHEDUIED SACEIFICE 5 

ACCIDENTALIY KIIIIC 

TEEMINAl SACEIFICf 39 35 42 36 

HISSING 3 

S_INCLUDES_AUTOLYZED_AJJIJJAi S _______ _ 

» NUMBER OF ANIdALS KITH TISSUE EXAMINED BICBOSCOPICJLLJ 

* tfUMBEE OF ANIMALS HECECESIED 

B-8
 
TABLE B2 (CONCLUDED)
 

LOW DOSE HIGH DOSE CONTRO L (UNTH ) CONTBOL (CNTR) LOS DOSE HIGH DOSE 

06-0030 06-0077 06-0025 06-0088 

1CTAL ANIMAL S WIT H ESIMAR Y TUMOBS * 17 22 1U 11 

TOTAL ERIMAB Y TUMOf S 21 32 15 11 

TOTAL ANIMAL S WIT H EENIG N TUMOR S 7 12 8

 

TCTAL BENIG N TUMOF S B 13 B 

TOTAL ANIMAL S KITH MALIGNANT TUMCBS 11 18 7 11 

TOTAL MALIGNAN T TUMOR S 13 19 7 11 

7CTAL ANIMALS 8I1H SICONDAPY TUMOHS* 1
 

TOTAL SECCNDAEY TUMORS 2
 

TOTAL ANIMALS HUH TUM05S UNCERTAINBENIGN CR MALIGNANT
 

TOTAL UNCKETAIN TUMORS
 

TOTAL ANIMALS WITH TUMOHS UNCIRTAISEEIHARY CR METASTATIC
 

TOTAL UNCERTAIN TUMORS
 

* PRIMARY TUM08S: ALL TUMORS EXCEPT SECONDARY TUHOBS
 

* SECONDARY TUMORS: MEIASTATIC TUMOHS OR TUMORS INVASIVE INTO AN ADJACENT ORGAN
 

APPENDIX C
 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC
 

LESIONS IN RATS TREATED WITH 2,4-DINITROTOLUENE
 

TABLE Cl

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN MALE RATS TREATED WITH 2,4-DINITROTOLUENE 

C

LOW DOSE 

ONTROL(UNTR ) 

HIGH DOSE CONTROL (UHTR) LOB DOSE HIGH DOSE 

01-0030 01-0081 01­0025 01-0087 

ANIMAL S INITIALL Y III STUDY 50 25 50 50 

ANIMAL S NECROPSIE S 46 25 49 49 

ASICAL S EXAMINE E HISTOPATHOLOGICALlY* * 4 5 25 49 49 

INTEGUMENTARY SYSTEM
 

»SKIN (46) (25) (49) 

IPIDEEMAI INCIUSICN CYST 1 (2X) 

COLLAGEN DISEASE <2X ) 

"IEFOSIS 1 (2*) 

NECROSIS, NOS 1 (4%) 

*SUBCUT TISSUE (46) (25) (49 ) (49) LPIDEEMAL INCLUSION CYST 

INFLAMMATION, CHRONIC 1 (2X) NECSOSIS, NOS 1 (2«) KELOIE 1 (2%) "IBROUS DYSPLASIA 1 (2*) 

BESPIRATORY SYSTEM
 

"1ASYNX (46 ) (25) (49) (49) INFLAMMATION ACUTE ANE CHRCSIC 1 (4X) INFLAMMATION, CHPONIC 7 (28X) 

ITSACHEA (45) (11) (44) (49) 

:;NFLA«HATICN, scs TYMPHOCYTIC INELAMMATOEY INFIITF 3 (7*) 

INFLAMMATION, ACUTE/CTRONIC 24 (53%) 29 (66%) 3 (6%) 

INFLAMMATION, CHRONIC 26 (53X) 

*LUNG/EEONCHUS (45 ) (25) (48) (49) 

ERCNCHIECTASIS 2 (8S) 1 <2X) 4 (8*) 

INFLAMMATION, FCCAL 1 (4*) 

INFLAMMATION, ACUTE/CHECNIC 1 (2X) 

HYPEEPLASIB, LYMPKOID 2 (4%) 3 (6X) 

• LUtlG/BECKCHICL t (25) (48) (49) 

I DUMBE B CF ANIMAL S WIT H TISSUE EXAMINE D MICEOSCOP1C ALL Y 

* KU'IBE H CF AKIMAL S NECFOESIE D 

**EXCLUDES PARTIALLY AUTOLYZED ANIMALS 

C-3
 
TABLE Cl (CONTINUED)
 

LOW DOSE CCNTBOL(UNTR) 

01-0030 

HIGH DOSE CONTECL (CUTS)

01-0081

 LOB DOSE 

 01-0025 

HIGH DOSE 

01-0087 

LYMEHOCYTIC INF I A FBATCRY INFIIT5 

INFLAMMATION, ACUTE/CHRONIC 

HYPERPLASIA, LYMPHOIC 

5 

3 

6 

(11*) 

(7*) 

(13*) 

2 (1*) 

1 <2«) 

ILDHG 

EMPHYSEMA, NOS 

ATELECTASIS 

EDEMA, NOS 

HEMOBPHAGE 

INFLAMMATION, INTERSTITIAL 

FRONCHOPNEUMONIA, ACUTE 

AESCESS, NOS 

PNEUMONIA, CHRONIC HURINE 

GRANULOMA, NOS 

FIEROSIS, DIFFUSE 

rERIVASCULITIS 

HYPERPIASIS, NOS 

HYPERELASIA, ACENOMATOUS 

HYPERPIASIA, AIVECIAR EPITHEIIDK 

(15) 

1 

1 

q 

2 

q 

1 

1 

(2%) 

(2*) 

(9*) 

(1*) 

(11%) 

(2%) 

(2*) 

(25) 

2 

1 

1 

11 

1 

(8*)

(HX) 

(KX) 

(HKX) 

(1*) 

(18) 

1 (2*) 

1 (2*) 

1 (2*) 

3 (6X) 

(19) 

3 

1 

2 

2 

(6*) 

(2*) 

(IX) 

(IX) 

HUNG/ALVEOLI 

CALCIFICATION, SCS 

(15) (25) (18) (19) 

1 (2*) 

HEBA-OPOIETIC SYSTEM 

«EONE flARRCW 

HEMORRHAGE 

KARYORRHF.XIS 

STBOPHY, NOS 

HYELOFIEPOSIS 

HYPERPLASI8, HEMATOPOIET1C 

ESYTHROPOIESIS 

MYELOPOIFSIS 

(15) 

1

1

1

1

1

 (2X) 

 (2%) 

 (9*) 

 (2*) 

 (2*) 

(25 ) 

2 (8%) 

(11) 

1

1

1

 (2*) 

 (2%) 

 <2X ) 

(16) 

ISPtEEK 

CONGESTION, NOS 

HEMORRHAGE 

HEMATOMA, NOS 

FIEROSIS 

HEMOSICEROSIS 

ATROPHY, NOS 

HYEEPFLASIA, HEMAIOPOIETIC 

HYPEFPLASIB, ERYTHPOIE 

HYPERPLASIA, RETICULUH CELL 

(15) 

1

1

 (2%) 

 (2X ) 

(25 ) 

1

1

1

 (IX) 

 (UX) 

 (<1X) 

(18) (19) 

1

1

1

 (2X) 

 (2X) 

(«*) 

(2X) 

 <2X) 

I NUI1BER C F ANIMAL S WIT H TISSU E EXAMINE D MICROSCOPICALL Y 

* HU'IBEB OF ANI1AL S ((ECPOESIE D 

C-4 
HENATCFOIE5I S

 

KRYTHROPOIESI S 

MYEIOFOIESI S 

*SP'.E!1IC CAFSUL E 

HEUCPSHAG E 

ILYSF H NCDE 

-LASMACYTCSI S 

•EA50TID LYFF H NOt E 

EYPEBF1ASIA , KC S 

Hr.ANDIBUlA P L . UOC E 

1YMFHANGIEC1ASI S 

(FEEIASTINA L L.NOt E 

1YPEHANGIECTASI S 

HEKOPPHAG E 

UlttYie S 

HYPEPFIASIA , IFITHELIA L 

CIEC'JIATCP Y SYSTT"1 

• HEAR T 

OIBIAETFPITI S PERIVASCDIITIS 

ilRYOCAPDIO M 

INFLABfATICN , ECCJL 

ItJFIAKKATICN , IN1E5STITIAL 

ISFLAPHATION , SCUTE/CHBONI C 

7IERCSIS 

FIEROSIS , POCA 1 

~IESOSIS, DIFFUSE 

DEGENESA^ION , NOS 

CALCIFICATION , NO S 

• ENDOCARDIU M 

IHFlAHfATICN , ACUTE/CHRONI C 

*ACRT A 

MINEP.AIIZATIC N 

_ ISPLA^MATION^BCUTEZCHSONIC 

» NDMBE R O F ANIflAL S WIT H TISSU E

* KU1EER Cf AtilMALS NECPOFSIED 

TABLECl (CONTINUED) 

LOW DOSE HIGH DOSE CONTRCl(USTH ) CCHTBOl (CSTR) 

01-0030 01-0081 

1 (2«) 

1 (2%) 

(U5) (25) 

1 (2% ) 

(11) (21) 

1 (H% ) 

(41) (2t) 

(U1) (2U) 

(U1) (21) 

(35) (22) 

1 (3%) 

(U5 ) (25) 

1 (UX ) 

2 (<*S) 

(U5 ) (25) 

1 (2%) 

3 (7*) 

1 (2«) 

1 (U*) 

13 (29%) 10 (U0%) 

(US) (25) 

(16) (25) 

 EXAMINE D MICROSCOPICALL Y 

LOS DOSE 

01-0025 

1 (2«) 

(i»8) 

(3V) 

(37) 

1 <3X) 

(37) 

2 (5X) 

(31) 

1 (3X) 

1 (3X) 

(26) 

(48) 

(48) 

2 (4X) 

2 (UX) 

2 (4X) 

1 (2X) 2 (4*) 

15 (31X) 

3 (6X) 

(48) 

1 (2X) 

(49) 

1 (2%) 

1_J2X1__ 

HIGH DOSE 

01-0087 

1 <2X ) 

1 (2X) 

(19) 

(44) 

(44) 

(44) 

(44) 

(34) 

(49) 

(49) 

2 (4X) 

2 <4X) 

1 (2X) 

1 <2X) 

(49) 

(49) 

C-5 
TABLE Cl (CONTINUED) 

1EDI8I CALCIFICATION
 

CALCIFICATION, FOCAL
 

*C030NABY ABTFBY
 

INFLAMMATION, ACUTE/CHRONIC
 

*FUI«ONABY APTF.EY
 

1EDIAI CALCIFICATION
 

*ANTEBIOE MEDIASTINAL
 

MEDIAL CALCIFICATION
 

'SPLENIC ARTEFY
 

1EDIAI CALCIFICATION
 

tSESSNTEBIC ABTFEY
 

MEDIAL CALCIFICATION
 

»BENAL AETEPY
 

HYPEBPLASIA, FOCAL
 

DIGESTIVE SYSTEM
 

IPASOTID GLAND
 

INFLAMMATION, INTERSTITIAL
 

ISUEMAXILLAEY GLANE
 

HYEIEfLASIA, FCCAL
 

IIITEB
 

CONGESTION, NOS
 

CONGESTICN, CHEONIC PASSIVE
 

HEMOPEHAGE
 

INFLAMMATION, FOCAL
 

INFLAMMATION, ACUTE FOCAL
 

CHOLANGIOFIEEOSIS
 

PEEIVASCULITIS
 

CEGENEEATION, NOS
 

NECROSIS, FOCAL
 

NECEOSIS, DIFFUSE
 

METAMOEPHOSIS FATTY
 

HYPEEPLASIA, NODDIAR
 

HYPEBPIASIA, NOS
 

HYPEEPLASIA, FOCAI
 

tllVEE/CENTRILOBULBB
 

(iECBCSISj_NOS
 

I NUMBE E 01 ANIMAL S WIT H TISSUE

* tiUMBE B OF ANIMAL S KECPCESIE D 

LOW DOSE HIGH DOSE CCNTBOL(UNIH) CONTBCL (CNTR) LOH DOSE HIGH DOSE 01-0030 01-0081 01-0025 01-0087 

2 (IX) 

1 (1*) 

(16) (25) (19) (49) 

1 (2X) 

(«6) (25) (19) (49) 

1 (2X) 

(16) (25) (19) (19) 

1 <2X) 

(16) (25) (19) (49) 

1 (2X) 

(16) (25) (19) (49) 

1 <2X) 

(16) (25) (19) (49) 

1 (2X) 

(13) 

1 (2t) 

(21) (13) (48) 

(13) (21) (13) (48) 

1 <2X) 

(15) (25) (49) (48) 

1 (2*) 

1 (<»*> 1 (2X> 2 (US) 

2 (1*) 1 (2*) 

1 (itX) 

1 (2X) 

2 (IX) 

1 (2*) 1 (1%) 1 (2X) 

1 <2X) 

3 <7«) « (16*) 21 (43%) 12 (25%) 

4 (8X) 

1 (2*) 

8 (13*) 25 (51X) 2 (IX) 

(15) (25) (19) (48) 

1 (2X1 

 EXJMINE D MICEOSCOPICftLL Y 

C-6 
TABLE Cl (CONTINUED) 

LOW DOSE 

CONTRO L (USTB) 

01-0030 

HIGH DOSE CCNTBOL (UNTE ) 

01-0081 

(•ET»MOBPHOSI £ FATT Y 1 (2%) 

niVEB/HEEATCCYTE S 

HYPHFFIASIA, FCCAL 

(US ) 

6 (13*) 

(25) 

*EILE EUC T 

INFLAMMATION , NCS 

HYPERPLASIA , MOS 

HYPERP1ASIA , FOCJI 

(tt6) 

5 (11X) 

11 (24*) 

(25) 

6 (2H*) 

•PANCREA S 

INFLAMMATION , tiCS 

INFIAMPATICN , INTF. BSTITIAL 

INFLAMMATION , ACUTE/CHRONIC 

SCAH 

EEHIABTEBITI S 

METAMORPHOSI S FATTY 

HYPERPLASTIC NODOLI 

(US) 

1

1

2

 (2*) 

 (2)S) 

 (US) 

(25) 

1 (4*) 

IFANC8EATI C DUCT 

HYPERP1ASIA, DCS 

(45 ) (25) 

1 («*) 

UtANCBIATIC ACINUS 

ATBCPHY, NCS 

ATROPHY, FCCAl 

13 (29X) 

2 (4%) 

(25) 

• ESOPHAGUS 

INFLAMfATICN, ACUTE FOCAL 1 (2*) 

(25) 

t=TC«ACH IPICEFHAI INCLDSICN CYST 

BYPHRKERATCSIS 

(45) 

1 (2%) 

(24) 

1 (IX) 

•GASTBIC CUCCSA CALCIFICATICN, DCS (15) (21) 

IGASTRIC MUSCOLAPIS CALCIFICATION MCS 

(45) (24) 

•SMALL INTESTINE 

HYPIRFIASIA, 1YCEHCIC 

(45) (21) 

•EEYHFS FATCH 

HYPEREIASIA, NCS HYPEEPIASIA, LYMIHOID 

(45) 

1 (2X) 

(21) 

2 (8X) 

IILIUM (45) (24) 

* NUMBE R C5 ANIMAL S WITH TISSUE EXIMINED MICROSCOPICA1LY
 

* NUMBE B CF ANIMA1S HICFCESIEP
 

C-7
 

LOS DOSE 

01-0025 

(U9) 

(U9 ) 

3 (6*) 

7 (1UX) 

7 (1UX) 

(15) 

1 (2X) 

5 (11*) 

2 (1*) 

1 (2*) 

(15) 

(15)
 

(42)
 

(45)
 

(45)
 

(15)
 

(45)
 

1 (2*)
 

(45)
 

(45)
 

.__J_J2X1_­

HIGH DOSE 

01-0087 

C»8) 

(19) 

(18) 

1 (2*) 

1 (2*) 

1 (2*) 

(18) 

(48)
 

(49)
 

(19)
 

(49)
 

2 (IX)
 

(19)
 

2 (4*)
 

(19)
 

(19)
 

(49)
 
TABLE Cl (CONTINUED) 

LOW DOSE HIGH DOSE CONTRO L (UNTR ) CONTFOL (DNTR ) 

01-0030 01-0081 

•COLON	 (UK) (214)
 

PARASITISM
 

URINARY SYSTBM
 

IKICNEY (15) (21)
 HYDRONEPHROSIS
 

CONGESTION, NOS 1 (2X)
 

GLOMEROLONEPHBITIS, NOS 31 (76*) 5 (21*)
 GLOMERULONEPHPITIS, FOCAL
 

INFLAMMATION, INTERSTITIAL 5 (11«)
 GLOMEEULONEPHRITIS, CHRONIC
 

NEPHROPATHY 1 (US)
 NEPHSOSIS, NOS 16 (67%)
 

GLOMEEULOSCLEHOSIS, NOS
 

CALCIFICATION, NOS
 

IKIDNEY/rEDULLA (U5) (24)
 MULTIPLE CYSTS 1 (2%)
 

IKIENEY/GLOMEFULUE («5) (21)
 INELAMflATICN, fECERANOU S 9 (20X)
 

•KICNEY/TUBULE	 (U 5) (24)
 CALCIFICATION, NCS
 

•UFINAEY	 ELACCEP (1*14) (23)
 

CALCUIUS, NCS 3 (13*)
 INFIAKBATICN, ACUTE/CHRONIC 1 (2%)
 

HYPERPIASIS, EPITKELIAL
 

ENDOCRINE SYSTH"
 

•PITUITARY	 (4U) (21)
 

CYST, NOS
 

CONGESTION, NOS 1 (2%)
 

HEMORRHAGE
 

HYPEPPLASIA, POCAI 6 (14X)
 

IPITUITABY/BASOPHII (44) (21)
 

NODULE 1 (5%)
 

(45) (25)
 

I NUMBE 9 CF ANIMALS WITH TISSUE EXJBINED MICEOSCOPICAIIY
 

* 1.UMBER CF ANIMAL S NECECFSIED
 

LOS DOSE 

01-0025 

(43)
 

4 (9%)
 

(48)
 

37 (77X)
 

2 (it*)
 

1 <2X)
 

1 (2X)
 

(48)
 

(U9)
 

4 (8*)
 

(48)
 

(44)
 

1 (2*)
 

(44)
 

1 (2X)
 

1 (2%)
 

5 (11*)
 

(44)
 

(46)
 

HIGH DOSE
 

01-0087
 

(46)
 

1 <2X)
 

(49)
 

1 (2*)
 

5	 (10*)
 

43 (88%)
 

1 (2*)
 

(49)
 

(49)
 

(49)
 

1 <2X)
 

(49)
 

(35)
 

2 (6*)
 

(35)
 

(45)
 

C-8
 
TABLEC1 (CONTINUED) 

LOW DOSE CCNTRCL(UNTB )

01-0030

 HIGH DOSE CONTBOL (tNTE )

 01-0081

HYPERELAEIA , FOCAL 

• ADRENAL CCFTEX

NODULI

METAMORPHOSIS FATTY 

HYPERTROPHY, FOCAL 

HYPERPLASIA, NOEULAP

HYPERPLASIA, NOS 

HYPERPLASIA, FOCAL

 (U5) 

1 (2%) 

1(2%) 

 7. (16%) 

(25) 

1 (IX) 

•ADFENAL MECt'LLA

HYFEPFLASIA, SCS

HYPEEPLASIA, FCCAL

ANGIECTASIS 

 (15) 

2 (1%) 

4 (9%) 

(25) 

•THYBOID

THYSOGLCSSAL DUCT CYST 

HYPEEP1ASIA, FCCAL

HYPEEPLASIA, C-CEIL

HYPERPLASIA, EOLLICULAR-CELL 

 (12) 

2 (5X) 

1 (2X) 

(23) 

IEANCEEATIC ISLETS 

HYPEEELASIA, 

HYPEEPLASIA, 

NCS 

FCCAL 

(15) 

2 (IX) 

(25) 

REPRODUCTIVE SYSTEM
 

*MAMMARY GLAHC (25)
 

CYST, NOS
 

HYPERPLASIA, NOS 3 (1%) 3 (12X)
 

LACTATION 7 (28X)
 

*EEIPUTIAI GLAND (16) (25)
 

ABSCESS, SOS 2 (US)
 

JEEOSTATE (15) (23)
 

INILABeATICN, BOS 1 (IX)
 

INFLAMMATION, ECCAL 1 (2%)
 

INFLAMMATION, ACUTE 10 (22*)
 

INFLAMMATION, SCUTE FOCAL U (9%)
 

INFLAMMATION, ACUTE/CHSONIC
 

DEGENEEATION, NOS 13 (29S)
 

ATROPHY, NOS 2 (IX) 1 (17X)
 

HYPEEPLASIA, FCCAL
 

I NUMBER CF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER Of ANIMALS NECEOSSIED
 

 LOB DOSE 

 01-0025 

(16)
 

2 (IX)
 

1 <2X)
 

3 (7X)
 

(16)
 

2 (IX)
 

(11)
 

1 (2X)
 

1 <2X)
 

(15)
 

1 (2%)
 

(19)
 

1 (2X)
 

1 (2X)
 

(19)
 

(15)
 

5 (11X)
 

11 (31X)
 

1 <2X)
 

1 (2X)
 

HIGH DOSE
 01-0087
 

<2X) 

(15) 

(15) 

(17)
 

1 (2X)
 

<2X) 

(18)
 

1 (2X)
 

1 (2K)
 

(19)
 

(18)
 

1 (2X)
 

C-9
 
TABLECl (CONTINUED) 

HYFEBELASIA, AIENCflATOUS
 

METAPLASIA, SQUAHOUS
 

*SE1INAL VESICLF
 

ATSOPHY, NOS
 

HYPERPLASIA, PAPILLARY
 

•COAGULATING GLAND
 

ATROPHY, NCS
 

(ITESTIS
 

EEGINEPATICN, NCS
 

CALCIFICATION, FOCAL
 

ATROPHY, NOS
 

HYPERPLASIA, INTERSTITIAL CEIL
 

ITESTIS/TUBUIE
 

DEGENERATICN, NCS
 

NERVOUS SYSTEM
 

IBRAIN
 

HEMCRFHAGE
 

CALCIFICATION, FOCAL
 

ICEBEBRAL CCRTEX
 

HEMORRHAGE
 

MALACTA
 

SPECIAL SENSE ORGANS
 

»EYE
 

SYNECHIA, POSTERIOR
 

CATARACT
 

MUSCULOSKELETAL SYSTEM
 

*SKELETAI MUSCLE
 

CALCIFICATION, FOCAL
 

BODY CAVITIES
 

NCN E
 

LOW DOSE
 CCfiTSOL(t)NTR)
 

01-0030
 

1 (2X)
 

(46)
 

26 (57*)
 

1 (2%)
 

(46)
 

3 (7%)
 

(45)
 

1 (2*)
 

(45)
 10 (22*)
 

(45)
 

(45)
 

1 <2X)
 

1 (2*)
 

(46)
 

1 (2%)
 

1 (2«)
 

(46)
 

HIGH~DOSE~~
 CONTEOL (CNTE
 

01-0084
 

(25)
 

1 (4%)
 

(25)
 

(24)
 

4 (17*)
 

12 (50*)
 

2 (8X)
 

(24)
 

(25)
 

2 (8*)
 

1 (4X)
 

(25)
 

(25)
 

(25)
 

1 (4*)
 

) LOU DOSE
 

01-0025
 

1 (2*)
 

(49)
 

1 <2X)
 

(49)
 

(46)
 

32 (70*)
 

2 (4*)
 

(46)
 

3 (7X)
 

(45)
 

(45)
 

(49)
 

(49)
 

HIGH DOSE
 

01-0087
 

(49)
 

(49)
 

(49)
 

5 (10X)
 

(49)
 

2 (4*)
 

(49)
 

(49)
 

(49)
 

(49)
 

* JiUMBEH CF ANIMALS WITH 1ISSUE EXAMINED BICEOSCOPICAILY
 

* BU»BEB CI AKIMALS NECEOFSIED
 

c-io
 
TABLE Cl (CONCLUDED) 

LOW DOSE HIGH DOSE CCNTBOL(UNTP) CONTBOL(CHTR) LQ» DOSE HIGH DOSE

 

01-0030 01-008U 01-0025 01-0087 

All OTHEP, 

ACIEOSE TISSUE
 

INFLAMMATION, ACUTE/CHRONIC
 

SPECIAL MOP-PHOLOGY SUPIMARY
 

AOTC/UICBCESY/HISIO PEHE 1
 

HUTC/UCBCESY/RC HISTO 1
 

ADTOLYSIS/NO NECBCPSY 1) 1
 

• HUKEES CF ANIMALS WITH IISSUE FXABINED HICBOSCOPICAIL?
 

* »U»!BEB OF ANIMALS NECPCPSIED
 

C-ll
 
TABLE C2

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN FEMALE RATS 

TREATED WITH 2.4-DINITROTOLUENE 

LOW DOSE HIGH DOSE CONTROL (UNTK ) CONTBCL (ONTE) LOB DOSE HIGH DOSE 

02-0030 02-0081) 02-0025 02-0087 

iSIMAL S IKITISLL Y I S STUD Y 50 25 50 50 

ANIMAL S NECRCFSIE D 48 23 49 50 

ANIMAL S EXAMINE D HISTOFATHOLOGICAII Y ** 47 23 49 50 

INTEGUMENTARY SYSTJM
 

*£UECUT TISSUE (18) (23) (49) (50)
 COLLAGEN DISEASE 1 (2%)
 

RESPIRATOR Y SYSTE M 

*NASAL SEPTUM (48) (23) (U9) (50) 

INFLAMMATION, CHRONIC 1 (2%) 

«IA1YNX	 (48) (23) (49 ) (50) 

INFLAMMATIO N ACUT E AN D CHRONIC 1 (4X) 

INFLAMMATION , CHRONI C 3 (13%) 

• (TRACHE A (U7 ) (5) (48) (50) 

LYMPHCCYTIC INFLAMMATORY INFIL1R 4 (9«) 4 (8X) INFLAMMATION, ACUTE/CHRONIC 18 (38X) 25 (52%) 1 (2%) 

INFLAMMATION, CHRONIC 20 (40%) 

ECLYE, INFLAMMATOR Y 1 (2« ) 

• IUNG/ERCNCHU S (47) (23) (49) (50) 

BRONCHIECTASIS 2 (4%)

 

INFLA^HATICN, ACUTE fOCAL 1 (2%)

 FIEROSIS 1 (2%)

 

tlUNG/EECNCHIOLF . (47) (23) (49) (50)

 

INFLAMMATION, KCS 4 (9%)

 

LYMPHOCYTIC INFLAMMATOR Y INFILTP 7 (15t) 4 (8%)

 

INFLAMMATION, JCUTE/C BRONIC 5 (11X) 

HYPEEPLASIA, LYMPHOID 2 (4%)

 

HUNG (U7 ) (23) (49) (50)

 

CONGESTION, NCS 2 (4%)

 

INFLAMMATION , FCCAL 1 (2% )

 

I NU1BE R CF ANIMAL S WIT H TISSUE EXJMINE D MICROSCOPICALL Y 

* NUMBE B CE ANIMAL S NECBOESIE D 

"EXCLUDES PARTIALLY AUTOLYZED ANIMALS 

C-12 
EFCNCHOPNEUMOUIA , ACUT E
 

PNEUMONIA, CHRONIC KUBINE
 

GBANULOMA , FOREIGN BODY
 

PERIVASCULITIS
 

CALCIFICATION, FOCAI
 

HYPERPLASIA, EPITHELIAL
 

HYPERPLASIA, ADF.NCMP.TOUS
 

HIUN3/ALVECLI
 

EETTHEIIAIIZATICK
 

HEMATOPCIETIC SYSIEf
 

(tEONE CAFECW
 

HYPOPIASIA, NOS
 

OSTEOSCLEBCSIS
 

HYEEEELASIA, B EM ATOPO IET 1C
 

((SPLEEN
 

CONGESTION, NCS
 

HEKATOPA, NOS
 

HEMOSIDEEOSIS
 

ATROPHY, NOS
 

HYEEFELASIA, H JM A1OPO IET 1C
 

HYPEFPLASIA, EHYTKEOIC
 

HEMATOPOIESIS
 

S3YTHF.OPOIESIS
 

IMANDIBULAR I. NODE
 

HYPFPFLASIA, NCS
 

•EANCREATIC	 L.NCCE
 

HYPEEEIASIA, NCS
 

KfESENIEFIC L. NOCE
 

HYPEPEIASIA, NCS
 

•THYMUS
 

HYPERFLASIA, FETICU1UM CELL
 

CIPCIILATOE J SYSTE M 

»HIA?T
 

PEEJAFTEEITIS__
 

« NUMBE R C E ANIMAL S WIT H TISSUE

* NUMBE R CI AtilfiAL S NECFCESIE D 

TABLE C2 (CONTINUED) 

LOW DOSE HIGH DOSE
 CONTROL (UNTR) COS1FCL (CNTB)


02-0030 02-0081


8 (35*)
 

1 (IX)
 15 (32%)
 

1 (IX)
 

1 (IX)
 

(HI) (23)
 1 (2%)
 

(H->) (22)
 1 (2%)
 

1 (2%) 1 (5X)
 

(U7) (23)
 1 (2%)
 

1 (IX)
 

2 <9X)
 

3 (13X)
 

1 (17X)
 2 (U*) 3 (13X)
 

(12) (21)


(12) (21)


(«2) (21)


(36) (20)


(HI) (23) 

 ZXJMIHE D MICROSCOPICALL Y 

C-13 

 LOB DOSE
 

 02-0025
 

1 (8X)
 

 (19)
 

 (18)
 

 (19)
 

 (11)
 

 (11)
 

1 (2X)
 

 (11)
 

 (27)
 

1 (IX)
 

(19) 

HIGH DOSE
 

02-0087
 

1 (2*)
 

1 <2X)
 

(50)
 

(15)
 

1 (2X)
 

(50)
 

1 (2X)
 

1 (2X)
 

1 <2X)
 

2 (IX)
 

(17)
 

1 (2%)
 

(17)
 

(17)


1 (2X)
 

(32)
 

(19) 
TABLE C2 (CONTINUED)
 

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

02-0030 02-0080 

?EBIVA£CUII1IS 1 (2%) HYPERTROPHY, NOS 1 (2%) 

*HYOCABCIUM (1*7) (23) INFLAMMATION, NOS 

INFLAMMATION, FCCAL 

INFLAMMATION, INTERSTITIAL 

INFLAMMATION, ACUTE/CHRONIC 

1 (2X) 

2 (4%) 

1 (ux) 

FIEROSIS, DIFFUSE 

DEGENERATION, NOS 2 («X) i) (17%) 

KINCOCAF.EIUM (17) (23) 

INFLAMSATICN, AC'JTE/CHBONIC 3 (6X) 

ICAFDIAC VALV J (17) (23) 

INFLAMMATION, ACOIF/CHBONIC 

*EENAL ABTEBY (18) (23) 

THROMBOSIS, NCS 

DIGESTIVE SYSTEM
 

HIVES (17) (23)
 CONGESTION, CHSCNIC PASSIVE 1 (IX)
 HEMOBBHAGS 1 (2X)
 INFLAMMATION, FOCAL 1 (2X)
 INFLAMMATION, CfRONIC DIFFUSE
 

SCLEROSIS 1 (2%)
 CHOLANGIOF1ESOSIS 1 (4X)
 PERIVASCULITIS 1 <2X)
 NECROSIS, FOCAL 1 (2%)
 NECROSIS, COAGULATIVE 1 (2X)
 METAMORPHOSIS FATTY U (9X) 2 (9X)
 BASOPHILIC CYTC CfJNGE 1 (17X)
 HYPERTROPHY, NOS 1 (2%)
 

HYPERPLASIA, MODULA R
 

HYPERPLASIA, NOS 1 (2%)
 HYPERPLASIA, FOCAL 21 (U5X) 3 (13X)
 

HYPERPLASIA, DIFFUSE 1 (2X)
 HYPERPLASIA, RETICOLOt! CELL
 

•IIVER/HFFATCCYTES	 (17) (23)
 

HYPERPLASIA, FCCAL 2 (4X)
 

»EILE EUC1 (18) (23)
 

INFLA«BATICN.t_SCS . - ­

* DUMBER CF ANIMALS WITH TISSUE EXAMINE D MICROSCOPICALLY
 

* BUWBER Cf ANIMALS NECPOSSIED
 

C-14
 

LOB DOSE
 

02-0025
 

(19)
 

1 (2X)
 

1 (2X)
 

5 (10X)
 

(t (8X)
 

(19)
 

(19)
 

1 <2X)
 

(19)
 

1 (2X)
 

(19)
 

1 (2X)
 

1 (2X)
 

8 (16X)
 

H (8X)
 

33 (67%)
 

1 <2X)
 

(19)
 

(19)
 

i_J2XJ .
 

HIGH DOSE
 

02-0087
 

(19)
 

2 <1X)
 

3 (6%)
 

(19)
 

(19)
 

(50)
 

(50)
 

1 <2X)
 

U (8X)
 

6 (12*)
 

(50)
 

(50)
 

1 (2X)_
 
INFLAMPATICN, ACUTE/CHRONIC
 

INFLAMMATION WITH FIBROSIS
 

HYPERPLASIA, NOS
 

HYPERPLASIA, FOCAL
 

IEAMCREAS
 

INFLAMMATION, INTERSTITIAL
 

"NFLAMBATICN, ACUTE/CHRONIC
 

ATROPHY, NOS
 

IFANCREATIC ACINUS
 

EEGENEEATICN, GPABULAR
 

ATROPHY, NCS
 

ATEOPHY, FOCAL
 

IST01ACH
 

INFLAMMATION , ACUTE FOCAL
 

ISMilLL INTESTINE
 

3YPERELASIA, IYPEHCID
 

HEEYESS PATCH
 

HYPERELASIA, NCS
 

tICCLCN
 

UICEP, ECCAI
 

PARASITISM
 

URINAPY SYSTEM
 

tKICSEY
 

GLCMF.PULCNEEHSITIS, NOS
 

'SLOMEEULONFPHPITIS, FCCAL
 

INFLAMMATION, INTERSTITIAL
 

PYELONEPHRITIS, ACOTE
 

PYELONEPHRITIS, CHRONIC
 

HEPHROSIS, NOS
 

POSTMORTEM CHANG E
 

CALCIFICATION, FOCAL
 

IPID1EY/CCRTEX
 

CYST, NOS
 

IKIESEY/GLOflERUL'JS
 

* NUMBE R C E ANIMAL S WIT H IISSUF

* liU«BEH C! ANIMAL S NECBOESIED 

TABLE C2 (CONTINUED) 

LOW DOSE HIGH DOSE
 CCNTBOL (UNTR) CONTEOL (tNTR)
 

02-0030 02-00814
 

1 <2X)
 

1 (2%)
 1 (8X) 2 (9X)
 

(146)	 (22)
 2 (UX)
 

1 (2X)
 

(146)	 (22)
 

1 (2%)
 

14 (9%)
 

1 (2%)
 

(146) (23)
 

(147)	 (23)
 1 (2%)
 

(147)	 (23)
 

14 (17%)
 

(146) (22)
 

1 (251)
 

2 (9%)
 

(147)	 (23)
 

32 (68%) •4 (17%)
 

2 («*)
 1 (14*)
 

1 (14%)
 

10 (143%)
 

1	 (14%)
 

(147)	 (23)
 

1 (2X)
 

(147) (23)
 

 FXSMINE D BICROSCOP1C ALL Y 

C-15 

LOW DOSE
 

02-0025
 

7 (1<4%)
 

7 (114%)
 

(147)
 

2 (U%)
 

5 (11%)
 

(147)
 

(49)


1 (2%)
 

(147)
 

2 (14%)
 

(147)
 

(146)
 

14 (9%)
 

(149)
 

19	 (39%)
 

3 (6%)
 

2 (14%)
 

1 (2%)
 

(149)
 

(U9)
 

HIGH DOSE
 

02-0087
 

1 (2%)
 

(148)
 

(148)
 

(148)
 

(50)
 

(50)
 

(149)
 

2 (14%)
 

(U9)
 

140 (82%)
 

(<49)
 

(19)
 
TABLE C2 (CONTINUED) 

LOW DOSE HIGH DOSE 

CCNTBOL(UNTR ) CONTROL<CNTR )

02-0030 02-0081

IKIDHEY/TUBUL E (47) (23)
 

NECROSIS, DOS 1 <<!*)
 

INDOCBINE SYSTEM
 

»EITUITARY (46) (21)
 

CYST, NOS'
 

HEMOBRHAGIC CYST 1 <5X)
 

HECSOSIS, FOCAI 1 (2*)
 

HYPERTROPHY, FOCBI 1 (2X)
 

HYPERPLASIA, FOCAI 6 (13%) 1 (5*)
 

IACPENAL (47) (23)
 

METAMORPHOSIS FATTY 1 (2*)
 

HYPERPIASIA, FOCAI 1 (2*)
 

IACSENAI. COP1EX (47) (23)
 

HEMORRHAG E 2 (4»)
 

NODULE 1 (9J)
 

DEGENEHATION, NOS 2 (U%)
 

NECROSIS, FOCAI 1 (2%)
 

METAMORPHOSIS FATTY 1 (2S)
 

PIGMENTATION, DOS 1 (2S)
 

HYPEPPIASIA, NOCUtAP 1 (2%)
 

HYPERPIASIA, PCCAI 9 (19S)
 

JJEFENA L BEDULLA (47) (23)
 

EYFFRPIASIA, NCDU1AR 1 (2*)
 

HYPEPPLAEIA, FCCAI 2 (!»«)
 

• THYROID (15) (21)
 

1YBFHCCYTIC INFI Af,1ATOHY INPUTS
 

HYPERPIASIA, FCCAI 3 (7X)
 

HYPERPLBSIA, C-CEH 1 (2%) 3 (11*)
 

REPRODUCTIVE SYSTEM
 

*«AMMAR Y GLAND (46) (23)
 

GALBCTOCELE 6 (13S) 1 («X)
 

MUITIPIE CYSTS
 

INFLAMMATION, AC01E
 

HYPERPIASIA, NOS 1 (2*) (4*)
 

I NUMBE R CF ANIMALS WITH TISSUK EXJMINED MICROSCOPICALLY
 

* NUMBE R CF ANIMAIS NECPOESIED
 

C-I6
 

 LOK DOSE 

 02-0025 

(49)
 

(45)
 

2 (IX)
 

1 (2%)
 

(49)
 

2 (4*)
 

(49)
 

2 (4%)
 

1 (2%)
 

(19)
 

(45)
 

1 (2%)
 

(49)
 

9 (18X)
 

1 (2X)
 

12 (24*)
 

HIG H DOSE 

02-0087 

(40)
 

(50)
 

(50)
 

(50)
 

(48)
 

1 (2*)
 

(50)
 

1 (2X)
 
TABLE C2 (CONTINUED) 

LOW DOSE CCNTSO L (OMIE )

02-003P

 HIGH DOSE CONTBOL ( CN1E) 

 02-0081 LOS DOSE 02-0025 

HIGH DOSE 

02-0087 

IACTATIC N 9 (39%) 

+CIITCPAI GIAND 

ABSCESS, SOS 

t-YPEHPLASIA, PAPIIIAP.Y 

1 (2*) 

(23) 

1 (2X) 

(50) 

*VAGIN A 

IOLYP 

(18) (23) (50) 

H U T >FU S 

F.YCSCM'Tt A 

INFLAM»ATTON , NCS 

TYOMETF A 

INFLAMMATION , ACUTE 

FIBBOSIS 

F.YFEEFlAfIA, AtENCMATOU S 

(U6) 

2 («) 

(23) 

3 (13X) 

2 (IX) 3 (6X) 

1 <2X) 

1 <2X) 

1 (2X) 

1 (2X) 

2 (IX) 

(njTEHUS/ENDCMET" 

CYST, NOE 

INFLAMMATION , 

INFLAMMATION , 

INFLAMMATION , 

INFLAMMATION, 

INFLAMMATION , 

INFLAMMATIO N , 

FYPEFTHOFFY, 

FYPEP, PLASIA , 

HYPEF.PLASIA, 

BYPEPPLASIA, 

IUM 

NCS 

SHPFUBATIV E 

ACCT E 

ACUTE FOCAL 

ACUTE/CHRONIC 

CHPCNIC 

NOS 

NOS 

FOCAI 

CYSTIC 

(K6) 

6 

1 

1 

1 

7 

(13*) 

(2%) 

(2*) 

(2%) 

(1S%) 

(23) 

1 (IX ) 

1 (IX) 

1 (IX) 

1 (IX) 

(17) 

3 

20 

1 

1 

7 

(6X) 

(13X) 

(2X) 

<2X) 

OSX) 

(19) 

7 (11X) 

t <ex) 

2 (»X) 

•CVAHY/CVIDUCT 

SETENTION Fill 

3NFLAMCATTCN , 

INFLAMMATION , 

I, ESCESS, NOS 

IF 

SUEEUEATIVF, 

ACUTE 

(16) 

1 

1 

1 

(2%) 

(2X) 

(2%) 

(23) 

1 (1%) 

1 (UX) 

(17) (19) 

tCVAB Y 

CYST, NCS 

FAEOVAPIP N CY 

HYPERPLASIA , 

ST 

NOS 

(U7) 

14 

1 

(9%) 

<2X) 

(22) 

3 (11X) 

(17) 

1 

1 (2X) 

(2X) 

(19) 

1 (2X) 

NEFVOU S SYSTEM 

iff!. IN (17) (18) (18) 

« NUM3E?, Of ANIMALS WITH TISSUE EXAMINED MI CBOSCOP ICALLY 

* KOMBE F OF AMMAL ? NECECFSIED 

C-17
 
TABLE C2 (CONCLUDED) 

LOW DOSE CCNTaCL(UNTR )

02-0030

 HIGH DOSE CCNTFOL (CNTR )

 02-0081

HEMCPFHAG I

CALCIFICATION , FOCAI

 1

 1

 (IX) 

 (IX) 

SPECIAL SENSE ORGAN S 

*EY E (16) (23)

CATARAC T 1 (2%)

*IYF/CCPNEA (US) (23)


TNFLAMMATICN, INTERSTITIAL


*EYE/RETINA (18) (23)


EEGENERATION, NOS 1 (2S)


MUSCULOSKEIETAL SYSTEf!
 

NCNE
 

BOCK CAVITIES
 

*ELE0B A (18) (23)


INFLAKBATICN, ACUTE/CHRONIC


HYPERPIASIJ, ADENCMATOUS


ALI OTHER SYSTEMS
 

*MULTIPLE ORGANS (18) (23)


ICSIMOFTEM CHANGI


SPECIAL MORPHOLOGY SUMMAR Y
 

AOTC/NECEOE£Y/NO HISTO 1
 

AUTOLYSIS/SC HECRCESY 2 2


» MIHBEB CE ANIMAL S KIIH 1ISSDE EJIJKINED KICPOSCOPICAIIY
 

* SUMBEF CE ANIMALS KECPCESIED
 

C-18
 

 LOS DOSE

 02-0025

 HIGH DOSE 

 02-0087 

 (19)

 1 (2% ) 

 (19)

 1 (2X) 

 (19)

 1 (2X) 

 (50) 

 (50) 

 (50) 

 (19)

 1 (2X) 

 (50) 

1 (2*) 

 (19)

 1 (2X) 

 (50) 

1 
APPENDIX D
 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC
 

LESIONS IN MICE TREATED WITH 2,4-DINITROTOLUENE
 

TABLE Dl

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN MALE MICE TREATED WITH 2,4-DINITROTOLUENE 

c LOW DOSE :OSTROL(UNTE)

05-0030

 HIGH DOSE CONTROL (UHTB)

 05-0077

 LOW DOSE

 05-0025

 HIGH DOSE 

 05-0088 

• SltJALS INITIALLY IN STCE Y 

ANIMAL S NECBOFSISD 

ANIHALS EXAMINED HISTOPATHOLOG ICA ILY ** 

50

46

46

 50 46

 45

 50 48

 48

 50 49 

 49 

INIEGOMENTAS Y SYSTEM 

*SKIN 

GSANIJLOBA, PYOGFNIC 

(46)

1 (2X) 

 (46) (48) (49) 

RESPIRATORY SYSTEM 

f LUNG/ERCNCHOS 

INFLAMMATION , ECCSL 

(46)

1 (2%) 

 (45) (48) (48) 

• LUNG 

F1FHYSBMA , NCS 

EDEMA , NOS 

HEMQRRHBG E 

INFLAMMATION , INTERSTITIAL 

5BTEEIOSCLESQSIS, NOS 

(46)

1 (2%) 

1 (21!) 

7 (15%) 

 (45)

1 <2X) 

 (48)

1 (2X) 

 (48) 

•lUNG/ALVFCLT 

INFLAMMATION , NCS 

(46)

1 <2X) 

 (45) (48) (48) 

H Ef ATOPOIFTIC SYSTEC 

ISFIEEK 

FIBROSIS 

HYPEBFLASIA , HFMATOEOI1TIC 

HYPEPPIASIS, EFYTHSOID 

HYPERPLASIA, RETICOLU1 CELL 

l!EM ATOPOIESIS 

(46)

1 (2%) 

1 (2%)

 (45)

1 (2X) 

3 (7%) I (2X)

 (47)

1 (2X) 

 (44) 

1 <2X) 

IIY^PH NCEE 

HEMOPEHAG E 

HYPEEPL1SIA, NOS 

(34)

1 <3X) 

1 <3X) 

 (35) (40) (34) 

<3X) 

•BANDIEULA E L. NODE 

HYPEPFIASIAi _HITICULUM_CELL_ 

(34 ) (35 ) (40) (34) 

I (iUIBE E ~.f ANirAL S WIT H TISSUE EXAMINE D

* BU1BE R C E AKIHAL S NECEOESIE D 

**EXCLUDES PARTIALLY AUTOLYZED ANIMALS 

 MICROSCOPICALL Y 

D-3 
TABLE Dl (CONTINUED) 

LOW DOSE 

CCNTBOL(ONTB ) 

05-0030 

HIGH DOSE COH1BOL (CNTE ) 

05-0077 

LOW DOSE 

05-0025 

HIGH DOSE 

05-0088 

IMESENTEEIC L. 

THBCMBOSIS , 

CONGESTION , 

HEMOBEHAG S 

HYPEBPLASIA , 

HYFEBFLAJIA , 

HYFEEELASIA , 

HYPEEPLSSIA , 

NODE 

N OS 

NCS 

NOS 

PIASM A CELL 

BETICULU M CELL 

LYMPHOI D 

(31) 

1

1

 (3*) 

 (3*) 

(35) (10) 

1 (3X) 

1 (3*) 

(31) 

1 (3*) 

2 <6X) 

1 (3X) 

1 <3X > 

CIBC'JLATOBY SYSTEM 

IENDOCABDIUM 

INFLAMMATION PBCLIFE5ATIVE 1 (2*) 

(148) (17) 

DIGESTIVE SYSTE1 

ISALIVAEY GLAND 

FEBIVASCT1LITIS 

(37) 

5 (114%) 

(13) (ID (17) 

*LIVEB 

INFLAB5ATICN, SECBCTIZING 

DEGENEBATICN, NOS 

NECROSIS, FOCAI 

NECPOSIS, HEHOEEHAGIC 

METAMOBPHOSIS FATTY 

HYPFPPLASIA, NODUIAP, 

HYPEPPLASIA, NOS 

HYPEBPLASIA, FOCAI 

(146) 

1 (2%) 

2 (IS) 

1 (2%) 

8 (17X) 

1 (2%) 

1 (2%) 

(15) 

1 (2*) 

3 (7*) 

(17)

1 (2X) 1 (2*) 

 (18) 

2 (1%) 

«IIVFB/PEEIJCBTAL 

INFLAMMATION, NOS 

(16) (15) 

1 (2*) 

(17) (18) 

ILIVEH/KUPFFFE CELL 

HYPFPELASIA, NCS 

(146) (15) 

2 (14*) 

(147) («8) 

»EILF tUCl 

INFLA("«ATICN, HOS 

LYMPHOCYTIC ISFLABMATOEY INFIITF 

(16) 

1 (2%) 

(146) 

1 (2%) 

(18) (19) 

IPANCEEAS 

CILATATICH/D'JCTS 

INFLAMMATION, FCCAL 

_ INFLAMM£TICN.t_JNTERSTITIAL__ 

(1414) 

2 (5«) 

. 1 J2%1 . . 

(141) 

_

(16)

1

 _

 (2*) 

 — —

 (12) 

_ — 

»

«

 NUSEE E

 NUIBEF .

 CF

 Cf

 ANIflAL S

 ANIHAL P

 WIT H TISSUE

 NECEOFSIE D 

 EXAMINE D KICFOSCOP1C ALL Y 

D-4 
TABLE Dl (CONTINUED) 

LOW DOSE CCNTBOItUNTF) 

05-0030 

HIGH DOSE COKTP01 (CNTE)

05-0077

INFLAMMATION , AC U1F/CHRCKIC 

HYPEBPLASIA, FOCAL 2 (5X) 

•FANCBEATIC DUCT 

INFLAMMATION, ACUTE FOCAL 

(44) (44)

tEANCSFATIC ACINUS 

ATBOPHY, FCCAL 

HYPERPLASIA, FCCAL 

(44) 

1 (2%) 

(14)

•STOMACH 

INFLAMMATION , NCS 

INFLAMMATION, ACUTE 

HYPERPLASIA, NOS 

HYPEEFLASIA, EPITHELIAL 

HYPEEPLASIA, FOCAL 

HYPEBPLASIf, ADEHCMATOUS 

(45) 

2 (<»*) 

1 (2%) 

1 (2%) 1 (2%) 

2 (IX) 

(12)

1 (2%) 

•GASTEIC SEBCSA 

PEEIABTESITIS 

(45) (12)

m?UM 

ABSCESS, NOS 

(46) (13)

'J BINAR Y SYSTEM
 

IHCNEY (46) (45)
 CALCULUS, NOS 20 (44X)
 TLOMEBULONEPHRITIS, NOS 3 (7X)
 

INFLAMMATION, INTERSTITIAL 7 (15S) 5 (11X)
 INFLAMMATION, BCUTE/CHSONIC
 

INFLAMMATION, CHEONIC 1 (2%)
 PSFIVASCULITIS 2 <4X)
 ABTEPIOSCLEBOSTS, NOS 1 (2%)
 NEPHROSIS, NOS 1 (2S)
 HYPEEPLASIA, TUEUIAP CELL 2 (4X)
 METAPLASIA , OSSEOUS 1 (2X)
 

IKIENEY/TUBULE (46) (45)
 EEGENEBATICN, NCS 1 (2X)
 METAKOPPHOSIS FATTY 9 (20X)
 

• !<ICNEY/EELVIS (46) (45)


INFLAMMATION, ACUTE/CHRONIC 3 <7X)
 

IUBINAEY ELAECEF (16) (11)


EE.E1VASCULA?_CUFEING
 

I NU"BE E CF ANIMAL S WIT H TISSUE EXAMINE D MICROSCOPICALLY 

* fiOMBE P CF AlilMAL S NECBOPSIE D 

D-5 

 LOH DOSE
 

 05-0025
 

2 (IX)
 

 (46)
 

1 (2X)
 

 (16)
 

1 (2X)
 

 (15)
 

 (15)
 

 (45)
 

1 (2X)
 

 (48)
 

 16 (33X)
 

1 (2X)
 

 (18)
 

 (48)
 

11 (29X)
 

 (16)
 -


HIGH DOS!
 

05-0088
 

(42)
 

(12)
 

(45)
 

1 <2X)
 

(45)
 

1 (2%)
 

(45)
 

(48)
 

(18)
 

(48)
 

(45)


1

 1221
 
HYPERELASIA, EPITHI1IAI
 

ENDOCRINE SYSTEM
 

•PITUITARY
 

CYST, NOS
 

HYPFRPLASIA, FOCAL
 

OACPINA L
 

NF.CPOSIS, FCCAI
 

HYPERPLASIA, FCCAI
 

»BDB ENAL CORTEX
 

HYPEPPLASIA, NOS
 

BYPERPLASTA, ECCAI
 

«FARATHYECID
 

CYST, NOS
 

REPRODUCTIVE SYSTEK
 

"PREPUTIAL GLAND
 

INFLAMMATION, ACUTE
 

I FRTSTATE
 

BYPERFIASIft, EPITHELIAL
 

K1ESTIS
 

•HH'RAIIZATION
 

DEGENERATION, NCS
 

•TESrlS/TUBULE
 

EHGENEPATICN, NCS
 

«FPICI:YCIS
 NECROSIS, DOS
 

NESVOOS SYSTEM
 

•ERAIN
 

1INFRALIZATICN
 

SPECIAL SEliSE C5GANS
 

NONB
 

I NUMBE R OF ftNIMALS WIT H TISSUE

* SU18ER CF ANIMALS UECEOPSIED
 

TABLE Dl (CONTINUED) 

LOW DOSE HIGH DOSE
 CCNTBOL (UNTE) CCN1BCL (I'NTR)
 

05-0030 05-0077
 

2 <!»%)
 

(39) (36)
 

1 (3*)
 

(UU) (43)
 

1 (2%)
 

(It) (13)
 

2 (5%)
 

1"4 (32%)
 

(17)	 (18)
 

1 (6%)
 

(K6) (16)
 

(U6) (11)
 1 (2*)
 

<U6) (15)
 

(«6) (15)
 

3 (7%) 2 (14%)
 

(16) (16)
 

(U€) (15)
 

 EXAPINE E HCHCSCOPICiLLY 

D-6
 

LOW DOSE
 

05-0025
 

(33)
 

1 (3%)
 

1 (3%)
 

(16)
 

2 (1%)
 

(16)
 

25 (51*)
 

(21)
 

(18)
 

(15)
 

(17)
 

1 <2X>
 1 (9S)
 

(17)
 

(48)
 

1 (2*1
 

(16)
 

18 (39%)
 

HIGH DOSE
 

05-CC88
 

(42)
 

(15)
 

(15)
 

(20)
 

(19)
 

1 (2*)
 

(17)
 

(17)
 

(47)
 

(49)
 

(45)
 
TABLE Dl (CONCLUDED) 

LOW DOSE HIGH DOSE CONTROL {UNTB) CONTBOL (t'NTR) LOU DOSE HIGH DCSE 

05-0030 05-0077 05-0025 05-0086 

HUSClILOSKEl^TA I SYSTEf! 

*SKILE1A I MUSCL E (46 ) (46) (18) (49) 

FE5IAFTEHITI E 1 (2*) 

EOE Y CAVITIE S 

*ABDOMINA L CAVIT Y (16) (46) (48 ) (49) 

STEATI1I S 1 (2%) 

NECROSIS , FAT 1 (2%) 

JLi OTHE 3 SYSTEM S 

*KU'TIPL E ORGAN S (46 ) (46) (48) (49) 

FCSTMCFTE M CHANG E 1 (2X) 

SPECIA L MOPPBOLOG Y SU«»AR Y 

N O LESIC1 FEfOETE E 2 8 1 28 

ACCIDEtiTA I DEAT H 3 

AI1TO/NEC3CPSY/EIST O PERF 1 2 

4UTC/»ICSCt£Y/N C HISTO 1 

AUTCLYSIS/S C SFCFCTS Y 1 4 2 1 

» tiU"BI P C f ANIMAL S HU H TISSU E EXSMINE D MICEOSCOPICAIL Y 

* NU^BE P C f ANIMAL S WECFOESIE D 

D-7 
TABLE D2

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN FEMALE MICE 

TREATED WITH 2,4-DINITROTOLUENE 

LOW DOSE HIGH DOSE c:ONTBOL(UNTR) CONTROL (ONTR) LOO DOSE HIGH DOSE 

06-0030 06-0077 06-0025 06-0088 

JNIMAL S INITIAII Y IK STUDY 50 50 50 50 

JBIMAL S KISSIN G 3 

»«IMAL S 8ICBOFSIED 47 46 16 50 

SBIHAIS EXAMINE D HISTCEATHCLOGICALL Y ** 47 16 16 50 

INTEGUMENTAR Y SYSTE M 

*SKTS (47) (1*6) (l»6) (50)

 

FIBROSIS 1 (2X)

 

FIEROSIS, FOCAI 1 (2%)

 

RESPIRATOR Y SYSTEM 

*LUNG/BP.ONCHU S (16) (15) (16) (18)

 

INFLAflfATICN , ACUTE FOCAI 1 (2»)

 

INFLAMMATICN , ACUTE/CHSOMIC 1 (2X )

 

INFLAMMATION , CFRONIC 1 (2X ) 

METAPLASIA , NOS 1 (2*)

 

HUNG (16) (15) (16) (18) 

CONGESTION, HOE 1 (2X) 

FDERA, NCS 1 (2X)

 

INFLAMMATION, FOCAL 1 (2X)

 

INFLAMMATION, INTEPSTITIAL 2 (1%) 

PERIARTEBITIS 1 (2t) 

I1UNG/ALVECLI (16) (15) (16) (18)

 

EMPHYSEMA, NOS 1 (2S)

 

HEBATOPCIETIC SYSTEB
 

IEONE KARFCW (45) (14) (45) (45)
 

HYPOPLASIA, NOS 1 (2*)
 KYELOFIBRCSIS 1 (2*) 1 (2*)
 

HYPEPPLASIB, HIMBTCPOIETIC 1 (2%)
 

ISPIEES (45) (43)
 

LYMPHCCYTCSIS 2 (H%)
 

__ HYPERPLAS|A. HIMftTOPOIETIC
 

I NUMBER OF ANI-1ALE HITH TISSUE E5IRKINF D MICROSCOPICALLY 

* NUMBE R OF ANIMAL S SfCRnFSIE D 

**EXCLUDES PARTIALLY AUTOLYZED ANIMALS 

D-8 
TABLE D2 (CONTINUED) 

LOW DOSE CCNTaOL(UNTB ) 

06-0030 

HIGH DOSE CONTFOL (CHTfi)

06-0077

 LOM DOSE 

 06-0025 

HYFEBELASia , BE1ICOHJ M CEL I 

HYPEPPLASIA , LYBPHOID 

HEMATOPOIEST S 

3 (7X) 

2 (1%) 

2 (5X) 

q (9X)

1 (2X)

 3 (7X) 

1 (2X) EBYTHEOPOIESI S 

•BAFDIEDLA S I. NOD E 

INFLABBATICN , ACUT E 

(27 ) (11) (12)

1 (2X) 

•PEEIASTINA L L.NCE E 

TNF1ABBATICN , ACUT F 

(27 ) (11) (12) 

1 <2X) 

•PANCREATI C L.NOD E 

HYPEEPIASIA , BETICULU M CELL 

(27) 

1 (qX ) (11) (12) 

IfESENTEPI C L . NOE E 

INFIABBATICN , ACUT E FOCAL 

INFLAMMATION , GB A NUL C BATCU S 

(27) CH) (12) 

1 (2X) 

1 <2X ) HYPEPPLASIA , PIASM A CEI L 

HYEEBELASIA , LYMPEOI D 

CIECULATOF Y SYSTE M 

«HEfET/ATBIO n («6) (15) (46) CALCIFICATION , FCCA L 1 (2%) 

IBYCCAEDIU M 

CALCIFICATION , FCCA L 

(16) (15)

1 (2X ) 

 (16) 

*EU!HCNAF Y AFTEF Y (17) (16) (16) hYPEPELASTA , DCS 1 <2X ) 

DIGESTIV E SYSTE M 

•SALIVAR Y GLAN D 

EEETVASCUIITI S 

(29) 

1 (3% ) 

(13) (11) 

»LIVE B 

MINEEAIIZATIO N 

(46) (15) (16) 

1 <2X ) INFIAFPATICN , KOS 

INFLAMMATION , G E AfiULOMATOU S 

NECEOSIS , FOCAL 

NECEOSIS , COAGULATIV E 

KETABOBPHOSI S FATT Y 

1 (2%) 

1 (2X) 

1 (2X) 

1 <2X ) 

1 (2X) 

1 (2X) 

HYPEEPLASIJ, NODULA R - _ 2 Ji*«l 

» NOBEE B CF AKIH&1 S WIT H TISSUE EXAMINE D SICPOSCOPICALIY 

* KUMBE B CF ANIMAL S NZCEOFSIE D 

HIGH DOSE 

06-0088 

1 <2X) 

3 (6X) 

(38) 

(38) 

(38) 

(38) 

1 OX) 

1 OX) 

(»7) 

<»7) 

(50) 

(18) 

(50) 

1 (2X) 

D-9 
HYPEEPLASIA, FCCJI
 

HYPEPPLASIA, DIFFUSE
 

ANGIECTASIS
 

•LIVEE/PEEIFOBTAL
 

INFLAMMATION , SOS
 

HYPERPLASIA, LYMEHOID
 

•IIVER/KUEFFE5 CELL
 

KYPF.RPLASIA, NCS
 

*EILE EUC1
 

INFLAMMATION , KS
 

INFLAMMATION, ACUTE/CHRONIC
 

INFLAMMATION, CBBCNIC DIFFUSE
 

•PANCREAS
 

INFLAMMATION, INTEFSTITIAL
 

INFLAMMATION, CHRONIC
 

FIEROSIE
 

ATROPHY, FOCAL
 

•PANCREATIC	 DUCT
 

INFLAP MATICN, NOS
 

•STCMACH
 

TNFLArCATLCti, UOS
 

ULCEB, FOCAL
 

INFLAMMATION, ACUTI FOCAL
 

INFLAMMATION, SCUTE/CHRONIC
 

HYPEfiPLASIA, EPITHELIAL
 

•SMALL	 INTESTINE
 

AMYLOICCSIS
 

IEEYFBS PATCH
 

PYPIF.PLASIA, NCS
 

•IUCEEKUH
 

ICTCPIA
 

•ILIUM
 

ABSCESS, NCS
 

XBF.CTUM
 PROLAPSE
 

TABLE D2 (CONTINUED)
 

LOW DOSE
 CCNTBOL (UNTK)
 

06-0030
 

1 (2X)
 

(16)
 

1 (2X)
 

(16)
 

(17)
 

(39)
 

(39)
 

1 (3X)
 

(15)
 

3 (7X)
 

(15)
 

(15)
 

1 <2X)
 

(15)
 

1 (2X)
 

(15)
 

(17)
 

.
 

HIGH DOSE
 CCSTBCL (CNTE)
 

06-0077
 

1 (2X)
 

1 (2X)
 

(15)
 

1 <2X)
 

(15)
 

1 (2X)
 

(16)
 

1 <2X)
 

(11)
 

(11)
 

(12)
 

(13)
 

(13)
 

(13)
 

(13)
 

(16)
 

« NUMBER OF ANIMALS MITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBE R OF ANI1ALS NECBOFSIED
 

D-IO
 

LOB DOSE
 

06-0025
 

1 (2X)
 

(16)
 

(16)
 

(16)
 

3	 (7X)
 

(11)
 

2	 (5X)
 

(11)
 

(11)
 

1 <2X)
 

1 <2X)
 

1 (2X)
 

1 (2X)
 

(12)
 

1 (2X)
 

(12)
 

(12)
 

(12)
 

1 (2X)
 

(16)
 

_. ... .
 

HIGH DOSE
 

06-0088
 

(50)
 

(50)
 

(50)
 

1	 (2X)
 

(11)
 

1 <2X)
 

1 (2%)
 

1 <2X)
 

(11)
 

(11)
 

(11)
 

(11)
 

1 (2%)
 

(11)
 

(11)
 

(50)
 

1 (2X1
 
TABLE D2 (CONTINUED)
 

LOW DOSE HIGH DOSE
 CCNTBCL (UNTR) CONTEOL (CNTE)
 

06-0030 06-0077
 

URINAB Y SYSTEM
 

1KIENEY (4E) (43)
 GLOMEBULCNIPHBITIS, NOS 3 (7*)
 

INFLAMMATION, INTERSTITIAL 9 (20X) 3 (7%)
 GLOMEEULONEPHBITIS, CBHONIC
 

PEBIVASCDLITIS 4 (9%)
 

HKIENEY/GIOMEBUIUS (15) (43)
 AMYLOIEOSIS 1 (2*)
 

IKIENEY/EELVIS (15) (43)
 INFLAKfATICN, (<CS
 

INFLAMMATION, ACUTE/CHBCNIC 1 (2%} 1 (2%)
 

•URINARY	 BLAEEEF (42) (11)
 INFLAMMATION, ACUTF/CHBONIC
 

HYPEEPLASIA, EPITHELIAL
 

ENDOCBINE SYSTEM
 

IPITUITAPY (37) (37)
 

HYPEKFLASIA, FOCAL
 

IAEPENAL (11) (13)
 

CONGESTICN, NCS
 

IAEF.ENAL COETEX (11) (13)
 NOCULE 1 (2*)
 HYPEEPLASIJ, NOS 2 (5%)
 HYPEEPLASIA, FOCAI
 

fTHYBOID (30)
 

COLLOID CYST
 

INFLAMMATION, ACDTE/CHBCNIC
 

HYPIEPLASIA, FOCAI
 

REPRODUCTIVE SYSTEM
 

ICTEBUS (13) (43)

 

HYEEOMETBA 1 (9X)

 

INFLAMMATION , SUEPUBATTV E 

INFLJMJLATJONi_ACUJE 

I NUMBER OF ANIMALS KITH TISSUE EXAMINED MICEOSCOPICALLY
 

* NUMBIB Of ANIMALS NECBOPSIED
 

LOK DOSE
 

06-0025
 

(46)
 

20	 (43X)
 

(16)
 

(46)
 

1 (2X)
 8 (17*)
 

(42)
 

9 (21X)
 

3 (7*)
 

(38)
 

7 (18X)
 

(12)
 

1 (2X)
 

(42)
 

3 (7S)
 

36 (86X)
 

(40)
 

1 (3X)
 

1 (3X)
 

1 (3*)
 

(11)
 

HIGH DOSE
 

06-0088
 

(46)
 

1 (2)1)
 

1 (2*)
 

(46)
 

(46)
 

(44)
 

(34)
 

(45)
 

(45)
 

(12)
 

(43) 

1 (2%) 

2 (5%) 

D-ll
 
TABLE D2 (CONTINUED)
 

LOW DOSE HIGH DOSE 

CONTROL (USTB) CODTEOL (CNIE)

06-0030 06-0077

ABSCESS , NC S 

NECROSIS , FOCA L 

METAPLASIA , SQUAMOO S 1 (2%) 

•UTERUS/ENDCMETFIU M 

CYST, NOS 

INFLAMMATICN , NO S 

(13)

1 (2% ) 

 (13)

2 <5X ) 

INFLAMMATION , SUPPUPATIV E 

INFLAMMATION , ACC1 E 

HYPERPLASIA , NO S 1 <2X) HYPEPPLASIA , FOCA L 

HYPEHPLASIA , CYSTIC 33 (77%) 35 (81*)

ICVAB Y 

CYST, NOS 

(11)

5 (11S) (11)1 (2X) HEMOHPHAGI C CYST 

INFLAMMATION , SUPFtJRATIVE 

INFLAMMATION , ACD1E 

INFLAMMATION , CBRCNIC 

NERVOUS SYSTEM
 

SPECIAL SENSE OBGANS
 

'IYI (17) (16) SYNECHIA , NC S 

CATARAC T 

*EYE/BETIN A (17) (16) 

DEGENERATION, NOS
 

*IYE/CBYSTALLINE IENS (17) (16) 

SYNECHIA, ANTEEIOB 

«tAFDERIAN GLAND (16) 

INFLAMMATION, ACUTE/CHRCNIC 

MUSCDLOSKELETA L SYSTE M 

*VERTEBR A (U7) (16) 

CST1C^S£LJIOSJJ _ 1_12*1 

I NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* fOBBEF 0? ANIMALS NECROESIED
 

 LOU DOSE 

 06-0025 

2 (5X) 

1 (2X> 

 (11) 

1 (2*) 

5 (11X) 

1 (2*) 

 31 (77*) 

 (11) 

5 (11X) 

1 (2X) 

1 (2*) 

1 (2*) 

(16) 

1 (2X) 

(16) 

1 (2*) 

(16) 

1 <2X) 

(16)
 

HIGH DOSE 

06-0088 

(13) 

1 (2X) 

3 (7*) 

1 (2X) 

1 (2*) 

23 (53X) 

(37) 

2 (5*) 

1 (3*) 

2 (5X) 

1 (11*) 

(50) 

(50) 

(50) 

(50) 

(50)
 

D-12
 
TABLE D2 (CONCLUDED) 

LOW DOSE CCNISOI(UNTS )

06-0030

 HIGH DOSE CONIBOL (tNTR )

 06-0077

 LOU DOSE

 06-0025

 HIGH DOSE 

 06-0086 

EOCY CAVITIE S 

*FLEUP,A

BYPE8PIASIA, LYPEHCID

 (1(7)

 1 (2%) 

 (1(6) (U6) (50) 

All OTHEP SYSTEMS 

*«ULTIP1E ORGAN S

ECSTBCFIEM CHANG E

 («7) (46) (16)

 1 (2*) 

 (50) 

SPECIAL MOPPHOLOGY SUMMAR Y 

NC LESION REPORTED

ANIflAI MISSING/NC NECROPSY

AUTO/NECBCPSY/HISIO PIHF

AUTOLY£I£/NO NECROPSY

I NUMEE R OF ANIMAL S SilTH TISSOJ EJiSCINE D

* SUKBER Cf ANIMAL S NBCPOPSIED 

1

 1

3

 MIC ECSCOPICAILY 

1

2 

 <t

 3 

1 

1 

6 

6 

D-I3
 

Review of the Bioassay of 2,4-Dinitrotoluene* for Carcinogenicity
 

by the Data Evaluation/Risk Assessment Subgroup of the
 

Clearinghouse on Environmental Carcinogens
 

January 18, 1978
 

The Clearinghouse on Environmental Carcinogens was
 

established in May, 1976 under the authority of the National
 

Cancer Act of 1971 (P.L. 92-218). The purpose of the Clear­ inghouse is to advise on the National Cancer Institute's
 

bioassay program to identify and evaluate chemical carcino­ gens in the environment to which humans may be exposed. The
 

members of the Clearinghouse have been drawn from academia,
 industry, organized labor, public interest groups, State
 

health officials, and quasi-public health and research
 

organizations. Members have been selected on the basis of
 

their experience in carcinogenesis or related fields and,
 collectively, provide expertise in organic chemistry, biochemistry, biostatistics, toxicology, pathology, and epidemiology. Representatives of various Governmental agencies
 

participate as ad hoc members. The Data Evaluation/Risk
 

Assessment Subgroup of the Clearinghouse is charged with the
 

responsibility of providing a peer review of NCI bioassay
 

reports on chemicals studied for carcinogenicity. In this
 

context, below is the edited excerpt from the minutes of
 

the Subgroup's meeting at which 2,^1-Dinitrotoluene was
 

reviewed.
 

The primary reviewer said that he agreed with the
 

staff's conclusion that, under the conditions of test,
 

2,4-Dinitrotoluene induced benign tumors in rats but showed
 

no evidence of carcinogenicity in either sex of mice. He
 

noted that a different strain of mouse was used for the pre-
 chronic study than for the chronic phase. After briefly
 

describing the experimental design, the primary reviewer
 

said that the study was deficient in that the maximum
 

tolerated dose was not adequately defined during the sub-


chronic studies. Therefore, it had to be approximated for
 

the chronic phase. He opined that the hemangiosarcomas in
 

the treated rats may be biologically significant. In summary,
 

the primary reviewer said that the tumors in the treated
 

rats must be viewed with concern, especially since the maximum
 

tolderated dose may not have been attained. He felt that the
 

data did not allow an assessment of human risk.
 

The secondary reviewer noted that 2,4-Dinitrotoluene
 

is an intermediate in the production of dyes. He added
 

47
 
that there may be considerable human exposure from residues
 

of 2,4-Dinitrotoluene in dye products. He continued that
 

there may be a potential for human risk because of the
 

increased tumor incidence seen in the treated rats. He
 

suggested that another study be done, possibly using another
 

species and' route of exposure.
 

A Program staff member commented that there was a
 

treatment-related biological effect produced by 2,4-Dinitro­ toluene in the rats. He added that the dose levels in the
 

mice and male rats approached the maximum tolerated dose,
 

as suggested by the growth curves.
 

A Subgroup member suggested that the biological activity
 

of 2,4-Dinitrotoluene may be due to its conversion to the
 

diamine compound. The rate of its enzymatic conversion may
 

limit its activity. He added that skin exposure would be a
 

more appropriate route if 2,4-Dinitrotoluene is to be retested.
 Another Subgroup member suggested that the staff consult
 

with Dr. Harris at North Carolina and Dr. Peters at Harvard,
 both of whom are doing epidemiologic studies among workers
 

exposed in the TDI process. The consultation would be
 

helpful in considering the need to retest 2,4-Dinitrotoluene.
 

It was moved that, in view of the significant number
 

of benign tumors in the treated rats and widespread human
 

exposure, 2,4-Dinitrotoluene be considered for retest. The
 

motion was seconded and approved unanimously.
 

Members Present Were:
 

Arnold Brown (Acting Chairman), Mayo Clinic
 

Lawrence Garfinkel, American Cancer Society
 

Joseph Highland, Environmental Defense Fund
 

Charles Kensler, Arthur D. Little Company
 

Verald K. Rowe, Dow Chemical, U.S.A.
 Sheldon Samuels, Industrial Union Department, AFL-CIO
 

Louise Strong, University of Texas Health Sciences Center
 

Sidney Wolfe, Health Research Group
 

Subsequent to this review, changes may have been made
 

in the bioassay report either as a result of the review
 

or other reasons. Thus, certain comments and criticisms
 

reflected in the review may no longer be appropriate.
 

48
 

IL.S . GOVERNMEN T PRIMIN G OFFICE : 1978-260-399/3055
 

DHEW Publication No. (NIH) 78-1360