National Cancer Institute 

CARCINOGENESIS 

Technical Report Series 

No. 146 

1979 

BIOASSAY OF 

NITHIAZIDE 

FOR POSSIBLE CARCINOGENICITY 

CAS No. 139-94-6 

NCI-CG-TR-146 

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

Public Health Service 

National Institutes of Health 

BIOASSAY OF
 

NITHIAZIDE
 

FOR POSSIBLE CARCINOGENICITY
 

Carcinogenesis Testing Program
 

Division of Cancer Cause and Prevention
 

National Cancer Institute
 

National Institutes of Health
 

Bethesda, Maryland 20014
 

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

Public Health Service
 

National Institutes of Health
 

DHEW Publication No. (NIH) 79-1702
 

REPORT ON THE BIOASSAY OF NITHIAZIDE
 

FOR POSSIBLE CARCINOGENICITY
 

CARCINOGENESIS TESTING PROGRAM
 

DIVISION OF CANCER CAUSE AND PREVENTION
 

NATIONAL CANCER INSTITUTE, NATIONAL INSTITUTES OF HEALTH
 

FOREWORD: This report presents the results of the bioassay of
 

nithiazide conducted for the Carcinogenesis Testing Program, Division of Cancer Cause and Prevention, National Cancer Institute (NCI),
 

National Institutes of Health, Bethesda, Maryland. This is one of
 

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

incidence of cancer than control animals, do not necessarily mean
 

the test chemical is not a carcinogen because the experiments are
 

conducted under a limited set of circumstances. Positive results
 

demonstrate that the test chemical is carcinogenic for animals under
 

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

The actual determination of the risk to man from animal carcinogens
 

requires a wider analysis.
 

CONTRIBUTORS: This bioassay of nithiazide was conducted by Litton
 

Bionetics, Inc., Bethesda, Maryland, initially under direct contract
 

to the NCI and currently under a subcontract to Tracor Jitco, Inc.,
 

prime contractor for the NCI Carcinogenesis Testing Program.
 

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

Weisburger (1,3). The principal investigators for the contract were
 

Dr. S. M. Garner (4,5) and Dr. B. M. Ulland (4,5). Mr. S. Johnson
 

(4) was the coprincipal investigator for the contract. Animal treatment and observation were supervised by Mr. R. Cypher (4), Mr. D. S.
 

Howard (4) and Mr. H. D. Thornett (4); Mr. H. Paulin (4) analyzed
 

dosed feed mixtures. Ms. J. Blalock (4) was responsible for data
 

collection and assembly. Chemical analysis was performed by Midwest
 

Research Institute (6) and the analytical results were reviewed by
 

Dr. N. Zimmerman (7).
 

Histopathologic examinations were performed by Dr. A. DePaoli
 

(4), Dr. P. Hildebrandt (4), Dr. R. Montali (4), Dr. H. Seibold (4),
 

and Dr. N. J. Wosu (4) at Litton Bionetics, Inc., the pathology narratives were written by Dr. A. DePaoli (4), and the diagnoses included
 

in this report represent the interpretation of these pathologists.
 

Histopathology findings and reports were reviewed by Dr. R. L.
 

Schueler (8).
 

111
 
Compilation of individual animal survival, pathology, and summary tables was performed by EG&G Mason Research Institute (9); the
 

statistical analysis was performed by Mr. W. W. Belew (7,10) and
 

Mr. R. M. Helfand (7), using methods selected for the Carcinogenesis
 

Testing Program by Dr. J. J. Gart (11).
 

This report was prepared at METREK, a Division of The MITRE Corporation (7) under the direction of the NCI. Those responsible for
 

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

(7), task leader Ms. P. Walker (7), senior biologist Mr. M. Morse
 

(7), biochemist Mr. S. C. Drill (7), and technical editor Ms. P. A.
 

Miller (7). The final report was reviewed by members of the participating organizations.
 

The following other scientists at the National Cancer Institute
 

were responsible for evaluating the bioassay experiment, interpreting
 

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

Cueto, Jr. (1), Dr. J. F. Douglas (1), Dr. D. G. Goodman (1,12), Dr.
 

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

T. W. Orme (1), Dr. R. A. Squire (1,13), Dr. S. F. Stinson (1), Dr.
 

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

1.	 Carcinogenesis Testing Program, Division of Cancer Cause and
 

Prevention, National Cancer Institute, National Institutes of
 

Health, Bethesda, Maryland.
 

2.	 Now with the U.S. Environmental Protection Agency, 401 M Street
 

S.W., Washington, D.C.
 

3.	 Now with the Naylor Dana Institute for Disease Prevention, American Health Foundation, Hammon House Road, Valhalla, New York.
 

4.	 Litton Bionetics, Inc., 5516 Nicholson Lane, Kensington, Maryland.
 

5.	 Now with Hazleton Laboratories America, Inc., 9200 Leesburg
 

Turnpike, Vienna, Virginia.
 

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

Missouri.
 

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

Boulevard, McLean, Virginia.
 

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

Maryland.
 

iv
 
9.	 EG&G Mason Research Institute, 1530 East Jefferson Street, Rockville, Maryland.
 

10.	 No wit w h The Solar Energy Research Institute, Cole Boulevard,
 

Golden, Colorado.
 

11.	 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.
 

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

Washington, D.C.
 

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

University, School of Medicine, Traylor Building, Baltimore,
 

Maryland.
 

SUMMARY
 

The bioassay of nithiazide for possible carcinogenicity was conducted using Fischer 344 rats and B6C3F1 mice. Nithiazide was administered in the diet, at either of two concentrations, to groups of 50
 

male and 50 female animals of each species. The high and low concentrations of nithiazide utilized were, respectively, 1250 and 625 ppm
 

for rats and 5000 and 2500 ppm for mice. Dosed rats received feed
 

containing nithiazide for 38 weeks, and as a result of a shortage of
 

nithiazide, the animals were not fed the dosed feed for the next 9
 

weeks. The dosed feed diet was then resumed and continued for 56
 

weeks, after which time a 1-week observation period followed. Dosed
 

mice received feed containing nithiazide for 61 weeks and, due to a
 

shortage of nithiazide, the animals were not fed dosed feed for the
 

next 9 weeks. The dosed feed diet was then resumed and continued for
 

33 weeks, followed by a 1-week observation period. Twenty animals of
 

each sex and species were placed on test as controls.
 

In both species, adequate numbers of animals survived sufficiently long to be at risk from late-developing tumors. There was
 

no significant positive association between dosage and mortality for
 

either rats or mice. Compound-related mean body weight depression
 

occurred in both sexes of each species.
 

Statistically significant incidences of hepatocellular adenomas
 

and carcinomas were found in high dose male mice but not in female
 

mice. Although the increased incidences of these tumors in dosed female mice were not statistically significant, the evidence presented
 

was strongly suggestive of carcinogenicity to the liver in female
 

B6C3F1 mice. Statistically significant increased incidences of a
 

combination of mammary and skin fibroadenomas and cystadenomas NOS
 

were found in the high dose female rats. No unusual tumors were
 

observed in either species.
 

Under the conditions of this bioassay, nithiazide was carcinogenic in male and probably female B6C3F1 mice, causing a combination
 

of hepatocellular carcinomas and hepatocellular adenomas. Nithiazide
 

was also carcinogenic in female Fischer 344 rats, causing an increase
 

in the incidence of mammary neoplasms. The compound was not carcinogenic in male Fischer 344 rats.
 

VI1
 

TABLE OF CONTENTS
 

I. INTRODUCTION	 1
 

II. MATERIALS AND METHODS	 3
 

A. Chemicals	 3
 

B. Dietary Preparation	 5
 

C. Animals	 5
 

D. Animal Maintenance	 6
 

E. Selection of Initial Concentrations	 8
 

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	 19
 

C. Pathology	 19
 

D. Statistical	 Analyses of Results 22
 

IV. CHRONIC TESTING RESULTS: MICE	 30
 

A. Body Weights and Clinical Observations	 30
 

B. Survival	 30
 

C. Pathology	 34
 

D. Statistical	 Analyses of Results 35
 

V. DISCUSSION	 41
 

VI. BIBLIOGRAPHY	 43
 

APPENDIX A SUMMARY OF THE INCIDENCE OF NEOPLASMS IN
 

RATS TREATED WITH NITHIAZIDE A-l
 

APPENDIX B SUMMARY OF THE INCIDENCE OF NEOPLASMS IN
 

MICE TREATED WITH NITHIAZIDE B-l
 

APPENDIX C SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC
 

LESIONS IN RATS TREATED WITH NITHIAZIDE C-l
 

APPENDIX	 D SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC
 

LESIONS IN MICE TREATED WITH NITHIAZIDE D-l
 
Figure Number
 

1
 

2
 

Table Number
 

LIST OF ILLUSTRATIONS 

Page 

CHEMICAL STRUCTURE OF NITHIAZIDE 2 

GROWTH CURVES FOR NITHIAZIDE CHRONIC STUDY 

RATS 20 

SURVIVAL COMPARISONS OF NITHIAZIDE CHRONIC 

STUDY RATS 21 

GROWTH CURVES FOR NITHIAZIDE

MICE 

 CHRONIC STUDY 

31 

SURVIVAL COMPARISONS OF NITHIAZIDE CHRONIC 

STUDY MICE 32 

PERCENT SURVIVAL OF NITHIAZIDE CHRONIC 

STUDY MICE 33 

LIST OF TABLES 

Page 

DESIGN SUMMARY FOR FISCHER 344 RATS— 

NITHIAZIDE FEEDING EXPERIMENT 10 

DESIGN SUMMARY FOR B6C3F1 MICE—-NITHIAZIDE 

FEEDING EXPERIMENT 11 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS 

AT SPECIFIC SITES IN MALE RATS TREATED WITH 

NITHIAZIDE 23 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS 

AT SPECIFIC SITES IN FEMALE RATS TREATED 

WITH NITHIAZIDE 26 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS 

AT SPECIFIC SITES IN MALE MICE TREATED WITH 

NITHIAZIDE 36 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS 

AT SPECIFIC SITES IN FEMALE MICE TREATED 

WITH NITHIAZIDE 38 
LIST OF TABLES (Concluded)
 

Table Number 

Al SUMMARY OF THE INCIDENCE OF NEOPLASMS IN 

MALE RATS TREATED WITH NITHIAZIDE A-3 

A2 SUMMARY OF THE INCIDENCE OF NEOPLASMS IN 

FEMALE RATS TREATED WITH NITHIAZIDE A-7 

Bl SUMMARY OF THE INCIDENCE OF NEOPLASMS IN 

MALE MICE TREATED WITH NITHIAZIDE B-3 

B2 SUMMARY OF THE INCIDENCE OF NEOPLASMS IN 

FEMALE MICE TREATED WITH NITHIAZIDE B-7 

Cl SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC 

LESIONS IN MALE RATS TREATED WITH NITHIAZIDE C-3 

C2 SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC 

LESIONS IN FEMALE RATS TREATED WITH NITHIAZIDE C-8 

Dl SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC 

LESIONS IN MALE MICE TREATED WITH NITHIAZIDE D-3 

D2 SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC 

LESIONS IN FEMALE MICE TREATED WITH NITHIAZIDE D-7 

I. INTRODUCTION
 

Nithiazide (Figure 1) (NCI No. C03792), an antiprotozoal compound
 

used in veterinary medicine, was selected for bioassay by the National
 

Cancer Institute because of its use and possible persistence in the
 

tissues and eggs of animals raised for human consumption.
 

The Chemical Abstracts Service (CAS) Ninth Collective Index
 

(1977) name for this compound is N-ethyl-N'-(5-nitro-2-thiazolyl)
 

urea. It is also called l-ethyl-3-(5-nitro-2-thiazolyl) urea.
 

Nithiazide is most commonly used against Histomonas maleagridas,
 

the organism which causes blackhead in fowl, particularly turkeys
 

(O'Neill et al., 1956; Rose and Rose, 1966).
 

Specific production data for nithiazide are not available; however, the exclusion of this compound from Synthetic Organic Chemicals:
 

United States Production and Sales, 1976 (U.S. International Trade
 

Commission, 1977) implies that nithiazide is not produced in commercial quantities (in excess of 1000 pounds or $1000 in value annually)
 

in the United States.
 

The potential for exposure to nithiazide is greatest among veterinary workers and workers in facilities which produce this compound.
 

The compound may persist in the tissues and eggs of treated poultry,
 

thereby constituting a potential for more widespread human exposure.
 

k
 

The CAS registry number is 139-94-6,
 
FIGURE 1

 

CHEMICAL STRUCTURE OF IMITHIAZIDE 

2 
II. MATERIALS AND METHODS
 

A. Chemicals
 

Nithiazide was purchased from Merck, Sharp and Dohme Research
 

Laboratory, a division of Merck and Company, Inc., Rahway, New Jersey.
 

Chemical analysis was performed by Midwest Research Institute, Kansas
 

City, Missouri. The range of the experimentally determined melting
 

point (223° to 230°C) included the reported literature value of
 

228°C (O'Neill et al., 1956). Thin-layer chromatography (TLC) was
 

performed utilizing two solvent systems (i.e., ethyl acetate:acetone
 

and chloroform:methanol). Each plate was visualized with 354 and 367
 

nm light and the reducing agent p-nitroso-dimethylaniline. The plate
 

developed with the first solvent system revealed one impurity, which
 

remained at the origin, while the plate developed with the other
 

solvent system indicated one motile and one nonmotile impurity. Elemental analysis was consistent with the molecular formula for nithiazide. High pressure liquid chromatography showed one homogenous peak.
 

The results af infrared (IR) and nuclear magnetic resonance (NMR)
 

analyses were consistent with those expected on a structural basis.
 

Ultraviolet/visible (UV/VIS) analysis revealed \max at 233, 352 and
 

429 nm with respective molar extinction coefficients (O of 73.7x
 

102, 13 x 10^ and 19.8 x 102. The results are suggestive of a high
 

purity compound.
 

A second batch of the compound was purchased from the same supplier. TLC was performed utilizing two solvent systems (i.e., ethyl
 
acetate racetone and benzene: 1,4-dioxane) . When visualized with ultraviolet light and p-nitroso-dimethylaniline/dimethylaminobenzaldehyde,
 

each plate revealed one nonmotile impurity. High pressure liquid
 

chromatography showed the presence of two minor and one major peak.
 

Elemental analysis was within 5 percent of the theoretical. The
 

range of the experimentally determined melting point (224° to 235°C)
 

once again included the literature value (O'Neill et al . , 1956). IR
 

and NMR analyses were consistent with those expected based upon the
 

structure of the compound. UV/VIS analysis showed ^max of 233, 353,
 

3 3 3


and 443 nm with e values of 8.33 x 10 , 11.8 x 10 , and 5.3 x 10 .
 

A third batch of the compound was purchased from the same supplier. TLC was performed utilizing two solvent systems (i.e., ethyl
 

acetate:acetone and benzene: 1,4-dioxane) . When visualized with ultraviolet light and p-nitroso-dime thy laniline/dimethy laminobenzaldehyde,
 

each plate revealed one impurity, remaining at the origin. High
 

pressure liquid chromatography showed the presence of one homogeneous
 

peak. Elemental analysis was within the acceptable limits of experimental variation (+_ 5 percent), based upon the molecular structure of
 

the compound. The determined point of thermal decomposition was in
 

general agreement with the point reported in the literature (O'Neill
 

et al., 1956). IR and NMR analyses were consistent with the results
 

expected based upon the structure of the compound. UV/VIS analysis
 

2
 

showed ^max °f 232.5, 352, and 436 nm with £ values of 73.7 x 10 ,
 

3 2
 

13.1 x 10 , and 6.75 x 10 , respectively.
 
Throughout this report the term nithiazide is used to represent
 

these compounds.
 

B. Dietary Preparation
 

The basal laboratory diet for both dosed and control animals consisted of Wayne Lab-Blox meal (Allied Mills, Inc., Chicago, Illinois).
 

Nithiazide was administered to the dosed animals as a component of
 

the diet.
 

The chemical was removed from its container and a proper amount
 

was blended with an aliquot of the feed using a mortar and pestle.
 

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

6 kg capacity Patterson-Kelley standard model twin-shell stainless
 

steel V-blender along with the remainder of the feed to be prepared.
 

After 20 minutes of blending, the mixtures were placed in double
 

plastic bags and stored in the dark at 4°C. The mixture was prepared once weekly.
 

C. Animals
 

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

through contracts of the Division of Cancer Treatment, National Cancer Institute. Rats were supplied by the Frederick Cancer Research
 

Center, Frederick, Maryland. Mice were supplied by Charles River
 

Breeding Laboratories, Inc., Wilmington, Massachusetts.
 

Rats and mice were approximately 4 weeks old when received.
 

Upon receipt, animals were examined for visible signs of disease or
 
parasites. Obviously ill or runted animals were culled. The remaining animals were quarantined for 2 weeks prior to initiation of test.
 

Animals which did not manifest clinical signs of disease were placed
 

on test at this time. Animals were assigned to groups and distributed
 

among cages so that the average body weight per cage was approximately
 

equal for a given species and sex.
 

D. Animal Maintenance
 

Animals were housed by species in rooms with a temperature range
 

of 22° to 26°C and a range in relative humidity of 45 to 55 percent.
 

Incoming air was filtered through HEPA filters (Flanders Filters,
 

McLean, Virginia) at a rate of 12 to 15 complete changes of room air
 

per hour. Fluorescent lighting was provided 8 hours per day (9:00
 

a.m. to 5:00 p.m.).
 

Rats were housed four per cage by sex and mice were housed five
 

per cage by sex. Throughout the study dosed and control animals of
 

both species were housed in polycarbonate cages (Lab Products, Inc.,
 

Garfield, New Jersey) suspended from aluminum racks. Racks were
 

fitted with a continuous stainless steel mesh lid over which a sheet
 

of filter paper was firmly secured. Filter paper was changed at
 

2-week intervals, when the racks were sanitized. Clean cages and
 

®
 bedding were provided twice weekly. Ab-sorb-dri hardwood chip
 

bedding (Wilner Wood Products Company, Norway, Maine) was used in
 

polycarbonate cages for the entire bioassay.
 
Acidulated water (pH 2.5) was supplied to animals in water bottles filled by an automated metering device that was checked daily
 

for diluting accuracy. Water bottles were changed twice weekly and
 

sipper tubes were washed at weekly intervals. During the period of
 

chemical administration, dosed and control animals received treated
 

®
 

or untreated Wayne Lab-Blox meal as appropriate. The feed was supplied in hanging stainless steel hoppers which were refilled three
 

times per week and sanitized weekly. Food and water were available
 

ad libitum for both species.
 

All dosed and control rats were housed in a room with other
 

rats receiving diets containing* 2,4-dimethoxyaniline hydrochloride
 

(54150-69-5) and 4'-(chloroacetyl)-acetanilide (140-49-8); and other
 

rats intubated with dosed solutions of trimethylphosphate (512-56-1).
 

All dosed and control mice were housed in a room with other
 

mice receiving diets containing 2,4-dimethoxyaniline hydrochloride
 

(54150-69-5); 4'-(chloroacetyl)-acetanilide (140-49-8); p-phenylenediamine dihydrochloride (624-18-0); 4-nitro-o-phenylenediamine (99­

56-9); and l-phenyl-3-methyl-5-pyrazolone (89-25-8); and other mice
 

intubated with dosed solutions of trimethylphosphate (512-56-1);
 

2-(chloromethyl)pyridine hydrochloride (6959-47-3); 3-(chloromethyl)
 

pyridine hydrochloride (3099-31-8); and pivalolactone (1955-45-9).
 

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

In order to establish the concentrations of nithiazide for administration to dosed animals in the chronic studies, subchronic toxicity
 

tests were conducted with both rats and mice. Rats were distributed
 

among thirteen groups, each consisting of five males and five females.
 

Nithiazide was incorporated into the basal laboratory diet and supplied ad libitum to eleven of the thirteen rat groups in concentrations of 464, 681, 1000, 1470, 2150, 3160, 4640, 6810, 10,000, 14,700
 

and 21,500 ppm. The two remaining rat groups served as control
 

groups, receiving only the basal laboratory diet.
 

Mice were distributed among six groups, each consisting of five
 

males and five females. Nithiazide was incorporated into the basal
 

laboratory diet and supplied ad libitum to five of the six mouse
 

groups in concentrations of 6800, 10,000, 14,700, 21,600 and 31,500
 

ppm. The sixth mouse group served as a control group, receiving
 

only the basal laboratory diet.
 

The dosed dietary preparations were administered for a period
 

of 7 weeks, followed by a 1-week observation period during which all
 

animals were fed the basal laboratory diet. Individual body weights
 

and food consumption data were recorded twice weekly throughout the
 

study. Upon termination of the observation period, all survivors
 

were sacrificed and necropsied.
 

At 21,500 ppm, one male rat died, while all female rats receiving
 

the same concentration died. At the end of the subchronic test, the
 

8
 
mean body weight gain of both male and female rats dosed with 1470
 

ppm was 21 percent less than the mean body weight gain of their respective controls. At a dietary concentration of 1000 ppm, the mean
 

body weight gain of male rats was 12 percent less than that of their
 

controls, while female rats receiving the same concentration displayed
 

a mean body weight gain 12 percent less than that of their controls.
 

At both of these concentrations yellow patches on the coat were observed. The high concentration selected for administration to dosed
 

rats in the chronic bioassay was 1250 ppm.
 

At a dietary concentration of 31,500 ppm, one male mouse died.
 

Two female mice died at a concentration of 21,600 ppm. At the end
 

of the subchronic test, the mean body weight gain of both male and
 

female mice dosed with 6800 ppm was 10 percent less than the mean
 

body weight gain of their respective controls. The high concentration selected for administration to dosed mice in the chronic bioassay was 5000 ppm.
 

F. Experimental Design
 

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

treated and untreated observation periods) are summarized in Tables
 

1 and 2.
 

All rats were approximately 6 weeks old at the time the test
 

was initiated and were placed on test simultaneously. The dietary
 

concentrations of nithiazide administered were 1250 and 625 ppm.
 
TABLE 1
 

DESIGN SUMMARY FOR FISCHER 344 RATS
 

NITHIAZIDE FEEDING EXPERIMENT
 

INITIAL 

GROUP NITHIAZIDE 

SIZE CONCENTRATION3 

MALE
 

CONTROL 20 0
 

LOW DOSE 50 625
 

0
 

625
 

0
 

HIGH DOSE 50 1250
 

0
 

1250
 

0
 

FEMALE
 

CONTROL 20 0
 

LOW DOSE 50 625
 

0
 

625
 

0
 

HIGH DOSE 50 1250
 

0
 

1250
 

0
 

Concentrations given in parts per million.
 

OBSERVATION PERIOD
 

TREATED UNTREATED
 

(WEEKS) (WEEKS)
 

0 104
 

38
 

9
 

56
 

1
 

38
 

9
 

56
 

1
 

0 104
 

38
 

9
 

56
 

1
 

38
 

9
 

56
 

1
 

10
 
TABLE 2
 

DESIGN SUMMARY FOR B6C3F1 MICE
 

NITHIAZIDE FEEDING EXPERIMENT
 

INITIAL 

GROUP NITHIAZIDE 

SIZE CONCENTRATION3 

MALE
 

CONTROL 20 0
 

LOW DOSE 50 2500
 

0
 

2500
 

0
 

HIGH DOSE 50 5000
 

0
 

5000
 

0
 

FEMALE
 

CONTROL 20 0
 

LOW DOSE 50 2500
 

0
 

2500
 

0
 

HIGH DOSE 50 5000
 

0
 

5000
 

0
 

j-J
 

Concentrations given in parts per million.
 

OBSERVATION PERIOD
 

TREATED UNTREATED
 

(WEEKS) (WEEKS)
 

0 104
 

61
 

9
 

33
 

1
 

61
 

9
 

33
 

1
 

0 105
 

61
 

9
 

33
 

1
 

61
 

9
 

33
 

1
 

11
 
Throughout this report, those rats receiving the former concentration are referred to as the high dose groups and those receiving the
 

latter concentration are referred to as the low dose groups. Dosed
 

rats were supplied with feed containing nithiazide for the first 38
 

weeks of the chronic study. Due to a shortage of nithiazide, dosed
 

diets were not available for the following 9-week period. Use of
 

dosed feed was then resumed and continued for 56 weeks, followed by
 

a 1-week observation period.
 

All mice were approximately 6 weeks old at the time the test
 

was initiated, and were placed on test simultaneously. The dietary
 

concentrations of nithiazide administered were 5000 and 2500 ppm.
 

Throughout this report, those mice receiving the former concentration are referred to as the high dose groups and those receiving the
 

latter concentration are referred to as the low dose groups. Dosed
 

mice were supplied with feed containing nithiazide for the first 61
 

weeks of the chronic study. Due to a shortage of nithiazide, dosed
 

diets were not available for the following 9-week period. Use of
 

dosed feed was then resumed and continued for 33 weeks, followed by
 

a 1-week observation period for males, and a 2-week observation
 

period for females.
 

G. Clinical and Histopathologic Examinations
 

Animals were weighed immediately prior to initiation of the experiment. From the first day, all animals were inspected twice daily
 

12
 
for mortality. Food consumption data were collected at monthly intervals from 20 percent of the animals in each group. Body weights were
 

recorded once monthly throughout this bioassay.
 

All moribund animals or animals that developed large, palpable
 

masses that jeopardized their health were killed. A necropsy was
 

performed on each animal regardless of whether it died, was killed
 

when moribund, or was killed at the end of the bioassay. The animals
 

were euthanized using carbon dioxide, and were immediately necropsied.
 

Gross and microscopic examinations were performed on all major tissues, organs, and gross lesions taken from sacrificed animals and,
 

whenever possible, from animals found dead.
 

Tissues were preserved in a 10 percent neutral buffered formalin
 

solution, embedded in paraffin, sectioned, and stained with hematoxylin and eosin prior to microscopic examination.
 

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

nodes, thymus, heart, salivary gland, liver, gallbladder (mice), pancreas, esophagus, stomach, small intestine, large intestine, kidney,
 

urinary bladder, pituitary, adrenal, thyroid, parathyroid, testis,
 

prostate, brain, tunica vaginalis, uterus, mammary gland, and ovary.
 

A few tissues were not examined for some animals, particularly
 

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

preclude histopathologic interpretation. Thus, the number of animals
 

13
 
for which particular organs, tissues, or lesions were examined microscopically varies and does not necessarily represent the number of
 

animals that were recorded in each group at the time that the test
 

was initiated.
 

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
 

in the form of graphs. Animals were statistically censored as of the
 

time that they died of other than natural causes or were found to be
 

missing; animals dying from natural causes were not statistically
 

censored. Statistical analyses for a possible dose-related effect
 

on survival used the method of Cox (1972) when testing two groups for
 

14
 
equality and used Tarone's (1975) extensions of Cox's methods when
 

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

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

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

The incidence of neoplastic or nonneoplastic lesions has been
 

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

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

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

histologically. However, when macroscopic examination was required
 

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

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

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

The purpose of the statistical analyses of tumor incidence is to
 

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

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

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

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
 

15
 
be less than or equal to 0.05/k. In cases where this correction was
 

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

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

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

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

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

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

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

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

from linear trend.
 

A time-adjusted analysis was applied when numerous early deaths
 

resulted from causes that were not associated with the formation of
 

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

tumor was observed were excluded by basing the statistical tests on
 

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

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

tumor was found, comparisons were based exclusively on animals that
 

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

was 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
 

16
 
observed tumor were computed as in Saffiotti et al. (1972). The week
 

during which animals died naturally or were sacrificed was entered
 

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

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

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

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

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

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 Pt/Pc where pt is the true binomial probability of the
 

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

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

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

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

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

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

larger proportion in the treated group than in the control.
 

The lower and upper limits of the confidence interval of the
 

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

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

17
 
of the risk in a treated group of animals to that in a control group
 

would be within the interval calculated from the experiment. When
 

the lower limit of the confidence interval is greater than one, it
 

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

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

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

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

the lower limit indicates the absence of a significant result while
 

the upper limit indicates that there is a theoretical possibility
 

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

detected under the conditions of this test.
 

18
 
III. CHRONIC TESTING RESULTS: RATS
 

A. Body Weights and Clinical Observations
 

For both male and female rats there was slight, although distinct, dose-related mean body weight depression (Figure 2).
 

No unusual clinical observations were reported.
 

B. Survival
 

The estimated probabilities of survival for male and female rats
 

in the control and nithiazide-dosed groups are shown in Figure 3.
 

For both males and females, the Tarone test for positive association
 

between dosage and mortality was not significant.
 

There were adequate numbers of male rats at risk from late-


developing tumors, as 33/50 (66 percent) of the high dose, 32/50 (64
 

percent) of the low dose and 16/20 (80 percent) of the control group
 

survived on test until termination of the study. For female rats,
 

with 42/50 (84 percent) of the high dose, 33/50 (66 percent) of the
 

low dose, and 17/20 (85 percent) of the control group surviving on
 

test until the termination of the study, there were adequate numbers
 

at risk from late-developing tumors.
 

C. Pathology
 

Histopathologic findings on neoplasms in rats are summarized in
 

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

summarized in Appendix C (Tables Cl and C2).
 

There was a variety of tumors in both the control and dosed
 

groups. Some types of neoplasms occurred with greater frequency in
 

19
 
TIM EON TEST (WEEKS) 

FIGURE 2 

GROWTH CURVES FOR NITHIAZIDE CHRONIC STUDY RATS 

20 
FIGURE 3

 

SURVIVAL COMPARISONS OF NITHIAZIDE CHRONIC STUDY RATS 

21 
dosed rats as compared with controls. The incidences of chromophobe
 

adenomas, mammary neoplasms and endometrial stromal polyps were
 

slightly elevated in the high dose female rats when compared to the
 

female controls.
 

In addition to the neoplastic lesions, a number of degenerative,
 

proliferative and inflammatory changes were encountered in the dosed
 

and control groups. Most of these nonneoplastic lesions are commonly
 

observed in aged Fischer 344 rats and were not considered to be
 

compound-related.
 

Based upon the results of this pathology examination, there
 

was no conclusive evidence for the carcinogenicity of nithiazide
 

in Fischer 344 rats under the conditions of this study.
 

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 is included for
 

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

tumors were observed in at least one of the control or nithiazidedosed groups and where such tumors were observed in at least 5 percent of the group.
 

For female rats the Cochran-Armitage test indicated a significant (P = 0.003) positive association between dosage and the incidences of a combination of fibroadenomas or cystadenomas NOS of the
 

skin, subcutaneous tissue, and mammary gland. This was supported
 

22
 
TABLE 3
 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS AT
 

SPECIFIC SITES IN MALE RATS TREATED WITH NITHIAZIDE£
 

TOPOGRAPHY: MORPHOLOGY
 

Hematopoietic System: Leukemia or Malignant
 

Lymphoma^
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

NJ
 

OJ Pituitary: Chromophobe Adenoma or Acidophil
 

Adenoma"
 

P Values0
 

Departure from Linear Trend6
 

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
 

CONTROL
 

4/20(0.20)
 

N.S.
 

87
 

6/15(0.40)
 

P = 0.007(N)
 

P = 0.018
 

—_
 

104
 

2/20(0.10)
 

N.S.
 

_ —
 

104
 

LOW
 

DOSE
 

8/50(0.16)
 

N.S.
 

0.800
 

0.250
 

3.327
 

90
 

2/36(0.06)
 

P = 0.005(N)
 

0.139
 

0.016
 

0.685
 

95
 

2/50(0.04)
 

N.S.
 

0.400
 

0.032
 

5.277
 

104
 

HIGH
 

DOSE
 

15/50(0.30)
 

N.S.
 

1.500
 

0.566
 

5.627
 

78
 

2/30(0.07)
 

P = O.Oll(N)
 

0.167
 

0.020
 

0.814
 

85
 

6/50(0.12)
 

N.S.
 

1.200
 

0.243
 

11.574
 

98
 
TABLE 3 (CONTINUED) 

TOPOGRAPHY: MORPHOLOGY CONTROL
 

Pancreatic Islets: Islet-Cell Adenoma or Islet-


Cell Carcinomab 1/19(0.05)
 

P Values0 N.S.
 

Relative Risk (Control) —_
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor 104
 

Testis: Interstitial-Cell Tumor 19/20(0.95)
 

P Values0 N.S.
 N3 

-PRelative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor 87
 

Body Cavities:, Mesothelioma NOS or Malignant
 

Mesothelioma 0/20(0.00)
 

P Values0 N.S.
 

Departure from Linear Trend P = 0.021
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

LOW
 

DOSE
 

3/50(0.06)
 

N.S.
 

1.140
 

0.101
 

58.635
 

100
 

47/50(0.94)
 

N.S.
 

0.989
 

0.922
 

1.168
 

86
 

4/50(0.08)
 

N.S.
 

Infinite
 

0.386
 

Infinite
 

52
 

HIGH
 

DOSE
 

0/46(0.00)
 

N.S.
 

0.000
 

0.000
 

7.707
 

48/50(0.96)
 

N.S.
 

1.011
 

0.942
 

1.149
 

0/50(0.00)
 

N.S.
 

78 
TABLE 3 (CONCLUDED)
 

Q
 

Treated groups received doses of 625 or 1250 ppm in feed.
 

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

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

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

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

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

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

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

group when P < 0.05.
 N>
 

Ul
 
___ 

___ 

TABLE 4
 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS AT
 

SPECIFIC SITES IN FEMALE RATS TREATED WITH NITHIAZIDE*
 

TOPOGRAPHY: MORPHOLOGY
 

Skin, Subcutaneous Tissue, and Mammary Gland:
 

Fibroadenoma or Cystadenoma NOS"
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Hematopoietic System: Leukemia or Malignant
 

Lymphoma"
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Pituitary: Chromophobe Adenoma or Acidophil
 

Adenoma"
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

CONTROL
 

1/20(0.05)
 

P = 0.003
 

93
 

3/20(0.15)
 

N.S.
 

94
 

5/18(0.28)
 

P = 0.034
 

93
 

LOW
 

DOSE
 

5/50(0.10)
 

N.S.
 

2.000
 

0.249
 

92.596
 

89
 

9/50(0.18)
 

N.S.
 

1.200
 

0.346
 

6.408
 

71
 

13/39(0.33)
 

N.S.
 

1.200
 

0.493
 

3.750
 

89
 

HIGH
 

DOSE
 

15/50(0.30)
 

P = 0.020
 

6.000
 

1.048
 

245.704
 

94
 

3/50(0.06)
 

N.S.
 

0.400
 

0.060
 

2.802
 

79
 

24/47(0.51)
 

N.S.
 

1.838
 

0.852
 

5.313
 

94
 
___ 

TABLE 4 (CONTINUED) 

TOPOGRAPHY : MORPHOLOGY
 

Thyroid: C-Cell Adenoma
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Pancreatic Islets: Islet-Cell Adenoma or Islet-


Cell Carcinomab
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Uterus: Endometrial Stromal Polyp
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

CONTROL
 

2/19(0.11)
 

N.S.
 

104
 

0/19(0.00)
 

N.S.
 

_—
 

1/19(0.05)
 

P = 0.039
 

104
 

LOW
 

DOSE
 

1/38(0.03)
 

N.S.
 

0.250
 

0.004
 

4.557
 

104
 

0/48(0.00)
 

N.S.
 

4/50(0.08)
 

N.S.
 

1.520
 

0.167
 

73.309
 

91
 

HIGH
 

DOSE
 

0/26(0.00)
 

N.S.
 

0.000
 

0.000
 

2.404
 

3/47(0.06)
 

N.S.
 

Infinite
 

0.254
 

Infinite
 

100
 

10/50(0.20)
 

N.S.
 

3.800
 

0.613
 

160.949
 

87 
TABLE 4 (CONCLUDED)
 

Treated groups received doses of 625 or 1250 ppm in feed.
 

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

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

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

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

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

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

N5
 

00
 
by a significant (P * 0.020) positive Fisher exact test comparing the
 

high dose group to the control group.
 

Based on these statistical results, nithiazide was carcinogenic
 

in female Fischer 344 rats under the conditions of this bioassay.
 

In female rats the Cochran-Armitage test for association between
 

dosage and incidence was significant for a combination of chromophobe
 

adenomas and acidophil adenomas of the pituitary (P = 0.034) and also
 

for endometrial stromal polyps of the uterus (P = 0.039). However,
 

in both cases, the Fisher exact tests comparing the high dose group
 

to the control and the low dose group to the control were not significant.
 

None of the statistical tests for any site in male rats indicated
 

a significant positive association between the administration of nithiazide and an increased tumor incidence.
 

The possibility of SL negative association between dose and tumor
 

incidence was noted in male rats for chromophobe adenomas or acidophil adenomas of the pituitary. The historical incidence of these
 

tumors in untreated male Fischer 344 rats from control data collected
 

by this laboratory for the NCI Carcinogenesis Testing Program was
 

23/188 (12 percent) as compared with the 6/15 (40 percent) incidence
 

in the controls of this bioassay.
 

29
 
IV. CHRONIC TESTING RESULTS: MICE
 

A. Body Weights and Clinical Observations
 

For both male and female mice there was a distinct and consistent
 

dose-related mean body weight depression (Figure 4).
 

No unusual clinical observations were reported.
 

B. Survival
 

The estimated probabilities of survival for male and female mice
 

in the control and nithiazide-dosed groups are shown in Figure 5. For
 

both males and females the Tarone test for association between dosage
 

and mortality was not significant.
 

The percentages of male and female mice surviving on test are
 

shown in Figure 6. Although 6 high dose and 3 low dose male mice
 

were missing by week 12, adequate numbers of males were at risk from
 

late-developing tumors as 39/50 (78 percent) of the high dose, 36/50
 

(72 percent) of the low dose and 15/20 (75 percent) of the control
 

group survived on test until the termination of the study.
 

There were also adequate numbers of female mice at risk from
 

late-developing tumors. Although 4 high dose, 8 low dose, and 1 control female mice were missing by week 12, 39/50 (78 percent) of the
 

high dose, 39/50 (78 percent) of the low dose, and 17/20 (85 percent)
 

of the control group survived on test until the termination of the
 

study.
 

30
 
TIME ON TEST (WEEKS) 

FIGURE 4 

GROWTH CURVES FOR NITHIAZIDE CHRONIC STUDY MICE 

31 
TIME ON TEST (WEEKS)

 

FIGURES 

SURVIVAL PROBABILITY COMPARISONS OF NITHIAZIDE CHRONIC STUDY MICE 

32 
TIME ON TEST (WEEKS) 

FIGURE 6 

PERCENT SURVIVAL OF NITHIAZIDE CHRONIC STUDY MICE 
C. Pathology
 

Histopathologic findings on neoplasms in mice are tabulated in
 

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

tabulated in Appendix D (Tables Dl and D2).
 

There was a variety of tumors in the control and dosed groups.
 

These lesions are commonly observed in aged B6C3F1 mice. Hepatic
 

tumors, however, did appear to be elevated in the high dose groups
 

when compared to controls, particularly among male mice as shown
 

below:
 

Males 

Low

Control Dose

 High 

 Dose 

Females 

Low

Control Dose

 High 

 Dose 

Liver 

Number of Animals with 

Tissues Examined 

Histopathologically (20) (46) (43) (18) (41) (43) 

Hepatocellular Adenoma 

Hepatocellular Carcinoma 

2 

2 

9 

6 

13 

12 

4 

0 

8 

4 

Grossly, the hepatic tumors varied from inapparent single or multiple
 

lesions to nodules, ranging up to 3 cm in diameter. The larger
 

lesions were usually malignant. Most of the hepatic tumors were
 

well-differentiated, expanding lesions. A spindle-cell component was
 

present in one hepatocellular carcinoma. Criteria employed to determine malignancy included: metastasis, local invasion, formation of
 

trabecular pattern, mitotic activity, anaplasia, and necrosis.
 

34
 
In addition to the neoplastic lesions a number of degenerative,
 

proliferative, and inflammatory changes were encountered in dosed and
 

control groups (Appendix C). Most of these nonneoplastic lesions are
 

commonly seen in aged laboratory B6C3F1 mice.
 

The results of this pathology examination suggest that nithiazide
 

was responsible for the observed increased number of hepatic tumors in
 

the high dose male and female B6C3F1 mice.
 

D. Statistical Analyses of Results
 

The results of the statistical analyses of tumor incidence in
 

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

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

tumors were observed in at least one of the control or nithiazidedosed groups and where such tumors were observed in at least 5 percent of the group.
 

For male mice the Cochran-Armitage test indicated a significant
 

(P = 0.002) positive association between dosage and the combined
 

incidence of hepatocellular carcinomas or hepatocellular adenomas.
 

Additionally, the Fisher exact test comparing the high dose group
 

to the control group was significant (P = 0.005). In female mice
 

an unusually large, though not statistically significant, number of
 

hepatocellular carcinomas or hepatocellular adenomas was also found
 

in the high dose group. It should be noted that in historical control data collected by this laboratory for the NCI Carcinogenesis
 

Testing Program 9/207 (4 percent) untreated female B6C3F1 mice had
 

35
 
___ 

___ 

TABLE 5
 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS AT
 

SPECIFIC SITES IN MALE MICE TREATED WITH NITHIAZIDE*
 

TOPOGRAPHY : MORPHOLOGY
 

Lung: Alveolar/Bronchiolar Adenoma
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Hematopoietic System: Malignant
 

Lymphoma*3
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Liver: Hepatocellular Carcinoma
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

CONTROL
 

3/20(0.15)
 

N.S.
 

«.»—.
 

99
 

4/20(0.20)
 

N.S.
 

98
 

2/20(0.10)
 

P - 0.037
 

100
 

LOW
 

DOSE
 

4/44(0.09)
 

N.S.
 

0.606
 

0.116
 

3.855
 

104
 

6/47(0.13)
 

N.S.
 

0.638
 

0.175
 

2.820
 

90
 

6/46(0.13)
 

N.S.
 

1.304
 

0.264
 

12.541
 

104
 

HIGH
 

DOSE
 

1/44(0.02)
 

N.S.
 

0.152
 

0.003
 

1.774
 

104
 

4/44(0.09)
 

N.S.
 

0.455
 

0.096
 

2.242
 

104
 

12/43(0.28)
 

N.S.
 

2.791
 

0.717
 

24.104
 

104
 
___ 

TABLE 5 (CONCLUDED)
 

LOW HIGH
 

TOPOGRAPHY : MORPHOLOGY CONTROL DOSE DOSE
 

Liver: Hepatocellular Carcinoma or
 

Hepatocellular Adenoma" 4/20(0.20) 15/46(0.33) 25/43(0.58)
 

P Values0 P = 0.002 N.S. P = 0.005
 

Relative Risk (Control) 1.630 2.907
 

Lower Limit 0.617 1.217
 

Upper Limit 6.077 9.903
 

Weeks to First Observed Tumor 99 104 87
 

Adrenal: Pheochromocytoma 2/17(0.12) 0/42(0.00) 0/41(0.00)
 

P Values0 P = 0.026(N) N.S. N.S.
 u>
 

—i
 Departure from Linear Trend P = 0.045
 

Relative Risk (Control) 0.000 0.000
 

Lower Limit 0.000 0.000
 

Upper Limit 1.353 1.385
 

Weeks to First Observed Tumor 104
 

Treated groups received doses of 2500 or 5000 ppm in feed.
 

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

£

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

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

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

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

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

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

group when P < 0.05.
 
___ 

___ 

TABLE 6
 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS AT
 

SPECIFIC SITES IN FEMALE HICE TREATED WITH NITHIAZIDE£
 

TOPOGRAPHY : MORPHOLOGY
 

Lung: Alveolar /Bronchiolar Adenoma
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Hematopoietic System: Malignant
 

oo Lymphoma^
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Liver: Hepatocellular Carcinoma
 

P Values0
 

Relative Risk (Control)
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

CONTROL
 

3/16(0.19)
 

N.S.
 

104
 

3/18(0.17)
 

N.S.
 

94
 

1/18(0.06)
 

N.S.
 

104
 

LOW
 

D^SE
 

1/42(0.02)
 

N.S.
 

0.127
 

0.003
 

1.474
 

104
 

7/42(0.17)
 

N.S.
 

1.000
 

0.267
 

5.520
 

99
 

0/41(0.00)
 

N.S.
 

0.000
 

0.000
 

8.171
 

HIGH
 

DOSE
 

2/43(0.05)
 

N.S.
 

0.248
 

0.023
 

2.011
 

104
 

8/45(0.18)
 

N.S.
 

1.067
 

0.300
 

5.761
 

4/43(0.09)
 

N.S.
 

1.674
 

0.186
 

80.455
 

80 

104 
TABLE 6 (CONCLUDED) 

LOW HIGH
 

TOPOGRAPHY rMORPHOLOGY CONTROL DOSE DOSE
 

Liver: Hepatocellular Carcinoma or
 

Hepatocellular Adenoma 3/18(0.17) 4/41(0.10) 12/43(0.28)
 

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

Relative Risk (Control) —_ 0.585 1.674
 

Lower Limit 0.114 0.536
 

Upper Limit 3.698 8.451
 

Weeks to First Observed Tumor 104 104 97
 

Treated groups received doses of 2500 or 5000 ppm in feed.
 

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

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

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

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

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

The 95% confidence interval on the relative risk of the treated group to the control group.
 
hepatocellular carcinomas or hepatocellular adenomas as compared with
 

the 3/18 (17 percent) combined incidence found in the control group
 

of this bioassay. Out of 11 female historical control groups observed, the incidence for the small control group in this bioassay
 

was the highest. Based upon these statistical results the administration of nithiazide was associated with the increased incidence
 

of liver neoplasms in male mice and possibly female mice under the
 

conditions of this bioassay.
 

The Cochran-Armitage test indicated a significant negative association between administration and the incidence of adrenal pheochromocytomas in male mice. The Fisher exact tests, however, were not
 

significant.
 

None of the statistical tests for any site in female mice indicated a significant positive association between the administration
 

of nithiazide and an increased tumor incidence.
 

40
 
V. DISCUSSION
 

There were no significant positive associations between the concentrations of nithiazide administered and mortality in either species.
 

Adequate numbers of animals in all groups survived sufficiently long
 

to be at risk from late-developing tumors. Slight dose-related mean
 

body weight depression was observed in both male and female rats.
 

Distinct and consistent dose-related mean body weight depression
 

occurred in male and female mice.
 

Among female rats there was a significant positive association
 

between dosage and the incidences of a combination of fibroadenomas
 

and cystadenomas NOS of the skin, subcutaneous tissue, or mammary
 

gland. This finding was supported by a significant high dose to
 

control Fisher exact comparison. No other tumors occurred in statistically significant increased incidences when dosed female rats
 

were compared to controls. None of the statistical tests indicated
 

a positive association between the administration of nithiazide and
 

increased tumor incidence in male rats.
 

In female mice, none of the statistical tests showed a significant positive association between increased tumor incidence and the
 

administration of nithiazide. For male mice the combined incidence
 

of hepatocellular carcinomas and hepatocellular adenomas (i.e., 4/20
 

in the controls, 15/46 in the low dose, and 25/43 in the high dose)
 

was significantly dose-related and the combined incidence of these
 

41
 
tumors in the high dose group was statistically significant when compared to the incidence in the male mouse control group. Furthermore
 

/
 

almost half of the hepatocellular neoplasms observed in the dosed
 

groups were carcinomas (i.e., 2/20, 6/46, and 12/43 in the control,
 

low dose and high dose groups, respectively).
 

In female mice an unusually large, though not statistically
 

significant, number of hepatocellular carcinomas or hepatocellular
 

adenomas was also found in the high dose group. It should be noted
 

that in historical data collected by this laboratory for the NCI Carcinogenesis Testing Program control female mice had 9/207 (4 percent)
 

heptocellular carcinomas or hepatocellular adenomas as compared with
 

the 3/18 (17 percent) combined incidence found in the control group
 

of this bioassay. Out of 11 female historical control groups observed, the incidence for the control group in this bioassay was the
 

highest. Although the increased incidences of these tumors in dosed
 

female mice were not statistically significant, the evidence was
 

strongly suggestive of carcinogenicity to the liver in female B6C3F1
 

mice.
 

Under the conditions of this bioassay, nithiazide was carcinogenic in male and probably in female B6C3F1 mice, causing a combination of hepatocellular carcinomas and hepatocellular adenomas.
 

Nithiazide was also carcinogenic in female Fischer 344 rats, causing
 

an increased incidence of a combination of skin and mammary neoplasms.
 

The compound was not carcinogenic in male Fischer 344 rats.
 

42
 
VI. BIBLIOGRAPHY
 

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, B.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.
 

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

Observations." Journal of the American Statistical Association
 

_53_:457-481, 1958.
 

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

"Carcinogenesis Bioassay Data System." Computers and Biomedical
 

Research 7:230-248, 1974.
 

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

Co., New York, 1966.
 

O'Neill, R.C., A.J. Basso, and K. Pfister, III, "l-Substituted-3-(5­

nitro-2-thiazolyl) ureas." U.S. Patent 2,755,285 (Merck and
 

Co., Inc.), July 17, 1956; Chemical Abstracts 51, 2873a.
 

Rose, A. and E. Rose, editors, The Condensed Chemical Dictionary,
 

7th edition. Van Nostrand Reinhold Company, 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.
 

43
 
Tarone, R.E., "Tests for Trend in Life-Table Analysis." Biqmetrika
 

162:679-682, 1975.
 

U.S.	 International Trade Commission Syntheti , c Organic Chemicals:
 

United States Production and Sales, 1976. USITC Publication
 

833, U.S. Government Printing Office, Washington, D.C., 1977.
 

44
 
APPENDIX A
 

SUMMARY OF THE INCIDENCE OF NEOPLASMS
 

IN RATS TREATED WITH NITHIAZIDE
 

TABLE Al

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN MALE RATS TREATED WITH NITHIAZIDE 

CONTROL (UNTf) LOW DOSS HIG H DOSE 

11-1315 11-1343 11-13141 

ANIMA.LS INITIALLY IN STUDY 20 50 50
 

ANIMALS NECBOPSIFD 20 50 50
 

ANIMALS EXAMINfcD HISTOPAIHOLOGICALLY ** 20 50 50
 

INTEGUM5NTABY SYSTEM
 

*SKIN (20) (50) (50) SEBACEOU S ADENOM A 1 (2X) KERATOACANTHOMA 1 (2%)
 

FIBPCMA 1 (2K)
 

*SDBi-UT TISSUE (20) (50) (50)
 

FIBROMA 1 (2%)
 

RESPIRATOR Y SYSTEM 

*LUNG (20 ) (50) (50) ISLET-CELL CAPCIMOMA , METASTATIC 1 (2%) 

AI.VBOLAB/BFONCHIOLA P ADENOM A 2 («*) 1 (2X) 

HEHATUPOIETIC SYSTEM 

*B"AiN (20) (50) (50) 

MaLIGNANT EniCULOSIS 1 (2% ) 1 (2*) 

•MULiIPLE ORGANS (20) (SO) (50) 

MALIGNANT 1YMPHOMA, NOS 1 <2X ) MALIG.LYMPHOMA, UNDIFFER-TYPE 1 (2%) 1 (2%) 

KALIG.LYMPHOMA, LYMPHOCYTIC TYPE 1 (2*) 

MALIGNANT LYMPHOMA, KIXED TYPE 1 (2X) 

LEUKEMIA,NOS 4 (20%) 6 (12%) 11 (22X) 

UNDIFFERENTIATED LEUKEMIA 1 (2S) 

CIRCULATOS Y SYSTE M 

* HUBbEB OF ANIMAL S WIT H TISSUE EXAMINE D MICROSCOPICALLY 

* NUflJER OF ANIMALS NECROPSIED 

"EXCLUDES PARTIALLY AUTOLYZED ANIMALS 

A-3 
TABLE A1 (CONTINUED) 

CONTROL (UNTR ) LOU DOSE 

11-1345 11-1343 

DIGESTIV E SYSTE M 

fLIVzf (20) (50) 

NEOPLASTIC NODUL E 1 (2*) 

#SMAi.L ISTESIINE (20) (49) 

NtUFOFIESCSArCOM A 

URINAR Y SYSTE M 

NON ^ 

ENDOCiilN E SYSTE M 

• PITUITAR Y (15) (36) CriROMOPHOF I ADENOM A 5 (33S) 2 (6%) Ai-IDOPHIL ADENOM A 1 (7«) 

CADPiNA L (20) (50) PHEOCHFOMCCYICM A 2 (10%) 2 (4%) 

*IHYaOI D (1 5) (30) FOLLICULAf-CEL L ADENOM A 

C-CELL ADL&C1 A 1 (7%) 1 (3%) 

*PAFATHYFOI 3 (6) (16) AJLNOKA , NCS

 

*PANCBEATI C ISLETS (19 ) (50) ISLET-CELL ADEN01 A 1 (5«) 2 ("»*) ISLET-CELL CARCINOM A 1 (2%) 

P^PtOuUCTIV E SYSTE M 

*MAMi"iASY GLAN D (20) (50) FibFOADENCB A 1 (5%) 1 (2%) 

SOREPHTIAL GLAN D (20 ) (50) SEBACEOU S ADENOM A

 

(TESTI S (20 ) (50) I.«TE"SriTIAL-CELL T'JfiOF . _ 19 195*1. _ . 47 JSU*X 

* Nl'MUa P O F ANIMAL S KIT H TISSUE EXAMINE D MICROSCOPICALL Y 

* NHH3E P OF AMMAL S KECao"slED 

HIGH DOSE 

11-1341 

(49) 

2 (»X ) 

(49) 

1 (2%) 

(30) 

2 (7X) 

(50) 

6 (12X) 

(21) 

1 (5X) 

(11)

1 (9X) 

(46 ) 

(50) 

1 <2X ) 

(50) 

1 (2%) 

(50) 

48 J96X1 

A-4 
NERVOUS SYSTEM
 

NONE
 

SPECIAL SENSE CBGSNS
 

*TA<>SAL GLANE
 

S1.3ACE00S ADENOMA
 

MUSCULOSKELETAL SYSTEM
 

NONi
 

BODY CAVITIES
 

*BODY CAVITIES
 

MESOTHELIOMA, MALIGNANT
 

*PEKITONEUM
 

MESOTHELICBA, NOS
 

MESOTHELIOMA, MALIGNANT
 

*PERIIONEAL CAVIIY
 

SARCOMA, NOS
 

*TUNiCA VAGINALIS
 

MESOTHELICKA, NOS
 

ALL OTHER SYSTEMS
 

*MIILIIPLE ORGANS
 

OSTEOSARCOMA
 

THORACIC CAVITY
 

SARCOMA,! NOS
 

TABLE A1 (CONTINUED)
 

CONTPOL (HNTF.) LOW DOSE iilGH DOSE 

11-13U5 11-13U3 11-1311 

(20) (50) (50) 

1 (2?) 

(20) (50) (50) 

1 (2%) 

(20) (50) (50) 

1 (2?) 

1 (27.) 

(20) (50) (50) 

1 (2%) 

(20) (50) (50) 

1 (2%) 

(20) (50) (50) 

1 (2«) 

1 

* NttMBEP OF JNIflALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBE R OF ANIMAL S NECROPSIE D 

A-5 
TABLE A1 (CONCLUDED)
 

CONTROL (UNTP) LOW DOSE HIGH BOSE
 

11-1345 11-1343 11-1341
 

ANIMlL DISEOSITICN SUMMAR Y
 

ANI1ALS INITIALLY IN STUDY 20 50 50
 

NATURA L uEA"H« 10
 

%urI3'!ND SACRIFICE 14 7
 

bk.FED''LED EACFIFICE
 ACCIDENTALLY KILLED
 i-FMIBAL SACRIFICE 32 33
 

AJIBAL P-SSING
 

AtTCLlZED
 

TQTiiL ANIMAL S 411-1 PRIMARY TUMORS* 20 50 50
 

TOTAL PHlCAR l T" 10P3 34 78 82
 

TOTAL ANIMAL S 1*11" 1-»'IGM T'lMO^S 20 49 49
 

TOTAL BENIGM TOiMOFS 30 61 62
 

TOlAL ANIMAL S illlh MALIGNAN T TI'MJRS 4 14 16
 

TOTAL KALIGNANT TUMORS 4 14 18
 

TOTAL ANIMAL S WITH SiCCNDiPY TUfiOPS* 

1JTAL SICOKDAR Y T'THOE S 

TOTnL ANIIAL b BIH T'HOES OMCLBTAIN ­ 3r»(j.GN OR MAIIGNAN T

 

TuHL IINCEFTAI N T'llOPS 

TOT«L ANIMA1 S Wild TU-jorS DNCE^TAIN ­ PPIMARY OR PETASTATIC

 

TOTAL 'JNCEETAI N TH10E b 

* BrIilA^Y IU10FS: ILL I'I10''S EXCi.°T SECONDAPY TUMORS
 * SECuNDAlY TUrOBS: METASTATIC TUMORS OE TUMOFS INVASIVE INTO AN ADJACENT ORGAN
 

A-6
 
TABLE A2

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN FEMALE RATS TREATED WITH NITHIAZIDE 

CONTROL (ONTE) 

11-13U 6 

L OU BOSE 

11-1311) 

HIGH DOSE 

11-1342 

ANIMALS INITIAILY IN STUDY 20 

ANIMALS NECPOESIID 20 

ANIMALS EXAMINED HI3TOPATHOLOGICALL Y ** 20 

50 

50 

50 

50 

50 

50 

IKIEGUMENTA=! Y SYSIE1 

*SKIS (20) (50) (50) 

SLBACEOU S A02NOCARCIMONA , ISVASI 1 (25S) 

FIBROM A 2 («) 

FIBROADENOMA 1 (2%)
 

*SUBCDT TISSUE (20) (50) (5C)
 

SARCOMA, NCS 1 (5%)
 

FIBPOADENCKA 1 (8X)
 

FESPIjJATORY SYSTEM
 

NONi
 

HEMATOPOIETIC SYSTEM
 

*MnLTIPLE OFGANS (20) (50) (50)
 

MALIGNANT LYMPHOMA, NOS 1 (2%)
 

LEUKEMIA,NOS 3 (157) 9 (IBS) 2 (li%)
 

CIPCULATORY SYSTEM
 

NONE
 

DIGESTIVE SYSTEM
 

NONf
 

iFINAhY SYSTEM
 

• KIDNEY (20) (50) (50)
 

TSAN3lTIOSALzCELL_CAFC:NOMA 1_I5S)
 

* NUMBER OF ANI1ALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* N"(i£)EP OF ANIMAL S NECROPSIE D 

**EXCLUDES PARTIALLY AUTOLYZED ANIMALS 

A-7
 
S'lP-hAP Y ELfDE- = 

TuANSIIIONHL-CIL L PAPILLOM A 

ENDOCr-IN, , SYSTIt t 

'PITUITAR Y 

Cr.ROMOEhOE E PCE'xOK A 

A^I'JOPHIl PDrsOM A 

*AJ'^NH L 

CJr-TICA L PI.-8C.1A 

tlHYKOI D 

C-CELL AEI1SCM A 

*PAN>.FFAIJ C ISLET S 

IjLET-CEL L SErSO/ A 

IbLET-CElL CA"CINO:i A 

CiS:ADENC^A , NO S 

FIBCOAD2NCM A 

• UTEfB S 

ENDOtlETI-HI STBOIAl , ^OLYi 3 

HuhANGECM H 

• C^^VI X "TE" I 

Si»rOt!A, NOS 

HLKANSTCSAFCCM A 

»UIta'JS/ENDCriT" > I'J K 

A-IilOCATIkCaA . NO S 

NEIVOJ S SYSTc K 

HoEr O F ANI'1Ai, S Wil d TISSU E

Kj"!= u F ANI1AL S » rC"OESIF D 

TABLE A2 (CONTINUED) 

CONTFO L (US1P) LOS DOSE 

11-131*6 11-1311 

(14 ) (37) 

(16) (39) 

5 (28".) 13 (33%) 

(20 ) (50) 

(19 ) (38) 

2 (11*) 1 (3%) 

(19 ) (18) 

(20 ) (50) 

1 (27,) 1 (5%) 1 (8%) 

(19) (50) 1 (57 ) 4 (8%) 

1 (57,) 

(19) (50) 1

 (57.) 

1 (2*) 

(19 ) (50) 

2 (4%) 

(20) (50) 

 EXA"Tig3 D MICPOSCOPICALL 1 

A-8 

HIGH DOSE 

11-1342 

(37) 

1 (3X) 

(17) 

23 (49"!) 

1 (2% ) 

(48) 

1 (2X ) 

(26 ) 

(17) 

1 (2% ) 

2 (4% ) 

(50) 

2 (4% ) 

8 (16%) 

(50) 

10 (20% ) 

(50) 

(50) 

(50) 
TABLE A2 (CONTINUED) 

CONTPOL(UNTF )

11-13*46

 LOW DOSE

 11-13UU

 HIG H DOSE 

 11-131(2 

MUSCULOSKELETAI SYSTEM 

NON E 

EOEY LAVITIES 

NONi. 

ALL OPHEE SYS1E1S 

NONE 

ANIMAi. DISEOSIIION SOMI1APY 

ANIaALS INITIALLY IN STUDY

NATURAL DEATHH

MOKIBUND SACRIFICE

SCHEDOLEE SACRIFICE 

ACCIDENTALLY KILLED 

TiPMINAL SACRIFICE

ANIMAL MISSING 

 20

 2

 1

 17

 50

 11

 6

 33

 50 

3 

5 

 42 

8 INCLUDES AOTCLYZED ANIMALS 

t NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY 

* NUHcEF OF ANI1ALS NECROPSIED 

A-9
 
TABLE A2 (CONCLUDED) 

CONTROL(UNTH ) LOW DOSE HIGH DOSE 

11-13116 11-13UU 11-1342 

1TJSOP SUMMAR Y 

TOTAL ANIMALS KITH PRIMARY TUMOPS* 13 28 38
 TOTAL PRIMAR Y TUMORS 16 38 57
 

TOTAL ANIMALS WITH BENIGN TUMOFS 7 22 37
 TOTAL EOICN TUMORS 10 25 52
 

TOTAL ASIMALS WITH MALIGNANT TDMOPS 12
 

TOTAL MALIGNANT TOHOES 13
 

TOT&L ANIMALS KITH SECONDARY TOMOP.S* 1
 

TOIAL SECONDARY TUMOES 1
 

TOTAL ANIBALS WITH TUMORS UNCERTAINEEN1GN OK MALIGNANT
 

TOTAL UNCERTAIN TUMOKS
 

TOTAL ANIMALS WITH TUMORS UNCEPTAINPRIMARY OF KITAS1ATIC
 

TOTAL UNCERTAIN TTMOFS
 

* PRIMAPY TUMCFS: ALL TUMORS EXCLPT SECONDARY TUMORS
 

I SECONDARY TUMORS: METASTATIC TDMORS OR TUMORS INVASIVE INTO AN ADJACENT ORGAN
 

A-10
 
APPENDIX B
 

SUMMARY OF THE INCIDENCE OF NEOPLASMS
 

IN MICE TREATED WITH NITHIAZIDE
 

TABLE Bl

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN MALE MICE TREATED WITH NITHIAZIDE 

CONTROL(HNTR)

22-2345

 LOW DOSE

 22-2343

 HIGH DOSE 

 22-2J41 

ANIMAiS INITIALLY IN STUDY

ANIMALS MISSING

ANIMALS NECROESISD

ANIMALS EXAMINID HISTOPATHOLOGICALY

 20

 20

 20

 50

 3

 47

 47

 50 

6 

 44 

 44 

INTEGUMENTARY SYSTEM 

*SKIN

NEOROFIBSCMA

 (20) (47) (44) 

1 (251) 

*SUB(JOT TISSMI

NlUROFIBFCPA

 (20) (47)

 1 (2%) 

 (44) 

PSSPIHATOBY SYSTEM 

NEOPLASM, NOS, METASTATIC

HEPATOCELLULAR CARCINOMA, MFTAST

AtVEOLAR/EBCNCHIOLAR ADENOMA

(20)

 1 (5%) 

3 (15?)

 (44)

 1 (2%) 

4 (9«)

 (44) 

1 (2X) 

HEMATOPOIETIC SYSTEM 

'MULTIPLE ORGANS 

MALIGNANT LYMPHOMA, NOS 

MALIG.LYMPHOMA, UNDIFFER-TYPE 

MALIG.LYMPHOMA, LYMPHOCYTIC TYPE 

MALIG.LYMPHOMA, HISTIOCYTIC TYPE 

(20)' 

1 (5X) 

(47) 

1 <2X) 

1 (2*) 1 (2%) 

(4(4) 

1 (2*) 

'ABDOMINAL CAVIT Y 

MALIG.LYMEtCMA, HISTIOCYTIC TYPE 

(20) 

1 (5*) 

(47) (44) 

#SPL£F.N 

NiOPLASM, SOS, METASTATIC 

MALIGNANT LYMPHOMA, NOS 

(19 ) (46) 

1

1

 (2",) 

 (2*) 

(43) 

IMESESTERIC I. NODE 

MALIG.LYMPHOMA, LYMPHOCYTIC TYPE 

(19) 

1 (5%) 

(43) (43) 

1 (2*) 

t NUMBER OP AMMAL S WITH TISSUE EXAMINE D

* NUMoE R OF ANIMAL S NECROPSIED 

**EXCLUDES PARTIALLY AUTOLYZED ANIMALS 

 MICROSCOPICALLY 

B-3
 
TABLE Bl (CONTINUED)
 

CONTROL (DNTP)

22-2345

 LOW DOSE

 22-2343

 HIGH DOSE 

 22-2311 

»HEART

MALIGNANT LYMPHOMA, NOS

 (19) (H2)

 1 (2%) 

 (12) 

*SMALL INTESTINE

MALIG.LYMPHOMA, LYMPHOCYTIC TYPE

 (19) (44) (43) 

1 (2*) 

CIPCOLATORY SYSTEM 

NONE 

DIGESTIVE SYSTIM 

#LIViP

NEOPLASM, NOS, METASTATIC

HhPATOCiLLnLAR ADENOKA

HEPATOCELLULAR CARCINOMA

 (20)

 2 (10*)

 2 (10*)

 (46)

 1 (2%) 

9 (20%)

 6 (13*)

 (43) 

 13 (30*) 

 12 (28X) 

UPINAnY SYSTEM 

*KID1»EY

NiOPLASM, NOS, METASTATIC

 (19) (46)

 1 (2*) 

 (42) 

«DRINARY PLAEIEB

HEMANGIOMA

 (16) (36)

 1 (3*) 

 (40) 

ENDOCKINE SYSTIM 

tADRjiNAL

NiOPLASM, NOS, MALIGNANT

CORTICAL AEENCM A

PHEOCHRCMCCYTOMA

 (17)

 2 (12*) 

 (42)

 1 (2X) 

1 (2*) 

 (41) 

tTHYSOID

FULLICULAF-CILL ADENOMA

 (14) (29)

 1 (3*) 

 (31) 

PfiPFODUCTIVE SYSTEM 

•TESIIS

INTfifiSTITIAL-CELL TUMO R

 (18) (44) (42) 

1 (2*) 

NERVOUS SYSTEM 

_NONi 

* NUMBER OF ANIMALS KITH TISSUE EXAMINED MICROSCOPICALLY 

* NUMbER OF ANIMALS NECROPSIED 

B-4
 
TABLE Bl (CONTINUED) 

CONTROL(OHTB) LOW DOSE HIGH DOSE 

22-2345 22-2343 22-2311 

SPECIAL SENSE ORGANS 

NONL 

MUSCULOSKELETAL SYSTEM 

NONi. 

ECEY CAVITIES 

*PLEURA (20) (U7) (l*t) 

MuSOTHELICf'A, MALIGNANT 1 (2%) 

ALL OTHER SYSTEMS 

NONn 

ANIMAL DISPOSITION SUMMARY 

ANIaALS INITIALLY IN STUDY 20 50 50 NATURAL DEATHS 10 MORIBUND SACRIFICE 1 

SCHEDULED SACRIFICE 

AcCIDENTALlY KILLED 

TiPMINAL SACRIFICE 15 36 39 ANIMAL MISSING 3 6 

NUMBER OF AMINAL S WITH TISSUE EXAMINE D MICROSCOPICALLY 

NUMBE R OF AMMAL S NECROPSIED 

B-5 
IUMO" SU1MABY
 

TOTAL ANIMALS WITH PRIMARY TIIMOHS*


TOTAL PPIFARY TUKOPS


TOTAL ANI.1ALS WITH BENIGN TUBOPS


TOTAL EEMCN TD10SS


TOTAL ANIMALS KITH MALIGNANT TU.10PS


TOTAL MALIGNANT TUMOFS


IDIaL ANIMAL S HITH SECONDARY TUMORS*


TOTAL SECOKDA^Y TUMOPS


TOTAL ANIMALS WITH TnMOPS HNCEP1AIN­ BENIGN OR MALIGNANT
 

TOTAL UNCEBTAIN TUIOTS
 

TOT«L ANIMALS KITH TUH03S UNCEBTAIN­ FEIKAPY OS METASTATIC
 

TOTAL UNCEFTAIN TTIMOFS
 

TABLE Bl (CONCLUDED)
 

CONTROL (DNTF.)


22-2345


 10


 13


 5


 7


 6


 6


 1


 1


 LOW DOSE HIGH DOSE
 

 22-2343 22-2341
 

 27 28
 

 31 32
 

 16 15
 

 17 16
 

 14 16
 

 14 16
 

1
 

4
 

* OPiaABY TUMCPS: ALL TtlMOPS EXCEPT SECONDASY TUMORS
 * SECONDARY TUMORS: METASTATIC TUMORS OR TUMOPS INVASIVE IKTO AN ADJACENT ORGAN
 

B-6
 
TABLE B2

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN FEMALE MICE TREATED WITH NITHIAZIDE 

(rONTEOL (HNTR) LOW DOSE
 

22-2316 22-23HU
 

ANIMALS INITIALLY IN ST'JDY 20 50
 

ANIMALS MISSING 2 8
 

ANI»ALS NECFOFSIED 18 12
 

ANIMALS SXAMIKED HISTOPATHOLOGICAL!, Y 18 i»2
 

INTEGUM3NTAFY SYSTEM
 

*SUBCUT TISSUE (18) (t2)
 

FIBROSAPCCBA
 

RESPIRATORY SYSTEM
 

ftL'JNo (16) (<42)
 

HEPATOCELLULAR CARCINOMA, METAST 1 (6*)
 

ALVEOLAR/EFCNCHIOLAP ADENOMA 3 (19*) 1 (2%)
 

HEMATOPOIETIC SYSTEM 

*MULTIPL15 OEGANS 

MALIGNANT IYHPHOMA, NOS 

(18) 

1 (6%) 

(42) 

2 (5",) 

MALIG.LYMPPOBA, tJNDIFFER-TYPE 1 (2%) 

MALIG. LYMPHOMA, LY1PHOCYTIC TYPh 

MALI3. LYHPHCMA, HISTIOCYTIC TYPE 1 (6%} 1 (2-5) 

*ABDOMINAL BAIL (18) (U2) 

MALIGNANT LYMPHOMA. NOS 1 (2%) 

*M£Sl.NTERIC 1. tiOCE (18) (38) 

MALI3. LYPPHCMA, HISTIOCYTIC TYPD 

*HEART (18) (36) 

MALIGNANT IY1PHOMA, NOS 1 (3«) 

*LIV£P 

MALIGNANT IYHEIIC.IA, NOS 

(18) 

1 (6%) 

(41) 

MALIG. LYMPHOMA, HISTIOCYTIC TYPE 

KUPFFER-CEIL SAPCCMA 

#SMAi-L INTESTINE (18) (40) 

MALIG. LYBPiCMA^ UNDIFFjJ>=-TYPE 1 .13*1 

# N01BFR OF 8MMAL S WIT H TISSUE EXAMINE 1 ) MICROSCOPICAL!Y 

* NOHcFS OF ANIMAL S NECSCPSIED 

**EXCLUDES PAJRTIALLY AUTOLYZED ANIMALS 

HIGH DOSE
 

22-2342
 

50
 

it
 

45
 

45
 

(45)
 

1 (2%)
 

(43)
 

2 (5%)
 

(45)
 

2 (4%)
 

1 (21)
 

1 (2%)
 

(15)
 

(40)
 

1 (3%)
 

(40)
 

(42)
 

2 (5%)
 

1 (2%)
 

(44)
 

B-7 
TABLE B2 (CONTINUED) 

CONTROL(DNTR ) LOW DOSE HIGh DOSE 

22-23146 22-231*4 22-2342 

• COLON (18) (38) (1*3) MrtLIG.LYMPHOMA , HISTIOCYTIC TYPE 1 (2X) 

CIBCULATOPY SYSTEM 

NONE 

DIGESTIVE SYSTEM 

*LIVr,R (18) (1*1) (1*3) 

HEPATOCELLULAR ADENOMA 2 (11%) 1* (10%) 8 (19«) 

HiPATOCELl'JLAR CARCINOMA 1 (6%) 4 (9!!) 

UFINAiY SYSTSF 

HUBIiAKY BLACCE3 (15) (32) (35) 

HEMANGIOM A 1 (3%) 

ESDOCnlHE SYS1E1 

• PITUITARY (5) (22) (ID) 

CH"OP!OPHOE! ADENOM A 1 (5%) 

BEPFOD'JCTIVE SYSTEM 

•UTFiU S (18 ) (42 ) (44) 

EuDOHETP.IS I STPOM1L POLY P 1 (2%) 1 (2«) 

^EnvOli S SYSTB M 

NON E 

SPECIAL SENS E CBGAN S 

*2YE/LACEIMA L GLAN D (18 ) (42 ) (45) 

PAPILLAR Y ADSNCM A 1 (2% ) 

MUSCULOSKELETA L 5YSTS M 

__NONji 

# SUMoE F O F ANIMAL S WIT H TISSUE EXAMINE D HICPOSCOPICALL Y 

* SUi'Jj^h?, OF AMFAL S NECSOPSIF D 

B-8 
TABLE B2 (CONCLUDED)
 

CONTROL (UNTR) LOW DOSS HIGH DOSE
 

22-2316 22-2344 22-2342
 

EOLY CAVITIES
 

NONc
 

A L L OIHER SYSTIK S 

NONi. 

ANIMAL DISEOEITICN SUMMAFY
 

ANIMALS INITIALLY IN STUDY 20 50 50
 NATURAL DEATHS 2 3 4
 

MORIBUND SACRIFICE 3
 

SCHEDULED SACRIFICE
 

Ai-CIDENTAIIY KILLED
 

TERMINAL SACRIFICE 16 39 39
 AuIMAL MISSING 2 8 4
 

3 INCLUDES AUTCLYZED ANIMALS
 

TUMOR SUMMAP1
 

TOTAL ANIMALS WITH PRIMARY TUMORS* 6 13 23
 

TOTAL PRIORY TUMORS 9 15 26
 

TOTAL ANIMALS KITH BENIGN TUMORS 4 7 11
 

TOTAL BFNIGN TUMORS 5 8 12
 

TOTAL ANIMAL S WITH MALIGNANT 1UMORS 1 7 14
 

TOTAL MALIGNANT TUMOFS 4 7 14
 

TOTAL ANIMALS WITH SECONDARY TUMORS* 1
 

TOTAL SECCkDAPY TUMOES 1
 

TOTiL ANIMALS WITH TUMORS ONCERTAINEFN1GN OH MALIGNANT
 

TOTAL UNCERTAIN TUMORS
 

TOTAL ANIMAL S WITH THHO'S UNCf,"TAIN-


PRIflARY OF CIIASTATIC
 

TOTAL UNCERTAIN TUMOFS
 

* PRIMARY lUBOPS: ALL TUMOaS EXCEPT SECONDARY TUMOFS
 

* SECONDARY TUMORS: METASTATIC TUMOSS OP TUMORS INVASIVE INTO AN ADJACJKT OPGAN
 

B-9
 

APPENDIX C
 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC
 

LESIONS IN RATS TREATED WITH NITHIAZIDE
 

TABLE Cl

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN MALE RATS 

TREATED WITH NITHIAZIDE 

CONTRO L (UNTR) LOU

11-13H5 11­

 DOSE

13U3

 HIGH

 11-1

 DOSE 

311 

ANIMALS INITIALLY IN STUDY

ANIMALS NECRCESIED M 20

ANIMALS EXAMINID HISTOPATHOLOGICAL!Y

 20

 20

 50

 50

 50

 50 

 50 

 50 

INTEGUMENTASY SYSTEM
 

NONZ
 

RESPIRATORY SYSTPM
 

tLUNo/BRONCHUS (20) (50) (50)
 

INFLAMMATION, NOS 1 (2X)
 

NECROSIS, NOS 1 '(2%)
 

*LUNG (20) (50) (50)
 

CONGESTION, NOS 1 (5%) 1 (21) 5 (10%)
 

PNEUMONIA, ASPIHATION 1 (2%)
 

BJJCNCHOFNZUMCNIA, ACUTiJ 2 (4%)
 

PiE'tNONIA, CHPONIC MOBINE 2 (10*) i» (6%) 3 (6X)
 

INFLAMMATION, GRANULOMATOUS 1 (2%)
 

HYPERPLASIA. ADENOMATOUS 2 (4%) •
 

tLUNG/ALVEOLI (20) (50) (50)
 

EjEMA, NOS 1 (2%)
 

HEMATOPOIETIC SYSTEM
 

*BONn KARPO H (17) (18) (46) 

HY°EEPLASIA , HEMATOPOIETI C 1 (21) 

»SPL£EN (20) (50) (19)
 

CONGESTION, CHFONIC 1 (5%)
 

NECROSIS, NOS 1 (2%)
 

HiMOSIDEPCSIS 1 (5%)
 

LYMPHOID EEPLETION 1 (2S)
 

#SFLiNIC CAFSULH (20) (50) (19)
 

» NUMBE R 0 ? ANIMAL S WIIH TISSUE EXAMINE D MICROSCOPICALLY 

* NUMJE E OF ANIMAL S NECROPSIE D 

"EXCLUDES PARTIALLY AUTOLYZED ANIMALS 

C-3
 
HIGH DOSE
 

11-1341
 

(49)
 

(50)
 

(50)
 

2 (4X)
 

(50)
 

1 (2%)
 

(50)
 

22 (44X)
 

(50)
 

(50)
 

2 (4%)
 

(50)
 

1 (2%)
 

(49)
 

1 (2%)
 

1 (2?!)
 

1 (2%)
 

1 (2*)
 

2 (4S)
 

1 (2%)
 

1 (2%)
 

•MESENTERIC	 L. NODE
 

NjiCBOSIS, NOS
 

CIFCULATOSY SYSTEM
 

• HEAclT
 

7HROMBUS, BUFAL
 

PiRIARTFlRIlIS
 

•HEAST/ATBIUM
 

ThROMBOSIS, NOS
 

THROMBUS, PUPAL
 

•AUPiCOLAP	 APPENDAGE
 

THROMBOSIS, SOS
 

•MYOCARDIUM
 

FIBROSIS
 

•ENDOCARDIUM
 

INFLAMMATION, NOS
 

*PtlLf,ONARY ABTEFY
 

MINERALIZATION
 

*HEPA1IC VEIN
 

TuFOMBOSIS, NOS
 

TIGESIIVE SYSTIM
 

*LIVr,F
 

INFLAMMATION, C1PONIC
 

HEPATITIS, IOXIC
 

NtCROSIS, NOS
 

NECROSIS, fOCAL
 

Ni.CP.OSIS, ISCHEMIC
 

H-TAMOSFHOSIS FATTY
 

BASOPHILIC CYTO CHANGE
 

FOCAL CELLDLAP CHANGE
 

ATYPIA. NOS
 

ATFOOHY, NCS
 

L..UKEMOID SSACTION
 

TABLECl (CONTINUED) 

CONTROL (UNTP) LOW DOSE
 

11-1315 11-1343
 

(20)	 (48)
 

1 (2%)
 

(20) (50)
 

1 (2%)
 

1 (5%)
 

(20)	 (50)
 

1 (2%)
 

(20) (50) 

1 (2%) 

(20) (50) 

12 (60%) 22 (44?) 

(20) (50) 

1 (2i5) 

(20) (50) 

2 («) 

(20) (50) 

(20) (30)
 

1 (5?)
 

1 (5%)
 

* NUMBER OF ANIMALS WITH TISSOH EXAMINED MICROSCOPICALLY
 

* NUMnEP OF AMRALS NFCROPSIED
 

C-4
 

1 (2*,) 

1 (2S) 

3 (6%) 

1 (2*!) 

1 (2%) 

1 J 2"il
 
TABLE Cl (CONTINUED) 

CONTROL (UNTE) LO* DOSE
 

11-1 3.«5 11-1313
 

HiMATOIOIESIS	 1 (5*)
 

»LIV ID/CENTSI LOBtJLAS (20) (50)
 

ATROPHY, NOS
 

*BIlt DUCT (20) (50)
 

FJ.BPOSIS 1 (5*) 1 (2*)
 

HYPERPLASIA, NOS	 1 (5)8) 5 (10*)
 

JPANC^EAS (19) (50)
 

PiRIARTERITIS 1 (2%)
 

•PANCREATIC	 ACINUS (19) (50)
 

A1ROPHY, NCS 5 (10X)
 

*STOMACH (18) (19)
 

ASYL3IDOSIS 3 (17S) 2 (IX)
 

*COLON	 (19) (16)
 

INFLAMMATION, ACUTE/CHRONIC
 

PARASITISM	 <l (21X) « (9K)
 

Ai'lYLOIDOSIS
 

IlFINAnY SYSTEM 

4KIDNEY 

HYDRONEPHROSIS 

INFLAMMATION, CHFONIC 

GRAN'JLCMA, NOS 

INFLAMMATION, FOCAL GPA NOLOMATOU 

NEPHROPATHY, TOXIC 

HiMOSIDEFOSIS 

(20) 

17 (85%) 

1 (5X) 

1 (5X) 

(50) 

1

35

 (2%) 

 (70S) 

»'IPI«'ARY BLADtEH 

EISTENTION (15) (33) 

HEMORRHAGE 

luFLAKMATICN, ACUTE 

HYPERPLASIA, EPITHELIAL 

ftlRFIHOA AdSCESS, NCS 

(20) (50) 

ENDOCRINE SYSTiM
 

fPITUITARY (15) (36)
 

H^aOSgHlGJC CYST
 

* NUMBE " OF ANIMAL S WIT H TISSUE EXAMINE D MICROSCOPICALLY 

* NOliJSR OF AMBAL S NECROPSIE D 

HIGH DOSE
 

11-1311
 

(19)
 

1 (2*)
 

(50)
 

5 (10*)
 

7 (11«)
 

(«6)
 

(«6)
 

2 (IX)
 

(16)
 

3 <7X)
 

(18)
 

1 (2X)
 

3 (6X)
 

1 (2X)
 

(50)
 

32 (6 IX)
 

2 (IX)
 

3 (6X)
 

2 (IX)
 

(37)
 

1 OX)
 

1 (3X)
 

1 <3«)
 

1 (3X)
 

(50)
 

1 (2X)
 

(JO)
 

J-13.Z1
 

C-5 
TABLECl (CONTINUED) 

CONTPOL (UNTR) LOU DOSE HIGH DOSE 

11-13U 5 11-1343 11-1341 

ANGIECTASIS 1 (3X) 

IADPENAL (20) (50) (50) 

MINERALIZATION 1 (2X) 

HEMORBHAGIC CYST 1 (2X) 

ANGIECTASIS 1 (2%) 

*ADRENAL COP1IX (20) (50) (5C) 

CYTOPLASMIC VACUOLIZATION 2 (OK) 

HiPEPPLASIA, NODULAR 1 (2%) 

REPRODUCTIVE SYSTEM 

*SEM^NAL VESICLE (20) (50) (50) 

INFLAMHATICN, ACUTE 1 (2*) 

ITESlIS (20) (50) (50) 

ATROPHY, NCS 1 (2^) 3 (6%) 

tTESj-IS/THBUL! (20) (50) (50) 

DEGENERATION. NOS 1 (2%) 

KERVOUS SYSTEM 

NONE 

SPECIAL SENS E C1GAN S 

NONE 

BUSC'JLOSKSLETAI SYSTE M 

NON E 

BODY I.AVITIES 

•ABDOMINA L CAVIT Y (20) (50) (50) 

THROMBOSIS , NOS 1 (2%) 

*HESLNTERY (20) (50) (50) 

» NUHBE H OF ANIMAL S WITH TISSUE EXAMINE D MICROSCOPICALLY 

* NUMBE R OF AMMAL S NECROPSIE D 

C-6 
TABLECl (CONCLUDED) 

CONT2OL (UHTR) 10» DOSE HIGH DOSE
 

11-1345 11-13«3 11-1311
 

PiRIARTEPIlIS 1 (2»)
 NECPOSIS, FAT 2 <<**) 3 (6*)
 

AiL OIHEP SYStJHS 

*B'JITIPi.E OFGANS (20) (50) (50)
 

LtUKEMOID tEACTION 1 (2*)
 

SPECIAL MOHPdOLOuY SUMMABY
 

NON1
 

« NDMBEE OF &HIHALS WITH TISSUE EXAKIMED MICROSCOPICALLY
 

* N'JMBFR OF ANIMALS NECROPSIID
 

C-7
 
TABLE C2

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN FEMALE RATS 

TREATED WITH NITHIAZIDE 

CONTROL(ONTP) LOW DOSE HIGH DOSE 

11-13116 11-13*4 11-1342 

ANIMALS INITIALLY IN STUDY 20 50 50
 

ANIHALS NECROPSIED 20 50 50
 

ANIHALS EXAMINED HISTOPATHOLOGICALLY** 20 50 50
 

INTEGUMFNTARY SYSTEM
 

NONE
 

RESPIRATORY SYSTEM
 

*LUNG (20) (50) (50) 

MINERALIZATION 1 (2%1 A1ELEC1ASIS 1 (SI) 1 (2X) 

EMBOLUS, SEPTIC 1 (2X) CONGESTION, NOS 1 (S%) 1 (2*) 2 (»«) EDEMA, NOS 2 <«) SDEMA, INTERSTITIAL 1 (5X) 

RuMOPRHAGE 1 (2*) BhONCHOFNEUMONIA, ACOTi 1 (2S) 

PNEUMONIA, CHFONIC IUJRINE 2 (10X) 8 (16%) 10 (20X) 

HYPERPLASIA, EPITHELIAL 1 (255) 

HYPERPLASIA, ADENOMATOUS 2 (U%) 

LEOKOCYTCSIS, NOS 1 (5%) 

HEMATOPOIETIC SYSTEM
 

JSPLEEN (20) (1*9) (50) 

CONGESTI01,, NOS 1 (2*) 1 (2X ) 

HfiMOSIDEFOSIS 3 (15X) 3 (6X) 1 (2%) 

HYPEETHOT>HI, NOS 1 (2)5) HiJlATOFOIISIS 1 (2«) 

*SPL£NIC CAPECL E (20) (50) 

FIBROSIS 1 (2*)

 

IMANUIBOLA P L. NOD E (19) (KB) 

* NUMBE R OF ANIMAL S rflTH IISSUH EXAMINE D MICROSCOPICALLY 

* NUMBhR OF ANIMAL S NECROPSIED 

"EXCLUDES PARTIALLY AUTOLYZED ANIMALS 

p.o V O 
TABLE C2 (CONTINUED) 

HIGH DOSE
 

11-1312
 

(18)
 

1 <2«)
 

(19)
 

(19)
 

2 (IX)
 

18 (37X)
 

1 (2X)
 

(19)
 

(50)
 

1 (2X)
 

1 (2X)
 

2 (IX)
 

1 (2X)
 

1 (2X)
 

7 (11X)
 

1 (2X)
 

2 (IX)
 

(5C)
 

1 (2X)
 

(50)
 

1 (2X)
 

(17)
 

#MES£NTERIC L* NODE
 

HcMOPRHAGI
 

CIRCULATORY SYSTEM
 

#AUSj.CULAR APfENDAGE
 

THROMBOSIS, NOS
 

#MYOl.ARDIUM
 

INFLAMMATION, NOS
 

FIBROSIS
 

FIBROSIS, FOCAL
 

DEGENERATION, GRANULAR
 

f ENDOCARDIUM
 

INFLAMMATION, NOS
 

DIGESIIVE SYSTIM
 

tLIVER
 

CONGESTION, NOS
 

INFLAMMATION, CHRONIC
 

GRANULOMA, NOS
 

DEGENERATION, HYDROPIC
 

N-iCPOSIS, NOS
 

NiCPOSIS, IOCAL
 

NiCPOSIS, ISCHEMIC
 

METAMORPHOSIS FATTY
 

BASOPHILIC CYTO CHANGE
 

EOSINOPHILIC CYTO CHANGE
 

ATYPIA, NCS
 

LaUKOCYTOSIS, NOS
 

HiMATOEOIISIS
 

ILIViR/CSNIPIIOSULAP
 

NkCIOSIS, NOS
 

ATROPHY, NCS
 

*BILi DUCT
 

FIBROSIS
 

HYPERPLASIA, NOS
 

*PANCPEAS
 

CONTROL (ONTR)
 

11-1316
 

(19)
 

(20)
 

(20)
 

9 (15X)
 

1 (5X)
 

(20)
 

(20)
 

1 <5X)
 

1 (5X)
 1 (5%)
 

1 (5X)
 

1 (5X)
 

(20)
 

(20)
 

1 (5X)
 

(19)
 

tOK DOSE
 

11-1311
 

(18)
 

(50)
 

1 (2X)
 

(50)
 

1 (2X)
 

16 (32X)
 

(50)
 

1 (2*)
 

(50)
 

1 (2X)
 

1 (2X)
 

1 (2X)
 

5 (10X)
 

2 (IX)
 

1 (2X)
 

(50)
 

1 <2X)
 

(50)
 

2 (IX)
 

2 (IX)
 

(18)
 

* NUMBER OF ANIMALS KITH TISSUE EXAMINED MICROSCOPICALLY
 

* NIIMdEB OF ANIMALS NECROPSIED
 

C-9
 
TABLE C2 (CONTINUED)
 

CONTROL (UNTR) LOB DOSE HIGH DOSE
 

11-1346 11-1344 11-1342
 

•PANCREATIC	 ACISUS (19) (48) (47)
 

MiTAHOPPHOSIS FATTY 1 (2X)
 

ATROPHY, NOS 1 <5X) 1 (2*) 4 (9*)
 

ATPOPHY, fCCAL 2 (11X)
 

•STOMACH	 (20) (50) (48)
 

AMYLOIDOSIS 2 (4X)
 

HYPERPLASIA, FOCAL 1 (2*)
 

•COLON	 (20) (49) (49)
 

INFLAMMATION, NECROTIZING 1 (2*)
 

INFLAMMATION, ACUTE 1 (2*)
 

PARASITISM 3 (15*) 9 (18*) 114 (29*)
 

DBINAfil SYSTIB
 

•KIDNEY	 (20) (50) (50)
 

MINERALIZATION 1 (2*) 2 (4*)
 

INFLAMMATION, SOS 1 (2X)
 

INFLAMMATION, CHRONIC 6 (30X) 7 (14*) 6 (12*)
 

FIBROSIS 1 (5*)
 

NiPHROPATHY, TOXIC 3 (6*) 1 (2«)
 

PIGMENTATION, NOS 1 <2X)
 

HiMOSIDEBOSIS 1 (2*)
 

ATROPHY, NOS 1 (5*)
 

HYPERPLASIA, TUBOLAB CEIL 1 (2X)
 

•KIDbEY/TUBUL!	 (20) (50) (50)
 DILATATION, NOS 1 (2X)
 

CAST, NOS 1 (2*)
 

DEGENERATION, NOS 1 (5*)
 

•KIDNEY/PELVIS	 (20) (50) (50)
 

INFLAMMATION, ACUTE 1 (2*)
 

•URINARY	 ELfEIER (14) (37) |37)
 

HYPERPLASIA, EPITHELIAL 1 (3*)
 

IHDOCBINE SYSTIB
 

(18) (39) (47)
 

£151* 121
 

* NOBBES OF ANISALS WITH TISSUE EXAMINED BICFOSCOPICALLY
 

« NUMBER OF AHIMALS NECROPSIED
 

C-10
 
TABLE c2 (CONTINUED) 

CONTROL (tJNTH) LOU DOSE HIGH DOSE
 

11-13H6 11-13«<t 11-131(2
 

HSKORRHAGIC CYST « (10X) 14 (91)
 

ANGIECTASIS 1 (21)
 

ttiURENAL (20) (50) («8)
 

CYTOLOGIC EEGiNE?ATION 1 (21)
 

*AD&hN&L CORTEX (20) (50) (48)
 

NiTAMOFPHOSIS FATTY 1 (21) 2 (tS)
 

PIGMENTATION, NOS 1 (21)
 

CYTOLOGIC EEGENESATION 1 (2%) 1 (21)
 

HiEEBPLASI*, FOCAL 1 (2%)
 

*THYKOID (19) (38) (26)
 

HYPERPLASIA, C-CELL 2 (5*) 2 (8*)
 

tITHYROID FOLIICLE (19) (38) (26)
 

DiGENERATICN, CYSTIC 1 (tl)
 

REPRODUCTIVE SYSTEM
 

•MAHdA''* GLAND (20) (50) (50)
 

CYSTIC DUCTS 1 (21)
 HYPERPLASIA, CYSTIC 1 (21)
 

tUTEaUS/ENDCKITRIUM (19) (50) (50)
 

INFLAMMATION, NOS 1 (21)
 HYPERPLASIA, FOCAL 1 (21)
 HYPERPLASIA, CYSTIC 1 (5S)
 

•OVUEY	 (19) (50) (18)
 

CYS1, NOS 1 (21) 1 (21)
 

PA"OVARIAN CYST 2 (111) 1 (21) 3 (61)
 

NEFVOUS SYSTEM
 

*BRAj.N (20) (U9) (U 8)
 INFLAMMATION, NOS 1 (21)
 

SPECIAL SENSE ORGANS
 

NONE
 

MnSClUOSKELFTAL SYSTEM
 

# NUMBER OF ANIMALS WITH TISSUE EXAHINED MICROSCOPICALLY
 

* KUKoES OF ANIMALS NECROPSIFD
 

C-ll
 
TABLE C2 (CONCLUDED) 

CONTROL (UNTR)

11-1346

 LO« DOSE

 11-13i»i»

 HIGH DOSE 

 11-13U2 

EODT CAVITIES 

*HES£NTEHY

NuCROSIS, fAT

 (20) (50)

 1 (2*)

 (5C) 

1 (211) 

Ail OTHER SYSTIMS 

NONI 

SPECIAL MOBPBCIOGY SUBBABY 

NO LESION EEPORTED

AUTO/NECPCESY/HISTO PERF

» SDHBER OF AK1MALS KITH TISSUE EXAHINED MICROSCOPICALLY 

* NUMBER OF ANIMALS NECROPSIED 

3

 2 

2 

C-12
 
APPENDIX D
 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC
 

LESIONS IN MICE TREATED WITH NITHIAZIDE
 

TABLED1

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN MALE MICE 

TREATED WITH NITHIAZIDE 

CONTROL (UNTB) LOR DOSE
 

22-23.45 22-2343
 

ANIMALS INITIAILY IN STUDY 20 50
 ANIMALS MISSING 3
 

ANIMALS NECPOPSIED 20 17
 ABIMALS EXAMIKID HISTOPATHOLOGICALLY ** 20 47
 

INTEGUMENTARY SYSTEM
 

NONE
 

BESPI8ATORY SYSTEM 

•LUNJ/BRONCHICLE 

INFLAMMATION, ACUTE FOCAL 

(20) (11) 

1 (2*) 

1LUNG 

THROMBOSIS, NOS 

CONGESTION, DCS 

HEMORRHAGE 

BiCNCHOINEUMONIA, NOS 

INFLAMMATICN, INTERSTITIAL 

AtiSCESS, NCS 

PNEUMONIA, CHRONIC MUPINE 

PiPIVASCIJLAR CUFFING 

HYPERPLASIA, ALVEOLAR EPITHELIUM 

(20) 

1 (5%) 

2 (10*) 1 (5%) 

(11) 

2

1

2

1

3

3

 (5X) 

 (2X) 

 (5X) 

 (2X) 

 (75!) 

 (7*) 

HEMATOPOIETIC SYSTEM
 

fSPLiEN (19) (16) 

FIBPOSIS 

i«c,CFOSIS. IOCAL 

HYPEBPLASIA , NODULA E 1 (2%) 

HYPERPLASIA , LYMPHOI D 2 (1*) 

HilMATOFOIESIS 1 (5*) 1 (2%) 

#LYBP H NODI (19 ) (13) 

HYPERPLASIA, LYMPHOID 2 <5«) 

*MESENTERIC I. NODE (19 ) (13) 

LYMPHANGJECTiSIS 

# NUMBE R OF ANIMAL S WIT H TISSUE EXAMINE D MICROSCOPICALLY 

* NOMBE P OF AMMAL S NPCFOPSIED 

**EXCLUDES PAKTIALLY AUTOLYZED ANIMALS
 

HIGH DOSE
 

22-2311
 

50
 

6
 

11
 

11
 

(41)
 

(44)
 

1 (2X)
 

2 <5X)
 

1 <2X)
 

2 (5X)
 

7 (16X)
 

1 (2X>
 

(43)
 

1 (2*)
 

1 (2X)
 

(43)
 

(43)
 

D-3
 
TABLE Dl (CONTINUED)
 

CONTROL (IJNTR) LOW DOSE HIGH DOSE
 

22-2345 22-2343 22-2341
 

INFLABBATICN. GRANULOBATOUS	 1 (2«)
 HYPERPLASIA, FETICULUfi CELL 1 (sr.)
 

CIRCULATORY SYSTIM
 

#MYOv-ARDIUB (19) (42) (42)
 

EOSINOPHILIC CYTO CHANGE 1 (2%)
 

DIGESTIVE SYSTEM
 

•LIVKR	 (20) (46) (43)
 

INFLAMMATION, NOS 1 (5)5)
 

AfaSCESS, DCS 1 (2%)
 NfiCROSIS, NOS 1 (5%)
 

NECROSIS. IOCAL 1 (5*)
 MuTAMOHFHCSIS FATTY 1 (2*)
 BASOPHILIC CYTO CHANGS 1 (2X)
 FOCAL CELLULAF CHANGl 1 (2%)
 INCLUSION. CYTOPLASMIC 1 (2*)
 

HYPERPLASIA, NOS 1 (2)5)
 

HYPERPLASIA, FOCAL 1 (2%) 6 (14*)
 

HiMATOEOIESIS 1 (5X)
 

*PASCREAS (19) (45) (43)
 

HiMORRHAGE 1 (5%)
 

HEMORRHAGIC CYST 1 (5*)
 

DEGENERATION, NOS 1 <5X)
 

IPANCPEATIC ACINUS (19) (45) (43)
 

ATROPHY. NCS 1 <2X)
 

ISTOBACH (19) (45) (44)
 

INFLAMMATION, FOCAL 1 (2%)
 

tPEYLRS PATCH (19) (44) (43)
 HYPERPLASIA, LYMPHOID 1 (2*) 3 (7«)
 

#COLCN (19) (42) (44)
 

PARASITISM 8 (42*) 14 (33*) 19 (43*)
 

URINAKY SYSTBM
 

• KIDNE Y (19) (46) (42)
 

«iOHALJZ21ICN_ 

* NUMBER OF ASIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANI1ALS NECROPSIED
 

D-4
 
HYDFCNF.PHEC5IS
 

IdFLAKSATICN. CHRONIC
 

*KIDSEY/CA?SULE
 

INFLAMMATION, ACUTE
 

»UBTtlARY ELftlEH
 

CiST, NOS
 

INFLAMMATION, CHROSIC
 

ENDOCf.INE SYSTZ1
 

*ADFcNAL
 

luFLAHMATICN, ACUTE
 

KADF^NAL COF1HX
 

HYPERPLASIA, NOS
 

HYPERPLASIA, FOCAL
 

itTHYrtOID
 

CiSIIC F01IICLSS
 

A1BOTHY, FCCAL
 

HiPERPLASIA, FCLLIC!ILA"-CELL
 

r-EPFOuUCTIVE SYSTF^
 

*SE1UNAL VESICLE
 

INFLASHAIICN, ACU1E
 

LIPOGRASDICHA
 

kOVOUS SYSTEM
 

KB^A^N
 

MINEBAIIZ^TICN
 

FSABM0.1A BODIES
 

SPECIAL SENSE CPGANS
 

NONn
 

MUSCUi.OSK'JLSTA l SYSTE M 

* N'JildER O F ANIMAL S WIT H TISSU E

* NUfldE? OF ASIMALS NTC"OPSIED 

TABLE Dl (CONTINUED) 

CONTROL (HNTR)
 

22-2345
 

2 (11X)
 

(19)
 

(16)
 

1 (6%)
 

(17)
 

(17)
 

(It)
 

(20)
 

(20)
 

7 (35%)
 

2 (10%)
 

LOW DOSE HIGH DOSE
 

22-23U3 22-2341
 

1 (2%) 1 (2%)
 

(16)	 (42)
 

1 (2*)
 

(36)	 (10)
 

1 (3%)
 

(12)	 (41)
 

1 (2*)
 

(42)	 (11)
 

1 (2%)
 

1 (2%)
 

(29)	 (31)
 

1 (31)
 

1 (3*)
 

1 (3«)
 

(47)	 (44)
 

1	 (2%)
 

1 (2X)
 

(46)	 (44)
 

11	 (24X) 4 (9%)
 

8 (17%) 20 (45*)
 

 EXAMINE D MICROSCOPICALLY 

D-5 
TABLE Dl (CONCLUDED) 

CONTROL (UNTP ) LOW DOSE HIGh DOSE 

22-23IJ5 22-23H3 22-2341 

BODY CAVITIE S 

*ABDOMINA L CAVIT Y (20 ) (U7 ) (<*U ) 

INFLAMMATION , NOS 1 (5*) 

NECROSIS. NOS 1 (5%) 

*MEShNTER Y (20) (47) (U« ) 

NiiCPOSIS, FAT 1 (5%} 

ALL OIHEE SYS1IMS
 

NONi
 

SPECIAL MOFEB0103Y SUMMAPY
 

NO LESION FEEOETED 5 2
 

ANIMAL MISSING/NO NECROPSY 2 6
 

» NUBdE1) OF AMMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMALS NECROPSIED
 

D-6
 
TABLE D2

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN FEMALE MICE 

TREATED WITH NITHIAZIDE 

CONTROL (USTS) LOW DOSE PIGh DOSE
 

22-234.6 22-2344 22-2342
 

ANIMALS INITIALLY IN STUEY 20 50 50
 

ANIMALS MISSING 2 8 4
 

ANIMALS NECROPSIED 18 42 45
 ANIMALS EXAMINED KISTOPATHCLOGICALLY ** 18 42 45
 

INTIGUMENTAF Y SYSTEM
 

*SKIN (18) (42) (.45)
 AoSCESS, CfcRCNIC 1 (2X)
 

RfcSPIhATOSY SYSTEM
 

*LUNli/BFONCHUS (16) (42) (43)
 luFLAMMATICN, ACUTE 1 (2X)
 

tLUNG (16) (42) (43)
 

THROMBUS, ORGANISED 1 (2%)
 

CCNGES1ICN, NOS 1 <2X)
 

INFLAMMATION, INTERSTITIAL 5 (12%) 1 (2%)
 

PKEUHONIA, CHFONIC MUPINE 1 (6%) 4 (10%) 13 (30X)
 

PfiPIVASCULA' CUFFING 3 (1%1 1 (2%)
 

FOAM-CELL 1 (2%)
 

"iGAKAFYCCYTOSIS 1 (2%)
 

HEMATOPOIETIC SYSTEM
 

fSPLnEN (17) (39) (43)
 

M^GAKARYCCYICSIS 1 (3%)
 HYPERPLASIA, LYMPHOID 7 (18X) 2 <5U)
 

»LYMPH NODS (18) (38) (4C)
 

HYPERPLASIA, LYMPHOID 3 (8%)
 

*MESiNT£RIC I. NODE (18) (38) (40)
 

HYPERPLASIA, LYMPHOID 1 (3%)
 

CIRCULATOR Y SYSTE M 

*MYOi_A'=DIU M (18) (36) (40) 

* NUMJ2 3 O F ANIMAL S WIT H TISSUE EXAMINE D MICROSCOPICALLY 

* NUMuE P OF AMMAL S NECROPSIE D "EXCLUDES PARTIALLY AUTOLYZED ANIMALS 

D-7 
TABLE D2 (CONTINUED) 

CONTROL (DNTH) 

22-2346 

LOW DOSE 

22-23«t 

HIGH DOSE 

22-2342 

*COEONARY APTI5Y 

Si-L£ROSIS 

(18) (42) 

1 (2X) 

(45) 

*EULfiONARY AP1ERY 

hlPERTHOFHY, NOS 

HYPERPLASIA, NOS 

(18) (42) 

2 (5%) 

2 (5%) 

(45) 

DIGESTIVE SYS1IM 

iLIVtIP 

INFLAMMATION, ACDTF FOCAL 

PERIVASCULAR CUFFING 

HYPERPLASIA, FOCAL 

hiMATOEOIESIS 

(18) 

1 (6*) 

(41) 

1 (2*) 

1 (2*) 

1 (2%) 

(43) 

2 (5%) 

tSMALL INTESTINE 

ABSCESS, NCS 

(18) (40) (44) 

1 (2%) 

*COION 

IMFLAtlMATICIJ, NECPOTIZING 

PARASITISM 

(18) 

2 (11%) 

(38) 

8 (21*) 

(43) 

1 (2X) 

12 (26%) 

UrlNAitY SYSTEP 

*KIDNEY 

M1NEBALIZA1ION 

HYDEONEPHBCSIS 

LYMPHOCYTIC I NFLAKM AlQi) Y INFILTR 

INFLAMMA1ICN, CHEONIC 

PtPIVASCULSH CUFFING 

(18) 

1

1

 (6%) 

 (6%) 

(41) 

1 (2%) 

1 (2%) 

1 (2",) 

(42) 

3 (7%) 

•UHINAPY ELJEIEP 

HYPERPLASIA, LYMPHOID (15) (32) (35) 

1 (3%) 

ENDOC.-dNE SYSTE.1 

ItADPENAL 

HiKOPPHAGE 

Ki-TAMOPFHCSIS FATTY 

lilfilDOSIS 

(18) 

._

(37) 

1 (31!) 

1 (3X) 

2 J5X) _ 

(36) 

I NUK3E P

* HUMBE"

 OF

 OF

 AMMAL S

 AMUAL S

 WIT H TISSUE

 NECPCPSIE D 

 EXAMINE D MICROSCOPICALL Y 

D-8 
TABLE D2 (CONTINUED) 

CONTRO L (ONTP ) L OU DOSE HIGH DOSE 

22-2346 22-23U4 22-2342 

REPRODUCTIV E SJSTI M 

tUTFR'IS (18) (U2 ) (44 ) 

HYDROMETR A 1 (2% ) 

CYST, SOS 1 (6% ) 3 (7%) 

HEMORRHAG E 1 <2t) 

*CE«VI X U1EF I (18 ) (K2 ) (44)

 

HYPERTROPHY , NO S 1 (2%) 

*UT!.hlIS/ENDC(fITRIOM (18) (U2 ) (44) 

INFLAMMATION , NO S 1 (6% ) 5 (12% ) 1 (2% ) 

HYPERPLASIA , NOS 7 (17%) 5 (11%) 

hYEEPPLASIA , CYSTIC 9 (50%) 23 (55%) 13. (30%) 

»OVAn Y (16) (36) (42 ) 

CiST, NOS 1 (3%) 3 (7%) 

FOLLICULAR CYS1, NOS 1 (6?) 6 (17%) 3 (7%) 

FAROVA8IA N CYST U (25*.) !» (11%) 4 (10X ) 

HYPERPLASIA , CYSTIC 1 (2%) 

KEPVOU S SYSTE K 

tBRAx K (16) (41 ) (45) 

MINERALIZATIO N 3 (19%) 10 (24%) 1 (2% ) 

PEFIVASCUIA R CUFFIN G 1 (2%) 

PSAMMOM A ECDIE S 2 (13%) 5 (12%) 6 (18%) 

SPECIA L SENS E CEGAN S 

*EYE (18) (42) (45) 

CATAPAC T 1 (2% ) 

MUSCULOSKELETA L SYSTE M 

NON E 

EOD Y LAVITIES 

*ABDOMINA L KAI L (18) (42) (45)
 

OFLAIll!AlIC.Ni_CHRONIC_ 

# NUMBER OF ANIMAL S WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMdFR OF ANIMAL S NECROPSIED
 

D-9
 
TABLE D2 (CONCLUDED) 

CONTFOL (ONTE) LOW DOSE HIGH DOSE 

22-2346 22-2314 22-2342 

•PERITONEAL CAVITY (18) (42) (45) 

PiPIAHlEFHIE 1 (2%) 

*«ESiSlE5Y (18) (42) (45) 

NiC^OSIS, FOCAL 1 (655) 

ALL OTHEP SYSTJMS 

•MULTIPLE OEGAN3 (IS) (42) (45) 

AhYLOIBCSIE 1 (2%) 

SPECIAL HOBFHCIOoY SDMMAF Y 

KG LESIO N FSFCPUD 3 2 2 

ASIMA L MISSING/N 2 O HECEOFS Y 8 4 

AUTOLYSIS/N O NECROPS Y 1 

* HHMBE R O F ANI1AL S WIT H TISSUE IXAMINE D HICEOSCOPICALL Y 

E^ OP ANIMAL S NEC?0?SIED 

D-10 
Review of the Bioassay of Nithiazide* for Carcinogenicity
 

by the Data Evaluation/Risk Assessment Subgroup
 

of the Clearinghouse on Environmental Carcinogens
 

October 25, 1978
 

The Clearinghouse on Environmental Carcinogens was established in
 

May, 1976, in compliance with DHEW Committee Regulations and the Provisions of the Federal Advisory Committee Act. The purpose of the
 

Clearinghouse is to advise the Director of the National Cancer Institute
 

(NCI) on its bioassay program to identify and to evaluate chemical
 

carcinogens in the environment to which humans may be exposed. The
 

members of the Clearinghouse have been drawn from academia, industry,
 

organized labor, public interest groups, and State health officials.
 

Members have been selected on the basis of their experience in carcinogenesis or related fields and, collectively, provide expertise in
 

chemistry, biochemistry, biostatistics, toxicology, pathology, and
 

epidemiology. Representatives of various Governmental agencies participate as ad hoc members. The Data Evaluation/Risk Assessment Subgroup
 

of the Clearinghouse is charged with the responsibility of providing a
 

peer review of reports prepared on NCI-sponsored bioassays of chemicals
 

studied for carcinogenicity. It is in this context that the below
 

critique is given on the bioassay of Nithiazide for carcinogenicity.
 

The reviewer for the report on the bioassay of Nithiazide said that
 

the results indicated the compound to be carcinogenic in treated male
 

mice but that the evidence for its carcinogenicity in females was "dubious."
 

Although a statistically significant increase in mammary tumors in treated
 

female rats was observed, he questioned its biological meaningfulness
 

because of the variability in incidence of the tumor type. The reviewer
 

concluded that Nithiazide was not carcinogenic in either sex of treated
 

rats. After briefly describing the experimental design, he noted a
 

nine week interruption in treatment due to the lack of Nithiazide.
 

Despite this shortcoming and the small number of control animals,'he
 

said that the study still appeared to be adequate. Based on the results
 

of the bioassay, the reviewer said that Nithiazide should be considered
 

to pose, at most, a slight carcinogenic risk to humans. There was no
 

objection to a recommendation that the report on the bioassay of Nithiazide
 

be accepted as written.
 

Clearinghouse Members Present
 

Arnold L. Brown (Chairman), University of Wisconsin Medical School
 

Joseph Highland, Environmental Defense Fund
 

William Lijinsky, Frederick Cancer Research Center
 

Henry Pitot, University of Wisconsin Medical Center
 

Verne A. Ray, Pfizer Medical Research Laboratory
 

Kenneth Wilcox, Michigan State Health Department
 

45
 
*	 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.
 

U.S. GOVERNMEN T PRINTIN G OFFICE : 1979-281-217/3065
 

46
 

DHEW (NIH) Publication No. 79-1702