1978 

National Cancer Institute 

CARCINOGENESIS 

Technical Report Series 

NO. 78 

BIOASSAY OF 

ICRF-159 

FOR POSSIBLE CARCINOGENICITY 

CAS No. 21416-87-5 

NCI-CG-TR-78 

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

Public Health Service 

National Institutes of Health 

BIOASSAY OF
 

ICRF-159
 

FOR POSSIBLE CARCINOGENICITY
 

Carcinogenesis Testing Program
 

Division of Cancer Cause and Prevention
 

National Cancer Institute
 

National Institutes of Health
 

Bethesda, Maryland 20014
 

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

Public Health Service
 

National Institutes of Health
 

DHEW Publication No. (NIH) 78-1328
 

BIOASSAY OF
 

ICRF-159
 

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
 

ICRF-159 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 greater incidence of cancer than control animals,
 

do not necessarily mean that the test chemical is not a
 

carcinogen, inasmuch as 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 that exposure to the chemical
 

is a potential risk to man. The actual determination of the risk
 

to man from animal carcinogens requires a wider analysis.
 

CONTRIBUTORS; This bioassay of ICRF-159 was conducted by
 

Southern Research Institute, Birmingham, Alabama, initially under
 

direct contract to NCI and currently under a subcontract to
 

Tracer Jitco, Inc., prime contractor for the NCI Carcinogenesis
 

Testing Program.
 

The experimental design and doses were determined by Drs. D. P.
 

Griswold1, J. D. Prejean1, E. K. Weisburger2, and J. H.
 

Weisburger2>3. MS. J. Belzer^- and Mr. I. Brown1 were responsible
 

for the administration of the chemical and the care of the
 

laboratory animals. Data management and retrieval were performed
 

by Ms. C. A. Dominick1. Histopathologic examinations were
 

performed by Drs. R. B. Thompson^ and J. C. Peckhaml, and the
 

diagnoses included in this report represent their interpretation.
 

iii 
Animal pathology tables and survival tables were compiled by EG&G
 

Mason Research Institute^. The statistical analyses were performed by Dr. J. R. Joiner-*, using methods selected for the
 

bioassay program by Dr. J. J. Cart". Chemicals used in this
 

bioassay were analyzed by Mr. A. R. Chamberlin', Mr. G. L. long',
 

and Mr. P. Lim^, and the results of the analyses were reviewed by
 

Dr. C. W. Jameson^.
 

This report was prepared at Tracer Jitco^ under the direction of
 

NCI. Those responsible for the report at Tracer Jitco were
 

Dr. Marshall Steinberg, Director of the Bioassay Program; Dr. L.
 

A. Campbell, Deputy Director for Science; Drs. J. F. Robens and
 

C. H. Williams, toxicologists; Dr. R. L. Schueler, pathologist;
 

Dr. G. L. Miller, Ms. L. A. Waitz, and Mr. W. D. Reichardt,
 

bioscience writers; and Dr. E. W. Gunberg, technical editor,
 

assisted by Ms. Y. E. Presley.
 

The statistical analysis was reviewed by members of the
 

Mathematical Statistics and Applied Mathematics Section of NCI6:
 

Dr. John J. Gart, Mr. Jun-mo Nam, Dr. Hugh M. Pettigrew, and Dr.
 

Robert E. Tarone.
 

The following other scientists at NCI were responsible for
 

evaluating the bioassay experiment, interpreting the results, and
 

reporting the findings:
 

Dr. Kenneth C. Chu
 

Dr. Cipriano Cueto, Jr.
 

Dr. J. Fielding Douglas
 

Dr. Dawn G. Goodman
 

Dr. Richard A. Griesemer
 

Dr. Harry A. Milman
 

Dr. Thomas W. Orme
 

Dr. Robert A. Squire8
 

Dr. Jerrold M. Ward
 

^•Southern Research Institute, 2000 Ninth Avenue South,
 

Birmingham, Alabama.
 

iv
 
^Carcinogenesis Testing Program, Division of Cancer Cause and
 

Prevention, National Cancer Institute, National Institutes of
 

Health, Bethesda, Maryland.
 

%ow with The Naylor Dana Institute for Disease Prevention,
 

American Health Foundation, Hammond House Road, Valhalla,
 

New York.
 

^EG&G Mason Research Institute, 1530 East Jefferson Street,
 

Rockville, Maryland.
 

-^Tracer Jitco, Inc., 1776 East Jefferson Street, Rockville,
 

Maryland.
 

"Mathematical Statistics and Applied Mathematics Section,
 

Biometry Branch, Division of Cancer Cause and Prevention,
 

National Cancer Institute, National Institutes of Health,
 

Bethesda, Maryland.
 

^Stanford Research Institute, 333 Ravenswood Avenue, Menlo Park,
 

California.
 

%ow with the Division of Comparative Medicine, Johns Hopkins
 

University, School of Medicine, Traylor Building, Baltimore,
 

Maryland.
 

v
 

SUMMARY
 

A bioassay of the experimental anticancer drug ICRF-159 for
 

possible carcinogenicity was conducted by administering the
 

compound by intraperitoneal injection to Sprague-Dawley rats and
 

B6C3F1 mice.
 

Groups of 35 rats and 35 mice of each sex were injected three
 

times per week with ICRF-159 in buffered saline at one of the
 

following doses, either 48 or 96 mg/kg body weight for the rats
 

and either 40 or 80 mg/kg body weight for the mice. Both rats
 

and mice were dosed for 52 weeks, then observed for 29-34
 

additional weeks. Untreated-control and vehicle-control groups
 

each consisted of 10 rats and 15 mice of each sex; pooled-control
 

groups consisted of the 10 vehicle controls of each sex of the
 

rats combined with 30 vehicle controls of each sex of rats from
 

similar bioassays of three other chemicals and the 15 vehicle
 

controls of each sex of the mice combined with 30 vehicle
 

controls of each sex of mice from similar bioassays of two other
 

chemicals. All surviving rats were killed at 81-86 weeks; all
 

surviving mice, at 86 weeks.
 

Mean body weights were depressed in rats and mice administered
 

ICRF-159, and mortality was dose related among male and female
 

rats and male mice. The high mortality among the male rats may
 

have been associated with inflammatory lesions observed in the
 

lungs, the liver, and the pleural and peritoneal cavities.
 

Sufficient numbers of female rats and of both male and female
 

mice were at risk for development of late-appearing tumors. In
 

the male rats, time-adjusted analysis of the incidence of tumors
 

was used for determining statistical significance.
 

In female rats, the incidence of uterine adenocarcinomas was
 

higher in the low- and high-dose groups (P < 0.001) than in the
 

pooled controls (controls 0/38, low-dose 10/33, high-dose 11/32);
 

the incidence was also dose related (P < 0.001). In male rats,
 

no tumors occurred in the dosed groups in a significantly
 

increased incidence.
 

vii
 
In female mice, the incidence of all hematopoietic neoplasms
 

(histiocytic lymphomas, lymphocytic lymphomas, or lymphocytic
 

leukemias), taken together, was higher in the low-dose group (P =
 

0.038) and in the high-dose group (P = 0.002) than in the pooled
 

controls (controls 1/45, low-dose 5/31, high-dose 9/34); the
 

incidence was also dose related (P = 0.002). In addition, the
 

incidence of these tumors in the high-dose group was higher (P =
 

0.026) than that in the vehicle controls (0/15), and the
 

incidence was dose related (P = 0.021) using the vehicle
 

controls. In male mice, lymphocytic neoplasms occurred only in
 

two low-dose and two high-dose animals.
 

It is concluded that under the conditions of this bioassay,
 

ICRF-159 was carcinogenic for female Sprague-Dawley rats,
 

producing uterine adenocarcinomas, and was also carcinogenic for
 

female B6C3F1 mice, producing lymphomas.
 

viii
 
TABLE OF CONTENTS
 

Page
 

I. Introduction 1
 

II. Materials and Methods 3
 

A. Chemical 3
 

B. Dosage Preparation 3
 

C. Animals 4
 

D. Animal Maintenance 4
 

E. Subchronic Studies 7
 

F. Designs of Chronic Studies 9
 

G. Clinical and Pathologic Examinations 12
 

H. Data Recording and Statistical Analyses 13
 

III. Results - Rats 19
 

A. Body Weights and Clinical Signs (Rats) 19
 

B. Survival (Rats) 19
 

C. Pathology (Rats) 22
 

D. Statistical Analyses of Results (Rats) 24
 

IV. Results -Mice 27
 

A. Body Weights and Clinical Signs (Mice) 27
 

B. Survival (Mice) 27
 

C. Pathology (Mice) 30
 

D. Statistical Analyses of Results (Mice) 32
 

V. Discussion 35
 

VI. Bibliography 37
 

APPENDIXES
 

Appendix A Summary of the Incidence of Neoplasms
 

in Rats Given Intraperitoneal Injections
 

of ICRF-159 39
 

Table Al Summary of the Incidence of Neoplasms
 

in Male Rats Given Intraperitoneal
 

Injections of ICRF-159 41
 

ix
 
Table A2


Appendix B


Table Bl


Table B2


Appendix C


Table Cl


Table C2


Appendix D


Table Dl


Table D2


Appendix E


Page
 

 Summary of the Incidence of Neoplasms
 

in Female Rats Given Intraperitoneal
 

Injections of ICRF-159 '44
 

 Summary of the Incidence of Neoplasms
 

in Mice Given Intraperitoneal Injections
 

of ICRF-159 -49
 

 Summary of the Incidence of Neoplasms
 

in Male Mice Given Intraperitoneal
 

Injections of ICRF-159 '.
 51
 

 Summary of the Incidence of Neoplasms
 

in Female Mice Given Intraperitoneal
 

Injections of ICRF-159 5'.
 4 

 Summary of the Incidence of Nonneoplastic
 

Lesions in Rats Given Intraperitoneal
 

Injections of ICRF-159 I57
 

 Summary of the Incidence of Nonneoplastic
 

Lesions in Male Rats Given Intraperitoneal
 

Injections of ICRF-159 !59
 

 Summary of the Incidence of Nonneoplastic
 

Lesions in Female Rats Given Intraperitoneal 1
 

Injections of ICRF-159 i64
 

 Summary of the Incidence of Nonneoplastic
 

Lesions in Mice Given Intraperitoneal
 

Injections of ICRF-159 i69
 

 Summary of the Incidence af Nonneoplastic
 

Lesions in Male Mice Given Intraperitoneal
 

Injections of ICRF-159 71
 

 Summary of the Incidence of Nonneoplastic
 

Lesions in Female Mice Given Intraperitoneal 1
 

Injections of ICRF-159 74
 

 Analyses of the Incidence of Primary
 

Tumors in Rats Given Intraperitoneal
 

Injections of ICRF-159 79
 

x
 
Table El
 

Table E2
 

Table E3
 

Appendix F
 

Table Fl
 

Table F2
 

Table 1
 

Table 2
 

Figure 1
 

Figure 2
 

Figure 3
 

Figure 4
 

Page
 

Analyses of the Incidence of Primary
 

Tumors in Male Rats Given Intraperitoneal
 

Injections of ICRF-159 , 81
 

Analyses of the Incidence of Primary
 

Tumors in Female Rats Given Intraperitoneal L
 

Injections of ICRF-159 . 84
 

Time-adjusted Analyses of the Incidence
 

of Primary Tumors in Male Rats
 

Given Intraperitoneal Injections of
 

ICRF-159 . 88
 

Analyses of the Incidence of Primary
 

Tumors in Mice Given Intraperitoneal
 

Injections of ICRF-159 , 91
 

Analyses of the Incidence of Primary
 

Tumors in Male Mice Given Intraperitoneal
 

Injections of ICRF-159 . 93
 

Analyses of the Incidence of Primary
 

Tumors in Female Mice Given Intraperitoneal L
 

Injections of ICRF-159
 

TABLES
 

Design of Chronic Studies of
 

ICRF-159 in Rats
 

Design of Chronic Studies of
 

ICRF-159 in Mice
 

FIGURES
 

Growth Curves for Rats Treated
 

with ICRF-159
 

Survival Curves for Rats Treated
 

with ICRF-159
 

Growth Curves for Mice Treated with
 

ICRF-159
 

Survival Curves for Mice Treated with
 

ICRF-159
 

, 95
 

. 10
 

. 11
 

. 20
 

. 21
 

. 28
 

xi
 

I. INTRODUCTION
 

ICRF-159 (CAS 21416-87-5; NSC 129943; NCI C01627) is an
 

experimental anticancer drug, developed by the Imperial Cancer
 

Research Fund in England and tested in Phase II clinical trials
 

in the United States. ICRF-159 has been administered to patients
 

with acute leukemias (Hellmann, 1970; Mathe" et al., 1970) and to
 

patients with tumors of the lung, breast, or colon (Carter and
 

Slavik, 1976). Unlike other cytotoxic drugs, ICRF-159 inhibits
 

cell growth only in the stages following DNA synthesis and
 

preceding mitosis (Sharpe and Field, 1970). It is most toxic to
 

cells that divide frequently, since they are likely to be found
 

in these stages. This drug has been shown to inhibit the
 

spontaneous metastasis of an experimental mouse tumor (Salsbury
 

et al., 1970).
 

ICRF-159 is one in a series of anticancer compounds that were
 

selected for testing in the Carcinogenesis Testing Program to
 

assess the possible carcinogenic effects of drugs that may be
 

administered to humans on a chronic basis.
 

1
 

II. MATERIALS AND METHODS
 

A. Chemical
 

The chemical (+)bis-4,4'-(l-methyl-l,2-ethanediyl)-2,6-piperazinedione, commonly called ICRF-159, was obtained for the chronic
 

study in a single batch (Lot No. PD/AS 5014/71) from the Imperial
 

Chemical Industries, LTD., Cheshire, England. The purity of this
 

batch was determined to be 98-99% by non-aqueous potentiometric
 

titration, chromatography, and elemental analysis (C,H,N,0) at
 

the Stanford Research Institute. The melting point was 229-233°C
 

with decomposition, similar to the value of 223°C reported
 

elsewhere (Wasserman et al. , 1973). Thin-layer chromatography,
 

comparing the test material with a mixture of hydrolysis
 

products, indicated 1-2% hydrolysis. Nuclear magnetic resonance,
 

infrared, and ultraviolet spectra were consistent with the
 

structure.
 

The chemical was stored in the presence of a desiccant
 

(Drierite®) at 5°C.
 

B. Dosage Preparation
 

Suspensions of ICRF-159 were prepared fresh each day that the
 

chemical was administered. The chemical was suspended in a
 

buffered saline vehicle by mixing in a Potter-Elvehjem tissue
 

3 
grinder. The buffered saline vehicle (pH = 6.9) contained 0.85%
 

NaCl, 0.40% NaH2P04, and 0.65% Na2HP04.
 

C. Animals
 

For the subchronic studies, female Sprague-Dawley rats and male
 

Swiss mice were obtained from Charles River Breeding
 

Laboratories, Inc., Wilmington, Massachusetts.
 

For the chronic tests, Sprague-Dawley rats and B6C3F1 mice of
 

each sex were obtained from Charles River Breeding Laboratories.
 

All animals were supplied through a contract with the Division of
 

Cancer Treatment, National Cancer Institute. Male rats were
 

received at the test laboratory at 29 days of age, female rats at
 

36 days of age, and male and female mice at 30 days of age. On
 

arrival at the laboratory, all animals were quarantined for
 

approximately 1 week. Animals with no visible signs of disease
 

were assigned to control or dosed groups and earmarked for
 

individual identification.
 

D. Animal Maintenance
 

The animals were housed in temperature- and humidity-controlled
 

rooms. The temperature range was 20-24°C, and the relative
 

humidity was maintained at 40-60%. The air was changed 15 times
 

per hour and passed through both intake and exhaust fiberglass
 

4 
roughing filters. In addition to natural light, illumination was
 

provided by fluorescent lighting for 9 hours each day. Wayne*51
 

Lab Blox animal feed (Allied Mills, Inc., Chicago, 111.) and
 

water were made available ad libitum and replenished daily.
 

Rats were housed five per cage and mice seven per cage in solid-


bottom stainless steel cages (Hahn Roofing and Sheet Metal Co. ,
 

Birmingham, Ala.). The rat cages were provided with Iso-Dri
 

hardwood chip bedding (Carworth, Edison, N.J.), and the cage tops
 

were covered with disposable filter bonnets, beginning at week
 

25; mouse cages were provided with Sterolit® clay bedding
 

(Englehard Mineral and Chemical Co., New York, N.Y.). Bedding
 

was replaced once per week; cages, water bottles, and feeders
 

were sanitized at 82°C once per week; and racks were cleaned once
 

per week.
 

The rats and mice were housed in separate rooms. Control animals
 

were housed with respective dosed animals. Animals administered
 

ICRF-159 were maintained in the same room in which the following
 

chemicals were on test:
 

RATS
 

Gavage Studies 

cholesterol (p-(bis(2-chloroethyl)amino)phenyl)acetate 

(phenesterin) (CAS 3546-10-9) 

estradiol bis((p-(bis(2-chloroethyl)amino)phenyl)acetate) 

(estradiol mustard) (CAS 22966-79-6) 

5 
Intraperitoneal Injection Studies
 

4'-(9-acridinylamino)methansulfon-m-aniside monohydrochloride
 

(MAAM) (NSC 141549)
 

acronycine (CAS 7008-42-6)
 

5-azacytidine (CAS 320-67-2)
 

beta-2'-deoxy-6-thioguanosine monohydrate (beta-TGdR)
 

(CAS 789-61-7)
 

1,4-butanediol dimethanesulfonate (busulfan) (CAS 55-98-1)
 

emetine dihydrochloride tetrahydrate (CAS 316-42-7)
 

3,3'-iminobis-l-propanol dimethanesulfonate (ester)
 

hydrochloride [IPD] (CAS 3458-22-8)
 

N,3-bis(2-chloroethyl)tetrahydro-2H-l,3,2-oxazaphosphorin-2­

amine-2-oxide (isophosphamide) (CAS 3778-73-2)
 

N-(2-chloroethyl)-N-(l-methyl-2-phenoxyethyl)benzylamine
 

hydrochloride (phenoxybenzamine hydrochloride) (CAS 63-92-3)
 

N-(l-methylethyl)-4-((2-methylhydrazino)methyl)benzamide
 

monohydrochloride (procarbazine) (CAS 366-70-1)
 

tris(l-aziridinyl)phosphine sulfide (thio-TEPA) (CAS 52-24-4)
 

2,4,6-tris(dimethylamino)-s-triazine (CAS 645-05-6)
 

MICE
 

Feed Studies
 

4-acetyl-N-((cyclohexylamino)carbonyl)benzenesulfonamide
 

(acetohexamide) (CAS 968-81-0)
 

anthranilic acid (CAS 118-92-3)
 

l-butyl-3-(p-tolylsulfonyl)urea (tolbutamide) (CAS 64-77-7)
 

4-chloro-N-((propylamino)carbonyl)benzenesulfonamide
 

(chlorpropamide) (CAS 94-20-2)
 

5-(4-chlorophenyl)-6-ethyl-2,4-pyrimidinediamine
 

(pyrimethamine) (CAS 58-14-0)
 

2,6-diamino-3-(phenylazo)pyridine hydrochloride (phenazopyridine
 

hydrochloride) (CAS 136-40-3)
 

L-tryptophan (CAS 73-22-3)
 

N-9H-fluoren-2-ylacetamide (CAS 53-96-3)
 

N-(p-1 oluenesulf onyl)-N'-hexame thyleniminourea
 

(tolazamide) (CAS 1156-19-0)
 

1-phenethylbiguanide hydrochloride (phenformin) (CAS 114-86-3)
 

pyrazinecarboxamide (pyrazinamide) (CAS 98-96-4)
 

4,4'-sulfonyldianiline (dapsone) (CAS 80-08-0)
 

4,4'-thiodianiline (CAS 139-65-1)
 

ethionamide (CAS 536-33-4)
 
Gavage Studies
 

cholesterol (p-(bis(2-chloroethyl)amino)phenyl)acetate
 

(phenesterin) (CAS 3546-10-9)
 

estradiol bis((p-(bis(2-chloroethyl)amino)phenyl)acetate)
 

(estradiol mustard) (CAS 22966-79-6)
 

Intraperitoneal Injection Studies
 

4'-(9-acridinylamino)methansulfon-m-aniside monohydrochloride
 

(MAAM) (NSC 141549)
 

acronycine (CAS 7008-42-6)
 

5-azacytidine (CAS 320-67-2)
 

beta-2'-deoxy-6-thioguanosine monohydrate (beta-TGdR)
 

(CAS 789-61-7)
 

1,4-butanediol dimethanesulfonate (busulfan) (CAS 55-98-1)
 

emetine dihydrochloride tetrahydrate (CAS 316-42-7)
 

3,3'-iminobis-l-propanol dimethanesulfonate (ester)
 

hydrochloride [IPD] (CAS 3458-22-8)
 

N,3-bis(2-chloroethyl)tetrahydro-2H-l,3,2-oxazaphosphorin-2­

amine-2-oxide (isophosphamide) (CAS 3778-73-2)
 

N-(2-chloroethyl)-N-(l-methyl-2-phenoxyethyl)benzylamine
 

hydrochloride (phenoxybenzamine hydrochloride) (CAS 63-92-3)
 

N-(l-methylethyl)-4-((2—methylhydrazino)methyl)benzamide
 

monohydrochloride (procarbazine) (CAS 366-70-1)
 

tris(l-aziridinyl)phosphine sulfide (thio-TEPA) (CAS 52-24-4)
 

2,4,6-tris(dimethylamino)-s-triazine (CAS 645-05-6)
 

E. Subchronic Studies
 

Subchronic studies were conducted using female Sprague-Dawley
 

rats and male Swiss mice to estimate the maximum tolerated doses
 

of ICRF-159, on the basis of which "low" and "high" doses were
 

determined for administration in the chronic studies. The rats
 

were administered doses of 1.2, 3, 6, 12, 24, 48, 96, 192, or 384
 

mg/kg body weight; the mice were administered doses of 2, 5, 10,
 

20, 40, 80, 160, 320, or 640 mg/kg body weight. Dosed animals
 

were injected intraperitoneally with ICRF-159 three times per
 

7
 
week for 45 days, then observed for an additional 45 days. Five
 

animals of each species were injected with the chemical at each
 

dose, 10 animals of each species were injected with the vehicle
 

alone (vehicle controls), and 10 animals of each species left
 

untreated (untreated controls).
 

In rats, death occurred by week 3 in 4/5 animals administered 384
 

mg/kg and by week 7 in 1/5 animals administered 192 mg/kg. No
 

animals died at any lower doses. At the end of the administration period, body weight gains were not affected in animals
 

administered 1.2 and 3 mg/kg. Weight gains in animals at higher
 

doses were variable, ranging from 52-86% of control values and
 

showing no dose-related trend. At the highest dose administered,
 

the mean weight gain was 78% of that of controls. All animals
 

having low weight gains at the end of the administration period
 

had mean weight gains comparable to those of controls by the end
 

of the study. No gross abnormalities were noted at necropsy.
 

The low and high doses for the chronic studies using rats were
 

set at 48 mg/kg and 96 mg/kg.
 

In mice, death occurred by week 3 in 2/5 animals administered 5
 

mg/kg and within the first 3 weeks of the study in all animals
 

administered 160 mg/kg and above. Mean body weight gains in the
 

surviving dosed animals were not affected except at a dose of 80
 

mg/kg, where the weight gains in the dosed animals were 40% lower
 

8
 
than that in the controls at the end of the administration
 

period. No gross abnormalities were found at necropsy. The low
 

and high doses for the chronic studies using mice were set 40
 

mg/kg and 80 mg/kg.
 

F. Designs of Chronic Studies
 

The designs of the chronic studies are shown in tables 1 and 2.
 

Since the numbers of rats and mice in the vehicle-control groups
 

were small, pooled vehicle-control groups also were used for
 

statistical comparisons. The groups of 10 vehicle-control rats
 

and 15 vehicle-control mice of each sex from the current bioassay
 

of ICRF-159 were combined with corresponding groups of 10
 

vehicle-control rats of each sex from similar bioassays of
 

procarbazine, thio-TEPA, and 3,3'-iminobis-l-propanol dimethanesulfonate (ester) hydrochloride [IPD] and of 15 vehicle-control
 

mice of each sex from similar bioassays of thio-TEPA and IPD to
 

give pooled groups of 40 vehicle-control rats and 45 vehicle-


control mice of each sex. The vehicle-control animals that were
 

used in the pooled-control groups were of the same strains
 

(Sprague-Dawley rats and B6C3F1 mice) and from the same supplier,
 

and they were examined by the same patholegists; furthermore, the
 

different control groups received the same vehicle for injection
 

9 
Table 1. Design of Chronic Studies of ICRF-159 in Rats
 

Sex and 

Test 

Group 

Initial 

No. of 

Animals3 

ICRF-159 

Doses 

(mg/kg)b 

Time 

Dosed 

(weeks ) 

on Study 

Observed 

(weeks ) 

Male 

Untreated-Control 

Vehicle-Control 

Low-Dose 

High-Dose 

10 

10 

35 

35 

0 

Oc 

48 

96 

52 

52 

52 

85 

33 

32 

2: 

Female 

Untreated Control 

Vehicle-Control 

Low-Dose 

High-Dose 

10 

10 

35 

35 

0 

Oc 

48 

96 

52 

52 

52 

85-86 

33 

32-33 

29 

a

Male rats were 35 days of age and female rats were 42 days of age
 

when placed on study.
 

"ICRF-159 was administered by intraperitoneal injection three
 

times per week in buffered saline, at a volume of 0.25 ml/100 g
 

body weight. Doses were based on individual weights.
 

c

Vehicle-control groups received only buffered saline solution,
 

at the same volume as dosed groups.
 

10 
Table 2. Design of Chronic Studies of ICRF-159 in Mice
 

Sex and 

Test 

Group 

Initial 

No. of 

Animals3 

ICRF-159 

Doses 

(mR/kg)b 

Time 

Dosed 

(weeks ) 

on Study 

Observed 

(weeks ) 

Male 

Untreated-Control 

Vehicle-Control 

Low-Dose 

High-Dose 

15 

15 

35 

35 

0 

Oc 

40 

80 

52 

52 

52 

86 

34 

34 

34 

Female 

Untreated Control 

Vehicle-Control 

Low-Dose 

High-Dose 

15 

15 

35 

35 

0 

Oc 

40 

80 

52 

52 

52 

86 

34 

34 

34 

a

All mice were 35 days of age when placed on study.
 

klCRF-159 was administered by intraperitoneal injection three
 

times per week in buffered saline, at a volume of 1 ml/100 g body
 

weight. Doses were based on the mean weight of the animals in
 

each cage.
 

c

Vehicle-control groups received only buffered saline solution,
 

at the same volume as dosed groups.
 

11
 
and were placed on study at starting times differing by no more
 

than 3 weeks.
 

G. Clinical and Pathologic Examinations
 

All animals were observed twice daily for signs of toxicity, and
 

those animals appearing moribund were killed and necropsied.
 

Rats were weighed individually each week for 2 months and every 2
 

weeks thereafter. Palpation for masses was carried out at each
 

weighing.
 

The pathologic evaluation consisted of gross and microscopic
 

examination of major tissues, major organs, and all gross lesions
 

from killed animals and from animals found dead. The following
 

tissues were examined microscopically: skin, muscle, lungs and
 

bronchi, trachea, bone and bone marrow, spleen, lymph nodes,
 

thymus, heart, salivary gland, liver, gallbladder and bile duct
 

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

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

parathyroid, mammary gland, prostate or uterus, testis or ovary,
 

brain, and sensory organs. Peripheral blood smears were prepared
 

from each animal whenever possible. Occasionally, additional
 

tissues were also examined microscopically. The different
 

tissues were preserved in 10% buffered formalin, embedded in
 

paraffin, sectioned, and stained with hematoxylin and eosin.
 

12
 
Special staining techniques were utilized when indicated for more
 

definitive diagnosis.
 

A few tissues from some animals were not examined, particularly
 

from those animals that died early. Also, some animals were
 

missing, cannibalized, or judged to be in such an advanced state
 

of autolysis as to preclude histopathologic evaluation. Thus,
 

the number of animals for which particular organs or tissues were
 

examined microscopically varies, and does not necessarily
 

represent the number of animals that were placed on study in each
 

group.
 

H. Data Recording and Statistical Analyses
 

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

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

clinical observations, survival, body weight, and individual
 

pathologic results, as recommended by the International Union
 

Against Cancer (Berenblum, 1969). Data tables were generated for
 

verification of data transcription and for statistical review.
 

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

13
 
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) for testing two groups for equality and Tarone's (1975)
 

extensions of Cox's methods for testing for 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 is 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
 

14
 
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) was used to compare the tumor incidence of
 

a control group with that of a group of dosed animals at each
 

dose level. When results for a number of dosed 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) requires that the
 

P value for any comparison be less than or equal to 0.05/k. In
 

cases where this correction was used, it is discussed in the
 

narrative section. It is not, however, presented in the tables,
 

where the Fisher exact P values are shown.
 

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

continuity correction (Armitage, 1971), was also used. Under the
 

assumption of a linear trend, this test determines 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 is a positive dose relation15
 
ship. This method also provides a two-tailed test of departure
 

from linear trend.
 

A time-adjusted analysis was applied when numerous early deaths
 

resulted from causes that were not associated with the formation
 

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

first tumor was observed were excluded by basing the statistical
 

tests on animals that survived at least 52 weeks, unless a tumor
 

was found at the anatomic site of interest before week 52. When
 

such an early tumor was found, comparisons were based exclusively
 

on animals that survived at least as long as the animal in which
 

the first tumor was found. Once this reduced set of data was
 

obtained, the standard procedures for analyses of the incidence
 

of tumors (Fisher exact tests, Cochran-Armitage tests, etc.) were
 

followed.
 

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

incidence of tumors. Curves of the proportions surviving without
 

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

The week during which an animal died naturally or was 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
 

16 
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 with its control was calculated
 

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

relative risk is defined as p^/Pc where pt is the true binomial
 

probability of the incidence of a specific type of tumor in a
 

dosed 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 dosed 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
 

dosed 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% of a large number of identical experiments, the true
 

ratio of the risk in a dosed 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 signifi17
 
cant result (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. RESULTS - RATS
 

A. Body Weights and Clinical Signs (Rats)
 

Mean body weights of both male and female dosed rats were lower
 

than those of either vehicle or matched controls, with the
 

weights of the high-dose rats being slightly lower than those of
 

the low- dose rats (figure 1). Fluctuation in the growth curves
 

may be due to mortality; as the size of a group diminishes, the
 

mean body weight may be subject to wide variation.
 

There were no other clinical signs recorded which were indicative
 

of drug-related toxicity in the rats.
 

B. Survival (Rats)
 

The Kaplan and Meier curves estimating the probabilities of
 

survival for male and female rats administered ICRF-159 by
 

intraperitoneal injection at the doses of this bioassay, together
 

with those of the untreated and vehicle controls, are shown in
 

figure 2.
 

The result of the Tarone test for positive dose-related trend in
 

mortality over the period of the bioassay is significant (P <
 

0.001) in each sex. In male rats, 10/10 (100%) of the untreated
 

controls, 9/10 (90%) of the vehicle controls, 24/36 (67%) of the
 

low-dose group, and 8/35 (23%) of the high-dose group lived at
 

19
 
UNTREATED CONTROL 

D VEHICLE CONTROL 

O LOW DOSE 

A HIGH DOSE 

TIME ON STUDY (WEEKS) 

Figure 1. Growth Curves For Rats Treated With ICRF-159 

20 
TIME ON STUDY (WEEKSr 

Figure 2. Survival Curves For Rats Treated With ICRF-159 

21 
least as long as week 52. In female rats, 10/10 (100%) of the
 

untreated controls, 9/10 (90%) of the vehicle controls, 32/34
 

(94%) of the low-dose group, and 26/35 (74%) of the high-dose
 

group were alive at week 52. Sufficient numbers of females were
 

at risk for development of tumors.
 

C. Pathology (Rats)
 

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 an increased incidence of uterine neoplasms in dosed
 

female rats.
 

FEMALE RATS
 

Untreated Vehicle Low High 

Site/Neoplasm Control Control Dose Dose 

Uterus 

Number of animals with tissues 

examined microscopically (10) (10) (33) (32) 

Adenocarcinoma, NOS* 

Endometrial stromal polyp 

0 

0 

0 

1 

10 

2 

11 

0 

Cervix
 

Number of animals with tissues 

examined microscopically (10) (10) (33) (32) 

Squamous-cell carcinoma 0 0 0 1 

*Not otherwise specified
 

The uterine tumors were primarily adenocarcinomas arising from
 

the endometrial glandular epithelium. These tumors were
 

22
 
characterized by neoplastic epithelial cells having large
 

vesicular nuclei, prominent eosinophilic nucleoli, and moderate
 

amounts of eosinophilic cytoplasm. The tumor cells formed
 

glands, ducts, and acini that were separated by a fibrovascular
 

stroma. The neoplastic glandular tissue originated in the
 

endometrium and projected into the uterine lumen and also
 

infiltrated the overlying muscle layers of the uterine wall. The
 

neoplastic cells had frequently penetrated the serosal surface of
 

the uterus and transplanted to multiple sites throughout the
 

abdominal cavity. Metastases were found at multiple sites in the
 

abdominal cavity of the females, but no tumors of the abdominal
 

cavity were observed in the dosed or control males. The
 

increased incidence in the females appears to be the result of
 

administration of the test chemical.
 

Endometrial glandular hyperplasias occurred in 1/32 (3%)
 

high-dose females and 1/33 (3%) low—dose females. Inflammatory
 

lesions of the uterus occurred in both dosed and control rats.
 

The incidence of suppurative uterine lesions were:
 

untreated-control females 5/10 (50%), vehicle-control females
 

1/10 (10%), low-dose females 20/33 (61%) and high-dose females
 

11/32 (34%).
 

With the exception of uterine tumors, neoplasms listed in
 

Appendix A occurred with approximately equal frequency in control
 

23
 
and dosed rats or occurred in insignificant numbers. All
 

neoplasms have been encountered previously as spontaneous lesions
 

in Sprague-Dawley rats.
 

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

proliferative, and inflammatory changes were encountered in
 

animals of the dosed and control groups (Appendix C). These
 

nonneoplastic lesions are commonly seen in aged Fischer 344 rats.
 

The small number of animals bearing tumors in the high-dose group
 

of male rats (4/30 [13%]) compared with the number in the low-


dose group (11/33 [33%]) may have been due to the decreased life
 

span in the high-dose males. The increased mortality in the
 

high-dose males may have been associated with inflammatory
 

lesions in the lungs, liver, and serous cavities (pleural and
 

peritoneal).
 

Based on the histologic examination, the administration of
 

ICRF-159 by intraperitoneal injection to Sprague-Dawley rats at
 

doses of 48 or 96 mg/kg was associated with uterine adenocarcinomas.
 

D. Statistical Analyses of Results (Rats)
 

Tables E1-E3 in Appendix E contain the statistical analyses of
 

the incidences of those primary tumors that occurred in at least
 

24
 
two animals in one group and with an incidence of at least 5% in
 

one or more than one group.
 

The untreated controls are not included in the tables and
 

analyses, because the test conditions of the vehicle controls
 

more closely resemble those of the dosed animals. Due to the
 

high mortality of the high-dose male rats, time-adjusted analyses
 

eliminating animals that died before 1 year on study are
 

performed; however, the first leukemia was observed at week 44,
 

and time-adjusted analyses on the incidence of this particular
 

tumor are based on animals that lived at least as long as week 44
 

on study. These time-adjusted analyses are shown in table E3 in
 

Appendix E, and the statistical narrative below on male rats is
 

based on the time-adjusted data only.
 

In male rats, neither the results of the Cochran-Armitage test
 

for positive dose-related trend in incidences of tumors nor the
 

results of the Fisher exact test for direct comparison of
 

incidences of tumors in the dosed groups with those in the
 

controls are significant. In females, the result of the
 

Cochran-Armitage test for dose-related trend in the incidence of
 

adenocarcinomas of the uterus is significant (P < 0.001) when the
 

pooled-control group is used, and the results of the Fisher exact
 

test show that the incidences of the tumor are significantly
 

higher (P < 0.001) in each of the dosed groups than that in the
 

25
 
pooled controls. The Fisher exact comparison of the incidences
 

of each of the dosed groups with that of the vehicle-control
 

group indicates probability levels that are above the 0.025 level
 

required by the Bonferroni inequality criterion when multiple
 

comparison is considered. The statistical conclusion suggests
 

that the incidence of adenocarcinomas of the uterus in female
 

rats is associated with administration of the test chemical. The
 

historical vehicle controls of 165 female Sprague-Dawley rats
 

compiled to date at this laboratory show no such tumor
 

occurrence.
 

A significant dose-related trend in the negative direction is
 

observed in the incidence of fibroadenomas of the mammary gland
 

in female rats when the pooled-control group is used. This
 

significant negative trend may be explained by the shortened
 

survival of the dosed animals, thus suppressing the possibility
 

of late tumor development in these groups.
 

26
 
IV. RESULTS - MICE
 

A. Body Weights and Clinical Signs (Mice)
 

Mean body weights of the high-dose mice were lower than those of
 

both the untreated and vehicle controls; body weights of the
 

low-dose mice were lower than those of the untreated controls,
 

but were comparable to those of the vehicle controls (figure 3).
 

Fluctuation in the growth curves may be due to mortality; as the
 

size of a group diminishes, the mean body weight may be subject
 

to wide variation.
 

There were no other clinical signs recorded which were indicative
 

of drug-related toxicity in the mice.
 

B. Survival (Mice)
 

The Kaplan and Meier curves estimating the probabilities of
 

survival for male and female mice administered ICRF-159 by
 

intraperitoneal injection at the doses of this bioassay, together
 

with those of the untreated and vehicle controls, are shown in
 

figure 4.
 

In male mice, the result of the Tarone test for positive dose-


related trend in mortality over the bioassay is significant (P <
 

0.001). Twenty-three out of 35 (66%) of the high-dose group,
 

26/35 (74%) of the low-dose group, 11/15 (73%) of the vehicle
 

27
 
>

0 AA 00 A A0 A 

OAOOO<> <>V « o°o«° o 

_ g n n o ogoaaaQD, ° 8 

D DOD QDA A A A D A AAA AA 

A 

3 

>­

o 

•

< 

LU 

 an 

MALE MICE 

A UNTREATED CONTROL 

D VEHICLE CONTROL 

O LOW DOSE 

A HIGH DOSE 

30 40 60 60 70

TIME ON STUDY (WEEKS) 

 80 90 100 

0 0 

o 

5 "H 

5 

( 

10­

FEMALE MICE 

0 UNTREATED CONTROL 

D VEHICLE CONTROL 

O LOW DOSE 

A HIGH DOSE 

0 20

Figure 3.

 30 40 50 60 70 80 90

TIME ON STUDY (WEEKS) 

 Growth Curves For Mice Treated With ICRF-159 

 100 110 

28 
TIME ON STUDY (WEEKS) 

Figure 4. Survival Curves For Mice Treated With ICRF-159 

29 
controls, and all (15) of the untreated controls were alive at
 

week 52. In females, the result of the Tarone test is not
 

significant. Twenty-one of 35 (60%) high-dose animals, 26/35
 

(74%) low-dose animals, and 12/15 (80%) of the vehicle or
 

untreated controls were alive at week 86. Sufficient numbers of
 

male and female mice were at risk for the development of tumors.
 

C. Pathology (Mice)
 

Histopathologic findings on neoplasms in mice are summarized in
 

Appendix B, tables Bl and B2; findings on nonneoplastic lesions
 

are summarized in Appendix D, tables Dl and D2.
 

A variety of tumors occurred in both the control and dosed
 

groups.
 

With the exception of neoplastic lesions of the lymphoreticular
 

system (tabulated below), the neoplasms listed in Appendix B
 

appeared with approximately equal frequency in dosed and control
 

mice or appeared in low numbers.
 

MICE 

MALE FEMALE 

Low High Low High 

Dose Dose Dose Dose 

Multiple Organs 

Number of Animals Necropsied (34) (30) (31) (34) 

Malignant lymphoma, histiocytic 

type 1 1 1 3 

Lymphocytic leukemia 1 1 4 3 

30
 
MICE 

MALE FEMALE 

Low High Low High 

Dose Dose Dose Dose 

Spleen
 

Number of animals with tissues
 

examined microscopically (34) (29) (31) (34)
 

Malignant lymphoma, histiocytic
 

type 0 0 0 2
 

Thymus
 

Number of animals with tissues
 

examined microscopically (34) (29) (31) (34)
 

Malignant lymphoma,
 

lymphocytic type 0 0 0 1
 

There was a slight increase in the incidence of neoplastic
 

lesions of the lymphoreticular system. These lesions were
 

confined to the dosed mice.
 

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

proliferative, and inflammatory changes were also encountered in
 

animals of the dosed and control groups (Appendix D). For the
 

most part, the nonneoplastic lesions are commonly seen in aged
 

mice and were not associated with increased mortalities or
 

decreased life spans.
 

Based on this histologic examination, the administration of
 

ICRF-159 by intraperitoneal injection to B6C3F1 mice at doses of
 

40 or 80 mg/kg was associated with an increase in lymphoreticular
 

neoplasms in the dosed females.
 

31
 
D. Statistical Analyses of Results (Mice)
 

Tables Fl and F2 in Appendix F contain the statistical analyses
 

of the incidences of those primary tumors that occurred in at
 

least two animals in one group and with an incidence of at least
 

5% in one or more than one group. The untreated controls are not
 

included in the tables and analyses, because the test conditions
 

of the vehicle controls more closely resemble those of the dosed
 

animals.
 

In male mice, neither the results of the Cochran-Armitage test
 

for positive dose-related trend in incidences of tumors nor the
 

results of the Fisher exact test for direct comparison of
 

incidences of tumors in dosed groups with those in the controls
 

are significant. However, a significant trend in the negative
 

direction is observed in the incidences of tumors of the liver,
 

where the incidences in the control groups exceed those in the
 

dosed groups. These significant negative results may be due to
 

the shortened survival of the dosed animals, which suppressed
 

late tumor development in these animals.
 

In females, the results of the Cochran-Armitage test on the
 

incidence of histiocytic lymphomas are significant when either
 

the pooled-control group (P = 0.006) or the vehicle-control group
 

(P = 0.041) is used. The results of the Fisher exact test show
 

32
 
that the incidence in the high-dose group is significantly higher
 

(P = 0.012) than that in the pooled controls. When all of the
 

tumors of the hematopoietic system are combined for analyses, the
 

statistical results show increased significance. Data for the
 

laboratory historical vehicle controls indicate an incidence of
 

1/111 (0.9%) of female mice with any type of hematopoietic tumor
 

(lymphomas or leukemias). The statistical analyses indicate the
 

possibility that hematopoietic tumors are associated with the
 

administration of ICRF-159.
 

33
 

V. DISCUSSION
 

Under the conditions of this bioassay, ICRF-159 administered by
 

intraperitoneal injection was toxic to both Sprague-Dawley rats
 

and B6C3F1 mice. Mean body weights of both species were
 

depressed by administration of the chemical, and tests for dose-


related trend in survival were positive for male and female rats
 

and for male mice. The high mortality among the high-dose male
 

rats may have been associated with inflammatory lesions in the
 

lungs, the liver, and the pleural and peritoneal cavities.
 

Sufficient numbers of female rats and of both male and female
 

mice were at risk beyond 52 weeks for development of tumors. For
 

the male rats, time-adjusted analysis of the incidence of tumors
 

was used.
 

In female rats, the incidence of uterine adenocarclnomas was
 

significantly higher in the low- and high-dose groups (P < 0.001)
 

than in the pooled controls (controls 0/38, low-dose 10/33,
 

high-dose 11/32); the incidence was also dose related (P <
 

0.001). Metastases of the adenocarcinomas were found in the
 

abdominal cavity of the females. In male rats, no tumors
 

occurred in the dosed groups in a significantly increased
 

incidence. In the high-dose males, tumors were observed in only
 

four animals. This small number may have been due to the
 

increased mortality among these animals.
 

35
 
In female mice, the incidence of all hematopoietic neoplasms
 

(histiocytic lymphomas, lymphocytic lymphomas, or lymphocytic
 

leukemias), taken together, was higher in the low-dose group (P =
 

0.038) and in the high-dose group (P = 0.002) than in the pooled
 

controls (controls 1/45, low-dose 5/31, high-dose 9/34); and the
 

incidence was significantly dose related (P = 0.002). In
 

addition, the incidence of these tumors in the high-dose group
 

was higher (P = 0.026) than that in the vehicle controls (0/15),
 

and the incidence was dose related (P = 0.021) using the vehicle
 

controls. In male mice, lymphocytic neoplasms occurred only in
 

two low-dose and two high-dose animals.
 

Chronic toxicity studies with ICRF-159 in rats (Hellman, 1972)
 

showed gross depletion of lymphoid elements in the spleen and
 

thymus but only slight depletion in the lymph nodes. Short-term
 

toxicity studies in dogs produced leucopenia and thrombocytopenia, together with cytotoxic changes in gastrointestinal, bone
 

marrow, and male reproductive tissues (Wasserman et al., 1973).
 

It is concluded that under the conditions of this bioassay,
 

ICRF-159 was carcinogenic for female Sprague-Dawley rats, producing uterine adenocarcinomas, and was also carcinogenic for female
 

B6C3F1 mice, producing lymphomas.
 

36
 
VI. BIBLIOGRAPHY

 

Armitage, P., Statistical Methods in Medical Research, John Wiley
 

& Sons, Inc., New York, 1971, pp. 362-365.
 

Berenblum, I., ed., Carcinogenicity Testing; A Report of the
 

Panel on Carcinogenicity of the Cancer Research Commission
 

of the UICC, Vol. 2. International Union Against Cancer,
 

Geneva, 1969.
 

Carter, S. K. and Slavik, M., Investigational drugs under study
 

by the United States National Cancer Institute. Cancer
 

Treatment Reviews 3;451 and 462-463, 1976.
 

Cox, D. R., Regression models and life tables. J. R. Statist.
 

Soc. B 34(2);187-220, 1972.
 

Cox, D. R., Analysis of Binary Data, Methuen & Co., Ltd., London,
 

1970, pp. 48-52.
 

Gart, J. J., The comparison of proportions: a review of
 

significance tests, confidence limits and adjustments for
 

stratification. Rev. Int. Statist. Inst. 39;148-169, 1971.
 

Hellmann, K., Effect of ICRF-159 on immune response. Proc. Roy.
 

Soc. Med. 65:264-265, 1972.
 

Hellmann, K. , Further Clinical Experiences with ICRF 159. In:
 

Advances in the Treatment of Acute (Elastic) Leukemias,
 

Mathe', G. , ed. , Springer-Verlag, Berlin, 1970.
 

Kaplan, E. L. and Meier, P., Nonparametric estimation from
 

incomplete observations. J. Am. Statist. Assoc. 53:457-481,
 

1958.
 

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

J., Carcinogenesis bioassay data system. Comp. and Biomed.
 

Res. 2:230-248, 1974.
 

Mathe', G. , Amiel, J. L. , Hayat, M., de Vassal, F.,
 

Schwarzenberg, L., Schneider, M., Jasmin, C., and Rosenfeld,
 

C., Preliminary data on acute leukemia treatment with ICRF
 

159. In: Advances in the Treatment of Acute (Elastic)
 

Leukemias. Mathe', G. , ed. , Springer-Verlag, Berlin, 1970,
 

pp. 54-55.
 

37
 
Miller, R. G., Jr., Simultaneous Statistical Inference,
 

McGraw-Hill, New York, 1966, pp. 6-10.
 

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

Kaufman, D. G., Respiratory tract carcinogenesis in hamsters
 

induced by different numbers of administrations of benzo (a)
 

pyrene and ferric oxide. Cancer Res. 32:1073-1081, 1972.
 

Salsbury, A. J., Burrage, K., Hellamann, K., Inhibition of
 

metastatic spread I.C.R.F. 159: selective deletion of a
 

malignant characteristic. Brit. Med. J. 4:344-346, 1970.
 

Sharpe, H. B. A. and Field, E. 0., Mode of action of the
 

cytostatic agent 'ICRF 159'. Nature 226:524-526, 1970.
 

Tarone, R. E., Tests for trend in life table analysis.
 

Biometrika 62(3);679-682, 1975.
 

Wasserman, T. H., Friedman, M. A., Slavik, M., and Carter, S. K. ,
 

(—) 1.2-Di(3,5-Dioxopiperazin-l-yl) Propane; Clinical
 

Brochure, Investigational Drug Branch, Cancer Therapy
 

Evaluation, Division of Cancer Treament, National Cancer
 

Institute, Bethesda, Maryland, December, 1973, pp. 1-53.
 

38
 
APPENDIX A
 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN RATS
 

GIVEN INTRAPERITONEAL INJECTIONS OF ICRF-159
 

39
 

TABLE A1.

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN MALE RATS 

GIVEN INTRAPERITONEAL INJECTIONS OF ICRF-159

 

UNTREATED 

CONTROL VEHICLE

CONTROL 

 LOW DOSE HIGH DOSE 

ANIMALS INITIAILY IN STUDY

ANIMALS NECROPSIED

ANIMALS EXAMINED HISTOPATHOLOGICALLY

 10 

 10 

 10 

10

10

10

 36

 33

 33

 35 

 30 

 30 

INTEGUMENTARY SYSTEM 

*SKIN

SiiUAMOUS CELL CARCINOMA 

 (10) (10) (33)

1 (3%) 

 (30) 

*SUBCUT TISSUE

SyUAMOUS CELL PAPILLOMA 

SARCOMA, SOS 

FIBR01A 

 (10) (10) (33)

2 (6X)

 (30) 

1 (3X) 

1 (3*) 

2 (1%) 

RESPIiiATORY SYSTEM 

NOSE 

HEMATOPOI^TIC SYSTEM 

*MULTIPLS ORGANS

GKANULOCYTHC LEUKFMIA

 (10) 

1 (10*) 

(10) (33)

2 <6X) 

 (30) 

CIRCULATORY SYSTEM 

NONE 

EIGESTIV* SYSTEM 

niViP

HJSPATOCELLOLAR CARCINOMA 

 (10) (10) (32)

1 (JX) 

 (28) 

fSMAj,L INTESIINJ

MUCINOUS ADENOCARCINOMA 

 (10) (10)

1 (10*) 

 (33) (30) 

#JEJUNUM

ALlENQCAPCI£CMAi_NOS 

 (10) (10) (33) (30) 

* NUMBER

* NUMBE R

 OF

 OF

 ANIMAL S

 ANIMAI S

 WITH TISSUE

 NECROPSIED 

 EXAMINED MICROSCOPICALLY 

41 
TABLE A1. MALE RATS: NEOPLASMS (CONTINUED) 

UNTREATED VEHICLE


CONTROL CONTROL
 

URINARY SYSTEM
 

NONi,
 

ENDOCRINE SYSTEM
 

*PITUITARY (9) (8)


CriKOMOPHOEE ADENOMA 1 (11X)


ChEOMOPHOEE CARCINOMA 1 (11%)


ITHYSOID (10) (9)


FOLLICULAS-CELL CARCINOMA
 

C-CELL ADEkCMA
 

tPANcKEATIC ISLETS (10) (10)


ISLET-CELL CARCINOMA 1 (1CX)
 

SEPHOOUCTIVE SJSTE1
 

NONh
 

MRVOUS SYST^I
 

NONJ
 

SPECIAL SENSF ORGANS
 

NONi
 

MUSCULCSKELE1AL 3YST£M
 

NONE
 

EOEY CAVITIES
 

*PEEITON?'UM (1C) (10)


FIBR01A 1 (10X)
 

*MESnNTERY (10) (10)


LlPO^il
 

#	 NUMBER OF AKIMAI S WITH TISSU" EXAMINED MICROSCOPICALLY
 

*	 NUMBER OF ANIMAL S NECROPSIED
 

42
 

 LOW DOSE HIGH DOSE
 

 (30)	 (22)
 

2 (7») 1 (5*)
 

1 (3%)
 

 (28) (25)
 

1 (1%)
 

1 (UJ6)
 

 (33) (30)
 

 (33) (30)
 

 (33)	 (30)
 

1 i3%l
 
TABLE A1. MALE RATS: NEOPLASMS (CONTINUED) 

UNTREATED VEHICLE LOW DOSE HIGH DOSE
 

CONTROL CONTROL
 

ALL OIHFR SYSTEMS
 

NONC
 

ANIMA L DISPOSITION SUMMAR Y
 

ANIMALS INITIALLY IN STUDY 10 11 36 35
 NATURAL DEATHS) 1 13 18
 MORIBUND SACRIFICE 1 9 10
 

SCHEDULED SACRIFICE
 

ACCIDENTALLY KILLED U
 TLRMINAL SACRIFICE 9 9 11 3
 

ANIMAL HISSING
 

3 INCLUDES AU1CLYZED ANIMALS
 

TUMOR SUMMAR Y
 

TOTAL ANIMAL S WITH PRIMARY TUMORS* 4 1 11 t
 

TOTAL PRIMARY TUMORS 5 1 12 6
 

TOTAL ANIMAL S WITH BENIGN TUMORS 2 € U
 

TOTAL BENIGN TUMORS 2 b t
 

TOTAL ANIMALS WITH MALIGNAN 2 T TUMORS 1 6 2
 

TOTAL MALIGNANT TUMOES 3 1 6 2
 

TOTAL ANIMALS WITH SECONDARY TUMORS*
 

TOTAL SECONDARY TUMOES
 

TOTAL ANIMAL S WITH TUMORS UNCERTAINBENIGN OR MAIIGNANT
 

TUTAL UNCERTAIN TUMOES
 

TOTAL ANIMALS WITH TUMORS UNCERTAIN­ PRIMARY OR BETASTATIC
 

TOTAL UNCERTAIN TUMOES
 

* PRIMARY TUMOES: ALL TUMORS EXCEPT SECONDARY TUMORS
 

# SECONDARY TDMQBJ3: METASTATIC TUMORS OE TUMOES INVASIVE INTO AN ADJACOT ORGAN
 

43
 
TABLE A2. 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN FEMALE RATS 

GIVEN INTRAPERITONEAL INJECTIONS OF ICRF-159 

UNTREATED

CONTROL

 VEHICLE

 CONTROL 

 LOW DOSE HIGH DOSE 

fNIHALS INITIALLY IN STUDY

ANIMALS NECBOEEIEE

ANIMALS EXAMINED HISTOPATHOLOG ICALLY

 10

 10

 10

 10

 10

 10

 31

 33

 33

 35 

 32 

 32 

INTEGUMENTARY SYSTEM 

*SKIN

SgUAMOUS CELL PAPILLOMA

sguAMOus CILL CARCINOMA

*SUBCOT TISSUE

SQUAMOUS CELL CARCINOMA

SARCOMA, NOS

FIBBOSAPCCKA

 (10)

 (10)

 (10)

 (10)

 (33)

 1 <3«) 

 (33)

 (32) 

1 (3%) 

 (32) 

1 (3%) 

1 (3«) 

1 (3X) 

FESPI8ATORY SYSTEM 

*LUNG

SiJUAMOUS CELL CARCINOMA

ADENOCARCINCMA, NOS, METASTATIC

ALVEOLAR/EFCNCHIOLAR ADENOMA

 (10) (10) (33)

 3 (9%) 

1 (3%) 

 (32) 

1 (3X) 

HEMATOPOIETIC SYSTEM 

*MULIIPLE ORGANS

LYMPHOCYTIC LEUKEMIA

 (10) (10) (33) (32) 

1 (3X) 

CIRCDLATORY SYSTEM 

NONE 

CIGESIIVE SYSTEM 

NONh 

URINA6Y SYSTEM 

NSNJ; 

# NDMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY 

* NUMBER OF ANIMALS NECSOPSIED 

44
 
TABLE A2. FEMALE RATS: NEOPLASMS (CONTINUED) 

UNTREATED VEHICLE 

CONTROL CONTROL 

ENDOCRIN E SYSTE M 

((PITUITARY (10) (9) 

CriEOMOPHOEE ADENOM A 2 (20*.) 1 (11X) 

f ADPiNA L (10) (10) 

PHEOCHROMCCYTOM A 

EEPEODUCTIV E SYSTE M 

*MAKMAB Y GLAN t 

ADENOCARCINCMA, NOS 

CYSTADENCCARCINOMA ,

FIBROSARCCK A 

FIBROADENCM A 

 NOS 

(10) 

3 (30%) 

(10) 

1 '(10X) 

2 (20*) 

IUTERU S 

ADENOCARCINCMA , NO S 

EMDOMSTBIA L STROMA L POLYP 

(10) (10) 

1 (10%) 

*CEEVI X UTE^ I 

SQUAflOO S CELL CARCINOM A 

(10) (10) 

#OVAa Y 

ADENOCARCINCMA , NO S 

(10) (10) 

NERVOU S SYSTE M 

NONii 

SPECIA L SENS E CEGAN S 

NON E 

MUSCULOSKELETA l SYSTE M 

NONi , 

EOD Y CAVITIE S 

*ABDOMINA L CAVIT Y (10) (10) 

ADENOCAECINOMft J t _NOS^_BETASTA T JC 

t NUIbER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMALS NECROPSIID
 

LOW DOSE 

(31) 

7 (23*) 

(33) 

1 (3*) 

(33) 

2 (6«) 

1 (3*) 

5 (15*) 

(33) 

1C (30*) 

2 (6S) 

(33) 

(33) 

(33) 

5_I27!|1__ 

HIGH DOSE 

(30) 

2 (7%) 

(32) 

(32) 

3 <9X) 

1 (3X) 

t (13*) 

(32) 

11 (31U) 

(32) 

1 (3X) 

(32) 

1 <3X) 

(32)

l_U2i__ 

45
 
TABLE A2. FEMALE RATS: NEOPLASMS (CONTINUED) 

UNTREATED 

CONTROL 

VEHICLE

CONTROL 

 LOW DOSE 

SARCOMA, I.CS 

MESOTHELICKA, MALIGNANT 

1

1

 (3%) 

 (3*) 

*PERITCNEUM

ADLNOCARCINCMA, NOS, METASTATIC 

 (10) (10) (33) 

1 (3X) 

*PELVIS

ADENOCARCINOMA, NOS, HETASTATIC 

 (10) (10) (33) 

*MESiNTERY

ADENOCARCINCMA, NOS, METASTATIC 

SARCOMA, NOS 

 (10) (10) (33) 

1 (33) 

ALL OIHEP SYSTIMS 

THOnAX 

SARC01A, NCS 

ANIMAL DISPOSITION SUMMARY 

UNIKALS INITIALLY IN STUDY

NATURAL DEATH3I 

aOEIBUND SACRIFICE

SCHEDULHE SACRIFICT? 

ACCIDENTALLY KILLED 

TERMINAL SACRIFICE

ANIMAL MTEEING 

 10 

1 

9 

10

2

8

 3K 

? 

it 

1 

24 

S INCLUDES AUTCLJZfD ANIMALS 

# NUMDER OF ANIMALS KITH TISSUE EXAMINED MICROSCOPICALLY 

* NUMBER OF ANIMALS NFCROPSIED 

HIGH DOSE
 

2 (6%)
 

(32)
 

(32)
 

1 <3X)
 

(32)
 

2 (6X)
 

1
 

35
 

9
 

16
 

10
 

46 
TABLE A2. FEMALE RATS: NEOPLASMS (CONTINUED) 

UNTREATED VEHICLE LOW DOSE HIGH DOSE
 CONTROL CONTROL
 

TUMOR SUMMAR Y 

TOTAL ANIMAIS WITH PRIMARY TUMOBS* TOTAL ERIHJHY TUMORS 4

 5

 4 

5 

24 

33 

22 

32 

TOTAL ANIMALS WITH BENIGN TUMORS TOTAL BHNIGN TUMORS 4

 5

 4 

1 

16 

1V 6 

6 

TOTAL ANIMAL S WITH MALIGNANT TUMORS

TOTAL MALIGNAN T TUMORS

 1 

1 

16 

16 

19 

26 

TOTAL ANIMALS WITH SECONDARY TUMORS* TOTAL SECONDARY TUMOES 

10 

13 

3 

it 

TOTAL ANIMALS WITH TUMORS UNCERTAIN­ BENIGN OR MALIGNANT TOTAL UNCEBTAIN TUMORS 

TOTAL ANIMAL S WITH TUMORS UNCERTAIN- PKIiiARY OR MITASTATIC 

TOTAL UNCEBTAIN TUMORS 

* PRIMARY TUMORS: ALL TUMORS EXCEPT SECONDARY TUMORS 

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

47
 

APPENDIX B
 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN MICE
 

GIVEN INTRAPERITONEAL INJECTIONS OF ICRF-159
 

49
 

TABLE B1.

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN MALE MICE 

GIVEN INTRAPERITONEAL INJECTIONS OF ICRF-159 

UNTREATED VEHICLE LOW DOSE HIGH DOSE
 

CONTROL CONTROL
 

ANIMALS INITIALLY IN STUEY 15 15 35 35
 ANIMALS NECROESIED 15 11 34 30
 

ANIMALS EXAMINE D HISTOP ATHOLOGIC ALLY 15 13 3U 29
 

INTEGUMENTARY SYSTEM
 

NONt
 

EESPIKATOHY SYSTEM
 

*LUNG (15) (13) (3U) (29)
 

ALVFOLAR/EFCNCHIOLAR ADENOMA 3 (20%)
 

ALVEOLAR/ESCNCHIOLAB CARCINOMA 1 (3%)
 

HEMATOPOIETIC SYSTEM
 

*MULTIPLE ORGANS (15) <1t) (31) (30)
 

MALIG.LYMPhCMA, HISTIOCYTIC TYPE 1 (3X) 1 <3X)
 

LYMPHOCYTIC LEUKEMIA 1 (3X) 1 (3X)
 

CIRCULATORY SYSTEM
 

NONE
 

DIGESTIVE SYSTEM
 

*LIVER (15) (13) (33) (29)
 

HEPATOCELLOLAH ADENOMA 1 (7X) 1 (8X)
 

HEPATOCELLDLAR CARCINOMA 1 (7%) 1 (8X)
 

URINAHY SYSTEM
 

NONE
 

INDOCBINE SYSTIM
 

NOSE.
 

* NUMBER OF ANIMALS WITH TISSDE EXAMINED MICROSCOPICALLY
 * NUMBER OF ANIMALS NECROPSIED
 

51
 
TABLE B1. MALE MICE: NEOPLASMS (CONTINUED) 

"
 

REPRODUCTIVE SYSTEM
 

NONE
 

NERVOUS SYSTEM
 

NONE
 

SPECIAL SENSE ORGANS
 

NONE
 

PUSCULOSKE1ETAI SYSTEM
 

NONi,
 

ECDY CAVITIES
 

NONE
 

AIL OTHER SYSTEMS
 

NONE
 

JNIHAi, DISPOSITION SUMMARY
 

ANIMALS INITIALLY IN STUDY
 

NATURAL DEATHS
 

MORIBUND SACRIFICE
 

SCHEDULED SACRIFICE
 

ACCIDENTAIIY KILLED
 

TERMINAL SACRIFICE
 

ANIMAL MISSING
 

a INCLUDES AUTOLYZED ANIMALS
 

UNTREATED VEHICLE LOW DOSE HIGH DOSE
 

CONTROL CONTROL
 

15 15 35 35 

5 12 13 

1 3 9 

1 

14 10 20 12 

# NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBEE OF ANIMAIS NECEOPSIED
 

52
 
TABLE B1. MALE MICE: NEOPLASMS (CONTINUED) 

UNTREATED 

CONTROL 

VEHICLE CONTROL  LOW DOSE HIGH DOSE 

TUMOR SUMMARY 

TOTAL ANIMALS WITH PRIMARY TUMORS* 

TOTAL PRIMARY TUMORS 

3 

5 

2

2

 2 

2 

3 

3 

TOTAL ANIMALS WITH BENIGN TUMOES 

TOTAL BENIGN TUMORS 

3 

4 

1 

1 

TOTAL ANIMALS WITH MALIGNANT TUMORS 

TOTAL MALIGNANT TUMORS 

1 

1 

1

1

 2 

2 

3 

3 

TOTAL ANIMALS WITH SECONDARY TUMORS 

TOTAL SECONDARY TUMOKS 

* 

TOTAL ANIBA1S WITH TUMORS UNCERTAIN BENIGN OR MAIIGNANT 

TOTAL UNCERTAIN TUMORS 

TOTAL ANIMALS WITH TUMORS UNCERTAIN 

PRIMARY OS BETASTATIC 

TOTAL UNCERTAIN TUMORS 

* PRIMARY TUMORS: ALL TUMORS EXCEPT SECONDARY TUMORS t SECONDARY TUMORS: METASTATIC TUMORS OR TUMORS INVASIVE INTO AN ADJACENT ORGAN 

53
 
TABLE B2.

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN FEMALE MICE 

GIVEN INTRAPERITONEAL INJECTIONS OF ICRF-159

 

UNTREATED CONTROL VEHICLE CONTROL 

 LOW DOSE HIGH DOSE 

ANIMALS INITIALLY IN STUDY ANIMALS MISSING ANIMALS NECRCESIED ANIMALS EXAMINID HISTOPATHOLOGICALLY 15 

 1U 

1 U 

15

15

15

 35

 31

 31

 35 

1 

 31 

 3U 

INTEGUMENTARY SYSTEM 

NONE 

RESPIRATORY SYSTEM 

NONE 

HEMATOPOIETIC SYSTEM 

*MDIIIPLE ORGANS

MALIG. LYBFHOMA, HISTIOCYTIC TYPE LYMPHOCYTIC LEUKEMIA 

#SPLEEN

MALIG. LYMPHOMA, HISTIOCYTIC TYPE 

 (1U) 

 (13) 

(15)

(15)

 (31)

1 (3X)

14 (13X)

 (31)

 (3U) 

3 (9*) 

3 (9X) 

 (3«) 

2 (6») 

*THYHOS

MALIG. LYHPHOMA, LYMPHOCYTIC TYPE (12) (15) (31) (34) 

1 (3X) 

CIRCULATORY SYSTEM 

NONE 

DIGESTIVE SYSTIM 

NONE 

URINAfiY SYSTEM 

NONE 

t NUMBER OF ANIMALS WITH TISSUE EXAMINED MICBOSCOPICALLY 

* NUMBER OF ANIMALS NECBOPSIED 

54
 
TABLE B2. FEMALE MICE: NEOPLASMS (CONTINUED) 

UNTREATED

CONTROL

 VEHICLE

 CONTROL 

 LOW DOSE HIGH DOSE 

ENDOCBINE SYS1IB 

NONt 

REPRODUCTIVE SYSTEM 

*HAMMARY GLANE ADENOMA, DCS

ADENOCA8CIKCMA, NOS

tUTEBUS

ADENOCAECINCBA, NOS

 (11)

 (13)

 (15)

 (15)

 (31)

 1 (3%) 

1 (3X) 

 (31)

 1 (3%) 

 (34) 

 (34) 

NERVOUS SYSTEM 

NONE 

SPECIAL SENSE ORGANS 

NONE 

MDSCULOSKELE1SI SYSTEM 

NO (id 

BODY CAVITIES 

*PERITONEnH

CARCINOMA, NOS

*MES£NTERY LIPOMA

 (1U)

 (14)

 (15)

 (15)

 (31)

 1 (3X) 

 (31)

 (34) 

 (34) 

2 (6X) 

AIL OTHER SYSTEMS 

NONE 

* NUMBER OF AMIMALS KITH TISSUE EXAMINED MICROSCOPICALLY 

* NUMBER OF ANIMALS NECROPSIED 

55 
TABLE B2. FEMALE MICE: NEOPLASMS (CONTINUED) 

UNTREATED

CONTROL

 VEHICLE 

 CONTROL 

LOW DOSE HIGH DOSE 

ANIMAL DISPOSITION SUMMARY 

ANIMALS INITIALLY IN STUDY NATURAL DEATHS

MORIBUND SACRIFICE SCHEDULED SACRIFICE ACCIDENTAL!! KILLED 

TERMINAL SACRIFICE ANIMAL MISSING 

 15

 3

 12

 15 

2 

1 

 12 

35

7

1

1 

26

 35 

 <• 

8 

 22 

1 

3 INCLUDES AUTCLYZED ANIMALS 

TUMOR SUMMARY 

TOTAL ANIMALS HITH PRIMARY TUMORS* 

TOTAL PRIMARY TUMORS 

8

9

 11 

 11 

TOTAL ANIMALS SITH BENIGN TUMORS 

TOTAL BENIGN TUMORS 

1

1

 2 

2 

TOTAL ANIMALS WITH MALIGNANT TUMORS 

TOTAL MALIGNANT TUMORS 

7

8

 9 

9 

TOTAL ANIMALS HITH SECONDARY TOTAL SECONDARY TUMORS 

 TUMORS* 

TOTAL ANIMALS HITH TUMORS UNCERTAIN­ BENIGN OR MALIGNANT 

TOTAL UNCERTAIN TUMORS 

TOTAL ANIMALS HITH TOMORS UNCERTAINPRIMABY OH BITASTATIC 

TOTAL UNCERTAIN TUMORS 

* PRIMARY TUMORS: ALL TUMORS EXCEPT SECONDARY TUMORS * SECONDARY TUMORS: METASTATIC TUMORS OR TUMORS INVASIVE INTO AN ADJACBKT ORGAN 

56 
APPENDIX C
 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN RATS
 

GIVEN INTRAPERITONEAL INJECTIONS OF ICRF-159
 

57
 

TABLE C1.

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN MALE RATS 

GIVEN INTRAPERITONEAL INJECTIONS OF ICRF 159

 

UNTREATED 

CONTROL 

VEHICLE 

CONTROL 

ANIMALS INITIALLY IN STUDY 

ANIMALS NECRCPSIED 

ANIMALS EXAMINED HISTOPATHOLOGICALLY 

10 

10 

10 

10 

10 

10 

INTEGUMENTARY SYSTEM
 

*SKIN (10) (10)
 EPIDERMAL INCLUSION CYST 1 (10X)
 

*SUBCUT TISSUE (10) (10)
 

ABSCESS, NCS 1 (10X)
 

INFLAMMATION, ACUTE/CHRONIC
 

INFLAMMATION, CHRONIC 1 (10X)
 

INFLAMMATION, CHRONIC FOCAL
 

INFLAMMATION HITH FIBFOSIS
 

EESPIhATOSY SYSTEM
 

•TRACHEA	 (10) (10)
 

INFLAMMATION, CHRONIC
 

INFLAMMATION, CHRONIC SDPPURATIV
 

iLUNG/BRONCHICLE (10) (10)
 

HYPERPLASIA, LYMPHOID
 

*LONt, (10) (10)
 

EDEMA, NOS
 

INFLAMMATION, INTERSTITIAL
 

BRONCHOEfcEUHONIA SUPPURATIVE
 

BRONCHOPNEUMONIA CHRONIC SDPPORA 1 OCX)
 

HEMATOPOIETIC SYSTEM
 

*BONt MARBOH (10) (9)
 ATPOPHY, NOS f (60S) 4 (44X)
 

*SPLtEN (10) (10)
 

INFLAMMAIICJLt_NQS


# NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMALS NECROPSIED
 

59
 

LOW DOSE


36
 

33
 

33
 

(33)
 

(33)
 

1 (3X)
 

1 (3X)
 

(32)
 

1 <3X)
 

1 (3X)
 

(33)
 

3 (9X)
 

(33)
 

1 (3%)
 

1 (3X)
 

11 (33X)
 

(29)
 

10 (345!)
 

(33)
 

 J_J3*1
 

 HIGH DOSE
 

35
 

30
 

30
 

(30)
 

(30)
 

1 (3«)
 

1 (3X)
 

(30)
 

(30)
 

2 (7%)
 

(30)
 

1 (3%)
 

3 (10X)
 

3 (10X)
 

4 (13%)
 

(28)
 

11 (39S)
 

(30)
 
TABLE C1. MALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

UNTREATED VEHICLE
 

CONTROL CONTROL
 

ATROPHY, NOS
 

HEMATOPOIIEIS
 

tCERVICAL LYMPH NODE (2)
 HYPERPLASIA, LYMPHOID 1 (SOX)
 

IPANCREATIC L.NOEE (2)
 INFLAMMATION, SOPPURATIVE 1 (SOX)
 

IMESENTERIC I. NODE (2)
 HYPERPLASIA, LYMPHOID
 

CIRCULATORY SYSTEH
 

IMYOCARDIUM (10) (10)
 

INFLAMMATION, INTERSTITIAL
 

*ENDOCARDIUM (10) (10)
 

INFLAMMATION, CHRONIC SDPPURATIT
 

*CORONARY ARTERY (10) (10)
 

MINERALIZATION
 

INFLAMMATION, CHRONIC
 

*CELIAC ARTEFY (10) (10)
 

NECROSIS, FIBHINOID
 

DIGESTIVE SYSTEM
 

tLIVER (10) (10)
 

INFLAMMATION, SUPPURATIVE
 INFLAMMATION, CHRONIC SUPPURATIV
 

INFLAMMATION, CHRONIC NECHOTIZIN
 

PfiRIARTERITIS 

DEGENERATION, NOS 

NECROSIS, FOCAL 

NECROSIS, COAGULATIVE 

tHEPATIC CAPSULE 

HEMORRHAGE 

(10) (10) 

1 (10X) 

tLIVER/CENTHILOBULAR 

NECROSIS^ SOAGULAJIVE __ . 

(10) (10) 

, 1 (10X) 

* NUMBER OF AHIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMALS NECKOPSIED
 

LOW DOSE
 

(3)
 

(3)
 

(3)
 

(33)
 

(33)
 

(33)
 

1 (3X)
 

(33)
 

1 <3X)
 

(32)
 

1 (3X)
 

1 <3X)
 

(32)
 

1 (3X)
 

(32)
 

HIGH DOSE
 

1 (3X)
 

7 (23X)
 

(3)
 

(3)
 

(3)
 

1 (33X)
 

(30)
 

1 (3X)
 

(30)
 

1 (3X)
 

(30)
 

1 (3X)
 

(30)
 

(28)
 

1 (<**)
 1 (4X)
 

1 (tX)
 

1 (IX)
 

1 C*X)
 

1 C*X)
 

(28)
 

1 («X)
 

(28)
 

1 (4X)
 

60
 
TABLE C1. MALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

LOW DOSE HIGH DOSE
 

(33)	 (30)
 

1 (3X)
 

2 (7X)
 

1 (3X)
 

(33)	 (30)
 

1 <3X)
 

(33)	 (30)
 

1 (3X)
 

1 (3X)
 

(33)	 (30)
 

1 (3%)
 

(33)	 (30)
 

2M (73X) 17 (57X)
 

(33)	 (30)
 

1 (3X)
 

(9)	 (17)
 

2(22%) 1 (6X)
 

(33)	 (30)
 

1 <3X)
 

(32)	 (28)
 

1 C»X)
 

1 (UX)
 

1 («X)
 

(33)	 (30)
 

1 (3X)
 

•PANCREAS
 

INFLAMMATION, SOPPURATIVE
 PERIARTERITIS
 

NECROSIS, FIERINOID
 

tSTOHACH
 

CALCIFICATION, METASTATIC
 

tSMALL INTESTINE
 

PERIARTERITIS
 

NECROSIS, FIBRINOID
 

*JEJUNUM
 

HEBORHHAGI
 

URINARY SYSTEM
 

f KIDNEY
 INFLAMMATION, CHRONIC
 

tORINARY BLAEDER
 

CALCOLaS, NOS
 

ENDOCRINE SYSTEM
 

IPARATHYROID
 

HYPERPLASIA, NOS
 

REPRODDCTIVE SYSTEM
 

tPROSTATE
 INFLAMMATION, SOPPURATIVE
 

ITESIIS
 INFLAMMATION, NECROTIZING
 

INFLAMMATION, CHRONIC SOPPORA .TIV
 

PjSRIARTERIlIS
 

*EPIDIDYMIS
 

INFLAMBATICN, SOPPURATIVE
 

NERVOUS SYSTEM
 

N0r)£
 

UNTREATED
 

CONTROL
 

(10)
 

(10)
 

(10)
 

(10)
 

(10)
 

7 (70%)
 

(10)
 

(7)
 

(10)
 

1 (10X)
 

(10)
 

(10)
 

VEHICLE
 

CONTROL
 

(10)
 

(10)
 

(10)
 

(10)
 

(10)
 

6 (60%)
 

(10)
 

(7)
 

1 <1«X)
 

(10)
 

(10)
 

(10)
 

t	 NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

*	 NOHBER OF ANIMALS NECROPSIED
 

61
 
TABLE C1. MALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

LOW DOSE HIGH DOSE
 

(33)	 (30)
 

1 (3X)
 

1 (3X)
 

(33)	 (30)
 

1 (3X)
 

(33)	 (30)
 

1 (3X>
 

(33) (30)
 

1 (3*)
 

4 (13*)
 

(33)	 (30)
 

2 (7*)
 

(33)	 (30)
 

1 (3*)
 

(33)	 (30)
 

2 (7*)
 

(33) (30)
 

(33)	 (30)
 

1 (3*)
 

3
 

SPECIAL SENSE ORGANS
 

*EYE/COENEA
 

MINERALIZATION
 

INFLAMMATION, FIBRINOUS
 

*EYELID
 HYPERKERA1CSIS
 

MUSCULOSKELETAL SYSTEM
 

*KNEE JOINT
 INFLAMMATION, CHRONIC SUPPDRATIV
 

EODY CAVITIES
 

*PERITONBUM
 

INFLAMBATICN, SOPPORATIVE
 

INFLAMMATION, NECROTIZING
 INFLAMMATION, CHRONIC
 

*PLEURA
 

INFLAMMATION, CHRONIC
 

*PERICARDIUM
 

INFLAMMATION, CHRONIC
 

*EPICARDIUB
 

INFLAMMATION, CHRONIC
 

*BESENTERY
 

MINERALIZATION
 

NJSCROSIS, FAT
 

ALL OTHER SYSTEMS
 

*MULTIPLE ORGANS
 CALCIFICATION, METASTATIC
 

SPECIAL MORPHOLOGY SUMMARY
 

NO_LESION_EEPCRTJD
 

UNTREATED
 

CONTROL
 

(10)
 

(10)
 

(10)
 

(10)
 

(10)
 

(10)
 

(10)
 

(10)
 

1 (10*)
 

1 (10*)
 

(10)
 

VEHICLE
 

CONTROL
 

(10)
 

(10)
 

(10)
 

(10)
 

1 (10X)
 

(10)
 

(10)
 

(10)
 

(10)
 

(10)
 

*	 NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

*	 NUMBER OF ANIMALS NECROPSIED
 

62 
TABLE C1. MALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

UNTREATED

CONTROL

 VEHICLE

 CONTROL 

 LOW DOSE HIGH DOSE 

AUTOLYSIS/NO NECROPSY 3 5 

* NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY 

* NUMBER OF ANIMALS NECROPSIED 

63
 
TABLE C2.

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN FEMALE RATS 

GIVEN INTRAPERITONEAL INJECTIONS OF ICRF-159 

UNTREATED CONTROL VEHICLE CONTROL LOW DOSE HIGH DOSE 

8NIHALS INITIALLY IN STUEY 

ANIMALS NECROESIED ANIMALS EXAMINED HISTOPATHO LOGICALLY 

10 

10 

10 

10 

10 

10 

3U 

33 

33 

35 

32 

32 

INTEGUMENTARY SYSTEM 

*SKIN 

ULCER, NOS 

(10) (10) (33) 

, 

(32) 

1 (3X) 

+SUBCUT TISSUE 

INFLAMMATION, CHRONIC 

(10) (10) (33) (32) 

1 (3X) 

RESPIRATORY SYSTEM 

*TRACHEA 

INFLAMMATION, SUPPURATIVE INFLAMMATION, CHRONIC 

(10) (10) (33) 

2 <6X) 2 (6X) 

(32) 

tLUNG/BRONCHDS 

INFLAMMATION, SUPPURATIVE INFLAMMATION, CHRONIC SDPPORATIV 

(10) (10) (33) 

1 (3X) 

1 (3X) 

(32) 

((LUNG/BRONCHIOLE 

HYPERPLASIA, LYMPHOID 

(10) (10) (33) 

2 (6X) 

(32) 

1 (3*) 

ILUNG 

INFLAMMATION, INTERSTITIAL BHONCHOENEUMONIA SUPPURATIVE PNEUMONIA INTERSTITIAL CHRONIC 

BRONCHOPNEDMONIA CHRONIC SUPPURA 

(10) (10) 

1 (10*) 

1 dOX) 

1 (10X) 

(33) 

1 <3X) 

6 (18X) 

(32) 

1 (3*) 

1 (3*) 

3 (9*) 

HEMATOPOIETIC SYSTEM 

tBONE MARROW ATROPHY, NCS 

(10) 

5 (50X) 

(10) 

3 (30X) 

(30) 

9 (30)!) 

(31) 

» (13*) 

#SPLEEN 

HE.MAT.OE2iE.SIS 

(10) (10) (33) 

Y (15*) 

(32) 

1t_JlSii 

* NUMBER OF ANIMALS WITH TISSUE EXAMINED * NUMBER OF ANIMALS NECROPSIED MICROSCOPICALLY 

64
 
TABLE C2. FEMALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

UNTREATED VEHICLE


CONTROL CONTROL
 

IMESENTERIC I. NODE
 

INFLAMMATION, GRANOLOMATOUS
 

*THYMUS (10) (10)


HYPERPLASIA, IYMPHOID
 

-IBCULATORY SYSTEM
 

•MYOCARDIUM	 (10) (10)


ISFLAMMATION, FIBRINOOS
 

CALCIFICATION, BETASTATIC
 

DIGESTIVE SYSTEM 

ILIVER 

CONGESTION, PASSIVE 

NECROSIS, COAGULATIVE 

LIPOIDOSIS 

(10) (10)

tLIVfiR/CENTRHOBOLAR 

NECROSIS, COAGOLATIVE 

CYTOLOGIC IEGENERATION 

(10) (10)

tPANCREAS 

INFLAMMATION, INTERSTITIAL 

INFLAMBATICN, CHRONIC 

(10) (10)

#GASTRIC MOSCULARIS 

CALCIFICATION, BETASTATIC 

(10) (10)

ORINAEY SYSTIB
 

*KID»EY (10) (10)


HYDRONEPHROSIS
 

INFLAMMATION, CHRONIC 2 (20X)


ENDOCRINE SYSTEM
 

IPITUITAHY (10) (9)


AJSCESS^ CHRONIC
 1 - ­

* NOMBEB OF AHIMALS WITH TISSOE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMALS NECROPSIED
 

65
 

 LOW DOSE
 

 (33)
 

1 (3X)
 

 (32)
 

1 (3X)
 

 (33)
 

1 (3X)
 

 (33)
 

1 (3X)
 

 (33)
 

1 (3%)
 

 (33)
 

1 <3X)
 

 (33)
 

1 (3*)
 

 12 (36X)
 

 (31)
 

HIGH DOSE
 

(3)
 

1 (33X)
 

(32)
 

(32)
 

1 (3X)
 

(32)
 

1 (3X)
 

1 (3X)
 

1 (3*)
 

(32)
 

1 (3%)
 

(31)
 

1 (3X)
 

(32)
 

(32)
 

17	 (53X)
 

(30)
 

1 J3SL
 
TABLE C2. FEMALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

UNTREATED VEHICLE
 

CONTROL CONTROL
 

1 ADRENAL (10) (10)
 

ANGIECTASIS 5 (50*) 1 (10X)
 

tPARATHYROID (6) (6)
 HYPERPLASIA, NOS
 

BEPBODUCTIVE SYSTEM
 

*MAHMARY GLARE (10) (10)
 

CYST, NOS 5 (SOS) 3 (30X)
 

IUTEHDS (10) (10)
 

INFLAHMATICN, CHRONIC SUPPURATIV
 

#DTEBUS/ENDOMETBIUI1 (10) (10)
 

INFLAMMATION. SUPPHEATIVE 3 (30X) 1 (10X)
 

INFLAMMATION, CHRONIC SUPPUEATIV 2 (20%)
 

INFLAMMATION, CHRONIC NECEOTIZIN
 

HYPEBPLASIA, SOS
 

HYPEEPLASIA. CYSTIC
 

*OVARY/OVIDOC1 (10) (10)
 

INFLAMMATION, CHRONIC SUPPURATIV
 

#OVAEY (10) (10)
 

CYST, NOS
 

INFLAMMATION, SUPPUHATIVE
 

INFLAMMATION, CHRONIC SOPPORATIV
 

METAPLASIA, SQOAMOUS
 

NERVOUS SYSTEM
 

NONE
 

SPECIAL SENSE CR6ANS
 

*EYE (10) (10)
 

HEMOEBHAGI
 

MUSCULOSKELETA1 SYSTEM
 

*JOINT (10) (10)
 

INFLAMMATION^ CHEONIC
 

# NUMBEE OF ANIMALS WITH TISSUE EXAMINED MICEOSCOPICALLY
 

* NUMBER OF ANIMALS NECROPSIED
 

LOW DOSE
 

(33)
 

5 (15*)
 

<2C)
 

(33)
 

<: (27*)
 

(33)
 

(33)
 

8 (21)5)
 

12 (36X)
 

1 (3«)
 

(33)
 

1 (3*)
 

(33)
 

1 0*)
 

(33)
 

(33)
 

HIGH DOSE
 

(32)
 

8 (25S)
 

(11)
 

1 (9X)
 

(32)
 

H (13*)
 

(32)
 

1 (3*)
 

(32)
 

1 (3%)
 10 (31*)
 

1 (3*)
 1 (3X)
 

(32)
 

(32)
 

2 (6*)
 

1 (3*)
 

8 (25*)
 

1 (3X)
 

(32)
 

1 (3*)
 

(32)
 1 _im_
 

66 
TABLE C2. FEMALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

*ABDOMINAL MUSCLE
 

ISFLAMMATICN, CHRONIC NECHOTIZIN
 

EOEY CAVITIES
 

*ABDOMINAL CAVITY
 

NLCROSIS, FAT
 

METAPLASIA, OSSEOCS
 

*PERITCNEUM
 

INFLAMMATION, SUPPURATIVE
 

INFLAMMATICN, CHRONIC
 

INFLAMMATICN, CHRONIC NECROTIZIN
 

*PLEURA
 

INFLAMMATICN, CHRONIC
 

*PERICARDIUM
 

INFLAMMATICN, FIBRINOUS
 

*EPICARDIOM
 

INFLAMMATICN, FIBHINOUS
 

*MEStNTERY
 

PiiRIARTERITIS
 

ALL OTHER SYSTEMS
 

ADIPOSE TISSUE
 

INFLAMMATICN, CHRONIC
 

SPECIAL MORPHC10GY SUMMARY
 

NO LESION FEPCBTED
 

AUTOLYSIS/NO NECROPSY
 

UNTREATED VEHICLE 

CONTROL CONTROL 

(10) (10) 

(10) (10) 

(10) (10) 

(10) (10) 

(10) (10) 

(10) (10) 

(10) (10) 

2 

LOW DOSE


(33)


(33)


(33)


2 (6%)
 

(33)


(33)


(33)


(33)


1 <3«)
 

1
 

1


 HIGH DOSE
 

 (32)
 

1 13X)
 

 (32)
 

1 (3«)
 

1 (3X)
 

 (32)
 

1 (3*)
 

1 (3X)
 

 (32)
 

1 (3*)
 

 (32)
 

1 (3%)
 

 (32)
 

1 (3%)
 

 (32)
 

1
 

3
 

* NUMBER OF ANIMAL S WITH TISSUE EXAMINED MICRCSCOPICALLY
 

* NUMBER OF ANIMAL S NECROPSIED
 

67
 

APPENDIX D
 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN MICE
 

GIVEN INTRAPERITONEAL INJECTIONS OF ICRF-159
 

69
 

TABLE 01.

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN MALE MICE 

GIVEN INTRAPERITONEAL INJECTIONS OF ICRF-159

 

UNTREATED 

CONTROL 

VEHICLE

CONTROL 

JNIHALS INITIALLY IN STODY 

ANIMALS NECBOPSIED 

ANIMALS EXAMINED HISTOPATHOLOGICALLY 

15 

15 

15 

15

14

13

INTEGUMENTARY SYSTEM
 

*SKIN (15) (T»)
 INFLAMMATICN, GEANDLOMATOUS 1 (7X)
 

*SOBCOT TISSUE (15) (10)
 EPIDERMAL INCLUSION CYST 1 (7X)
 INFLAMMATICN, CHRONIC FOCAL 1 (7*)
 

RESPIEATORY SYSTEM
 

ILUNG (15) (13)
 BRONCHOPNEUMONIA SOPPUEATIVE
 

HYPEEPLASIA, LYSPHOID
 

HEMATOPOIETIC SYSTEM
 

tSPLEEN (15) (13)
 INFLAMMATION, HEMOEEHAGIC
 

HYPEEPLASIA, HEMATOPOIETIC
 

HYPERPLASIA, IYMPHOID
 

HEMATOPOIESIS 3 (20X)
 

tMESENTEEIC L. NODE (1) (2)


CONGESTION, NOS 1 (100*)
 INFLAMMATICN, SOPPDRATIVE 1 (100*) 2 (100X)
 

HYPEEPLASIA, LYMPHOID
 

fAXILLABY LYMPH NODE (1) (2)
 HYPEEPLASIA, LYMPHOID
 

CISCULATOBY SYSTEM
 

_M&£
 

t	 NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

*	 NUMBER OF ANIMALS NECROPSIED
 

71 

 LOW DOSE HIGH DOSE
 

 35 35
 

 31 30
 

 3<4 29
 

 <3«) (30)
 

 (3<1) (30)
 

 <3U) (29)
 

1 <3X)
 

2 (6X) 1 (3X)
 

 (3<4) (29)
 

1 (3X)
 

1 (3X)
 

1 (3X) 1 <3X)
 

2 (6X)
 

 (3)	 (3)
 

2 (67X)
 

1 (33*)
 

 (3)	 (3)
 

1 (33*)
 
TABLE 01. MALE MICE: NONNEOPLASTIC LESIONS (CONTINUED) 

DIGESTIVE SYSTEM
 

fLIVEE
 CYST, NOS
 

NECBOSIS, COAGULATIVE
 

HYPEEPLASIA, NODULAR
 

ICOLON
 INFLAMMATION, NECROTIZING
 

DBINABY SYSTEM
 

IKIDNEY
 HYDRONEPHEOSIS
 

PYELONEPHEITIS SOPPUEATIVE
 

INFLAMMATION, CHRONIC
 

HYPOPLASIA, NCS
 

HYPEEPLASIA, LYMPHOID
 

#UEINAEY BLfCtER
 CALCULUS, NOS
 

INFLAMMATION, CHRONIC
 

ENDOCRINE SYSTEM
 

fTHYEOID
 

HYPERPLASIA, FOLLICULAB-CELL
 

REPRODUCTIVE SYSTEM
 

tPROSTATE
 INFLAMMATION, CHRONIC SUPPUEATIV
 

ITESIIS
 

INFLAMMATION, SUPPDRATIVE
 

tTUNICA ALBOGINEA
 MINERALIZATION
 

NERVOUS SYSTEM
 

ICEBEBRUM
 INFLAMMATION, CHRONIC SUPPURATIV
 

UNTREATED VEHICLE LOW DOSE HIGH DOSE 

CONTROL CONTROL 

(15) (13) (33) (29) 

1 <3X) 

1 (8X) 1 (3X) 

7 (21X) 1 (3X) 

(15) (13) (33) (29) 

1 <3X) 

(15) (13) (34) (29) 

2 (6X) 

1 (3X) 

1 (8X) 1 (3X) 

1 <3X) 

1 (3X) 1 (3X) 

(15) (9) (3K) (29) 

1 <3X) 

1 (3X) 

(11) (11) (32) (22) 

2 <6X) 

(15) (13) (33) (29) 

1 (3X) 

(15) (13) (32) (29) 

1 <8X) 

(15) (13) (32) (29) 

3 (9X) 

(11) (13) (33) (28) 

1 (3X) 

t NUMBER OF ANIMALS KITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMAIS NECROPSIED
 

72
 
TABLE 01. MALE MICE: NONNEOPLASTIC LESIONS (CONTINUED) 

UNTREATED

CONTROL

 VEHICLE

 CONTROL 

 LOW DOSE HIGH DOSE 

SPECIAL SENSE ORGANS 

NONE 

MDSCUiOSKELETAL SYSTEM 

NONE 

ECDY CAVITIES 

*PERITONEBB

HEMORRHAGE INFLAMMATION, SUPPUHATIVE

 (15) (1U)

 1 (7X) 

 (3«) (30) 

1 (3X) 

*PELVIS

CYST, NOS

METAPLASIA, OSSEOUS

 (15) (14) (34)

 1 (3%) 

1 (3X) 

 (30) 

ALL OTHER SYSTEMS 

NONE 

SPECIAL MORPHCIOGY SUMMARY 

NO LESION REPORTED

ACCIDENTAL DEATH

NECROPSY FERF/NO HISTO PERFORMED

AUTO/NECROESY/HISTO PERF

AUTO/NECRCFSY/NO HISTO AUTOLYSIS/NO NECROPSY

 8 7

 1 

1 

1

 8

 1

 18 

1 

1 

4 

# NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICAL!Y 

* NUMBER OF ANIMALS NECROPSIED 

73
 
TABLE D2.

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN FEMALE MICE 

GIVEN INTRAPERITONEAL INJECTIONS OF ICRF-159

 

UNTREATED VEHICLE

CONTROL CONTROL 

ANIMALS INITIALLY IS STUDY 15 15

ANIMALS MISSING 

ANIMALS NECEOPSIED 14 15

JNINALS EXAMIMHD HISTOPATH0106ICALLY 14 15

INTEGUMENTARY SYSTEM
 

*SKIN (14) (15)


ULCEH, CHECNIC 1 (7*)
 

RESPIRATORY SYSTEM
 

*LUNG/BBONCHIOLE (13) (15)


PLASMA-CELL INFILTRATE
 

*LUNG (13) (15)


INFLAMMATION, INTERSTITIAL 1 <8X) 1 (7X)


PNEUMONIA INTEBSTITIAL CHRONIC
 

BhONCHOENIOMONIA CHRONIC SDPPHRA 1 (7X)


HYPE8PLASIB, PLASMA CELL 1 (7%)
 HYPERPLASIA, LYMPHOID
 

HEMATOPOIETIC SYSTEM
 

*BONE HARROW (13) (15)


ATROPHY, NOS
 

HYPERPLASIA, HEMATOPOIETIC
 

»SPL£EN (13) (15)


INFLAMMATION, SUPPURATIVE 1 (7X)
 

ATROPHY, NOS
 

HYPERPLASIA, HEMATOPOIETIC
 

HYPERPLASIA, LYMPHOID
 

Hi.MATOPOIESIS 3 (23X)
 

»MESiiNTERIC L. NODE (1)
 

LYMPHANGIECTASIS 1 (100X)
 

HIMORRHiGE 1 1100S1
 

* NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* DUMBER OF ANIMALS NECROPSIED
 

74 

 LOW DOSE HIGH DOSE
 

 35	 35
 

1
 

 31 3<t
 

 31 3D
 

 (31)	 (34)
 

 (31)	 (34)
 

1 (3*)
 

 (31)	 (34)
 

1 (3%)
 

1	 (3X)
 

1 (3X)
 

1 (3%)
 

 (29)	 (33)
 

1 (3%)
 

1 (3X)
 

 (31) (34)
 

1 (3%)
 

1 (3X)
 

2 (6%)
 

3 (10X) 3 (9X)
 

(4) (9)
 
TABLE D2. FEMALE MICE: NONNEOPLASTIC LESIONS (CONTINUED) 

UNTREATED 

CONTROL 

VEHICLE

CONTROL 

INFLAMMATION, SUPPUBATIVE 

INFLAMMATION, ACUTE SUPPDRATIVE 

HYPERPLASIA, LYMPHOID 

tTHYMUS 

HYPERPLASIA, HEMATOPOIETIC 

(12) (15)

CIRCULATORY SYSTEM
 

NONE
 

EIGESTIVE SYSTEM
 

tLIVER (13) (15)


MINERALIZATION
 

THROMBOSIS, NOS 1 (7%)
 

CONGESTION, NOS 1 (8%)
 

NECROSIS, COAGULATIVE
 

HYPERPLASIA, NODULAR 1 (7%)
 

ANGIECTASIS 1 {7%)
 

tHEPATIC CAPSULE (13) (15)


FIBROSIS, FOCAL
 

tPANCREAS (13) (15)


INFLAMMATION, ACUTE SUPPURATIVE
 

INFLAMMATION, PYOGRANULOMATOUS 1 (7X)
 

tCOLON (13) (15)


NEHATODIASIS 1 (7%)
 

URINARY SYSTEM
 

tKIDNEY (13) (15)


HYPERPLASIA, LYMPHOID
 

tUBIHARY BLADDER (13) (15)


MINERALIZATION 1 (7%)
 

ENDOCRINE SYSTEM
 

HO£E_
 

« NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMALS NECROPSIED
 

 LOW DOSE HIGH DOSE 

1 (25X) 

1 (25%)

 (31)

1 (11X) 

3 (33%) 

 (3i») 

1 (3X) 

 (31) (34) 

1 (3%) 

1 <3X) 

 (31)

 (31)

1 (3X) 

 (34) 

1 (3%) 

 (34) 

 (31) (33) 

 (31)

 (31)

 (34) 

1 (3%) 

 (3U) 
TABLE D2. FEMALE MICE: NONNEOPLASTIC LESIONS (CONTINUED) 

UNTREATED CONTROL VEHICLE CONTROL 

LOW DOSE HIGH DOSE 

REPRODUCTIVE SYSTEM 

#UTEHUS/ENDCHETRIOM

INFLAMMATICN, SUPPURATI7E 

INFLAMMATICN, NECROTIZING 

INFLAMMATION, CHBONIC SDPPURATIV

HYPERPLASIA, CYSTIC

 (13)

 7 (SIX)

 (15) 

3 (20*) 

 11 (73X) 

(31)

5 (16X)

1 (3«) 

19 (61*)

 (3U) 

3 (9«) 

 18 (53X) 

NERVOUS SYSTEM 

NOME 

SPECIAL SENSE ORGANS 

NONE 

MOSCULOSKELETAL SYSTEM 

NONE 

BODY CAVITIES 

*PERITONEOM INFLAMMATICN, CHRONIC

IHFLABHATICN, CHRONIC FOCAL 

INFLAMMATION, FOCAL GRANULCMATOD 

INFLAMMATICN, PYOGRANULOMATOOS

 (14)

 1 (7X) 

 (15) 

1 (7%) 

(31)

1 (3*) 

 (34) 

1 (3*) 

*PLEURA

INFLAMMATICN, FOCAL GRANOLCMATOO 

CYTOMEGALY 

 (14) (15) (31)

1 (3X) 

1 (3X) 

 (3«) 

*M£SENTERY

NECROSIS, fOCAL

 (1«)

 1 (7%) 

 (15) (31) (34) 

ALL OTHER SYSTEMS 

NONE 

t NUMBER OF ANIMALS WITH TISSUE EXAMINED

* NUMBER OF ANIMALS NECROPSIED 

 MICROSCOPICALLY 

76 
TABLE D2. FEMALE MICE: NONNEOPLASTIC LESIONS (CONTINUED) 

UNTREATED

CONTROL

 VEH 

CON' 

ICLE

TROL 

 LOW DOSE HIGH DOSE 

SPECIAL MORPHOLOGY SUMMARY 

NO LESION BEPOHTED 

ANIMAL BISSING/NO NECROPSY 

NO NECROPSY PERFORMED AUTOLYSIS/NO NECROPSY 

3 

1 

3 

1 

3 

8 

1 

t NUMBER OF ANIMALS WITH TISSUE EXAMINED

* NUMBER OF ANIMALS NECROPSIED 

 MICROSCOPICALLY 

77 

APPENDIX E
 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS IN RATS
 

GIVEN INTRAPERITONEAL INJECTIONS OF ICRF-159
 

79
 

__ 

81

 

Table El. Analyses of the Incidence of Primary Tumors in Male Rats
 

Given Intraperitoneal Injections of ICRF-159a
 

Pooled Vehicle Low High
 

Topography: Morphology Control Control Dose Dose
 

Integumentary System: Fibroma'3 1/40 (3) 0/10 (0) 2/33 (6) 2/30 (7)
 

P Values0 »d N.S. N.S. N.S. N.S.
 

Relative Risk (Pooled Control)f 2.424 2.667
 

Lower Limit 0.132 0.145
 

Upper Limit 138.563 151.896
 

Relative Risk (Vehicle Control)f Infinite Infinite
 

Lower Limit 0.099 0.109
 

Upper Limit Infinite Infinite
 

Weeks to First Observed Tumor 86 78 57
 

Hematopoietic System:
 

Leukemiab 0/40 (0) 0/10 (0) 2/33 (6) 0/30 (0)
 

P Valuesc'd N.S. N.S. N.S. N.S.
 

Departure from Linear Trend6 P = 0.038
 

Relative Risk (Pooled Control)f Infinite
 

Lower Limit 0.361
 

Upper Limit Infinite
 —
 

—
 

Relative Risk (Vehicle Control)f Infinite
 

Lower Limit 0.099
 —
 

—
 Upper Limit Infinite
 —
 

Weeks to First Observed Tumor 44
 —
 
82

 

Table El. Analyses of the Incidence of Primary Tumors in Male Rats 

Given Intraperitoneal Injections of ICRF-1593 

(continued) 

Pooled Vehicle Low High
 

Topography; Morphology Control Control Dose Dose
 

Pituitary: Chromophobe 

Adenoma or Carcinoma" 3/37 (8) 0/8 (0) 3/30 (10) 1/22 (5) 

d P Values0

» N.S. N.S. N.S. N.S. 

Relative Risk (Pooled Control)f 1.233 0.561 

Lower Limit 0.177 0.011 

Upper Limit 8.542 6.399 

Relative Risk (Vehicle Control)f Infinite Infinite
 

Lower Limit 0.185 0.022
 

Upper Limit Infinite Infinite
 

Weeks to First Observed Tumor 86 78 75
 

a

Dosed groups received 48 or 96 mg/kg.
 

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

c

Beneath the incidence of tumors in a control group is the probability level for the Cochran-


Armitage test when P < 0.05; otherwise, not significant (N.S.) is indicated. Beneath the
 

incidence of tumors in a dosed group is the probability level for the Fisher exact test for
 

the comparison of that dosed group with the vehicle-control group (*) or with the pooled-


control group (**) when P < 0.05 for either control group; otherwise, not
 

significant (N.S.) is indicated.
 
Table El. Analyses of the Incidence of Primary Tumors in Male Rats 

Given Intraperitoneal Injections of ICRF-1593 

(continued) 

^A negative trend (N) indicates a lower incidence in a dosed group than in a control group.
 

e

The probability level for departure from linear trend is given when P < 0.05 for any comparison.
 

95% confidence interval of the relative risk between each dosed group and the specified
 

control group.
 

83 
Table E2.


Topography: Morphology


Integumentary System:
 

Squamous-cell Carcinomab


P Valuesc»d


Relative Risk (Pooled Control)f
 

Lower Limit
 

Upper Limit
 

Relative Risk (Vehicle Control)f
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Pituitary: Chromophobe
 

Adenomab


P Valuesc»d


Relative Risk (Pooled Control)f
 

Lower Limit
 

Upper Limit
 

Relative Risk (Vehicle Control)f
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor


 Analyses of the Incidence of Primary Tumors in Female Rats
 

Given Intraperitoneal Injections of ICRF-159a
 

Pooled
 

 Control
 

 0/38 (0)
 

 N.S.
 

 9/37 (24)
 

 N.S.
 

 84
 

Vehicle Low 

Control Dose 

0/10 (0) 0/33 (0) 

N.S. N.S. 

__ 

— 

— __ 

— 

__ _— _ 

1/9 (11) 7/31 (23) 

N.S. N.S. 

0.928 

0.331 

2.454 

2.032 

0.336 

88.007 

85 84 

High
 

Dose
 

2/32 (6)
 

N.S.
 

Infinite
 

0.355
 

Infinite
 

Infinite
 

0.102
 

Infinite
 

81
 

2/30 (7)
 

N.S.
 

0.274
 

0.031
 

1.194
 

0.600
 

0.038
 

34.226
 

73
 

84

 
85

 

Table E2. Analyses of the Incidence of Primary
 

Given Intraperitoneal Injections of
 

(continued)
 

Pooled Vehicle
 

Topography: Morphology Control Control
 

Mammary Gland: Adenocarcinoma
 

or Cystadenocarcinoma, NOSb 2/38 (5) 1/10 (10)
 

P Valuesc»d N.S. N.S.
 

Relative Risk (Pooled Control)f
 

Lower Limit
 

Upper Limit
 

Relative Risk (Vehicle Control)f
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor 85 85
 

Mammary Gland: Fibroadenomab 12/38 (32) 2/10 (20)
 

P Valuesc»d P = 0.031(N) N.S.
 

Relative Risk (Pooled Control)f
 

Lower Limit
 

Upper Limit
 

Relative Risk (Vehicle Control) ^
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor 79 85
 

Tumors in Female Rats
 

ICRF-1593
 

Low
 

Dose
 

2/33 (6)
 

N.S.
 

1.152
 

0.088
 

15.075
 

0.606
 

0.037
 

34.683
 

59
 

5/33 (15)
 

N.S.
 

0.480
 

0.148
 

1.292
 

0.758
 

0.160
 

7.306
 

83
 

High
 

Dose
 

4/32 (13)
 

N.S.
 

2.375
 

0.365
 

24.769'
 

1.250
 

0.152
 

59.496
 

44
 

4/32 (13)
 

N.S.
 

0.396
 

0.103
 

1.158
 

0.625
 

0.114
 

6.349
 

76
 
Table E2. Analyses of the Incidence of Primary 

Given Intraperitoneal Injections of 

(continued) 

Pooled Vehicle


Topography: Morphology Control Control


Uterus: Adenocarcinoma, NOSb 0/38 (0) 0/10 (0)


P Valuesc»d P < 0.001 N.S.


Relative Risk (Pooled Control)f
 

Lower Limit
 

Upper Limit
 

Relative Risk (Vehicle Control) f
 

Lower Limit
 

Upper Limit
 CO 

ON 

Weeks to First Observed Tumor —
 

Uterus: Endometrial Stromal
 

Polypb 2/38 (5) 1/10 (10)


P Valuesc'd N.S. N.S.


Relative Risk (Pooled Control)
 

Lower Limit
 

Upper Limit
 

Relative Risk (Vehicle Control)f
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor 79 85


Tumors in Female Rats 

ICRF-1593 

 Low
 

 Dose
 

 10/33 (30)
 

P = 0.048*
 

P < 0.001**
 

Infinite
 

3.477
 

Infinite
 

Infinite
 

1.012
 

Infinite
 

74
 

 2/33 (6)
 

 N.S.
 

1.152
 

0.088
 

15.075
 

0.606
 

0.037
 

34.683
 

 84
 

High
 

Dose
 

11/32 (34)
 

P = 0.030*
 

P < 0.001**
 

Infinite
 

4.008
 

Infinite
 

Infinite
 

1.166
 

Infinite
 

73
 

0/32 (0)
 

N.S.
 

0.000
 

0.000
 

3.957
 

0.000
 

0.000
 

5.791
 

—
 
87

 

Table E2. Analyses of the Incidence of Primary Tumors in Female Rats
 

Given Intraperitoneal Injections of ICRF-1593
 

(continued)
 

a

Dosed groups received 48 or 96 mg/kg.
 

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

c

Beneath the incidence of tumors in a control group is the probability level for the Cochran-


Armitage test when P < 0.05; otherwise, not significant (N.S.) is indicated. Beneath the
 

incidence of tumors in a dosed group is the probability level for the Fisher exact test for
 

the comparison of that dosed group with the vehicle-control group (*) or with the pooled-


control group (**) when P < 0.05 for either control group; otherwise, not
 

significant (N.S.) is indicated.
 

^A negative trend (N) indicates a lower incidence in a dosed group than in a control group.
 

e

The probability level for departure from linear trend is given when P < 0.05 for any comparison.
 

95% confidence interval of the relative risk between each dosed group and the specified
 

control group.
 
Table E3. Time-adjusted Analyses of the Incidence of Primary Tumors
 

in Male Rats Given Intraperitoneal Injections of ICRF-159a
 

Topography: Morphology


Integumentary System: Fibroma (52)b


P Valuesc»d


Relative Risk (Vehicle Control)f
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor


Hematopoietic System: Leukemia (44)b


P Valuesc»d


Relative Risk (Vehicle Control)f
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor


Vehicle Low High 

 Control Dose Dose 

 0/9 (0) 2/22 (9) 2/7 (29) 

 N.S. N.S. N.S. 

Infinite Infinite 

0.136 0.434 

Infinite Infinite 

— 78 57 

 0/9 (0) 2/22 (9) 0/7 (0) 

 N.S. N.S. N.S. 

__ 

Infinite 

0.136 — Infinite — 

— 44 — 

88

 
89

 

Table E3. Time-adjusted Analyses of the Incidence of Primary Tumors
 

in Male Rats Given Intraperitoneal Injections of ICRF-1593
 

(continued) 

Vehicle Low High 

Topography: Morphology Control Dose Dose 

Pituitary: Chromophobe Adenoma
 

or Carcinoma (52)b 0/8 (0) 3/21 (14) 1/6 (17)
 

P Values0>d N.S. N.S. N.S.
 

Relative Risk (Vehicle Control)f Infinite Infinite
 

Lower Limit 0.265 0.079
 

Upper Limit Infinite Infinite
 

Weeks to First Observed Tumor 78 75
 —
 

a

Dosed groups received 48 or 96 mg/kg.
 

"Number of tumor-bearing animals /number of animals examined at site (percent), based on the number
 

of animals that lived at least as long as the number of weeks shown in parenthesis after the
 

description of morphology.
 

c

Beneath the incidence of tumors in the control group is the probability level for the Cochran-


Armitage test when P < 0.05; otherwise, not significant (N.S.) is indicated. Beneath the
 

incidence of tumors in a dosed group is the probability level for the Fisher exact test for
 

the comparison of that dosed group with the vehicle-control group when P < 0.05; otherwise,
 

not significant (N.S.) is indicated.
 

^A negative trend (N) indicates a lower incidence in a dosed group than in a control group.
 

e

The probability level for departure from linear trend is given when P < 0.05 for any comparison.
 

95% confidence interval of the relative risk between each dosed group and the control
 

group.
 

APPENDIX F
 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS IN MICE
 

GIVEN INTRAPERITONEAL INJECTIONS OF ICRF-159
 

91
 

93

 

Table Fl. Analyses of the Incidence of Primary Tumors in Male Mice
 

Given Intraperitoneal Injections of ICRF-159a
 

Topography: Morphology


Hematopoietic System: Lymphoma
 

or Lymphocytic Leukemia"


P Valuesc»d


Relative Risk (Pooled Control)^
 

Lower Limit
 

Upper Limit
 

Relative Risk (Vehicle Control)f
 

Lower Limit
 

Upper Limit
 

Pooled Vehicle Low High 

 Control Control Dose Dose 

 1/42 (2) 0/14 (0) 2/34 (6) 2/30 (7) 

 N.S. N.S. N.S. N.S. 

2.471 2.800 

0.134 0.152 

141.342 159.477 

Infinite Infinite 

0.130 0.147 

Infinite Infinite 

Weeks to First Observed Tumor 85 69 64 

Liver: Hepatocellular 

Adenoma or Carcinoma^ 3/42 (7) 2/13 (15) 0/33 (0) 0/29 (0) 

P Valuesc » d N.S. P = 0.028(N) N.S. N.S. 

Departure from Linear Trend6 P = 0.045 

Relative Risk (Pooled Control)f 

Lower Limit 

Upper Limit 

0.000 

0.000 

2.086 

0.000 

0.000 

2.362 

Relative Risk (Vehicle Control) f 

Lower Limit 

Upper Limit 

0.000 

0.000 

1.303 

0.000 

0.000 

1.475 

Weeks to First Observed Tumor 86 86 — 
94

 

Table Fl. Analyses of the Incidence of Primary Tumors in Male Mice
 

Given Intraperitoneal Injections of ICRF-159a
 

(continued)
 

a

Dosed groups received 40 or 80 mg/kg.
 

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

c

Beneath the incidence of tumors in a control group is the probability level for the Cochran-


Armitage test when P < 0.05; otherwise, not significant (N.S.) is indicated. Beneath the
 

incidence of tumors in a dosed group is the probability level for the Fisher exact test for
 

the comparison of that dosed group with the vehicle-control group (*) or with the pooled-


control group (**) when P < 0.05 for either control group; otherwise, not
 

significant (N.S.) is indicated.
 

"A negative trend (N) indicates a lower incidence in a dosed group than in a control group.
 

e

The probability level for departure from linear trend is given when P < 0.05 for any comparison.
 

95% confidence interval of the relative risk between each dosed group and the specified
 

control group.
 
95

 

Table F2. Analyses of the Incidence of Primary Tumors in Female Mice
 

Given Intraperitoneal Injections of ICRF-159a
 

Topography: Morphology


Hematopoietic System:
 

Lymphoma, Histiocytic Typeb


P Valuesc»d


Relative Risk (Pooled Control)f
 

Lower Limit
 

Upper Limit
 

Relative Risk (Vehicle Control)f
 

Lower Limit
 

Upper Limit
 

Pooled Vehicle Low High 

 Control Control Dose Dose 

 0/45 (0) 0/15 (0) 1/31 (3) 5/34 (15) 

P = 0.006 P = 0.041 N.S. P = 0.012** 

Infinite Infinite 

0.078 1.680 

Infinite Infinite 

Infinite Infinite 

0.027 0.594 

Infinite Infinite 

Weeks to First Observed Tumor 86 74 

Hematopoietic System: 

Lymphocytic Leukemia or 

Lymphoma, Lymphocytic Type" 1/45 (2) 0/15 (0) 4/31 (13) 4/34 (12) 

P Valuesc'd N.S. N.S. N.S. N.S. 

Relative Risk (Pooled Control)f 

Lower Limit 

Upper Limit 

5.806 

0.609 

275.610 

5.294 

0.554 

252.309 

Relative Risk (Vehicle Control)f 

Lower Limit 

Upper Limit 

Infinite 

0.479 

Infinite 

Infinite 

0.436 

Infinite 

Weeks to First Observed Tumor 86 27 64 
__ 

96

 

Table F2. Analyses of the Incidence of Primary Tumors in Female Mice
 

Given Intraperitoneal Injections of ICRF-1593
 

(continued) 

Pooled Vehicle Low High 

Topography: Morphology Control Control Dose Dose 

Hematopoietic System:
 

All Neoplasms*3 1/45 (2) 0/15 (0) 5/31 (16) 9/34 (26)
 

P Values0 >d P = 0.002 P = 0.021 P = 0.038**	 P = 0.026*
 

P = 0.002**
 

Relative Risk (Pooled Control)f 7.258 11.912
 

Lower Limit 0.866 1.778
 

Upper Limit 330.475 501.635
 

Relative Risk (Vehicle Control)f Infinite Infinite
 

Lower Limit 0.653 1.243
 

Upper Limit Infinite Infinite
 

Weeks to First Observed Tumor 86	 27 64
 

Mesentery: Lipomab 0/45 (0) 0/15 (0) 0/31 (0)	 2/34 (6)
 

P Valuesc'd	 N.S. N.S. N.S. N.S.
 

Relative Risk (Pooled Control)f Infinite
 

Lower Limit 0.393
 

Upper Limit	 —
 Infinite
 —
 

Relative Risk (Vehicle Control)f Infinite
 —
 Lower Limit	 0.138
 —
 Upper Limit	 Infinite
 —
 

Weeks to First Observed Tumor — 86
 —	 —
 
97

 

Table F2. Analyses of the Incidence of Primary Tumors in Female Mice
 

Given Intraperitoneal Injections of ICRF-159a
 

(continued)
 

a

Dosed groups received 40 or 80 mg/kg.
 

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

c

Beneath the incidence of tumors in a control group is the probability level for the Cochran-


Armitage test when P < 0.05; otherwise, not significant (N.S.) is indicated. Beneath the
 

incidence of tumors in a dosed group is the probability level for the Fisher exact test for
 

the comparison of that dosed group with the vehicle-control group (*) or with the pooled-


control group (**) when P < 0.05 for either control group; otherwise, not
 

significant (N.S.) is indicated.
 

^A negative trend (N) indicates a lower incidence in a dosed group than in a control group.
 

e

The probability level for departure from linear trend is given when P < 0.05 for any comparison.
 

95% confidence interval of the relative risk between each dosed group and the specified
 

control group.
 

Review of the Bioassay of ICRF-159* for Carcinogeniclty
 

by the Data Evaluation/Risk Assessment Subgroup of the
 

Clearinghouse on Environmental Carcinogens
 

March 7, 1978
 

The Clearinghouse on Environmental Carcinogens was
 

established in May, 1976, in compliance with DHEW Committee
 

Regulations and the Provisions of the Federal Advisory
 

Committee Act. The purpose of the Clearinghouse is to
 

advise the Director of the National Cancer Institute (NCI)
 

on its bioassay program to identify and to evaluate chemical
 

carcinogens in the environment to which humans may be
 

exposed. The members of the Clearinghouse have been drawn
 

from academia, industry, organized labor, public interest
 

groups. State health officials, and quasi-public health and
 

research organizations. Members have been selected on the
 

basis of their experience in carcinogenesis or related fields
 

and, collectively, provide expertise in chemistry, biochemistry,
 biostatistics, toxicology, pathology, and epidemiology.
 

Representatives of various Governmental agencies participate
 

as ad hoc members. The Data Evaluation/Risk Assessment Subgroup
 

of the Clearinghouse is charged with the responsibility of
 

providing a peer review of reports prepared on NCI-sponsored
 

bioassays of chemicals studied for carcinogenicity. It is in
 

this context that the below critique is given on the bioassay
 

of ICRF-159 for carcinogenicity.
 

The primary reviewer agreed that the compound induced
 

uterine adenocarcinomas In female rats and lymphomas in
 

female mice. Although the intraperitoneal route of exposure
 

was a limitation of the study, he said that the bioassay
 

was adequate enough to support the conclusion regarding the
 

carcinogenicity of ICRF-159• The primary reviewer pointed
 

out a number of other tumors observed among the treated
 

animals.
 

The secondary reviewer noted that the tumor incidence
 

at some organ sites in treated animals was lower than in
 

controls. He attributed this to the shorter lifespan of
 

the treated animals. He added, however, that the life-
 shortening effect did not interfer with the conclusion
 

regarding the carcinogenicity of ICRF-159.
 

It was moved that the report on the bioassay of ICRF159 be accepted as written. The motion was seconded and
 

approved unanimously.
 

99
 
Members present were:
 

Gerald N. Wogan (Chairman), Massachusetts Institute of
 

Technology
 

Arnold Brown, Mayo Clinic
 

E. Cuyler Hammond, American Cancer Society
 

Joseph Highland, Environmental Defense Fund
 

Henry Pitot, University of Wisconsin Medical Center
 

George Roush, Jr., Monsanto Company
 

Michael Shimkin, University of California at San Diego
 

*	 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.
 

ft U. S. GOVERNMENT PRINTING OFFICE : 1978—260-899/3154 

100 

DHEW Publication No. (NIH) 78-1328