Supplement to the Carcinogenic Potency Database (CPDB): Results of AnimalBioassays Published in the General Literature in 1993 to 1994 and by the NationalToxicology Program in 1995 to 1996Lois Swirsky Gold,1'2 Neela B. Manley,1 Thomas H. Slone,1'2 and Lars Rohrbach1

'Division of Biochemistry and Molecular Biology, University of California, Berkeley, California USA; 2Life Sciences Division, E.O. LawrenceBerkeley National Laboratory, Berkeley, California USA

The Carcinogenic Potency Database (CPDB) is a systematic and unifying analysis of results ofchronic, long-term cancer tests. This paper presents a supplemental plot of the CPDB, including 513experiments on 157 test compounds published in the general literature in 1993 and 1994 and inTechnical Reports of the National Toxicology Program in 1995 and 1996. The plot standardizes theexperimental results (whether positive or negative for carcinogenicity), including qualitative data onstrain, sex, route of compound administration, target organ, histopathology, and author's opinion andreference to the published paper, as well as quantitative data on carcinogenic potency, statisticalsignificance, tumor incidence, dose-response curve shape, length of experiment, duration of dosing,and dose rate. A numerical description of carcinogenic potency, the TD50, is estimated for each set oftumor incidence data reported. When added to the data published earlier, the CPDB now includesresults of 5,620 experiments on 1,372 chemicals that have been reported in 1,250 published papersand 414 National Cancer Institute/National Toxicology Program Technical Reports. The plot presentedhere includes detailed analyses of 25 chemicals tested in monkeys for up to 32 years by the NationalCancer Institute. Half the rodent carcinogens that were tested in monkeys were not carcinogenic,despite usually strong evidence of carcinogenicity in rodents and/or humans. Our analysis of possibleexplanatory factors indicates that this result is due in part to the fact that the monkey studies lackedpower to detect an effect compared to standard rodent bioassays. Factors that contributed to thelack of power are the small number of animals on test; a stop-exposure protocol for model rodentcarcinogens; in a few cases, toxic doses that resulted in stoppage of dosing or termination of theexperiment; and in a few cases, low doses administered to monkeys or early termination of theexperiment even though the doses were not toxic. Among chemicals carcinogenic in both monkeysand rodents, there is some support for target site concordance, but it is primarily restricted to livertumors. Potency values are highly correlated between rodents and monkeys. The plot in this papercan be used in conjunction with the earlier results published in the CRC Handbook of CarcinogenicPotency and Genotoxicity Databases [Gold LS, Zeiger E, eds. Boca Raton FL:CRC Press, 19971 andwith our web site (http.//potency.berkeley.edu), which includes a guide to the plot of the database, acomplete description of the numerical index of carcinogenic potency (TD50), and a discussion of thesources of data, the rationale for the inclusion of particular experiments and particular target sites,and the conventions adopted in summarizing the literature. Two summary tables permit easy accessto the literature of animal cancer tests by target organ and by chemical. For readers using the CPDBextensively, a combined plot on diskette or other format is available from the first author. It includesall results published earlier and in this paper, ordered alphabetically by chemical. A SAS database isalso available. Key words: animal cancer test, carcinogenic potency, database, human carcinogen,monkey neoplasm, TD50. - Environ Health Perspect 1 07(suppl 4):527-600 (1999).http.//ehpnetl.niehs. nih.gov/docs/1999/suppl4/527-600gold/abstract.html

BackgroundThe Carcinogenic Potency Database (CPDB)is a systematic and unifying analysis of thepublished results of the diverse literature ofchronic, long-term animal cancer tests onindividual chemicals. The CPDB standardizesthe experimental results and creates an acces-sible resource widely used to address a varietyof research and regulatory issues in carcino-genesis. All results in the CPDB prior to thispaper are presented in an easily readable plotformat in the 1997 Handbook ofCarcinogenicPotency and Genotoxicity Databases (1), whichincludes all experimental results publishedin several papers in Environmental HealthPerspectives beginning in 1984 (2-7). Thispaper is a supplement to the CPDB, report-ing bioassay results published in the general

literature in 1993 to 1994 and in TechnicalReports of the National Toxicology Program(NTP) in 1995 to 1996. Our analyses arepresented in the same plot format as the ear-lier plots in Environmental Health Perspectives(2-7) and in the CRC handbook (1). Dataare reported for 513 new experiments on 157chemicals. When added to the data publishedearlier, the CPDB now includes results of5,620 experiments on 1,372 chemicals thathave been reported in 1,250 published papersand 414 National Cancer Institute (NCI)/NTP Technical Reports.

In the CPDB, detailed information andanalyses important in the interpretation ofbioassays are reported on each experiment(whether positive or negative for carcino-genicity), including qualitative data on strain,

sex, target organ, histopathology, and author'sopinion, as well as quantitative informationon carcinogenic potency, statistical signifi-cance, tumor incidence, dose-response curveshape, length of experiment, dose rate, andduration of dosing. Each set of experimentalresults references the original published paper,and a series of appendices defines the codes inthe plot. A numerical description of carcino-genic potency, the TD50 (8), is estimated foreach set of tumor incidence data reported inthe CPDB, thus providing a standardizedquantitative measure for comparisons. In asimplified way, TD50 may be defined as thatdose rate in mg/kg body wt/day which, ifadministered chronically for the standard lifespan of the species, will halve the probabilityof remaining tumorless throughout thatperiod. Stated differently, TD50 is the dailydose that will induce tumors in half of the testanimals that would have remained tumor freeat zero dose. We estimate TD50 using a one-hit model (8). TD50 is analogous to LD50,and a low value of TD50 indicates a potentcarcinogen, whereas a high value indicates aweak one. TD50 is often within the range ofdoses tested and does not indicate anythingabout carcinogenic effects at low dosesbecause bioassays are usually conducted at ornear the maximum tolerated dose (MTD).The range of TD50 values is at least 10 mil-lionfold for carcinogens in each sex of rat ormouse (1,9).

The CPDB is exhaustive in that it includesall published results of experiments that meeta set of inclusion criteria designed to measurepotency; however, because many tests do not

Address correspondence to L.S. Gold, NIEHSCenter, 401 Barker Hall, University of California,Berkeley, CA 94720-3202. Telephone: (510) 486-7080.Fax: (510) 486-6773. E-mail: lois@potency.berkeley.eduhttp://potency.berkeley.eduWe thank the many researchers who provided addi

tional experimental results for the CPDB, sometimesby having to go back to early pathology reports. U.Thorgeirsson, D. Dalgard, and S. Sieber have providedus with information for many years about the bioassaysof nonhuman primates. We also thank B.N. Ames, J.Ward, L. Bernstein, and D. Freedman for their manyyears of advice on the CPDB project. This work wassupported through the University of California,Berkeley, by National Institute of Environmental HealthSciences Center grant ES01896 and through theLawrence Berkeley Laboratory by U.S. Department ofEnergy contract DE-AC-03-76SFO0098.

Received 16 March 1999; accepted 17 June 1999.

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GOLD ET AL.

meet the criteria, not all cancer tests areincluded. No attempt has been made to per-form an evaluation of whether a compoundinduced tumors in any given experiment;rather, the opinion of the published authors ispresented as well as the statistical significanceof the TD50 calculated from their results.

In presenting this supplement to theCPDB, our intention is that the material beused as follows, in conjunction with the CRCHandbook of Carcinogenic Potency andGenotoxicity Databases (10) and/or with ourfully searchable, 100-page web site on theWorld Wide Web (http://potency.berkeley.edu), which has been accessed by individualsfrom 111 countries:* Complete results for each experiment in

the CPDB to date are obtainable by usingthe plots in the CRC handbook (1) andthe present paper.

* Detailed descriptions of the methods usedto develop the CPDB and a guide on howto read the plot format are given in theCRC handbook and on the web site. Themethods page (http://potency.berkeley.eduItext/methods.html) describes inclusion rules,sources of data, statistical methods on esti-mate TD50, selection of tissue and tumortypes to indude, estimation of the averagedaily dose rate, and extrapolation of TD50to the standard life span of each species.The guide to the plot (http://potency.berkeley.edu/text/guide.html) uses a sampleexperiment to describe each field of theplot format; special analysis and standardi-zation methods for the NCI/NTP bioas-says are also reported.

* A complete list of references that are thesource of bioassay results in the CPDB isprovided on the web site, includingpapers in the plot in this present paper(http://potency. berkeley. edultextiCPDBreferences.html).

* Two summary tables on our web site havebeen updated to include the experimentalresults presented in this paper. TheSummary Table by Chemicals in theCPDB (http://potency.berkeley.edu/chemi-calsummary.html) is organized alphabeti-cally by chemical name and summarizesresults for each sex-species group of rats,mice, hamsters, dogs, and monkeys onpositivity, target organ, and potency(TD50). Chemicals with no evidence ofcarcinogenicity are included. Informationis reported on each chemical aboutmutagenicity in Salmonella, whetherpotency values vary greatly within aspecies, and whether there is more thanone positive test in each species in theCPDB. The Summary Table of TargetOrgans in the CPDB (http://potency.berke-ley. edulpathology. html) organizes resultsfrom the CPDB by target organ so that

researchers can quickly determine allchemicals that induce tumors in eachspecies at a given site, e.g., mammarygland or liver. This table includes onlypositive chemicals; results are organizedby target organ, species, and mutagenicityin Salmonella. The table indicates whethereach carcinogen has tests in another speciesin the CPDB and whether the chemical ispositive in another species. In combinationwith the plots of the CPDB, these twotables provide a comprehensive summaryof the field of long-term chronic bioassaysand give full citations to each publicationon each chemical and each target site.For readers using the CPDB extensively, acombined plot on diskette or other formatis available from the first author thatincludes all results published earlier and inthis paper, ordered alphabetically by chem-ical. A SAS database is also available (seehttp://potency.berkeley.edu/ordeform. html).

CPDB Plot in This SupplementThis plot of the CPDB analyzes results of 513long-term, chronic experiments on 1 57chemicals, including tests in rats, mice, ham-sters, and monkeys. Many chemical classesand compounds with a variety of uses areincluded: a) drugs, e.g., fluvastatin, codeine,ritalin, phenolphthalein, tamoxifen, dehydro-epiandrosterone (DHEA), and potassiumbicarbonate; b) industrial chemicals, e.g., ace-tonitrile, methyl tert-butyl ether (MTBE),nickel sulfate hexahydrate, sodium dichro-mate, and isoprene; c) pesticides and foodadditives, e.g., captan, captafol, piperonylbutoxide, ethyl acrylate, citric acid; andd) naturally occurring components of food,e.g., phenethyl isothiocyanate, 2-amino-i-methyl-6-phenylimidazo [4,5-b] pyridine(PhIP), catechol, L-ascorbic acid, and ethylalcohol. Eighty-three of the 157 chemicalswere already included in the database, and wehave flagged these names in the plot withtriple asterisks (***). The TD50 values for thecompounds in this plot fall within the 10millionfold range reported earlier. The posi-tivity rate among chemicals reported here,based on the original author's evaluation, is94/157 (60%), which is similar to the rate forthe database as a whole.

The experimental results reported herereflect recent changes in the field of animalcancer testing. For NTP bioassays, for exam-ple, the standard bioassay protocol nowincludes three dose groups and a control; incontrast, in NCI/NTP Technical Reportspublished before 1994, only 8% of the experi-ments had more than two dose groups. In thegeneral literature, over time fewer experimentshave only one dose group, and in this plot42% of the experiments have more than twodose groups and a control. In the 1990s there

has also been an increase in the proportion ofexperiments that restrict histopathology toknown target sites, indicating investigation ofcocarcinogenic effects in parallel studies or ofmechanisms of carcinogenesis. Of the 139papers in this plot, 16 include data on celldivision rates (indicated by the notecode "C"on the plot). We have consulted with nearlyhalf the published authors and have receivedclarification of results or data in addition tothat which appeared in their published papers;these citations are marked in the plot with"pers.comm." to indicate personal communi-cation. For some literature papers we havebeen able to obtain from the authors fulllifetable data and have used those results toestimate TD50 values, e.g., tamoxifen, MTBE.

Analyses That Use the CPDBDuring the past 15 years we have publishedmany papers based on results in the CPDB,including investigations of methodologicissues such as reproducibility of bioassayresults and constraints on potency estimation,questions about extrapolation of carcinogenic-ity between species, estimation of regulatoryrisk, and providing a broad perspective oncancer risk estimates by ranking possible haz-ards at typical and average human exposuresto rodent carcinogens. A variety of exposuresare ranked, including natural chemicals in thediet, pharmaceuticals, workplace exposures,pesticide residues, and synthetic pollutants.Recently, we presented an overview of thoseresults and included the principal findings intabular form (9,11). We have now updatedeach of the analyses to include the experi-mental results presented in this paper. Thefindings in all cases are similar to the earlierpublished results, and we therefore refer thereader to the earlier overviews (9,11,12;http://potency.berkeley.edu), which also providecitations to the original papers on each topic.We continue to find that about half of allchemicals tested are carcinogenic in at leastone experiment; about half are also positivefor several subsets of the data, induding natu-rally occurring and synthetic chemicals (Table1). We have discussed in several papers the

Table 1. Proportion of chemicals evaluated as carcino-genica for several data sets in the CPDB.

Chemicals tested in both rats and miceChemicals in the CPDB 350/590 (59%)Naturally occurring chemicals 79/139 (57%)Synthetic chemicals 271/451 (60%)

Chemicals tested in rats and/or miceChemicals in the CPDB 702/1348(52%)Natural pesticides 37/71 (52%)Mold toxins 14/23 (61%)Chemicals in roasted coffee 21/30 (70%)

CPDB, Carcinogenic Potency Database.'A chemical is classified as positive if the author of at least onepublished experiment evaluated results as evidence that thecompound is carcinogenic.

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plausible explanations for the high positivityrate, induding various high-dose effects (9,11;http://potency. berkeley. edu).

Comparison of Carcinogenicityin Monkeys and RodentsThis plot includes analyses of 25 NCIcarcinogenicity studies in rhesus andcynomolgus monkeys, which lasted up to 32years. Some of the experimental results werereported in earlier plots of the CPDB (theseare marked on the plot with a "j" notecode);however, we have performed new lifetableanalyses using the final control data that haverecently become available (13). Results foreight chemicals tested in monkeys by NCI arereported for the first time. Some of the indu-sion rules and methods of the CPDB havebeen relaxed for these monkey tests; they aredescribed in Appendix 1.

Comparison of PositivityAs assessment of human cancer risk is usuallybased on studies in rodents, one ideally wantsto know whether rodent carcinogens arehuman carcinogens; however, human data arerarely available. One major goal of this seriesof experiments in monkeys was to determinewhether rodent carcinogens would also becarcinogenic in nonhuman primates (as mon-keys are more closely related to humans thanare rodents) and thus provide evidence insupport of interspecies extrapolation in car-cinogenesis (14). Conversely, if rodents andmonkeys are different with respect to carcino-genicity under laboratory conditions, thenthis casts doubt on the validity of ex-trapolations from rodents to humans. Thechemicals selected by NCI for use in monkeystudies primarily have strong evidence of car-cinogenicity in rodents, i.e., many positiverodent experiments with high tumor inci-dence rates, carcinogenic effects at dosesbelow the MTD, or short latency periods formalignant tumor induction (10,15,16). Thestrong evidence in rodents for these chemicalssuggests that when compared to other rodentcarcinogens they would be likely to be car-cinogenic in monkeys. Only one of thesechemicals, arsenic, is not a rodent carcinogenin standard bioassays; however, it is a humancarcinogen (17).

Table 2 describes positivity for the 25chemicals tested in monkeys that are reportedin detail in the plot presented here. Several ofthe test agents are model rodent carcinogens,some are nitrosamines or chemotherapeuticagents with strong evidence in rodents, and afew are synthetic food additives or pesticides.Two sweeteners, saccharin and cyclamate,have weaker evidence of carcinogenicity inrodents than the other chemicals tested.Several of the test agents have been evaluatedby the International Agency for Research on

Cancer (IARC) as human carcinogens(aflatoxin, arsenic, azathioprine, cyclophos-phamide, melphalan) (17); of these, onlyaflatoxin and melphalan were evaluated ascarcinogenic in these monkey studies.

Table 2 indicates that only 1 1 of 25 chemi-cals were evaluated by NCI as monkey car-cinogens under the conditions of these studies;3 had equivocal results, and 11 were not car-cinogenic (13,18). The NCI evaluations arebased on malignant tumors only. For monkeycarcinogens the tumor yields were higher (seeplot) and the latency period shorter than forchemicals that were not carcinogenic. For 8 ofthe 11 monkey carcinogens, the first tumoroccurred in animals that died before 5 years ontest and for the other 3 carcinogens by 13years. Nearly all chemicals tested in monkeysare mutagens in Salmonella and about half arenot carcinogenic in monkeys. A few chemicalswere not mutagenic, and none of them werecarcinogenic in monkeys, e.g., DDT, sodiumsaccharin, sodium cyclamate, sodium arsenate.

Possible explanatory factors for lack ofevidence of carcinogenicity in monkeys forthese rodent and human carcinogens includea) a true species difference including species-specific mechanism of action; b) a small num-ber of monkeys on test, making it difficult todetect an effect; c) an experiment in onlycynomolgus or rhesus monkeys but not both(Appendix 1); d) dose levels that may havebeen too low; e) dose levels that may havebeen too high (thus producing toxic effectsthat reduced life span and therefore the powerto detect an effect); J) early termination ofexperiment before natural death; and g) otheraspects of experimental design such as routeof administration or length of dosing.

Table 2 indicates that experimentaldesigns varied widely across this series of testsin monkeys with respect to whether bothcynomolgus and rhesus were tested, numberof monkeys on test, length of dosing andlength of experiment, and average daily life-time dose rate compared to rodent averagedaily lifetime dose rate. Detailed results oneach experiment are given in the plot, indud-ing route of administration, dose rate, tumortypes and incidence, TD50 and its confidencelimits, and shape of the dose response.

We have investigated factors that mightaccount for the lack of carcinogenic effects inmonkeys, while noting that because of thesmall number of chemicals, the small numberof animals, and the wide variation in proto-cols, it is difficult to conclude which factorsmay be most important. First, the number ofdosed rhesus or dosed cynomolgus monkeyswas generally fewer than 15 (Table 2). Thusone cannot rule out the possibility that a pos-itive result might have been obtained with alarger number of animals. Second, the spon-taneous tumor rate in the large colony control

at NCI is low (13,19); thus, if a tumoroccurred in even one or two of the smallnumber of dosed animals, the results are diffi-cult to interpret. Compared to rodents, whichhave a high spontaneous tumor rate, the life-time spontaneous tumor rate in the NCImonkey colony is low: in rhesus controls,19/120 (16%) had tumors (seven malignant),and in cynomolgus monkeys, 4/106 (4%)had tumors (three malignant). No type oftumor occurred spontaneously in more thantwo animals, except uterine leiomyoma,which occurred in 6 rhesus monkeys and 1cynomolgus monkey.

For the model rodent carcinogens (14),the testing protocol used a short dosingperiod (5 years, i.e., one-fourth the standardlife span of 20 years) (Table 2). Under thesestop-exposure conditions only one chemical(urethane) was carcinogenic, even though thestudies were continued for 20-32 years. Incontrast, all of the model carcinogens inducetumors in rodent experiments lasting lessthan 6 months, i.e., less than one-fourth thestandard life span of 2 years (15,16). Underthe conditions of the 5-year dosing protocol,the following model rodent carcinogenswere not carcinogenic in monkeys: 2-acetyl-aminofluorene, 2,7-acetylaminofluorene,N,N-dimethyl-4-aminoazobenzene, 3'-methyl-4-dimethylaminoazobenzene,3-methylcholanthrene (18). Urethane had theweakest evidence of carcinogenicity amongthe monkey carcinogens: the latency periodwas longer than those of other monkey car-cinogens, and only one tumor of any type wasinduced, i.e., no site was a target site for morethan one animal in a monkey experiment.Thus, the stop-exposure design was less sensi-tive than the chronic dosing protocol (whichalso gives a higher total dose and higher aver-age daily dose rate) that is used in standardrodent bioassays and in most of the monkeyexperiments. A true species difference cannotbe ruled out, however, because monkeys weredosed for 5 years of a 20-year life span, i.e.,one-fourth and this is the same proportionthat produced carcinogenic effects in rodentsfor these chemicals, i.e., 6 months of a 2-yearlife span.

A difference in route of chemical adminis-tration between monkey tests and the positivetests in rodents does not appear to be impor-tant in explaining why many rodent carcino-gens are not carcinogenic in monkeys. Routesof administration were more often differentfor the rodent carcinogens that were positivein monkeys than for the rodent carcinogensthat were not carcinogenic in monkeys.We examined whether the number of

animals on test or the dose rate (mg/kg/day)may have been factors affecting positivity inmonkeys. Our analysis first found the medianvalue for the combination of negative and

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GOLD ET AL.

Table 2. Protocol characteristics and carcinogenicity of 25 chemicals tested in monkeys by the National Cancer Institute.

Rhesus monkeys Cynomolgus monkeysAuthor's opinion of Starting Exposure Experiment Dose Starting Exposure Experiment Dosecarcinogenicity in monkeys number (years) (years) ratioa number (years) (years) ratioa

NOT CARCINOGENIC2-Acetylaminofluorene

2,7-AcetylaminofluoreneArsenate, sodiumCyclamate, sodiumDDT

N,N-Dimethyl-4-aminoazobenzene3'-Methyl-4-dimethylaminoazobenzene3-MethylcholanthreneN-Nitrosodimethylamine

N-Methyl-N'-nitro-N-nitrosoguanidineSaccharin, sodium

EQUIVOCALAdriamycin

AzathioprineCyclophosphamide

CARCINOGENICAflatoxin B1Cycasin mixture (diet)(IPJ)

IQ

Melphaland

N-Nitroso-N-methylurea

N-Nitrosodiethylamine (diet)(IPJ)

N-NitrosodipropylamineN-Nitrosopiperidine (diet)

(IPJ)

Procarbazine.HCI

Sterigmatocystin

Urethane

10 5 26 0.5 (R)0.1 (M)

7 5 32 ND

9 24 24 0.3 (M)11 11 25 0.4(R)

0.3 (M)6 5 20 0.7 (R)

13 5 24 0.8 (R)9 5 26 0.9 (R)6 10 10 0.4 (R)

2 (M)18 23 23 0.2 (R)7 23 23 0.009 (R)

656

1813

18105

25

101511

15188

913161613

15248

NDNDND1 (M)2 (R)0.8 (M)

0.4 (R)NDcNDc

8 16 19 0.2 (R)0.3 (M)

19 18 22 25 (R)0.7 (M)

14 22 22 6 (R)82 20 20 2 (R)4 3 3 4 (R)7 12 12 22 (R)

21 (M)6 20 20 0.6 (R)

0.5 (M)22 16 23 0.4 (R)

0.7 (M)

11 5 25 2 (R)0.2 (M)

7199

13

5

6

5489

176

36

12

19

1661

5

17

30

6

5142411

5

23

5101511

1718

9

16

18

1616

12

16

17

5

29142425

ND_b

0.3 (M)0.4(R)0.3 (M)

27 0.7 (R)

23 0.009 (R)

15151613

NDND1 (M)2 (R)1 (M)

17 0.5 (R)24 NDC

9 1 (R)0.4 (M)

18 0.2 (R)0.3 (M)

22 20 (R)0.6 (M)

16 7 (R)16 2 (R)

12 32 (R)29 (M)

19 0.4 (R)0.8 (M)

17 0.09 (R)0.07 (M)

23 2 (R)0.2 (M)

Abbreviations: CPDB, Carcinogenic Potency Database; IPJ, intraperitoneal injection; IQ, 2-amino-3-methylimidazo[4,5-flquinoline; M, mice; ND, no rodent tests meet inclusion rules of the CPDB, but posi-tive in other rodent test; R, rats.aRatio from CPDB of highest monkey daily lifetime dose rate (mg/kg/day)/highest rodent daily lifetime dose rate (mg/kg/day). bNo positive rodent test. Ratio of the negative monkey dose in CPDB to thehighest negative rodent dose is 0.0009 (rats). CMonkeys received a mixture of cycad flour, cycasin, and methylazoxymethanol acetate. There are no rodent tests of this mixture; cycad flour is carcinogenicin rats. dMelphalan was not carcinogenic in rhesus monkeys.

positive monkey experiments for the numberof monkeys tested and for the ratio of themonkey dose rate compared to the positiverodent dose rate. For the number of animalswe took the total number of monkeys thatstarted on test for both monkey species com-bined, as the NCI evaluation of carcinogenic-ity for a chemical did not distinguish betweenthe two species (18). The median value of thenumber of monkeys on test for the 24 rodentcarcinogens in Table 2 was 19. We found asignificant difference between the monkey pos-itives and negatives. All of the chemicals inTable 2 that were NOT CARCINOGENIC

in monkeys were below the median number ofanimals on test, except DDT. Only a few ofthe CARCINOGENIC chemicals were belowthe median. The three EQUIVOCAL chemi-cals were all above the median, thus suggestingthat the greater power provided by more ani-mals may be a factor in the evaluation ofequivocal compared to not carcinogenic. All ofthe model rodent carcinogens had the stop-exposure protocol, and also had fewer thanthe median number of monkeys on test, thusproviding a protocol with little power.

To assess whether the dose in monkeys(mg/kg/day) was low compared to the positive

doses in rodents (mg/kg/day), we used theratio (Table 2) of the average daily dose ratein monkeys to that in rodents, and againfound the median for all chemicals regardlessof carcinogenicity result in monkeys. Forrodents, the dose was taken from a positiveexperiment with the same route of adminis-tration as the monkey experiment, whereverpossible. Table 1 reports the ratio of the high-est daily average lifetime dose rate in monkeys(mg/kg/day) to the highest in rats or in micefrom the CPDB (mg/kg/day). The medianfor all chemicals was 0.4, indicating that forhalf the chemicals tested, the monkey dose

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was about half the highest dose that inducedtumors in rodents. The monkey doses were

generally close to the rodent doses, even whenthey were lower. We repeated this analysisthree ways, using the ratio in Table 2, theratio using the lowest positive high dose ineach rodent species, and the lowest dose at

which tumors were significantly increased ineach rodent species. These measures giveincreasing weight to how low a dose inrodents was tumorigenic, and therefore thesecond and third ones produce highermonkey-to-rodent ratios than the value ofthe first, which is presented in Table 2.Regardless of which dose ratio was selected,there was no difference between the chemicalsthat were carcinogenic in monkeys and thosethat were not carcinogenic. The overall differ-ence in positivity between monkeys androdents does not appear to be due to doselevel, as the monkey doses (mg/kg/day) were

generally not much below the doses thatinduced tumors in rodents. For the 5 humancarcinogens the monkey doses were similar to

the human carcinogenic doses even for the 3chemicals that were not evaluated as carcino-genic in monkeys (arsenic, azathioprine,cyclophosphamide) (20-22).

Some aspects of the conduct of thesemonkey experiments contributed to a lack ofsensitivity for some chemicals that were not

carcinogenic in monkeys. All but one of thepositive chemicals were tested in both rhesusand cynomolgus monkeys, i.e., there were at

least five cynomolgus and five rhesus on test.

For 6 of the chemicals that were not carcino-genic, however, only one monkey species hadfive animals on test. Among the 5 human car-

cinogens, only aflatoxin and melphalan were

evaluated as carcinogenic in monkeys; how-ever, for the others (azathioprine, cyclophos-phamide and arsenic) the experiments lackedpower to detect a carcinogenic effect becausethey were terminated early (by 16 years) even

though the doses were not toxic.Because the evaluations of carcinogenicity

in monkeys were based on malignant tumors

only, whereas evaluations in rodents usuallyconsider both malignant and benign tumors,it is possible that if benign tumors had beenconsidered, more monkey studies might havebeen evaluated as positive. This was not thecase, however, as there were few benigntumors in tests of chemicals that were not

carcinogenic, and nearly all were tumor typesthat also occur in control monkeys.

As a second general approach to under-standing the discordance between rodentsand monkeys, we searched on a case-by-casebasis for idiosyncratic factors that may explainwhy some rodent carcinogens were not car-

cinogenic in monkeys. In a few cases, dosingwas stopped or animals died because of toxic-ity, and this may be the reason for a negative

result. For example, for DDT the dose wasneurotoxic to rhesus monkeys and dosing hadto be stopped at 10 years. In cynomolgus,however, the dose was not neurotoxic, thusproviding a short exposure that may havebeen less effective than the exposures for mostchemicals that were carcinogenic in monkeys.

Of the 4 nitrosamines that were positive inmonkeys, all were tested at doses more than 3times the rat carcinogenic dose (Table 2). Bothnitrosamines that did not induce tumors inmonkeys were tested at less than half the ratcarcinogenic dose. For N-nitrosodimethy-lamine, which did not induce tumors, theadministered doses produced toxic hepatitis,and all animals died by 10 years; thus, the ani-mals may not have lived long enough todevelop tumors. For N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) a published studyindicates that it is carcinogenic in cynomolgusmonkeys (23). In that positive cynomolgusexperiment (23), MNNG induced stomachtumors in just 1 year; MNNG was given bygavage 3 times per month at a 10-fold higheradministered dose than the negative 23-yearNCI rhesus experiment in which administra-tion was by diet 5 times per week. Thus, ahigher administered dose given less frequentlyby a different route to a different monkeyspecies, induced tumors. One or a combina-tion of these factors likely contributed to thedifference in result from the NCI monkey test.

For the two sweeteners, sodium saccharinand sodium cyclamate, doses were selected onthe basis of human consumption (14). Thesaccharin dose was equivalent to 5 cans of dietsoda daily, and the cyclamate dose was equiva-lent to 30 cans daily (18). The average dailydose rate of sodium saccharin gave the lowestdose ratio of all chemicals (0.009) (Table 2),and sodium cyclamate was also relatively low(0.3). Neither chemical was carcinogenic inmonkeys. In rats, induction of bladder tumorsfrom sodium saccharin appears to require ahigh dose and is related to development of acalcium phosphate-containing precipitate inthe urine (24). At the low dose administeredto monkeys, there was no effect on the urineor urothelium, and no evidence of increasedurothelial cell proliferation or of formation ofsolid material in the urine (25). Thus, onewould not expect to find a carcinogenic effectunder the conditions of the NCI study inmonkeys. Additionally, however, there may bea true species difference, because primateurine has a low concentration of protein andis less concentrated (lower osmolality) than raturine (25). Human urine is similar to monkeyurine in this respect (24).

Six of the chemicals tested were antineo-plastic and immunosuppressive drugs.Monkeys were administered doses within afactor of 2 of the human therapeutic doses,which are also similar to doses that induced

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tumors in rodents (Table 2). Three of thesedrugs were carcinogenic to monkeys (melpha-lan, N-methyl-N-nitrosourea, procarbazine),and three were evaluated as EQUIVOCAL(adriamycin, azathioprine, cyclophospha-mide). Experiments with all three of theEQUIVOCAL drugs were less sensitive thana standard life span study because they wereterminated by 16 years (adriamycin becauseof cardiotoxicity). For azathioprine andcyclophosphamide the doses were not toxic.

We note that most of the chemicals thatare carcinogenic in monkeys are naturallyoccurring chemicals (Table 2). Among 12 nat-ural chemicals tested in monkeys, 8 are posi-tive (67%), whereas among 13 syntheticchemicals only 3 are positive (23%). Twochemicals, both naturally occurring, inducedtumors rapidly in monkeys: N-nitrosodi-propylamine (in 3 years) and 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) (in 8years). These results in monkeys are consistentwith the finding that among the agents thatIARC has evaluated as human carcinogens,64% (35/55) are naturally occurring (15,17).

Comparison of Target Sitesand PotencyConcordance in target sites according to thepublished author's opinion is reported inTable 3 for 11 chemicals that are positive inmonkeys and rodents in the CPDB. Thisanalysis is complicated by two facts: first, theevaluation of carcinogenicity for the monkeystudies was based on malignant tumors only(18), whereas in rodents, evaluations of car-cinogenicity often include benign tumors.We have indicated with an asterisk (*) inTable 3 those target sites for which theauthor's evaluation was based on a tumorincrease that included benign tumors. A sec-ond complication of this analysis is the factthat there are frequently many positive exper-iments in rodents for a chemical; repeatedtesting provides an increased chance offinding a target site in common betweenmonkeys and rodents.

The liver is the most common target siteof chemically induced cancer in monkeys androdents (Table 3), even among the chemicalsthat were noncarcinogenic in monkeys.Unlike rodents, monkeys had no spontaneousliver tumors in these lifetime studies (13). Wenote that among chemicals that are positive inboth monkeys and rodents, there is a targetsite in common between monkeys and at leastone rodent species for all chemicals exceptmelphalan, which was evaluated as carcino-genic in monkeys on the basis of "all malig-nant tumors" (13). As expected, the liver isthe most frequent site in common betweenmonkeys and each rodent species. Only 3 car-cinogens have a target site other than liver incommon between rodents and monkeys

531

GOLD ET AL.

Table 3. Comparison of target sitesa for eleven chemicals that are carcinogenic in monkeys and rodents.

Chemical agent Rhesus monkeys Cynomolgus monkeys Rats Mice

Aflatoxin B, Liver, gall bladder/bile Liver, gall bladder/bile Liver, *colon, kidneyduct, vascularc duct, vascularc

Cycasin mixture Liver, kidney Liver Liver, large intestinedIQ Liver Liver, clitoral gland, large and small Liver, *forestomach, *lung

intestine, mammary gland, oral cavity,*pancreas, *preputial gland, *sebaciousgland, skin, *urinary bladder, *vascular,cZymbal's gland

Melphalan - "All malignant tumors" Peritoneum *Lung, lymphosarcomaNNitrosodiethylamine Liver Liver Liver, esophagus, *kidney, oral cavity,

*stomach, vascularcN-Nitrosodipropylamine Liver Liver, *esophagus, nasal cavityNNitroso-Nmethylurea Upper gastrointestinal tract Upper gastrointestinal tract Forestomach, *lung, *nervous system Stomach, vascularN-Nitrosopiperidinee Liver Liver Liver, esophagus, nasopharynx *Liver, forestomach, *lungProcarbazine.HCI Leukemia Leukemia Leukemia,d brain, lymphoma, mammary Leukemia,d brain, *lung, lymphoma,

gland lymphosarcoma, uterusSterigmatocystin Liver Liver, vascularc VascularUrethanebe Liver, vascular,cjejunum Lung, vascular,c pancreas 'All malignant tumors" Liver, *lung, vascular,c reticulum

cell sarcoma

IQ, 2-amino-3-methylimidazo[4,5-flquinoline.*When tumors were induced in monkeys and rodents at the same target site, the site is listed first for each species and is italicized. In monkeys, opinions are based on malignant tumors. In rats and mice,asterisk (*) indicates that the author's opinion for a target site included benign tumors. hOnly one monkey in each indicated species had a tumor at the target site. cVascular tumors in the liver. dFor cycasin,results in rats are from a bioassay of cycad flour 115). For procarbazine.HCI, leukemias were induced in bioassays that did not meet inclusion criteria in the Carcinogenic Potency Database ( 15). N-Nitrosopiperidine and urethane were also tested in hamsters. For Nnitrosopiperidine, target sites in hamsters were similar to those in rats and mice (liver, digestive tract, respiratory system); for urethane,the sites in hamsters were different.

(Table 3): N-nitroso-N-methylurea (gastroin-testinal), procarbazine.HCI (leukemia), andurethane (lung and liver hemangiosarcoma).Thus, while there is some support for targetsite concordance, it is primarily in liver.

The predominance of liver as a target sitein laboratory studies in rodents and monkeysis noteworthy because in the United States,human liver cancer is rare. Chronic inflamma-tion of the liver caused by Hepatitis B and Cviruses is a common cause of liver cancer inAsia and Africa (26). The rodent liver may bea common site in carcinogenesis bioassaysbecause the administered doses are high, andthe liver is the primary site for detoxification.High doses can frequently cause increasedliver cell proliferation by inducing cytotoxicityand regeneration This effect is similar to whatis seen in the chronic hepatitis-inducedinflammation in humans. High doses can alsoincrease cell proliferation by other means suchas peroxisome proliferation, inhibiting apop-tosis with an accumulation of cells (especiallyin foci), inducing apoptosis with consequentregeneration, or by other mechanisms (27).

Carcinogenic potency values are highlycorrelated between monkeys and rodents forthe chemicals that are carcinogenic to both(r = 0.79 for log TD50). This result isexpected, given the similarity in doses thatwere administered to rodents and monkeys(Table 2: "Dose ratio") and the fact thatpotency estimates are constrained to a narrowrange about the high dose tested (28,29).Given the wide range of administered dosesacross chemicals, the correlation in potencyfollows statistically (28).

Half the 22 rodent carcinogens in theCPDB that have been tested by NCI in mon-keys were not carcinogenic in monkeys,according to the published authors. Thisoverall result does not provide strong evi-dence of the validity of extrapolation of car-cinogenicity between species. Our analysisindicates, however, that this result is due atleast in part to the fact that the monkeystudies lacked power to detect an effect com-pared to standard rodent bioassays. Factorsthat contributed to the lack of power are thesmall number of animals on test, a stop-expo-sure protocol for model rodent carcinogens,in a few cases toxic doses that resulted instoppage of dosing or termination of theexperiment, and in a few cases low dosesadministered to monkeys or early terminationof the experiment even though the doses werenot toxic. Among chemicals carcinogenic inboth monkeys and rodents, there is some sup-port for target site concordance, but it is pri-marily restricted to liver tumors. Potencyvalues are highly correlated as expected.

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