### EPA/630/R-03/003F

March 2005



# Supplemental Guidance for Assessing Susceptibility from

Early-Life Exposure to Carcinogens

Risk Assessment Forum

U.S. Environmental Protection Agency

Washington, DC 20460

### AUTHORS AND CONTRIBUTORS

This document was prepared by a Technica l Panel of the Risk Assessm ent Forum

.

AUTHORS

Hugh Barton, National Health and Environm en tal Effects Research Laboratory, OR D

Jim Cogliano, National Center for E nvironm ental Assessm ent, ORD

Michael P. Firestone, Office of Children's Health Protection, OA

Lynn Flowers, National Center for E nvironm ental Assessm ent, ORD

R. W oodrow Setzer, National Health and Envi ronm ental Effects Research Laboratory,

ORD

Larry Valco v ic, Nation a l Center f o r Environm ental Assessment, ORD

Tracey W o odruff, Office of Policy, Econom ics, and Innovatio n, OA

CONTRIB U TORS

Neepa Choksi, Fellow to the Office of Solid W a ste and Em ergency Response

Resha M. Putzrath, Risk Assessm ent Forum Staff, National C e nter for Environm ental

Assessm ent, ORD

W illiam P. Wood, Risk Assessm ent Forum Staff, National Center for Environm ental, ORD

ii

### ACKNOWLEDGME N TS

We would especially like to thank D a vid Bennett f o r his ear lier work in le ading the initia l

efforts for this work, Bill W ood for his support, a nd Julian Preston for his helpful review of the

docum ent. Special than ks also go to Rebecca Brown, Rosemary Castorin a, Ellen Phelps, and

Linda Poore for their efforts in pullin g together the underlying inform ation.

iii

### CONTE N T S

PREFACE

........................................................................................................................ ............... v

1. I NTRODU CTION



### .............................................................................................................................. 1

2. P RO CE DU RE S



### ............................................................................................................................... .. 6

2.1. DATA SOURCES FOR ANIMAL STUDIES

................................................................... 6

2.2. EVAL UATING T H E MODE OF ACTION OF CARCINOGE NS

................................... 8

2.3. QUANTITATIVE METHODS

.......................................................................................... 9

2.4. IONIZING RADIATION

................................................................................................. 12

3. R ESULTS



### ............................................................................................................................... ....... 13

3.1. QUALI T ATIVE E VALUATI ON OF THE DAT ABASE

............................................... 13

3.2. QUANTITATIVE EVALUATI ON OF THE DATABASE

............................................ 15

3.2.1. Carcinogens with a Mutagenic Mode of Action

........................................................ 16

3.2.1.1. Early Postnatal, Juvenile, and Adult Re peated Dosing Studi es o f

Chemicals

with a Mutagenic Mode of Action

......................................................................... 16

3.2.1.2. Acute Dosing Studies of Chemicals with a Mutagenic Mode of Action

........ 17

3.2.2. Carcinogens with Modes of Action Other Than Mutagenicity

.................................. 19

3.2.3. Ionizing Radiation

...................................................................................................... 21

4. D ISCUSSI ON



### ............................................................................................................................... .. 23

5. G UIDANCE FOR ASSESSING CANCER RI SKS FROM EARLY



### - LIFE EXPOSURE



### .................................... 30

6. C OM B I N I NG 

### L IFESTAGE 

### D IFFERENCE S IN 

### E XPOSU R E AND 

### D OS E 

### -R ESPONSE 

### W HEN 

### A SSE SSING



### C ARCINO GEN 

### R ISK 

### - S OME 

### E XAM P LES FOR 

### C ARCINOGENS 

### T HAT 

### A CT 

### T HRO U GH A 

### M UTAGENIC



### M ODE OF ACTION



### ........................................................................................................................ 36

6.1 C ALCUL A TING 

### L IFET IM E 

### R ISKS 

### A SSOC I ATED WITH 

### L IF ETI M E 

### E XPOSU R ES



### ........................... 36

6.2 C ALCUL A TING 

### L IFET IM E 

### R ISKS 

### A SSOC I ATED WITH 

### L ESS THAN 

### L IFETIM E 

### E XPOSU R ES



### ....... 368

A PPENDIX 

### A: T AB LES



### ................................................................................................................... A-1

Table 1a. C h em icals that have been found to have carcinogenic effects from prenatal or postnatal

exposure in anim als as identified in different review articles........................................A-1

Table 1b. L i st of chem icals considered in this an alysis. (These are chem icals for which there are

both early-life and adult exposure reported in the sam e ani m a l experim e nt.)......... ...........A-3

Table 2. Methodological inform ation and tum o r incidence for anim al studies with early

postnatal and juvenile and adu lt repeated exposures..................................................A-5

Table 3. Methodological inform ation and tum o r incidence for anim al studies with early

postnatal and juvenile and adult acute exposure...................................................A-17

Table 4. Ratio of early-life to adult cancer potencies for studie s with repeated exposures of

juvenile and adult anim a l s to carcinogen s with a mutagenic mode of action.....................A-44

Table 5: Ratio of early-life to adult cancer po tencies for studies with repeated exposures of

juvenile and adult anim a l s to chem icals with other than m u tagenic m odes of action.......A-45

iv

### Table 6. Ratio of early-life to adult cancer potencies for st udies with acute exposures of

juveniles and adult animals to chem icals with m u tage nic m o des of action.....................A-46

Table 7. Ratio of early-life to a dult cancer potencies for studies with lifetim e exposures starting

with juvenile and adult...................................................................................A-57

Table 8. Summary of quantitative estim ates of ratio of early- life to adult cancer

potencies...................................................................................................A-60

Table 9. Excess Relative Risk (ERR) estim a te s for cancer incidence from Life Span Study

(Japanese survivors)......................................................................................A-61

Table 10. E x cess Relative Risk estim ates for incidence of thyroid cancer from Life Span

Study........................................................................................................A-62

Table 11. Coefficients for the Revised Methodo logy m o rtality risk m odel (from U.S. EPA,

1999)........................................................................................................A-63

R EFER ENC E S



### ............................................................................................................................... .... R-1

LIST OF F I GURES

Figure 1. Flow chart for early -life risk assessm ent using m ode of action fra m ework

............... 42

Figure 2. Posterior, unweighted geom etric m eans and 95% confidence intervals for the ratios of

juvenile to adult cancer potency for carci nogens acting prim arily through a m u tagenic

mode of action............................................................................................................. 43

Figure 3. Study designs

........................................................................................................ ...... 44

v

### PREF ACE

U.S. Environm ental Pro t ection Agency (EPA or the Agency ) cancer risk assessm ents

m a y be conducted differently th an envisioned in this Supplem ental Guidance for m a ny reasons

including, for exam ple, new infor m ation, new scientific understanding, or different science

policy judgm e nt. The practice of risk assessm ent with respec t to accounting for early-life

exposures to toxicants conti nues to develop, and specific com ponents of this Supplem ental

Guidance may becom e outdated o r may otherwis e require m o dification in individual settings. It

is EPA’s intent to use, to the extent practi cab le an d consistent with Agency statu t es an d

regulations, the best available sc ience in its risk assessm ents and regulatory actions, and this

Supplem ental Guidance is not inte nded to provide any substantiv e or procedural obstacle in

achieving that goal. Therefore, the S upplem en tal Guidance has no binding effect on EPA or on

any regulated entity. Where EPA does use the approaches in the Supplemental Guidance in

developing risk assessm ents, it will b e because E P A has decided in the context of that risk

assessm ent that the approaches from the Suppl em ental Guidance are suitable and appropriate.

This judgm ent will b e tested th rough peer review , and the ris k assessm ent will b e m o dified to

use different approaches if appropriate.

This Supplem ental Guidance is intended fo r guidance only. It does not establish any

substantive “rules” under the Ad m i nistrative P r ocedure Act or any other law and has no binding

effect on EPA or any regulated entity, but instead represents a non-binding statem ent of policy.

The Supplem ental Guidance addresses a num ber of issues pertaining to cancer risks

associated w ith early-life exposures genera lly, but provides specific guidance on potency

adjustm e nt only for carcinogens acting through a m u tagenic mode of action. This guidance

recomm ends for such chem icals, a default appro ach using estim ates from chronic studies (i.e.,

cancer slope factors) with appropr iate m odifications to address th e potenti al for di fferent i a l ri sk

of early-lifestage exposure. Default adjustm e nt factors are m eant to be used only when no

chem ical-sp ecific data are availab l e to asses s directly cancer suscep tibility f r om early -lif e

exposure to a carcinogen acting thro ugh a m u tagenic m ode of action.

The Agency considered both the adv a ntag es and disadvantages of extending the

recomm ended, age dependent adjustm e nt factors fo r carcinog enic potency to carcinog enic agen ts

for which the m ode of action rem a ins unknown. EPA recomm ends these factors only for

carcinogens acting through a m u tagenic m ode of action based on a com b i n ation of analysis of

available data and long-s t anding s c ience policy p o sition s that set out the A g ency’s overall

approach to carcinogen risk assessm e n t, e.g., th e use of a linear, no threshold extrapolation

procedure in the absence of data in order to be he alth protective. In gene ral, the Agen cy prefers

vi

### to rely on analyses of data ra ther than on general defaults. W h en data are available for a

susceptible lifestage, they should be used directly to evaluate ri sks for that chem ical and that

lifestage on a case-by-case basis. In the case of nonm utagenic carcinogens, when the mode of

action is unknown, the data were judged by EPA to be too limited and th e modes of action too

diverse to use this as a category for which a gene r a l def a ult ad justm e nt f actor approach can be

applied. In this situation per the Agency’s Guidelines for Carcinogen Risk Assessment , a linear

low-dose extrapolation m e thodology is recommended. It is the Agency’s long-standing science

policy position that use of the linear low-dos e extrapolation approach (without further

adjustm e nt) provides adequate publ ic health conservatism in th e absence of che m ical-specific

data indicating differential ea rly-life susceptibility or when the m ode of action is not

mutagenicity.

The Agency expects to p r oduce add i tional supplem e ntal guidance for other m odes of

action, as data from new research and toxicity testing indicate it is warranted. EPA intends to

focus its research, and to work collaboratively w ith its federal pa rtners, to im prove understanding

of the im plications of earl y life exposure to carcinogens. Developm ent of guidance for

estrogenic agents and chem icals acting through othe r processes resulting in endocrine disruption

and subsequent carcinogenesis, for exam ple, m i ght be a reasonable priority in light of the hum an

experience with die t hylstilb esterol a nd the existing early -lif e anim al studies. I t is wo rth noting

that each m o de of action for endocrin e disrup tion will prob ably require separate analy s is.

As the Agency exam ines additional carcinog enic agents, the age groupings m a y differ

from those recomm ende d for assessing cancer risks from early-life exposure to chem i cals with a

mutagenic mode of action. Pubert y and its associated biological changes, for exam ple

, involve

m a ny biological processes that coul d lead to chan ges in susceptibility to the effects of som e

carcinogens, depending on their m o de of action. The Agency is intere sted in identifying

lif estag e s th at m a y be particula r ly se nsitiv e or ref r acto r y f o r c a rcinog enesis, and believ es tha t the

mode of action fra m ework described in the Agency’s Guidelines for Carcinogen Risk

Assessment is an appropriate m echanism for elucid ating these lifestag e s. F o r each add i tiona l

mode of action evaluated, the various age grouping s determ ined to be at differential risk m a y

differ from those described in this Supplem enta l Guidance. For exam ple, the age g r o upings

selected for the age-dependent adjustm e nts were initially sele c t ed based on the ava ilab l e data,

i.e., for the laborato r y an im al age ran g e represen tative of birth to < 2 years in hum ans. More

lim ited data and inform a tion on human biology are being used to determ ine a science-inform ed

policy regarding 2 to < 16 years. D a ta were not av ailable to refine the la tter age group. If m o re

data becom e available regarding carcinogens wi th a m u tagenic m ode of a c tion, consideration

m a y be given to further refinem e nt of these age groups.

vii

### Access to data and other inform ation relating to the Cancer Guidelines (U.S. EPA, 2005)

and this Supplem e ntal Guidance will be th rough EPA' s Risk Assessm ent Forum website, under

Publica tions , Guideline s

, Guideline s f o r Cancer Risk Assessm e nt. The URL is

http://www.epa.gov/cancerguidelines . The data and results o f analyses are available in

spreadsheets.

viii

### 1. INTRO DUCTION

Cancer risk to children in the context of the U.S. Envi ronm ental Pro t ec tio n Agency’s

cancer guidelines (U.S. EPA, 2005) includes both early -life exposures that m a y result in the

occurrence of cancer duri ng childhood and early-lif e exposures that m a y contribute to cancers

later in life. The National Research Counc il (N RC, 1994) recomm ende d that “EPA should

assess risks to infants an d child ren whenever it a ppears that their risks m i ght be greater than

those of adults.” This docum e nt foc u ses on can cer risks from early-life exposure compared with

those from e xposures occurring later in lif e. Evaluating childhood cancer and childhood

exposures resulting in cancer later in life are relate d, but separable, issues.

Historically, the focus on cancer has been as a disease associated with aging, resulting

from extended exposure duration with prolonged la tency periods before the cancers appear.

Because m u ch of cancer epidem iology addresse s occupational exposures and because rodent

cancer studies are d e sig n ed to last approxim a tely a lifetim e (two years) beginning af ter sexual

m a turity, the cancer database used by EPA and other agencies for risk ass e ssm ent focuses on

adults. However, extensive literature dem onstrates that exposures early in life (i.e.,

transp lacen tal or in utero , early postnatal, lactational) in anim als can result in the developm ent of

cancer (reviewed in Toth, 1968; Della Porta a nd Terracini, 1969; Druc kery, 1973; Rice, 1979;

Vesselinovitch et al., 1979; Ri ce and W a rd, 1982; Vesselovitch et al., 1983; Anderson et al.,

2000). Thus, one elem ent in extending analyses to children is to evaluate the extent to which

exposures early in life would alte r the incidence of cancers observed later in life, compared with

the incidence observed with adult-only expos ures (Anderson et al., 2000; NRC, 1993).

The causes of cancer en com p ass a variety of possible risk factors, including genetic

predisposition (Tom linson et al., 1997), diet , lif estyle, associations with congenital

m a lfor m a tions (Bosland, 1996), and exposure to bi ological and physical agents and chem icals in

the environm ent. In som e cases, tumors in adults and children have been com p ared (Anderson et

al., 2000; Ginsberg et al., 2002). Children and ad ults generally develop the sam e spectrum of

tum o rs whe n they have inherited gene and chromosom a l mutations, such as Li-Fraum eni

syndrom e (Birch et al., 1998). W ith ionizing radiation, which operates through a m u tagenic

mode of action, both the young and the old deve lop m a ny of the sam e tumors, with the

difference being that children are m o re sus ceptible for a num ber of tum o r types (NRC, 1990;

U.S. EPA, 1994; UNSCEAR, 2000). Studies with anticancer drugs (cytotoxic and

immunosuppressive) demonstrate a si m ilar spectrum of tum o rs (Hal e et al., 1999; Kushner et al.,

1998; Larson et al., 1996; Nyandoto et al., 1998). Various viral infections, such as E p stein Barr

and hepatitis B, lead to lym phom a a nd liver cancer , respectively, in both age groups (Lindahl et

1

### al., 1974; Mahoney, 1999). These observations in hu m a ns indicate that the m ode of action for

these agents would be the sam e or sim ilar for adults and children.

Although there are sim i larities between childhood and adult tum o rs, significant

differences are also known to exist (Gruffe r m an, 1998; Israel, 1995). Tumors of childhood

generally consist m o re of e m bryonic cell tum o rs, while adults have m o re carcinom as.

Leukem i as, brain and other nervous system tum o rs, lym phomas (lym ph node cancers), bone

cancers, soft tissue sarco m as, kidney cancers, ey e cancers, an d adrenal gland cancers are th e

most common cancers of children, w h ile skin, pros tate, breast, lung, and co lorectal cancers are

the m o st common in adults (Ries et al., 1999; U. S. Cancer Statistics W o rking Group, 2002).

Som e tum o rs are unique to the young , including several with well es tablished genetic bases, such

as tum o rs of the kidney (W il m s ’ tumor) or eye (retinoblastom a) (Anderson et al., 2000; Israel,

1995).

The relative rarity in the in cidence of childhood cancers and a lack of anim al testing

guidelines w ith perinatal 1 

### exposure im pede a full assessm ent of children’s cancer risks f r om

exposure to chem icals in the environm ent. Un equivocal evidence of childhood cancer in hum ans

occurring from che m ical exposures is lim ited (A nderson et al., 2000). Established risk factors

for the developm ent of childhood cancer in clude radiation an d certain ph arm aceutical agents

used in chemotherapy (Reise, 1999). There is som e evidence in hum ans for adult tumors

resulting from perinatal exposure. P h arm acol ogical use of diethylstilbesterol (DES) during

pregnancy to prevent m i scarriages induced clea r cell adenocarcinom a of the vagina in a few

daughters exposed in utero though this tum o r was not observed in exposed m o thers (H atch et al.,

1998; Robboy et al., 19 84; Vessey, 1989). In ad d ition to the lim ited human data, there are

exam ples of transp lacen tal carcinoge ns in anim al studies, such as recent studies with nickel and

arsenic (Diwan et al., 1992; W aalkes et al., 2003 ), as well as studies suggesting that altered

developm ent can affect later su scep tibility 2 

### to cancer induced by expos ure to other chem icals

(Anderson et al., 2000; Birnbaum a nd Fenton, 2003).

Infrequently, perinatal exposure in anim als has been shown to induce tumors of different

types th an th ose observed with adult exposures. Studies with saccharin (Cohen et al., 1995;

Whysner and W illiam s , 1996; IARC, 1999) and as corbate (C ohen et al., 1998; Cohen et al.,

1995; NTP, 1983) found cancer when exposures were initiated in the peri natal period. In

1 Peri nat a l i s de fi ne d as t h e t i m e aro u n d bi rt h a n d m a y i n clude b o t h p r e n a t al (pri or t o bi r t h) a n d post n at al

(after birt h) pe rio d s.

2 Su scep tib ility is d e fi n e d h e re as an in creased lik elih oo d o f an ad v e rse effect, often d i scussed in term s o f

rel a t i ons hi p t o a fact o r t h at ca n be use d t o de scri be a h u m a n su bp o pul at i o n (e. g

. , l i f est a ge , dem o g r ap hi c feat ure

, or

g e n e tic ch aract eristic). Th e term s “su s cep tibilit y” an d “s ensitiv ity” are u s ed with a v a riety o f d e fi n itio ns in

p u b lish e d literatu re m a k i n g it essen tial th at read ers are aw are o f t h ese d i fferen ces in termin o l og y acro ss d o c u m en ts.



### 2

### contrast, studies subm itted to the Food and Drug Adm i nistration of approxim a tely a dozen other

food additiv es and colo rings that were not adult carcinog ens d i d not ind i cate cancer, ev en when

perinatal exposures occurred (U.S. EPA, 1996). W h en observed, the differences between

childhood and adult cancers suggest the im portan ce of evaluating the im pa cts of m a ternal

exposures during pregnancy as well as exposures to children (Anderson et al., 2000). The effects

of m a ternal exposures and tran splacental carcinogens require separate evaluation and are not

quantitatively evaluated in th e analysis presented below.

The lim ited hum a n inform ation described br iefly above is supported by a num b er of

anim al bioassays that include both perinatal and a dult exposures to chem icals. Standard anim al

bioassays generally begin dosing after the anim als are 6-8 weeks old, when m a ny organs and

system s are alm o st fully developed, though substa ntial growth in body size continues thereafter

(as m o re fully discussed in Hattis et al., 2005). The literature can be divid e d roughly in to three

types of exposure scenarios: those that include repeated exposures for the early postnatal to

juvenile period, as com p ared w ith chronic later-life dosing; lifetime (i.e., com b ined perinatal and

adult) exposure as com p ared w ith chronic later-life dosing; and those that include m o re acute

exposures, such as a single intrap eritoneal (ip) or subc utaneous injection, for both early-life and

later-life dosing. In the early-life exposure stud ies that are available, perinatal exposure usually

induces higher incidence of tum o rs later in lif e tha n the inc i de nce seen in s t andard b i oa ssays

where adult anim als only were exposed; som e ex am ples include diethyln itros am ine (DEN) (Peto

et al., 1984), benzidine (Vesseli novitch et al., 1979), DDT (Ve sselinovitch et al., 1979), and

polybrom inated biphenyls (PCBs) (Chhabra et al., 1993a). Reviews comparing early-life

carcinogenesis bioassays with standard bioassay s for a lim ited num b er of chem icals (McConnell,

1992; Miller et al., 2002; U.S. EPA, 1996) have concluded:

• The sam e tumor sites usually are observed fo llow i ng either perinatal or adult exposure.

• Perinatal exposure in conjuncti on with adult exposure usually increases the incidence of

tum o r bearing anim als or reduces the latent period before tumors are observed.

There is lim ited evid ence to inf o rm the m ode(s ) of action leading to differences in tum o r

type and tumor incidence following early-life exposure and exposure la ter in life. Differences in

the capacity to m e tabolize and clear chem icals at different ages can result in larger or sm aller

internal doses of the active agent(s), either in creasing or decreasing risk (Ginsberg et al., 2002;

Renwick, 1998). There is reason to surm ise that som e chem ic als with a mutagenic m o de of

action, which would be expected to cause irreversible changes to DNA, would exhibit a greater

effect in early-life versus later- life exposure. Several studies have shown increased susceptibility

3

### of weanling anim als to the for m ation of DNA a dducts following exposure to vinyl chloride (Laib

et al., 1989 ; Morinello et al., 2002a ; Morinello et al., 2002b). Add itionally, even thou gh not used

quantitatively in the an alyses in th is docum ent, a recen t analy s is of in vivo transp lacen tal

m i cronucleus assays indicated that fetal tissues generally are more sensitive than m a ternal tissues

for induction of m i cronuclei from mutagenic ch em icals (Hayashi et al., 2000), providing

qualitative support for the early-l ife susceptibility. Sim ilarl y, the neonatal mouse m odel for

carcinogenesis, which uses two doses prior to we aning follow e d by observation of tumors at one

year, shows carcinogenic responses f o r m u tagenic agents (Flammang et al ., 1997; McClain et al.,

2001). These results are consistent w ith the current understanding of biological processes

involved in carcinogenesis, which leads to a reas onable expectation that children can be m o re

susceptible to carcinogenic agents than adul ts (Anderson et al., 2000; Birnbaum and Fenton,

2003; Ginsberg, 2003; Miller et al ., 2002; Scheuplein et al., 2002) . Som e aspects potentially

leading to childhood susceptibility include the following issues.

• More frequent cell division during developm ent can result in enhanced fixation of

mutations d u e to the red u ced tim e availab l e for repair of DNA lesions and clonal

expansion of m u tant cells gives a larger population of m u tants (Slikker et al, 2004).

• So m e e m bryonic cells, such as brain cells, lack key DNA repair enzym e s.

• Som e com p onents of the imm une system ar e not fully functional during developm ent

(Holladay and Sm ialowicz, 2000; Holsapple et al., 2003).

• Hor m onal system s operate at different levels during different lifes tages (Anderson et al.,

2000).

• Induction of developm ental abnormalities can result in a predisposition to carcinogenic

effects later in life (Ande rson et al., 2000; Birnbaum and Fenton, 2003; Fenton and

Davis, 2002).

The m e thodology that has been generally used by the U.S. EPA to estim ate cancer risk

associated w ith oral expo sures relies on estim a tio n of the lifetim e average daily dos e, which can

account for differences between ad ults and children with respect to exposure factors such as

eating habits and body weight. However, suscep tibility differences with respect to early

lif estag e s ar e not tak e n into consid eration becaus e cancer slo p e factors 3 

### are based upon effects

3 Cancer slope factor – An u pper b oun d esti mate o f th e i n cr eased can cer ris k from a lifetime expos u re t o an

agent

. T h i s est i m at e, usual l y e x p r esse d i n uni t s of p r op ortio n (o f a p o p u lation ) a ffecte d pe r unit e x p o s u re (e. g

. ,

m g / kg-day or ug/ m 3 ), i s ge ne ral l y reser v ed f o r

use i n t h e l o w- do se r e gi on of t h e d o se- r es po nse rel a t i o ns hi p. It i s

o f

ten th e statistical u p p e r boun d on t h e p o t en cy and t h er efo r e th e r i sk. “U pp er b oun d ” in th is con t ex t is a p l au sib l e



### 4

### observed following exposures to adult hum ans or se xually m a ture anim als. Since a m u ch larger

database exists for chemicals i nducing cancer in adult hum ans or se xually m a ture animals, it is

necessary to determ ine whether adjustm e nt of s u ch adult-base d cancer slope factors would be

appropriate when assess ing cancer risks associated with exp o sures ea rly in life. The analys is

undertaken here addresses this i ssue, focusing upon studies that de fine the potential duration and

degree of increased susc eptibility that m a y arise from chil dhood, defined as early-life (typically

postnatal and juvenile anim al) exposures. Som e of these analyses, along with a m o re com p lete

description of the procedures us ed, have been published (Bart on et al., 2005). The analysis

presented in this Supplem ental Guidance and in the published article form the basis for

developing Supplem ental Guidance for evaluating can cer susceptibility associ ated with early-life

exposures.

u p p e r limit to th e tru e p r ob ab i lity.



### 5

### 2. PROCEDURES

This sec tion describ e s th e steps taken to asses s po tential su sce p tibility to e a rly- lif e

exposure to carcinogenic com pounds com p ared with adult and whole-life exposure. The readily

available literatu re was reviewed to identify anim al studies that comp ared tum o r incidence

between early-life and ad ult-only exposures or between ear ly -lif e-and-adu l t and adult-o n ly

exposures. S t udies were categor ized by length of exposure; th ose studies with quantitative

inform ation to estim a te tum o r incidence over time for early-life and adult exposures w e re

identified. These studies provi ded the basis for quantitativel y estim ating the dif f

e ren ce in

susceptibility between early-life and adult exposure s, as described below. Finally, summaries of

available hum a n data for radiation exposure were reviewed in the cont ext of tum o r incidence

from early-life versus later-in-life exposure.

2.1. DATA SOURCES FO R ANI MAL STUDIES

Studies in the literature included in this analysis are t hose that report tum o r response from

experim e nts that included both early-life and a dult exposure as separate experim e ntal groups.

Initial studies for consideration were identifie d through review articles and a search of the

National Toxicology Program (NTP ) database. Reviews of the li terature regarding cancer

susceptibility from early-life exposure in anim als include Mc Connell (1992), Ginsberg (2003),

Anderson et al. (2000), Miller et al. (2002) and U.S. EPA (1 996). A literature search was

conducted utilizing key words and MeSH headings (Medline) from studies identified in the

available reviews. The list of chem icals include d in this ana l ysis f o r qua ntitativ e eva l uation is

shown in Table 1a and 1b.

Abstracts or papers were re viewed to determ ine if a st udy provided information that

could be used for quantitative analysis. The criter ia used to decide if a study could be included

in the quan t itative ana l ys is were:

• Exposure groups at different post-natal ages in the sam e study or sam e laboratory, if not

concurrent (to control for a large number of potential cross-laboratory experim e ntal

variables including pathological exam inations),

• Sa m e strain/species (to e lim inate strain-specific responses confounding age-dependent

responses),

• Approxim a tely the sam e dose within the lim its of diets and drinking water intakes that

obviously can vary with age (to elim inat e dose-dependent responses confounding agedependent responses),

6

### • Sim ilar latency period f o llowing exposures of different ages (to control f o r confounding

latency period for tum o r expression with age-de pendent responses), arising from sacrifice

at >1 year for all groups exposed at differe nt ages, where early-lif e exposure can occur up

to about 7 w eeks. Variations of arou nd 10 to 20% in latency period are acceptab l e,

• Postnatal exposure for juvenile rats and m i ce at ages younger than the standard 6 to 8

week start for bioass ays ; prenatal ( in utero ) exposures are not part of the current analysis.

Studies that have postnatal e xposure were included (without adjustm e nt) ev en if they also

involved prenatal exposure,

• “Adult” rats and m i ce exposure beginning at appr oxim a tely 6 to 8 weeks old or older, i.e.

com p arable to the age at initiation of a st andard cancer bioassay (McConnell, 1992).

Studies with anim als only at young ages do not provide appropriate com p arisons to

evaluate age-dependency of response (e.g., the m a ny neonatal mouse cancer studies).

Studies in other species were used a supporting evidence, beca use they are relatively rare

and the determ ination of the approp riate com p aris on ages ac ro ss species is not sim p le,

and

• Num b er of affected animals and total num b er of anim als examined are available o r

reasonably reconstructed for control, young, a nd adult groups (i.e., studies reporting only

percent response or not includi ng a control group would be excluded unless a reasonable

estim ate of historical background for the strain was obtainable).

Tables 2 and 3 include infor m ation on the m e thods and results from the anim al studies

identified in Table 1b. P e rtinent inform a tion on species, sex, dosing regim e n, and tumor

incidence is given. Additionall y, the “Notes” colum n includes general inf o rm ation about the

relationship between tumor inciden ce, anim al age at first dosing, and sex. The data in Tables 2

and 3 were used for the calcu la tions, described below, for esti m a ting pote n tia lly in cre a sed canc e r

risk from early-life exposure.

The available literature includes a wide range of exposure scenarios. This range is due in

part to the lack of a defined pr otocol for early-life testing and th e difficulty of standardizing and

adm i nistering doses preweaning. As noted previ ously, the literature can be divided roughly into

three types of exposure scenarios: those that include repeated expos ures f o r the early postnatal to

juvenile period, as com p ared with chronic later-life dos ing; lifetim e (i.e., com b ined perinatal and

adult) exposure as com p ared with chronic later-life dosing; and those that include m o re acute

exposures, such as a single intrap eritoneal (ip) or subc utaneous injection, for both early-life and

later-life dosing. Table 2 in cludes the studies that had early post natal to juvenile exposures, adult

chronic exposures, and lifetim e exposures. Tabl e 3 includes studies with acute exposures. A

discussion of the im plications of the different ex posure scen arios is in clu d ed in Section 3.

7

### Studies were identified for m o re than 50 ch em icals not included in Tables 2 and 3 that

dem onstrated carcinogenesis follow i ng perinata l exposure, but did not directly com p are

exposures at different ages. A large num b er of studies address in utero exposures only. More

than 100 chem icals (with both negative and positiv e findings) have been studied in the neonatal

mouse assay, but this assay doe s not have a com p arable adul t exposure (F lammang et al., 1997;

McClain et al., 2001; Fu jii, 1991 ). S t udies acros s la boratories often varied in their us e of anim al

strains (e.g., for AZT studies, Diwan et al ., 1999 used CD-1 m i ce, while NTP, 1999 used

B6C3F 1 

### m i c e ). Studies of tam oxi fen use two Wist a r-de r ived stra ins a nd had very dif f

erent

periods for tum o r expression, i.e., sacrifice at 20 months for adult- exposed rats and natural death

up to 35 m o nths for juvenile-exposed rats, with uterine tum o rs observed in anim als dying after

22 m onths (Carthew et al., 2000; Carthew et al., 1996; Carthew et al., 1995). Due to these

factors, the chem icals that bel ong to this group w e re not evalua ted quantitatively. In addition,

there were studies ass e ssing radi ation in anim als (Covelli et al., 1984; Di et al., 1990 ; Sasaki et

al., 1978). The radiation data were not analyzed in depth, in pa rt because there are recognized

differences in toxicokinetics and toxicodynam i cs between radiation and chem icals with a

mutagenic mode of action for carcinogenesis. Even though the data on A-bom b survivors

provide inform ation for m a ny different cancer si tes in hum ans with a single exposure involving

all ages, a num b er of national and internati onal comm ittees of expert s have analyzed and

modeled these data to develop risk estim ates for various specif i c applicatio ns. Furtherm ore, lack

of unifor m ity regarding radiation doses, gestational age at exposure, and the anim al strains used

m a ke it diffi cult to m a ke com p arisons across studies (Pre ston et al., 2000).

2.2. EVALUATI N G THE MODE OF ACTION OF CARCINOGENS

Evaluation of the m ode of action of a car cinogen was based upon a weight-of-evidence

approach. Multip le m odes of action are asso cia t ed with the chem icals in this da tabas e , but a

num b er are associated w ith m u tagenicity (i.e ., benzo(a)py ren e , benzid ine, dibenzan thracene,

diethylnitrosam i ne, di m e thylbenz(a)anthracen e, dim e thylnitrosam ine, ethylnitrosourea, 3-

m e thylcholanthrene, m e thylnitrosour ea, safrole, urethane, and vinyl chloride). Determination of

carcinogens that are operating by a m u tagenic m o de of action entails evaluation of short-term

testing results for genetic endpoints, m e tabo lic profiles, physicochem i cal properties, and

structure-activity relationship (S AR) analyses in a weight-of-evide nce approach (Dearfield et al.,

1991; U.S. EPA, 1986, 1991; W a ters et al., 1999), as has been done for several chemicals (e.g.,

Dearfield et al., 1999; Mc Carroll et al., 2002; U.S. EPA, 2000a) . Key data for a m u tagenic m ode

of action m a y be evidence that the carcinogen or a m e tabolite is DNA r eactive and/or has the

ability to bind to DNA. Also, such carcinogens usually produce positive effects in m u l tiple test

8

### system s for different genetic endpoints, particular ly gene mutations and structural chrom o som e

aberrations, and in tests perform e d in vivo which generally are supported by positiv e tests in

vitro . Additionally, carcinogens m a y be identified as operating via a m u tagenic m ode of action

if they have sim ilar properties and S A R to established m u tagenic m ode of action.

2.3. QUANTITATIVE M E THODS

To estim ate the poten tial difference in sus ceptibility between early -life and adult

exposure, w e calculated the estim ated ratio of the cancer potency from e a rly-life exposure

com p ared to the estim ated cancer potency fr om a dult exposure. The cancer potency w a s

estim ated from a one-hit m odel, or a restricted form of the Weibull m odel, which is commonly

used to es tim a te cum u l a tive incid e nce for tum o r onset. The general form of the equation is:

P(dose) = 1-[1-P(0)]exp(-cancer potency*dose)

The ratio of juvenile to a dult can cer potencies were ca lculate d by f itting this m odel to the

data for each age group. The m odel fit depe nded upon the design of the experim e nt that

generated the data. Two designs should be handl ed separ a te ly: exper i m e nts in which anim als are

exposed eith er as juv e niles or as adu l ts (with eith er a sing le o r m u ltiple d o se in each period), and

experim e nts in which exposure begins either in the juvenile or in the adult period, but once

begun, continues through life.

For the firs t case, th e m o del equation s are:

(1) 

# ( )

( ) 

# ( ) 0 0

e

0 0 

### 1 ( 1 e )

1 1 e A A

A J m

A

m

J 

### P P P

P P P λ δ

δ −

− 

### = +

− −

= +

− −



### where:

subscrip ts A and J refer to the adult and j uvenile period, respectively,

λ is th e natu ral logar ith m of the juvenile:adult cancer potency ratio,

P 0 

### is the f r a c tion of control an im als with the particular tum o r type being modeled,

P

x 

### is the fraction of animals exposed in age period x with the tum o r,

m

A 

### is the rate of accum u lation of “hits” pe r unit o f tim e for adults, i. e., the cancer

potency, and

δ

x 

### is the duration or number of exposures during age period x

.

For a substantial num ber of da ta sets (acute exposures), δ J 

### = δ A 

### = 1. W e are in tere sted in

9

### determ ining λ

, which is the logar ithm of the estim ated r a tio of juvenile to adult can cer potencies,

a m easure of potential suscepti bility f o r early-life exposure.

For the second kind of design, the model e quations should take into account that

exposures that were initiated in the juvenile period continue th rough the adult peri od. The m odel

equations for the fraction of anim als exposed only as adults with tum o rs in this design are the

sam e as in the first design, but the f r action of anim als whose first exposure occurred in the

juvenile period is:



# ( ) ( ) 

# ( )

e

0 0 

### 1 1 e A J A A A m m

J 

### P P P λ δ δ δ

− − − 

### = + +

− 

### (2)

All sym bols in (eq. 2) have the sam e interpre tation as their counterparts in (eq. 1), but

now δ J 

### includes the duration of exposure during the juvenile period as well as the subsequent

adult period.

Param e ters in these m odels were estim ated using Bayesian methods (see, for exam ple,

Carlin and L ouis, 2000), and all inferences about the ratios were based on the m a rginal posterior

distribution of λ . Som e of these analyses, including a m o re com p lete description of the

procedures (including the potenti al effect of alternative Baye sian priors that have been

exam ined) have been pu blished (Barton et al., 20 05). The data for estim ating each ratio were in

the form of num b ers of anim als tested and number affected for each of control, juven ile-exposed,

and adult-ex posed anim als, and duration of expos ure for each of the juvenile-exposed and adultexposed groups. A few data sets had separate control groups fo r the juvenile-exposed and adultexposed gro ups, and equ a tions 1 and 2 were m odi fied accord ingly. The likelihood for the

param e ters in the m odel was the product of th ree (or four, if there were two control groups)

binom ial probabilities: f o r the num ber of anim als with tum o rs in the control group (s), f o r the

juvenile-exposed group, and for the adult-exposed group. The prior for P 0 

### (the fraction of

control anim als with a particular tumor) was right triangular (right angle at the origin), based on

the assum p tion that control incidences should be re latively low. (The base of the distribution is

one, as P 0 

### can not exceed one. As this is a prob ab ility distribu tion, th e area of the trian g le is on e.

Therefore, its height at the orig in m u st be 2.) The effect of exposure in adults is quantified by

the extra ris k, Q, where the probab i lity that an an im al has a tu mor is P 0 

### + ( 1 – P 0 

### )Q

. 

### So, from

equations 1, 1 e A A m 

### Q δ

−



### = − , Q was given a uniform prior on th e inte rval (0, 1 ), ref l ec tin g tota l

ignorance about the extra risk of ad ult exposure. Finally, the prior for λ w a s Gaussian with m ean

0 (corresponding to a m e dian or geom etric m ean ratio of one) and standard deviation 3. The

prior f o r th e log ra tio of juvenile to adult cancer potency ha s som e influe nce over the posterior

estim ates for the ratio of juvenile to adult po tency. The m a gnitude of th at influence depends on

10

### the am ount of support in the data for different valu es of the log ratio. The prior also ef fectively

downweights extrem ely large or sm a ll va lues f o r the juven ile to a dult potency ratio. T h ree priors

for the standard deviation were evaluated (Barto n et al., 2005, see Appendix) , with the intent of

finding the largest prior, i.e., one that would contain the leas t inf o rm ative assum p tion f o r the

prior. A standard deviation of 9 was tried, but som e of the intervals would not converge. A

standard deviation of 3 worked well, allowed ratio es tim ates to be derived, with all of the data of

inte rest. An inte rm ediate value of 6 was also exa m ined to ascerta in if a le ss inf o rm ative prior

could be used. W h ile the intervals converged, a sens itivity an alysis showed that this v a lue f o r the

standard deviation resulted in sufficient down-we ighting of the ratios with lim ited inform ation

that these data would not influe nce the result. This was considered an unreasonable bias, so a

standard deviation of 3 was used for the further analyses. A further discussion of these analyses

can be found in Barton et al. (2005).

The posterior distribution for the unknown param e ters in these m odels is the product of

the likelihood from the data and the prio rs (the “unnorm a lized” prior), divided by a

norm a lization constant that is th e integral of the unnorm a lized prior over the ranges of all the

param e ters. This norm a lization constant was co mputed using num e rical integration, as were

posterior m eans and variances and m a rginal posterior quantiles for the log-ratio λ . A ll num erical

com putations were carried out in the R sta tistical programm i ng la nguage (version 1.8.1; R

Developm ent Core Team, 2003).

This m e thod produced a posterior m ean rati o of the early-life to adult cancer potency,

which is an estim ate of the potential susceptibi lity of early-life exposure to carcinogens. If the

ratio was greater than one, this indicated th at the experim e nt found that there was greater

susceptibility from early-life expos ure. If the ratio was less than one, this indicated that the

experim e nt found that there was less susceptibilit y from early-life exposure. Summ aries of the

individu al ratios from each of the dose groups from the different exp e rim e nts for different

groupings w e re also calculated (for exam ple for a ll acute exposures of chem icals that are

carcinogenic by a m u tagenic m ode of action). T h e summ ary ratios were constructed from the

individu al ratios with in a group, by varian ce-weigh ting th e means of each ratio. The individual,

posterior m eans were weighted by using reciprocals of their p o sterior va rian ce. This weighting

procedure is commonly used becaus e it g i ves greater weight to those s t u d ies for which the

varian ces, i. e., the uncertain ties, are sm aller. Because th e ratios were calculated as lo g ratios (se e

eq. 1), exponentiating the resul ting inverse-variance-weighted m ean yielded inverse-varianceweighted geom etric m eans of ratios.

11

### 2.4. IONI Z I NG RADI ATION

A supporting role was assigned to the avai lable hum a n radiation data, where cancer

incidence in adults who were children at th e tim e of the atom ic bom b (A-bom b) exposure was

com p ared with cancer incidence in adults who were older at the tim e of e xposure. Although there

are recognized differences in toxicokinetics a nd toxicodynamics between radiation and chem ical

carcinogens with a m u tagenic m ode of action, the data on A-bom b survivors provide infor m ation

for m a ny different cancer sites in hum a ns with a singl e exposure involving a ll ages. In addition

to the richness of the data, a num ber of nationa l a nd inte rnatio nal comm ittees of experts have

analyzed an d m odeled these data to develop risk estim ates for various specific applications.

The repor t o f the United Nations Scie n tif ic Committee on the Ef f

ects of Atom ic

Radiation (UNSCEAR, 2000, with Scie ntific Annexes) lists m o re th an 80 studies, in addition to

the reports of the Japanese A-bom b s u rvivors, in which at least one typ e o f cancer was m easured

in hum ans who were exposed either intentionall y or accidentally to som e form of ionizing

radiation. H o wever only the A-bom b survivor repo rts have relevant inform ation on incidence of

early -life exposures. On e of the m o r e recen t pap e rs cited in the UNSCEAR report, b y Thom pson

et al. (1994), contains detailed data on the incidence of 21 di ffe rent cancers in 37,270 exposed Abom b survivors (42,702 unexposed). Also, EPA has used data from the A-bom b survivors to

develop age-specific relative risk coefficients using various m e thods for transporting the risk

from the Japanese population to the U.S. populati on (U.S. EPA, 1994). It is beyond the scope of

this ef f

o rt to present all of th e radiation data or a discussion of the various analyses and m odeling

efforts. Rather, inform ation relevant to com p aring cancer risks from juvenile versus adult

exposure from UNSCEAR (2000) an d U.S. EPA (1994; 1999) is presented as representative

findings to determ ine whether the radiation data are sim ilar qu alitatively to the chem ical

findings. More detailed data on the A-bom b survivors can be found in Delongcham p et al.

(1997) and Preston et al. (2000).

As previously noted, several st udies have assessed radiation in anim al studies (Covelli et

al., 1984; Di et al., 1990; Sasaki et al., 1978). However, lack of unifor m ity regarding radiation

doses, gestational age at exposure, and the anim al stra ins used m a ke it difficult to com p are the

experim e ntal data on cancer i nduction after prenat al irradiation (Preston et al., 2000).

12

### 3. RESULTS

3.1. QUALITATIVE EVALUA TION OF THE DATABASE

The question addressed in this analysis wa s whether, and how, available quantitative

scien tif ic da ta could inf o rm risk asse ssm ent po licy choices for adjusting cancer slope factors

when they are used in the assessm ent of cancer risk from childhood exposure. Cancer slope

factors are, with few exceptions, based on adult hum a n epidem iology or standard chronic adult

rodent bioassays, which do not address the im pact s of early-life exposures . Thus, the critical

data are either hum an epidem iological data on childhood exposures resulting in adult cancer or

research stu d ies with ro dents involving early postna tal exposures. The m a jor hum a n data

available are from radiation exposures (studies summarized in Tables 9- 11), with very lim ited

data available for hum a ns exposed during childh ood to chemicals (reviewed in Anderson et al.,

2000; Miller et al., 2002).

A review of the litera ture identified several hundred re ferences reporting more than 50

chem icals that have been shown to be able to cause cancer following pe rinatal exposure (Table

1a) (reviewed in Toth, 1968; Della P o rta a nd Terracini, 1969; Druckery, 1973, Rice, 1979;

Vesselinovitch et al., 1979; Rice and W a rd, 1982; Vesselovitch et al.; 1983; Fujii, 1991;

Anderson et al., 2000). S t udies (or groups of studies from a single laboratory on a given

chem ical) th at directly provide d quantitative data on carcinog enesis follo w ing early p o stnatal

exposures and adult exposures to chem icals in anim als were identified for 18 chem icals, listed in

Table 1b, 2, and 3. Of the iden tified studies, there were 11 chem icals involving repeated

exposures during early postnatal and adult lifes tages (Table 1b) and 8 chem icals using acute

exposures (typically single doses) at different ag es (Table 1b). Som e of the studies evaluated

single tiss u es or organs f o r tum o rs (e.g., only liv e r), while o t h e rs evaluated m u ltiple tissues and

organs (Tab les 2 and 3). Mice, r a ts, o rboth spec ie s and som e tim es multiple stra ins wer e tes t ed.

These stud ies serve as th e basis for the quantit ative analys es presented later in the res u lts.

In addition to the studies identified in Table 1b, studies were identified with early

postnatal and early-life e xposures that were evaluated qualitat ively but not quantitatively. Som e

of these studies are notable a nd provide im portant supporting info r m ation. Two recent studies

used tran sge n ic m ouse models f o r hu m a n tum o rs . Incre a sed multiplicity of colon tumors was

observed following earlier versus later azoxym ethane exposures (Pau lsen et al., 2003).

Shortened m a mmary tumor latency following es tradiol exposure occu rred when exposures

occurred between 8 and 18 weeks as opposed to earl ier or later, which is generally consistent

with the incidence resu lts analyzed for DMBA (Yang et al., 2003). Several notable exam ples

exist of developm ental windows l eading to cancer susceptibilities that were not observable in

13

### adults. Several potent estrogenic chem icals including DES, tam oxifen, a nd genistein produce

uterine tum o rs with early postnatal exposures of m i ce, though there also appear to be straindependent differences in the tum o r sites in a dult m i ce (Gass et al., 1964 ; Greenm a n e t al., 1990;

Newbold et al., 1990, 1997, 1998, 2001). Developm enta l susceptibilit ies are believed to play a

key role in effects observed with saccharin (Cohen et al., 19 95; W hysner and W illiam s , 1996)

and ascorbate (Cohen et al., 1998; N T P, 1983), w ith bladder tum o rs arising when early-life

exposures occurred. Studies with several species, incl uding rat, m ouse, and opossum

, indicate

that nervous system s tumors associated with exposures to E NU and several other chem icals

appear to be highly dependent upon exposures occurring within certain windows, particularly

prenatal ones (Rice, 1979; Rice a nd Ward, 1982; Jurgelski et al., 1979).

Analyses of the difference in cancer risk fr om exposures during different lifetim e periods

ideally should address both the period of pot entia l suscep tib ility and the m a gnitude of the

susceptibility. Available studies used a variety of study designs (see Tables 2 and 3), which can

be valuab le because they provide d i fferent inform ation (Figure 1). However, variations in study

design can result in a lack of co m p arability acros s chem icals, and can limit inform atio n on the

consistency of effects with di fferent chem icals acting through different modes of action. The

acute dos ing (larg e ly s i n g le dose) stu d ies (Tab le 3) are v a luable becau se they invo lve identical

exposures w ith explicitly defined doses and tim e periods dem onstrating that differential tum o r

incidences arise exclusively from age

- dependent susceptibility. These studies address both the

period and m a gnitude of susceptibility. They w e re not as appropr iate for quantitative

adjustm e nts for the cancer poten cy estim ates beca use of their lim itations, including that m o st

used subcutaneous or ip injec tion that historically have not been considered quantitatively

relevant routes of environm ental exposure fo r hum a n cancer risk assessm ent by EPA, and that

these routes of exposure are expected to have only partial or a com p lete absence of first pass

m e tabolism that is likely to af f

ect potency estim a tes.

The repeated dosing studies with exposures during early postnatal or adult lifetim e

provide usef ul inform ation on the relative im pact of repeated exposures at different lif estages

and m a y be more likely to have exposure occur du ring a window of susceptibility, if there is one.

One notable difference in study designs was that studies with repeated early postnatal exposure

were includ ed in the analysis ev en if they al so inv o lved earlier m a ternal an d/or pren atal

exposure, w h ile studies addressi ng only prenatal exposure were not otherwise a part of this

analysis. Another notable differe nce am ong studies involved the ti ssues that were evaluated for

tum o rs: som e studies focused on a single tissue, particularly liver, wh ile others evaluated

multiple tis s u es.

Com p arisons within a single repeated dosi ng study m a y have lim itations f o r evaluating

14

### differential s u sceptibility because exp o sures to the chem ical can differ during the different

lifestages, particularly w h en dietary or drin king water exposures are involved. A notable

exam ple is the PCB study (Chhabra et al., 1993 a), in which mobilization of such lipid-soluble

chem icals in to m o ther’s m ilk would be expected to resu lt in inf a nts rece iving m u ch larger

exposures than other lifestages. W h ile lactational transf er is just as relevant to hum an nursing

offspring, this difference in exposure obscures the extent to which the early lifestage is

quantitatively m o re susceptible (i.e ., part of the increas ed early-lif e cancer risk arises from higher

exposure than during the adult period). Maternal m e tabo lism of com pounds such as

diphenylhydantoin (DPH) (Chhabr a et al., 1993b) also m a y resu lt in lower exposure during

lactation, potentially underestim a ti ng the early-lif estage risk, if the parent com pound i s the active

for m of the chem ical. Sim ilar issues exist due to norm a l age-dependent changes in food and

water consumption. Ascribing differential effects observed in anim al studies solely to lifestage

susceptibility m u st be done carefully as there ma y also be differences in the exposures. There

are substantial and clear benefits , therefore, from experim e ntal consistency when comparisons

are m a de directly within a study (e.g., sam e species and strain, cons istent pathological

evaluation).

One issue to note is the r a tiona le f o r the organization of the ava ilable data. It was

observed that the results across a broad range of chem icals with a vari ety of m odes of action

were som e what variable. Theref ore, consistent w ith th e appro ach of the EPA cancer g u idelines

(U.S. EPA, 2005), an approach based on m ode of action appear ed to be a common fra m e w ork

for analysis. Variability in lifes tage-dependent susceptibility and susceptibility across a range of

modes of action was further s upported by theoretical analyses using m u ltis tage and two-stage

models of carcinogenesis (Goddard and Krewski, 1995; Murdoch et al., 1992).

3.2. QUANTITATI VE EVALUA TION OF THE DATABASE

As described in the Section 2.3, the potential difference in suscepti bility between earlylife and adult exposure w a s calcula ted as the estim a ted ratio of cancer potency from e a rly-life

exposure over the cancer potency from adult expo sure. Tables 4-7 present the results of the

quantitative analysis using the st ud ie s that wer e d e term ined quali tatively to have appropriate

study designs (Tables 2 and 3) cont aining sufficient inform ation to analyze. Based on the studies

available, the calculations were organized into four tables : (1) com pounds acting through a

prim arily mutagenic m o de of action, where th e com pound was adm i nistered by a chronic dosing

regim e n to adults and repeated dosing in the early postnatal period (T able 4); (2) compounds

acting through a prim arily nonm utagenic m ode of action, where the compound was adm i nistered

by a chronic dosing regim e n to adults and repeated dosing in the early postnatal period (Table 5);

15

### (3) com pounds acting through a prim arily m u tage nic m ode of action, where the compounds we re

adm i nistered by an acute dosing regim e n (Table 6); and (4) com pounds acting prim arily through

either a m u tagenic or nonm utagenic mode of ac tion with chronic adult dos ing and repeated early

postnatal dosing (Table 7). In these tables, the 2.5% and 97.5% ar e percentiles of the posterior

distribution. For a Bayesian di stribution, these percentiles func tion in a m a nner sim ilar to the

95% confidence lim its for other type s of statis tical analyses. T h e results are discussed below,

followed by a description of results f r om analyses of studies of hum a ns e xposed to radiation.

3.2.1. Carcinogens w i th a Mutagenic Mode of Action

The m o st inform ative database on early-lifest age susceptibility exists for chem icals with

a well-accep ted m u tagenic m ode of a c tion (e.g., d i ethy lnitrosam i ne, vinyl chloride). T h is

database includes both single-dose studies and repeated-dose studies involving periods of

postnatal and/or chronic exposure. T h ese st udies help define the periods of increased

vulnerability and the m a gnitude of the susceptib ility. The acute dosing studies dem onstrate that

the age-dependent responses are not due to di fferences in exp o sure, becau s e these stud ies

explic itly co ntrol the exp o sure.

3.2.1.1. Early Postnata l, Juvenile, a n d Adult Repeated Dosing Studies o f

Chemica l s with a

Mutagenic Mode of Action

Studies com p aring repeated dosing f o r early-life, adult, or lif etim e exposures exist for six

carcinogens with a m u tagenic m ode of acti on [benzidine, diethylnitrosam ine (DEN), 3-

m e thylcholanthrene, safrole, uretha ne, and vinyl chloride]; DEN also had acute dosing studies.

Lifetim e (i.e., com b ined juvenile and adult) com p ared to adult exposure studies were analyzed

for DEN, sa frole, and urethane, while studies comparing juvenile with adult exposures were

analyzed for benzidine, 3-m e thylcholanthrene, sa frole, and vinyl chloride. These chem icals all

requir e m e tabolic a c tiv ation to th e ac tive c a rc ino g enic f o rm . Analysis of the tum o rs arising pe r

unit tim e of exposure found that ju venile exposures with each chem ical could be m o re effective

than adult exposures were at i nducing tum o rs (Tables 4 and 7; Fi gure 2, a graphic representation

of the posterior, unweighted geom etric m eans and their 95% confidence intervals, for the ratios

of juvenile to adult cancer potency for carci nogens acting through a m u ta genic m ode of action).

The weighted geom etric m ean for repeat and lif etim e exposures is 10.4; for acute exposures the

weighted geom etric m ean value is 1.5. For be nzidine and saf r ole, there w a s a notable sex

difference, with high liver tum o r incidence obser ved for early postnatal exposures of m a le, but

not f e m a le, m i ce. For both the a c ute and the r e p eated /lif e time data, the 9 5 th 

### percentile of the

individual, unweighted geom etri c m e ans is above 10 (Figure 2).

16

### This analysis focused upon the duration of e xposure as a surrogate for dose, essentially

assum i ng that the do ses anim als received during the different periods of th ese stud ies were

sim ilar. This assum p tion is a lim itation of the an alysis b ecau se these stud ies invo lved exposures

via lactation (i.e., dosing the m o ther prior to we aning), drinking water, di et, or inhalation, which

have the potential to deliver diffe rent doses at different lifestag es. However, the range of the

m a gnitudes of the tum o r incidence ratios of juvenile to adult exposures is sim ilar (Table 8) for

the repeated dosing studies (0.12 – 111, weight ed geom etric m ean 10.5, 42% of ratios greater

than 1), lifetim e dosing studies (0.18 – 79, weighted geom etric m ean 8.7, 67% of ratios greater

than 1), and acute dosing studies (0.01 – 178, weighted geometric m e an 1.5, 55% of ratios

greater than 1), suggesting that these differences in dosing are not the sole determ inant of the

increased in cidence of early tum o rs, i.e., uncer tainty and variability rem a in. Because these

com p arisons include d i fferent chem icals with diffe rent tissue specif i c ities, it m a y be inf o rm ative

to consid er liver as a target organ affected by all of these chem icals. The range of the

m a gnitudes of the liver tum o r incidence ratios of juvenile to adult expo su res is sim ilar f

o r the

repeated dosing studies (0.12 – 111, weighted geom etric m ean 41.8, 86% of ratios greater than 1,

Table 4), lifetim e dosing studies (0.47 – 79, weighted geometric m ean 14.9, 80% of ratios greater

than 1, Table 7), and acute dosing studies (0.1 – 40, weighted geom etric mean 8.1, 77% of ratios

greater than 1, Table 8). Thus, the repeated dose studies support th e conc ept tha t ea rly-lif es tage

exposure to carcinogenic chem icals with a m u tageni c m ode of action would lead to an increased

tum o r incidence com p ared with adult e xposures of a sim ilar duration and dose.

3.2.1.2. Acute Dosing Studies of C h emicals with a Mutagenic Mode of Action

Acute dosing studies are availa ble for eight carcinogens with a m u tagenic mode of action

that were ad m i nistered to m i ce or rats [b enzo[a]pyrene (BaP), dibenzan thracene (DBA),

Diethylnitrosam ine (DEN), dim e thylbenzanthra cene (DMBA), dim e thylnitrosam ine (DMN),

e t hyl ni t r os our e a ( E NU) , me t hyl nitrosourea (NMU), and urethane (also known as ethyl

carbam a te)] (Table 1b). Except for ENU and NM U, these compounds require m e tabolic

activation to their active carcinogenic for m s. The s e acute dosing studies generally compared a

single exposure during the first fe w weeks of life with the identical or sim ilar exposure in young

adult anim als (Tables 3 and 6). Many of these studies com p ared exposures during the

preweaning period (i.e., approxim a tely day 21 for rats and m i ce) with effects around w e ek 6,

which is app r oxim a tely the age at wh ich typ i cal chronic bioassays begin dosing anim als. These

studies largely were by subcutaneous or ip injection, which histori cally have not been considered

quantitatively relevant routes of environm enta l exposure for hum a n cancer risk assessm e nt by

EPA. For purposes of com p aring age-dependent susceptibilities to tum o r developm ent, thes e

17

### data are highly relevant. The in jection route typically alters th e pharm acokinetic time courses of

the parent com pound and the m e tabolites com p ared with oral or other ex posures due to altered

kinetics of absorption and m e tabolism. Howeve r, for these com pounds and the system ic organ

effects observed, there are several ph arm acokineti c reasons to believe that the age-dependent

trends would be sim ilar with ot her routes of exposure. Thes e com pounds are expected to be

reasonab l y well absorb ed orally, co mparable with injection routes, and la rgely require m e tabolic

activation, so partial or com p lete absence of fi rst pass m e tabolism in the injection studies would

be sim ilar to or underestim ate m e tabolic activ ation when compared with oral exposure.

The early exposures often resu lted in higher incidence of tumors than later exposures,

with increased early su sceptibilities up to 178-fo ld (unweighted ratios in Table 6 range from

0.011 to 178, with a weighted geometric m ean of 1.5, and 55% of ratios greater than 1, Figure 2,

Table 8). E x am ples of t h e general age-dependent decline in susceptibi lity of tum o r response

include BaP (liver tum o rs), DEN (liver tum o rs), ENU (liver and nervous system tum o rs), and

urethane (liver and lung tum o rs). W h ile genera lly the Day 1 and Day 15 tim e points were higher

than la ter tim e points, in severa l cas es sim ila r tu mor inciden ce was obser ved at both these early

tim es (e.g., ENU-induced kidney tumors, Tables 6 and 8).

W h ile the de gree of susc eptib ility ge nerall y declines during the early postnatal period

through puberty into early adult hood, there are exceptions due pe rhaps to pubertal periods of

tissue developm ent (e.g., m a mm ary tissues) or very early developm ent of xenobiotic

m e tabolizin g enzym e s. One such exception was th e increas ed inciden ce of m a mmary tum o rs in

5-8 week old rats given DMBA, compared with older or younger rats (Meranze et al., 1969;

Russo et al., 1979). Meranze et al. (1969) re ported 8% m a mmary tum o rs following a single dose

of DMBA at less than tw o weeks, 56% if give n once to anim a l s between 5 and 8 weeks old, and

15% when given once to 26 week old rats. T hus, a ratio of 7.1 is obtained when com p aring

susceptibilities of 5–8 week and 26-week-old rats (Table 6) compared to a ratio of 0.2 when

com p aring the exposure at 2 weeks versus 26 weeks. A sim ilar effect was observed by Russo et

al. (1979); see Table 3. This observation corre sponds well with pubertal developm ent of the

m a mmary tissue, with ovarian function commenci ng between 3 and 4 weeks (after the < 2 week

tim e point in the Meranze et al., 1969 study) , and m a mm ary ductal growth and branching

occurring such that it is approxi m a tely two-thirds com p lete by week 5, consistent with the 5–8

week susceptible period of Meran ze et al. (Silberstein, 2001 ). W h ile this diffe rs from the general

trend pr evio usly discu s s e d, it indica tes suscep tib i lity later in the juvenile period r a the rthan

earlier. Another exam pl e of deviation from the ge neral trend toward an age-dependent decline is

DEN-induced lung tum o rs that were som e what lower in incidence following exposure on day 1

than observed for the day 15 or day 42 exposures (Vesselinovitch et al., 1975) (Tables 3 and 6).

18

### There are su bstantial d i fferences in th e early-life susceptibility of different tissues observed in

the acu te stu d ies (Tab le 8). It s hould be noted that th e targ et tissues vary with chem ical, so th e

num b er of chem icals for which data are availab l e varies for each tissue. Several tissues have

weighted geom etric m ean ratios of greater than 1 including kidney, leukem i a, liver, lymph,

ma mmary, n e rve, reticular tissue, thym ic lym phom a , and uterus/vagin a. Som e of these, such as

the nerve and m a mm ary tum o rs, appear to have a very specific window of susceptibility, as

noted above, and the ratios were m u c h higher if the exposure occurred during this w i ndow.

Tissues with weighted mean ratio s less than 1 include forestom ac h, harderian gland, ovaries, and

thyroid. Lung has a weighted geometric m ean of 1. Many of the studies produced very high

lung tum o r responses regardless of age, so the re sults are difficult to inte rpret, as illustrated by

the dose-response data with urethane in Rogers ( 1951) in w h ich the increased early susceptibility

is only apparent when the dose is low. The large num bers of studies with high lung tum o r

responses at all ages contribute to the differences in the weight ed geom etric m e ans for the acute

and for the repeated dosing studies.

Overall, the acute dosing studies support th e concept that early- lifestage exposure to

carcinogenic chem icals with a m u tagenic m ode of ac tion would lead to an increased incidence of

tum o rs compared with adult exposures of a si m ilar dose and duration. These studies generally

use the sam e dose and duration at all ages, and thus do not have th e type of issues discussed for

the repeated dosing studies. On the other ha nd, the acute dosing studies have lim itations that

were sufficient to decide that th ey should not be included in the quantitative adjustm e nt of cancer

potency. First, as m e ntioned in the previous paragraph, the large num ber of studies of lung

tum o rs with alm o st 100% response observed at all doses and all ages would significantly bias the

m e dian ratio toward unity for a reason ba sed on study design rather than biology. Second,

cancer potency estim a tes are usually derived from chronic exposures. T h erefore, any adjustm e nt

to those potencies should be, if possible, from si milar exposures. Third, m o st exposures of

concern to the Agency are from repeated or ch ronic exposures rather th an acute exposures.

Finally, m a ny of the acute studies used ip exposures , which is not the usual route of exposure for

environm ental chem icals

. Thus, the repeated a nd lif etim e stud ies ar e m o re appropriate for the

purpose of this analysis.

3.2.2. Carcinogens With Modes of Ac tion Oth er Than Mutagenicity

Studies com p aring tum o rs observed at the sam e sites following early postnatal and

chronic adult exposures in a single protocol we re available for six ch em icals that do not act

through a m u tagenic m o de of action [a m itrole, di chlorodiphenyltrichloroethane (DDT), dieldrin,

ethylene thiourea (ETU), diphenyl hydantoin (DPH), polybrom inated biphenyls (PBB)] (Table 5).

19

### These chem icals cause tu mors throug h several di f f

erent, not n ecessarily well defined, modes of

action. For exam ple, thyroid horm one disruption by ETU causes thyroid tumors; som e PBBs act

through ary l hydrocarbo n (Ah) recep tors, while o t hers are phenobarbital-like pleio t rop h ic

inducers of liver enzym e s and liver tum o rs. Thr ee of these studies evaluated only m ouse liver

tum o rs (am i trole, DDT, dieldrin), w h ile the othe r three evalu a ted a large num b er of tissues in

both m i ce and rats (ETU, DPH, PBB). These st udies generally included a com b ined perinatal

and adult exposure as w e ll as the separate perinata l or adult-only groups. It should be noted that

no acute perinatal dosing studies of carcinogenesis were identif ied for these agents; such

protocols are generally consid ered largely non-responsive for m odes of action other than

mutagenicity and potent es trogenicity (e.g., DES).

For five chem icals (amitro l e, DDT, diel drin, PBB and DPH) , the sam e tumors were

observed from early and/or adult exposures, t hough the studies for am itrole, DDT, and dieldrin

only evalu a tes the an im als for liv er tum o rs. W ith ETU, no tum o rs in m i ce or rats were observed

f

o llowing pe rina tal expo sure alon e (e xcept a sm all, not-s ta tis tically -sign i f i cant in crea s e in m a le

rat thyroid tum o rs), while thyr oid tumors were observed in a dult rats and thyroid, liver, and

pituitary tumors in adult m i ce. Analysis of th e incidence of tum o rs pe r tim e of expos ure shows

early -lif esta ge susceptib ilitie s. The r a nge of the m a gnitudes of the tum o r inciden ce ra tios of

juvenile to adult exposures is sim ilar for the repeated dosing studies (0.06–13.3, weighted

geom etric mean 2.2, 27% of ratios greater than 1, Tables 5 and 8) and lifetim e dosing studies

(0.15–36, weighted geom etric m ean 3.4, 21% of ratios greater than 1, Tables 7 and 8). These

ranges and m eans are sim ilar to those for chem i cals with a mutagenic m o de of action, though the

m eans and m a xi m u ms are som e what lower. A g ain, liver tu mors are co mmon to these chem icals

.

The range of the m a gnitudes of liver tum o r inciden ce ratios of juvenile to adult exposures also is

sim ilar for the repeated dosing studies (0.06–13. 3, weighted geom etric mean 2.6, 43% of ratios

greater than 1, Tables 5 and 8) and lifetim e dosing studies (0.15–36, weighted geom etric m ean

5.8, 33% of ratios greater than 1, Tables 7 and 8).

The m a jor factor that co mplicates th e interpre tation of the results is that these studies,

except with DDT and dieldrin, invol ved dietary feeding initially to the m o ther, which potentially

could increase or decrease the dose received by the pups. Due to the m a ternal dosing during

pregnancy and lactation, the extent to which offs pring received sim ila r doses during different

early and ad ult lif e stage s is par ticu l ar ly uncer ta in for DPH, ETU, and PBBs. Oral gavage doses

in young anim als were selected to approxim a te th e average daily dose in adult dietary studies

based on standard estimates of feed consum ption in the s t ud ies with DDT and dield r in, while the

am itrole study involved dietary f eeding postnatally to the m o t h er so the young were dosed via

lactation. In addition, DDT, dieldr in, and som e PBBs are m o r e persistent in the body than are

20

### most chem icals, leading to a prolonged exposur e even following lim ited dosing. Thus, these

studies provide evidence that earl y lifestages can be m o re suscep tible to exposures to chem icals

causing cancer through a variety of modes of action other than m u tagenicity. However, the

studies with ethylene thiourea, which acts via thyroid disruption, indicate that this is not

necessarily the case for all m odes of action.

3.2.3. Ionizing Radiation

As m e ntioned previously, th e UNSCEAR, Anne x I (2000) in cludes inform ation derived

from a wide range of both intentional (gener ally diagnostic or th erapeutic m e dical) and

acciden t al radiation expo sures. Only infor m ati on derived fro m the Japan e se population (referred

to as the Life Span Study in the UNSCEAR Annex I) is presented here. A statistically significant

excess cancer m o rtality associated w ith radi ation has been found am ong the bom b survivors for

the following types of cancer: esophagus, stom ach, colon, liver, lung, bone and connective

tissue, skin, breast, u r in a r y tra c t, and leukem i a. Tables 9 and 10 ar e ex tra c ted f r om the tables in

UNSC E AR, Annex I. The excess rela tive risk (ERR) is the inc r eased cancer rate relativ e to an

unexposed population; an ERR of 1 corresponds to a doubl ing of the cancer rate. Because of the

low num b ers of cancers in individual sites with in narrow age groups, the ERRs for the various

solid tum o rs and leukem i a were presented only as le ss than o r greater than 20 years o f age at the

tim e of exposure. The larger num ber of thyroid tu mors enable a m o re detailed breakout shown in

Table 10. Most sites show greater risks in the younger than in the older ages.

The U.S. EPA (1994) docum ent presents a m e thodology for estim ation of cancer risks in

the U.S. population due to low-LET (linear energy transfer) radiation exposures using data from

the Atom ic Bom b Survivor Study (ABSS) as we ll as from s e lected m e dical exposures. The

report developed m o rtality risk co efficients using several m o dels th at took into accoun t age and

gender dependence of dosim etry, radiogenic risk, and com p eting causes of death as well as

transporting of risks across populat ions. The risk projections we re updated using m o r e recent

vital statistics in a repo rt that also included an uncertainty analysis (U .S. EPA, 1999). Details of

the derivatio n of these co efficients are available at

http://www.epa.gov/radiation/docs/rad_risk.pdf

.

Table 11 co ntains the calculated age-speci fic risk coefficients derived from the

application of the various m odels to the ABSS data. For m o st of the site s in the tab l e, the risk

coefficients are higher in the earlier age groups; liver, bone, ski n, and kidney coefficients are

age-independent and only esophageal cancer coef fici ents increase with incr easing age. Also of

note is that the coefficients gene r a lly are high er f o r f e m a les. Sim ilar to th e inf o rm ation f r om the

UNSCEAR (2000) Annex, m o st sites show greater risks in the younge r ages than the older ages.

21

### However, a com p arison of the two tables seem s to show reversal of risks for som e sites as a

function of age at exposure. W h ile the high sam p li ng variability in the epidem iological data for

som e ages m a y contribute to this apparent reve rsal, the cho i ce of risk m odels and as sociated

param e ters also is a factor.

22

### 4. DISCUSSION

The challenge for this analysis was how to use the existing, but lim ited, scientific

database on early postnatal and juve nile expo sur e s to ca rcino g ens to inf o r m a science policy

decision on whether, and if so how, to assess th e risk from c h ildhood exposures to chem icals for

which we have evidence of carcinogenicity only in adult hum ans or sexually m a ture laboratory

anim als. The database overall is of lim ited size (particularly com p ared with th e num ber of

chem icals that have been studi ed in a dult occup a tional ep ide m iological studies or chronic

bioassays). The m a jority of th e human data involves exposures to ionizing radiation or DES

(Anderson et al., 2000). More than 50 chem ical s have been dem onstrated to cause cancer

following perinatal exposures in anim als (without adult exposure s), but only a subset of the

chem icals have com p arative s t udies across ag es. The com p ar ative experim e ntal studies used 18

chem icals, 12 of which had m u tagenic m odes of ac tion and 6 of which had data from repeated or

lifetim e exposures. Other analyses of si m ilar data have found sim ilar results (Hattis et al. 2005),

but have focused on other aspects of the data, e.g., gender differences.

Previously published or internal U.S. EPA analyses have concluded that the standard

anim al bioassay protocols usually do not m i ss ch em icals that would have been identified as

carcinogens if perinatal exposures had been undertaken (McC onnell, 1992; Miller et al., 2002;

U.S. EPA, 1996). Given the increased com p lexity and costs of chronic bi oassays with perinatal

exposures, a lim ited number of such studies have been perform e d. Howe ver, these are the

studies that largely constitute th e av ailab l e database for this anal ysis. In addition to the chronic

bioassays with perinatal exposures , there are studies with acute dos ing at different lifestages and

a large number of studies with perinatal exposur es without a directly com p arative adult study.

Two other kinds of inform ation can contri bute toward developi ng a sc ientif ically

inform ed policy : theo retical anal yses and analyses of stop studies. 4 

### Theoretical analyses suggest

that the dif f

e rential su sce p tibilit y would depend in part on the m ode of action (i.e., at what step

in the cancer process(s) the chem ical was acting) and that the use of th e average daily exposure

prorated over a lifetim e m a y underestim ate or over estim a te the cancer risk when exposures are

tim e-dependent (Goddard and Krewski, 1995; Mu rdoch et al., 1992). Evidence for oldage-depend e n t prom otion of basophilic f o ci in ra ts by perox i s o m e prolif er ators app ear s to

provide a concrete example consis tent with these theo retical analyses (Ca ttley et al., 1991;

Kraupp-Grasl et al., 1991). Th e stop studies perform e d by the National Toxicology P r ogram

began exposure at the standard post-weaning age, but stopped exposure afte r varying periods of

months. Other groups of anim als we re exposed f o r a full two years; all anim als were evaluated

4 St o p st udi es a r e st u d i e s i n w h i c h ex p o s u re i s hal t e d a f t e r a pre d et erm i ned peri od

.



### 23

### for tum o rs a t the end of two years regardless of the duration of exposure (Halm e s et al., 2000).

Related data also are available from t h e stop st udies with vinyl chloride (Drew et al., 1983).

Analysis by Halm es et al. (2000) showed that, for six of the eleven chem icals and half the tum o r

sites, the assum p tion that the cancer risk would be equal when the produc t of concentration and

tim e (i.e., C x T) was constant was incorrect, and us ually underestim ated risk, as m o re of the risk

cam e from the beginning of the exposure rather th an the end. This dependence of risk on both

duration and intensity of exposure did not appear to be correlated with m u tagenicity. It should

be noted that these stop studies all involved exposures ea rly in the life of the anim al (as opposed

to a lim ited num b er of cancer st udies that looked at later periods of life; e.g., Drew et al., 1983),

but the extent to which the differences in tum o r outcom e result from increased susceptibility in

these early periods or the extended period for expression of the cancer cannot be evaluated.

These stop s t udies also u s ed doses as high as or higher than the highe st dose used in the two-year

exposure. T h is latter factor clearly had a signific ant effect for two chem icals, causing tum o rs at

higher doses that were not observed at lower dos es. These results suggest that pharm a c okinetic

or other dose-rate dependencies can m a ke the effects of expo sure s at high doses different from

those exposures at lower doses. W h ile not dir ectly inform ative about early childhood exposures,

these studies provide a perspective on the comm on cancer ris k assessm ent practice of averag ing

exposures over a lifetim e, especially those that include earlier lifestages. Thus, alternative

m e thods for estim ating risks from short-term exposures during childhood should be considered.

Inf o rm ation on dif f

e rent lif estag e sus ceptib il ities to cance r r i s k s f o r hum a ns exists f o r

ionizing radiation. The effects of chem ical m u ta gens at different lifest ages on cancer induction

are derived f r om laboratory anim al studies. W h ile the induction of cance r by ionizing radiation

and the induction of cancer by chem ical m u tagens are not identical processe s, both involve direct

dam a ge to DNA as critical caus a l s t eps in the p r o cess. In bo th cases, th e impacts of early

exposure can be greater than the im pacts of la ter exposures, probably due to som e combination

of early-lifestage susceptibility a nd the longer periods for observation of effects. As indicated in

Tables 9 and 10, A-bom b survivor s exhibited different lifestage dependencies at different tum o r

sites, though the total radiation- related incidence of tu m o rs showed a general slow decline with

age at exposure. However, as previously note d, there are app a rent d i fferences at som e sites

between the two tables. In addition to the sa m p ling and m odel i ng differences, the excess risk

values in Table 9 are based on Japanese baselin es while th e coefficients in Table 10 reflect

UNSCEAR’ s effort to transport the risks from the Japanese population to that of the U n ited

States. However, it is cle a r tha t the to tal radia tion -related tumor inc i denc e showed a general slow

declin e with age at expo sure.

The studies in rodents of chem icals with m u tagenic m odes of action sim ilarly support a

24

### general decline in induced cancer risk with age at exposure a nd sim ilarly show som e differences

for individual tum o r sites. In general, the earl iest two or three postnatal weeks in m i ce and ra ts

appeared to be the m o st susceptible, though some degree of increased susceptibility through

puberty in rats (beginning ar ound 5–7 weeks) and m i ce (beginni ng around 4–6 weeks) for som e

types of tumors exists.

All the acute dosing studies that dem o nstrated carcinogenicity with anim als of different

ages used ch em icals with a m u tagenic m ode of action (Tables 4 and 6). These studies provide

the clearest dem onstrations of periods of differen tial susceptibility becaus e the exposure rate is

constan t at the different ages. The repeated dose studies also include several of the m o st

inform ative studies for assessing perinatal carcin ogenesis, notably those on vinyl chloride and

DEN (Tables 2 and 4). T h e vinyl chloride studies by Malton i and colleagu e s are p a rt o f a large

series of studies on this com pound t h at include d exposures to different concentrations for

varying durations, including som e at early lifestages (Maltoni et al., 1984). The DEN study by

Peto et al. (1984) used a unique chronic study de sign in which groups of rats were exposed to

multiple d r inking water concentratio ns star ting at 3, 6, or 20 weeks of lif e. This des i gn provides

inform ation on the susceptibility of early exposur e periods within a nearly lifetim e exposure.

Beyond the analysis described here, there are conceptual biological rationales that w ould

suggest DNA-dam a ging agents would have greater im pacts on early lifestages. Growth involves

substantial levels of cel l replication, even in organs that in adults are only very slowly

replicating, thus increasing the likelihood that a cell will undergo division before the DNA

dam a ge caused by the m u tagen has been repaired. Increased replication also can lead to a

greater div i s i on of initiated cells, le a d ing to a lar g er num ber of initia ted c e lls p e r spe c i f i ed dose.

These periods of cell replication can vary for di fferent tissues. For example, DMBA appears to

be m o re effective at initiating m a mmary tum o rs in 6-8 week old rats, which are undergoing

developm ent of that tissue, than during earlier or later periods (Mer anze et al., 1969). W h ile

tum o r promotion processes can be very depe ndent upon the duration of prom otion, initiation

processes can occur in relative ly brief periods (e.g., the sing le-dose studies in anim als or

radiation exposure in hum a ns). Most tum o rs take extended periods to develop, m a king dam a ge

that occurs earlier in lif e more likely to result in tumors prio r to death than w ould exposures that

occur later in life. W h ile som e of these observ a ti ons m a y also pertain to other m odes, all of them

(with som e differences am ong tum o r sites) appear to be potentially relevant to a greater

susceptibility to m u tagenic m odes of action duri ng early -life stages (vs. later-life stag es).

The inform a tion on lifestage susceptibility for chem icals inducing cancers through m o des

of action other than direct DNA interaction is m o re varied, sh owing an increase in tumor

incidence during perinatal exposure versus expos ures of m a ture anim als (e.g., polybrom inated

25

### biphenyls induced liver tum o rs), no tum o rs from perinatal exposure (e .g., ethylene thiourea

induced thyroid tum o rs), no effect of com b ined perinatal and adult e xposure (e.g., DP H liver

tum o rs in rats and fe m a le m i ce), and different tu mors f r om perina tal exposure versus adult

exposure (e.g., DES, ascorbate). These variations are likely a result of the m odes of action of

these ch em i cals and the pharm acokinetic d i fferences in doses during different periods of life. No

studies were evaluated that were d i re ctly com p arable to the s i ngle-dose studies with mutagens,

which clearly show significant differences in tumor responses after explicitly controlled doses at

different lifestages.

Som e evidence for an effect of early

- lifestage exposures on tum o r incidence was

observed in studies with polybrom inated biphenyls, am itrole, DDT, dieldrin, and

diphenylhydantoin. These studies show increased incidence of tum o rs in m i ce from perinatal

exposure, though only those for polybrom i nated biphe nyls w e re statistically significant. (A

nonstatistically significant incr ease also was observed in m a le rats with polybrom inated

biphenyls.) Com b ined perinatal and adult exposures generally gave sta tis tica lly s i gnif i cant

increases, th ough not necessarily for each sex an d species (rat and m i ce) in the

diphenylhydantoin and polybr om inated biphenyl studies.

There are important demonstr ations of che m icals acting th rough m odes of action other

than m u tagenic to cause different tumor types with early-lifestage exposures com p ared with

exposures for adults, e.g., tam oxifen and DES (C arthew et al., 2000; Carthew et al., 1996, Gass

et al., 1964; Newbold et al., 1990, 1997, 1998). In addition, studies with in utero exposure to

atrazine (Fenton and Davis, 2002) , DES, and arsenic (W aalkes et al., 2003) indicate that earlylife exposures to com pounds can alter suscep tib ility of endocrine and reproductive o r g a ns. Three

of these compounds (i.e., DES, genistein, and tam oxifen) bind to the estrogen receptor. Ongoing

studies on ethinyl es trad iol, nonylphenol, and ge n i stein by the National To xicology Program will

add to this database for estrogens (Laurenzana et al., 2002; N e wbold et al., 2001). These studies

will evaluate cancer incidence in offspring expos ed in utero , during lactation, and through

adulthood via diet. A study with genistein f ound uterine tum o r developm ent to be dependent

upon early-lifestage exposures (New bold et al., 2 001). Another recent stu dy of estrogen found a

shorter latency for m a mm ary tum o rs in m i ce e xposed at 8 and 12 weeks as com p ared to m i ce

exposed at 4 or 18 weeks, indicating a suscep tible period between 8 to 12 weeks of exposure

(Yang, 2003). Thus, there is an actively growing database from which to consider issues of

childhood exposure and cancer for com pounds acting through the estrogen receptor or other

m echanis m s of endocrine disruption.

The ability to estim ate with any accu racy th e juv e nile to adult cance r pote n cy ratio

depends very m u ch on the experim e ntal design us ed. The lif etim e design has less ability to

26

### distinguish increased su sceptibility from early-lif e exposure than the other types of designs.

Consider tw o different experim e ntal designs. In the first, the “lifetim e” design, a group of

anim als are exposed starting as juveniles, a nd exposure continues through adulthood. A second

group are exposed only in adulthood, and the juve nile:adult ratio results from a com p arison of

tum o r incidences in the two groups. In the se cond, the “repeated” de sign, one group of anim als

is exposed only during the juvenile period, a nd is then followed through adulthood to assess

tum o r incidence, and a second group of anim als is exposed only through adulthood. The lifetim e

design turns out to be a particularly insensitiv e design for estim a ting the juvenile:adult ratio.

The following exam ple dem onstrates the m a gnitude of the problem : Suppose the risk per

day of exposure of a chem ical is ten fold greater in the juvenile period as in the adult period, and

anim als exposed through adulthood at a particular dose level have an extra risk of 60% for

having at least one tum o r, while 1% of control anim als have tumors. The adult exposure period

is 94 weeks, while the juvenile exposure period is 4 weeks. Thus, in the lifetim e design, the

group of anim als exposed as juven iles will receiv e a total of 98 weeks of exposure, (4 in juvenile

and 94 in adult), while those receiving the adult- only exposure receive 94 weeks of exposure. In

the repeated design, animals expos ed as juveniles receive only 4 weeks of exposure, w h ile the

adults receive 94 weeks, just as in the lifetim e design. Each group starts with 50 anim als. Under

these assumptions, using equations (1) and (2) from Section 2.3, the expected num ber of anim als

with tum o rs in the three treatm e nt groups (cont rol, juvenile-exposed, adult-exposed groups) in

the two designs is:

Number of animals w i th tumors

Control Early-life exposure Adult exposure

Lifetime 1 3 6 3 0

Repeated 1 1 6 3 0

Notice that in the “lifetim e” design, only six m o re juvenile-exposed anim als have tumors

than in the adult-exposed group, whereas in th e “repeated” design, 16 ju venile-exposed anim als

have tum o rs. The data in the lifetime design are consistent w ith the hypothesis of no tum o rs

being induced during the juveni le period: the ratios 36/50 an d 30/50 are not statistically

significantly different. In other words, the da ta f r o m the lif etim e design are sta tis tica l ly

consistent w ith the hypothesis of no risk at all during the juvenile peri od, even though the real

response is a 10 tim es greater risk from early-lif e exposure. T h e difference between the results

from the two different study designs is due to th e one-hit m o del: each add ition a l week of a long

exposure contributes less than th e previous week to th e total num b er of an im als with tum o rs.

27

### Note that, even if the one-hit m odel is not co rrect, chronic exposure pr obably results in a nonstatistically significant increase for the lifetim e exposure including juveniles as com p ared with

only adult exposure.

The proper m easure of relative potency of an exposure in the juveni le period relative to

an exposure in the adult period is the ratio of doses in the tw o periods that give the sam e

incidence of tum o rs. However, m o st of the data sets used in this repo rt contained only one noncontrol dose, precluding the ex tensive dose-response m odeling that would be required to

estim ate this ratio of doses. However, this do cu m e nt largely considered chem icals for which a

mutagenic mode of action has been establis hed and for which a linear, no-threshold doseresponse function is assum e d for the low-dose range being considered for risk assessm ent. In the

case of the linear dose-response f unction, the analysis of the relative response from the sam e

dose will p r oduce the s a m e value as ratio of doses that p r odu ces the s a m e inciden ce o f tum o rs.

For a one-hit dose- respo n se equation , the probab i lity of deve loping a tum o r af ter the

sam e dose and duration in the juvenile or adult period is



# ( )

( ) x m

j x m

a

j a



### e P P e P P

− −



### − − = − − =

0 0



### 1 1 1 1

for dose x . Suppose we want to calculate the dose D

a 

### or D j 

### th at resu lts in a given incidence of

tum o rs after an adult or juven ile exposure. From equation 1, D

a 

### and D j 

### equal:

j c

j a c

a



### m P P

D m P P

D

⎟ ⎟

⎠ ⎞

⎜ ⎜

⎝ ⎛

− −

−

= ⎟ ⎟

⎠ ⎞

⎜ ⎜

⎝ ⎛

− −

−

=

0 0



### 1 1 ln 1 1 ln

Thus, the ratio D

a 

### / D

j 

### = m j 

### /m

a 

### , the r a tio calcu lated in this do cu m e nt.

In summ ary, this analysis supports the conclu sion that there can be greater su sceptibility

for the developm ent of tumors as a result of exposures to chemicals acting through a m u tagenic

mode of action, when the exposures occur in early lifestages as com p ared with later lifestages.

Thus, this S upplem ental Guidance recommends for chem icals with a m u tagenic m ode of action

for carcinogenesis when chem ical -specific data on early-life exposure are absent, a default

28

### approach us ing estim ates from chronic studies (i.e., cancer slo p e factors ) with approp riate

modifications to address the potential for differential risk of early-lifestage exposure. For

chem icals acting through a non-m u tagenic m ode of action, e.g., horm onally m e diated

carcinogens, the available data suggest that othe r approaches m a y need to be developed for

addressing cancer risk estim ates from childhood exposures. T h is is a p a rticular concern because

the tum o rs arising from horm onally active chem i cals appear to involve different sites when

exposure is during early-life vers us adulthood, an effect that has been observed relatively

infrequently. Development of such approaches w ould require additional re search to provide an

expanded scientific basis for their support, in cluding additional res earch and the possible

developm ent of new toxicity testing protoc ols that consider early lifestage dosing.

The current data do also not al low analysis of so m e issues of potential interest for risk

assessm ent, e.g., potential increased risk of childhood cancer, from in uter o or childhood

exposures. Assessing the role of environm ental exposures on childhood cancers is dif f icult, but

additional research could include epidem iological studies or experim e ntal studies with anim als

genetically designed to expre ss cancers analogous to hum an childhood cancers. Rigorous

quantification of exposure doses at different lifestages and in rode nt pups in experim e ntal studies

would be useful for evaluating whether th ere is greater child hood susceptibility.

Pharm acokinetic m odeling could bett er def i ne th e inte rnal do ses to im prove determ ination of the

m a gnitude of increased s u sceptibility

.

29

### 5. GUIDANCE FOR ASSE SSI NG CANCE R RISKS

FROM EARLY-LIF E EXPOSURE



### Consisten t with the app r oach and reco mmendations of the U.S. EPA cancer risk

assessm ent guidelines (U.S. EPA, 2004), any asse ssm ent of cancer susceptibility will begin with

a critical analysis of th e available inform ation. Figure 3 show s the proposed steps in the process.

The potential for increased susceptibility to cancer from early-life exposure, relative to

com p arable exposure later in life, generally warr ants exp licit considerat io n for each assessm ent.

W h en developing quantitative es tim ates of cancer risk, th e Agency recom m e nds

integration of age-specific values for both expos ure and toxicity/potency where such data are

available and appropriate. Children, in general, are expected to have som e exposures that differ

from those of adults (either higher or lower), due to differe nces in size, physiology, and

behavior. F o r exam ple, childre n are generally assum e d to eat m o re food and drink m o re water

relative to their body weight than adults. Children’s norm a l activitie s, such as putting their hands

into their m ouths or playing on the ground, can result in exposures to contam inants that adults do

not encounter. Moreover, children and adults exposed to the sam e concentration of an agent in

food, water, or air m a y receive d i fferent (h igher o r lower) internal doses d u e to differences, for

exam ple, in intake, m e ta bolism

, or absorption ra tes. Childr e n are less like ly than adu l ts to be

exposed to p r oducts typically u s ed in industrial settings and often have m o re lim ited diets th an

adults. W h en assess ing r i sks, if the d a ta a r e ava i lable and rele vant, it is im portan t to in clude

exposure that is m easured or m odeled for a ll lifestages, including exposures during childhood

and during adulthood. E P A continues to deve lop better tools for assessing childhood exposure

differences, such as the Child-Specific Exposure Factors Handbook (U.S. EPA, 2002a ), and

models, such as Stochastic Hum a n Exposure a nd Dose Sim u l a tion (SHEDS) and Consolidated

Hu m a n Activity Database (CHAD) (McCurdy et al., 2000; Zartarian et al., 2000)

Mode-of-action studies can be a source of data on quantitative differences between

children and adults (Figure 3, Box 1). If the av ailable inf o rm ation is suf f i cient to establish the

agent’s m ode of action f o r ear ly- lif e and adult ex posures, th e n the im plic ations f o r e a r ly- lif e

exposure of that m ode of action are used to develop separate risk estim a tes for childhood

exposure. P e rtinent inform ation can be obtained both from a g ent-specific studies and from other

30

### studies that investigate the gene ral properties of the particular m ode of action. All data

indicating q u antitative d i fferences b e tween chil d r en and adu l ts are consid ered in d e veloping

those portion(s) of the risk es tim ates for exposure estim ates that include childhood exposure.

Som e examples in clude the poten tial for children to have a d i fferent internal dose of the active

agent or a change in a k e y precu rsor event (see S ection 2.4.3.4 of the Guideline s for Cancer Risk

Assessment ).

When the mode of action cannot be established (Figure 3, Box 2), the policy choice

would be to use lin ear ex trapo l at ion to lower doses such that risk estim ates are bas e d on a

lifetim e average daily ex posure with out furthe r adjustm e nt. No general adjustm e nt is

recomm ended at th is tim e. This policy cho i ce is consis tent with past U. S. EPA practice that h a s

been favorably evalu a ted over the y ears. The result would be expected to produce plausible

upper bound risk estim ates, based on th e use of linear extrapolation as a default in the absence of

inform ation on the likely shape of the dose-response curve.

When a m o de of action other than m u tagenicity is estab lished, if it is non linear (Figu r e 3,

Box 3) or linear (Figure 3, Box 4), no general ad justm e nt is recomm ende d at this time. Although

the ava ilab l e studies (dis cussed prev iously) ind i c a tes that h i g h er or lower cancer risk s m a y result

from early-life exposure, there is insufficient inform ation or an alyses currently available to

determ ine a general adjustm e nt at this tim e. As other m odes of action becom e better understood,

this inform ation m a y include data on quantitativ e differences between children and ad ults. If

such data are available, an analys is of the differences could be us ed to a d just r i sk es tim a tes f o r

childhood exposure. EPA expects to expand this Supplem ental Guidance to specifically address

modes of action other than m u tagenicity when sufficient data are availab l e and analyzed.

W h en the data ind i cate a mutagenic mode of action, 5 

### the available s t udies (discus s ed

5 Determ ination of chem icals that are operati n g by a m u ta geni c m ode o f ac t i on e n t a i l s eva l uat i on o f t e st r e sul t s f o r

genet i c e n dp oi nt s, m e t a bol i c pr ofi l e s, p h y s i c ochem i cal pro p ert i e s, a n d st r u ct u r e-act i v i t y anal y s es i n a w e i ght

- o f

evi d e n ce a p pr o ach ( W at e r s et al ., 1 9 9 9 )

. Est a bl i s he d pr ot oc ol s are u s ed t o gene rat e t h e da t a (C im i no, 2 0 01; OEC D

, 19 9 8 ; U

. S. EP A, 2 0 0 2 b ); h o w eve r

, i t i s rec o g n i zed t h at ne wer m e t hods a n d t ech nol ogi e s suc h as t h ose ari s i n g fr om

gen o m i cs can pr o v i d e use f ul dat a an d i n si g h t s t o a m u t a gen i c m ode of act i o n

. C a rci n oge ns act i n g t h r o u g h a

m u tag e n i c m o d e o f actio n g e n e rally in teract with DN A and can produce s u ch effects as DNA a d ducts and/ or

b r eak a g e . Carcin og ens with a m u tag e n i c m o d e o f actio n oft e n p r od u c e po sitiv e effects i n m u l tip le test syste m s fo r

di ffe re nt ge net i c en dp oi nt s

, pa rt i c ul arl y ge ne m u t a t i ons a n d st ruct u r al c h r o m o so m e aberra t i ons, an d i n t e st s

per f o r m e d in vivo

, whi c h gen e ral l y are s u p p o rt e d by t h ose per f o r m e d in vitro . Th is m o de of action is ad dressed i n

m o re det a i l i n Sect i on 2. 3. 5 o f EP A’ s ca ncer guidelines (U .S. EPA

, 200 5)

.



### 31

### above) ind i cate high er cancer risks resulting from a given exp o sure occu rring early in life when

com p ared with the sam e am ount of e xposure during adulthood. However, chem ical-specific data

relating to mode of ac tion (e.g., toxicokinetic or toxicodyna mic inform ation) m a y suggest that

even though a com pound has a m u tagenic m ode of action, higher cancer risks m a y not result.

Such data should be considered before appl ying the age-dependent adjustm e nt factors.

If the available, chem ical-specific inf o rm ation includes an epidem iologic study of the

effects of childhood exposure or an anim al bioass ay involving early-life exposure (Figure 3, Box

5), then thes e studies are analyzed to develop risk estim ates (i. e ., cancer slope factors ) that

specifically address any potential f o r

dif f

e rentia l potency in e a rly lif estag e s. An exam ple is the

IRIS assessm e nt of vinyl chlo ride (U.S. EPA, 2000b; c).

In the absence of early-life st udies on a specific chem ical under consideration (Figure 3,

Box 6), the extrapo l atio n from th e point of departure to lower dos es em ploys linear ex trapo l ation

(see Section 3.3.1 of the U.S. EPA [2005] cancer guidelines). This choi ce is based on mode-ofaction data indicating that m u tagens can give rise to can ce rs with an appa rently low-d o se lin ear

response. A d justm e nts to the resultant risk estimates are specified with regard to childhood

exposures. This approach is adop ted because ri sk estim ates based on an average d a ily exposure

prorated over a lifetim e do not consider the pote ntial for hig h er cancer risks from ear ly-life

exposure.

The adjustm e nts described below ref l ect the potential for early-life exposure to m a ke a

greater con t ribution to cancers ap p ear ing la ter in lif e . The 10-f o ld adjus t m e nt rep r esen ts an

approxim a tion of the weighted geom etric m ean tu m o r incide nce ra tio f r o mjuvenile o r adult

exposures in the repeated dosing st udies (see T a ble 8). This adjustm e nt is applied for the first 2

years of life, when toxicokinetic and toxicodyna m i c differences between children and adults are

greatest (Ginsberg et al., 2002; Renwick, 1998). Toxicokinetic di fferences from adults, which

are greatest at birth, reso lve by appro x im ately 6 months to 1 year, wh ile higher grow th rates

extend f o r lo nger per i ods . The 3-f o ld adjustm e nt repres ents a n inte rm ediate lev e l of a d justm e nt

that is applied after 2 years of age through <16 years of age. This upper age lim it represents

m i ddle adolescence following the period of rapi d developmental changes in puberty and the

conclusion of growth in body height in NHANE S data (Hattis et al., 2005). Efforts to m a p the

approxim a te start of m o use and rat bioassays (i.e ., 60 days) to equivalent ages in hum ans ranged

from 10.6 to 15.1 years (Hattis et al., 2005). Data are not available to ca lculate a specific doseresponse adjustm e nt factor for the 2 to <16- year age range, so EPA selected the 3-fold

32

### adjustm e nt because it reflects a m i dpoint, i. e., a pproxim a tely half the difference between 1 and

10 on a logarithm i c scale (10 1/2 

### ), between the 10-fold adjustm e nt f

o r the f i rst two ye ars of lif e

and no adjustm e nt (i.e., 1-fold) for adult expos ure. EPA also recognizes that exposures

occurring near the end of life m a y ha ve little effect on lifetime cancer risk , but lack s adequate

data a t pres e n t to prov ide an adjus t m e nt f o r this "wasted dose" effect. Sim ilarly, since m o st of

the studies involved only one latenc y period, the potential effect of early-life exposure on latency

for the observed tum o rs could not be evaluated. The lack of data on effect on latency also

lim ited the types of analyses that could be perform e d, e.g., more com p l e x dose-response

f unctions, su ch as m u lti-s t age or clon al expans ion m odels, cou l d not be ev aluated. Thus, the

potential effects of early-life expos ures on latency were not evaluate d. Finally, as the adjustm e nt

f

actors a r e d e rived f r om a weighted g e om etric m e an of the data evalua ted, these ad justm e nt will

both over-estim ate and under-estim at e the potential potency for ear ly-life exposure for chem icals

with a m u tagenic m ode of action for carcinogenesis. An exam ination of the data in the tables

dem onstrates that som e of the ratios were less th an one, while others exceeded 10. For this

reason, the Supplem ental Guidance em phasizes that chem ical-specific data should be used in

preference to these default ad justm e nt fact ors w h enever su ch data are av ailab l e.

The following adjustm e nts rep r esen t a practical approach th at reflect s the results of th e

preced ing analysis, which concluded that cancer risks gen e rally are higher from early-life

exposure than from si m ilar expos ure durations later in life:

• For exposures before 2 years of age (i.e., span ning a 2-year tim e interval from the first

day of birth up until a ch ild’s second birthd ay), a 10-fold adju stm e nt.

• For exposures between 2 and <16 years of age (i .e., spanning a 14-year tim e interval from

a child ’s second birthd ay up until their sixt eenth b i rthd ay), a 3

- fold adjus t m e nt.

• For exposures after turning 16 years of age, no adjustm e nt.

Clearly other age groups, such as an ag e group experiencing pubertal changes in

physiology, or approxim a tely ages 9 - 15, m a y ex perience changes in biological processes that

could lead to m odifications in the susceptibil ity to the effects of so m e carcinogens, depending on

the m ode of action. This Supplem ental Guidan ce focuses on carcinogens with a m u tagenic m ode

33

### of action. F o r any m ode of action, th e Agency is inte rested in identif y ing lif estag e s th at m a y be

particularly sensitive or refr actory for carcinogenesis, and beli eves that the m ode of action

fra m ework as described by EPA’s cancer guide lines (U.S. EPA, 2005), is an appropriate

m echanism for elucidating these lifestages. In g e ne ral, the Agency’s an alyses of lifestages that

m a y be susc eptible will depend on three factor s: (1) establishing th e m ode of action for

carcinogenesis; (2) using knowledge about the biological and toxico logical key events in that

mode of action that are likely to be affected by lif estages; and (3) the availability, or

developm ent, of data that allow analysis of the effects of chem icals actin g by that m ode of action

during th e relevant ages. For each m o de of action evaluated, th erefore, th e various ag e groupings

determ ined to be at a differen tial risk, which m a y di ffer significantly from those proposed for the

mutagenic mode of action, are expected to be ev aluated independently of other m odes of action.

When data, including well establis hed m ode of a c tion data, are availab l e that allow sp ecific

evaluation of lifestage differences in toxicokinetics or toxicodynam i cs that would lead to lesser

or greater susceptibility f r om early-lif e exposures to carcinogens, then thos e data should be used,

as generally discussed in EPA’ s cancer guidelines (U.S. EPA, 2005) , in preference to the default

procedures described in this Supplem ental Guidance.

The 10-fold and 3-fold adjustm e nts in slope f acto r are to be com b ined with age-specific

exposure estim a tes when estim ating cancer risks f r om early life exposure to carcinogens that act

through a m u tagenic m o de of action. It is im portant to em phasize that these adjustments are

com b ined with co rrespo nding age-s p ecific es tim a tes of exposure to assess cancer risk. For

exam ple, for a 70-year lifetim e, wher e there are data showing negligible exposure to children,

the estim ated cancer risk from childhood exposure would be also negligible and the lifetim e

cancer risk would be reduced to th a t resu lting f r om the rele vant num ber of years of adult

exposure (in the absence of specific infor m ation, 55 years). Where there are data (m easured or

modeled) for childhood exposures, the age-group specific exposure values are used along with

the corresponding adjustm e nts to the slope factor. W h ere there are no relevant data or m odels

for childhood exposures and only lifetim e averag e exposure data are available, the lifetim e

exposure data are used with th e adjustm e nts to th e slope facto r for each ag e segm ent.

It is recognized that, when the exposure is fairly uniform ove r a lifetim e, the effect of

these ad justm e nts on estim ated lifetim e cancer risk are sm all rela tive to th e overa ll un certainty of

34

### such estim a tes. These adjustm e nts can be appl ied when estim a ting the can cer risk resulting from

childhood exposure. These adjustments are appl ied when developing risk estim ates from

conventional anim al bioassays or epidem iologic studies of effects of adult exposure. Som e

exam ples f o llow in th e n e xt sec tion.

The Agency has also carefully considered both the advantages and disadvantages to

extending the default potency adju s t m e nt factors to carcinog enic chem icals for which the m ode

of action rem a ins unknown. It is the Agency’s l ong-standing science polic y position that use of

the linear low-dose extrapolation approach (without further adju stm e nt) provides adequate public

health conservatism in the absence of chem ical -sp ecific data indicati ng dif f

erentia l ea rly-lif e

susceptibility. At the present tim e , therefor e, EP A is recomm ending these age-d e pendent

adjustm e nt factors only for carcinogens acting through a m u ta genic m ode of action based on a

com b ination of analysis of available data and the above-m entioned science policy position. In

general, the Agency prefers to rely o n analyses o f data, rath er th an general defaults. When data

are availab l e for a suscep tible lifes tag e , they shou ld be used directly to ev aluate risk s for that

chem ical and that lifestage on a case-by-case basis. In this analysis, the data for non-mutagenic

carcinogens, when the mode of action is unknown, were judged to be too lim ited and the m odes

of action too diverse to use this as a category f o r which a gen e ral d e f a ult adjustm e nt f actor

approach can be applied.

35

### 6 COMBINING LIFES T AGE DIF F ERE N CE S IN EXPO SURE AND DOSERESPONS E WHEN ASSESSI NG CARCINO GEN RISK - SOME EXAMPL ES FOR

CARCINO GENS TH AT ACT T H ROUGH A MUTAG E NIC MODE OF ACTION



### It is im portant for the risk assessor to consid er lif estage differences in both exposure and

dose-response when assessing cance r risk resulting from early-lif e exposures. As discussed in

Section 5, age dependent adjustm e nts factors (A DAFs) in dose response (i .e., slope factors) are

com b ined with age specific exposure estim ates when assessing cancer risks. This is a departure

from the way cancer risks have historically been based upon the prem ise that risk is proportional

to the daily average of lifetim e dose. This Supplem ental Guidance recomm ends an integrative

approach th at can be u s ed to a sse ss tota l lif etim e risk resulting f r om lif etim e or less-than-lif etim e

exposure during a specif ic portion of a lifetim e.

The following exam ples can help demonstrate how to apply this guidance by integrating

potential lifestage d i fferences in exposure and/or dose-response (potency), and also demonstrate

what the resulting im pacts are on calculated risks. These hypothetical examples consider risks

from both lifetim e, as well as le ss-than-lifetim e oral exposures. Risks associ ated with inhalation

exposure to carcinogens that act vi a a m u tagenic mode of action are calculated in sim ilar fashion

by applying the appropriate ADAF(s ) along with the corresponding i nhalation unit risk estim ate,

using pertinent estim ates of exposure concentration.

Note again, ADAFs are only to be u s ed for agen ts with a m u tagenic m o de of action for

carcinogenesis when chem ical-specif i c data are abse nt. For all m odes of action, when chem icalspecific data are available for early-lif e exposure, those data should be used.

6.1 CALCULATING LIFETIME RISKS ASSO CIATED WITH LIFETIME EXPOSURES

Example 1 : 

### Consider a scenario of exposure to a carcinogen with a nonmutagenic 

### mode of

action. Suppose the oral cancer slope factor derived from a t ypical animal study (i.e., where

dosing begins after puberty) is estim a ted to be 2 per m g /kg-d, and the exposure rate rem a ins

constant throughout life at 0.0001 mg /kg-d (this is equivalent to saying the daily average of

lifetim e dose rate is equal to 0.0001 mg/kg-d). T h e risk from lifetim e exposure is calculated by

multiply ing the slop e factor and the exposure rate:

Risk = (2 per m g /kg-d) x (0.0001 m g /kg-d)

36

### = 2 x 10 -4



### Example 2 : 

### Now consider the sam e exposur e scenario for a carcinogen with a muta genic 

### m ode

of action for which the oral cancer slope fact or, derived from a typical anim al study where

dosing begins after puberty, is also estim ated to be 2 per m g /kg-d. In this case, ADAFs are used,

as f o llows.

a. To calculate lifetim e risk for a population with average life exp ectancy of 70 years,

sum the risk associated with each of the th ree relevant tim e periods :

• Risk during the first 2 years of lif e ( w here the A DAF = 10);

• Risk for ages 2 through < 16 (ADAF = 3); and

• Risk for ages 16 until 70 years (ADAF = 1).

Thus, risk equals the sum of:

• Risk for birth through < 2 yr = (2 per m g /kg-d) x 10 (ADAF) x (0.0001 m g /kg-d)

x 2yr/70yr

= 0.6 x 10 -4



### • Risk for ages 2 through < 16 = (2 per m g /kg-d) x 3 (A DAF) x (0.0001 m g /kg-d)

x (13yr/70yr)

= 1.1 x 10 -4



### • Risk for ages 16 until 70 = (2 per m g /kg-d) x 1 (A DAF) x (0.0001 m g /kg-d)

x (55yr/70yr)

= 1.6 x 10 -4



### Risk = 0.6 x 10 -4 

### + 1.1 x 10 -4 

### + 1.6 x 10 -4



### = 3.3 x 10 -4



### b. If exposure varies with age, then such differences are also included. N o w suppose the

sam e exa m ple as imm e di ately above, except that exposure for ages 1 through <12 was

twice as high as exposure for all other ages. In this case, sum the risk associated with

each of the five relevan t tim e periods in wh ich ex posure rates and/or po ten c ies (slope

37

### factors) v a ry :

Risk equals the sum of :

• Risk for birth through < 1 yr (1yr) = (2 per m g /kg-d) x 10 (ADAF) x 0.0001 m g /kg-d

x 1yr/70yr

= 0.3 x 10 -4



### • Risk for ages 1 through < 2 (1yr) = (2 pe r m g /kg-d) x 10 (ADAF) x 0.0002 m g /kg-d

x 1yr/70 yr

= 0.6 x 10 -4



### • Risk for ages 2 through < 12 (10yr) = (2 per m g /kg-d) x 3 (A DAF) x 0.0002 m g /kg-d

x 10yr/70yr

= 1.7 x 10 -4



### • Risk for ages 12 through < 16 (4yr) = (2 per m g /kg-d) x 3 (A DAF) x 0.0001 m g /kg-d

x 4yr/70yr

= 0.3 x 10 -4



### • Risk for ages 16 until 70 years (55y r) = (2 per m g /kg-d) x 1 (ADAF) x 0.0001 m g /kg-d

x 55yr/70yr

= 1.6 x 10 -4



### Risk = 0.3 x 10 -4 

### + 0.6 x 10 -4 

### + 1.7 x 10 -4 

### + 0.3 x 10 -4 

### + 1.6 x 10 -4



### = 4.5 x 10 -4



### 6.2 CALCULATING LIFETIME RISKS ASSO CIATED WITH LESS THAN LI FETIME

EXPOSURES



### If exposure only occurs for a lim ited num b er of years (for exam ple, consider a fam ily that

lives n ear a source of ex posure for a five-year p e riod of tim e before m ovi ng away), it is critical

to com b ine lifestage differences in exposure and dose-response for the relevant tim e interval.

The exam pl es presented below demonstrate how adjusting potency and/or exposure can affect

the assessment of cancer risk.

38

### Example 3 : 

### If exposure to a carcinog en with a mutagenic mode of action with an oral slope

factor equal to 2 per m g /kg-d occurs during ad ulthood for only 5 years, the daily average of

lifetim e dose is tim e weighted to apportion risk f

o r the num ber of years of exposure by a factor

of 5/70:

Risk = (2 per m g /kg-d) x (0.0001 m g /kg-d) x (5yr/70yr)

= 1.4 x 10 -5



### Example 4 : 

### If this 5-year exposure occurs during chil dhood, the risk calculations are adjusted to

consider the potential for higher potency from early-lif e exposure. Assessors should rem e mber

that the age dependent adjustm e nt factors for ca rcinogens with a m u tagenic m ode of a c tion are

applied only to exposure periods occurring up to age 16.

a. For a child exposed betw een ages 5 and 10, only a 3-fold ADAF is applied because

the exposure occurs entirely be tween ages 2 and <16 years:

Risk = 3 (ADAF) x (2 per m g /kg-d) x (0.0001 m g /kg-d) x (5 yr/70 yr)

= 4.3 x 10 -5



### b. For an exposure between ages 13 and <18, a 3-fold ADAF i s applied only to the

3-year portion occurring before age 16:

Risk equals the sum of :

• Risk for ages 13 through < 16 (3yr) = 3 ( ADAF) x (2 per m g /kg-d) x (0.0001 m g /kg-d)

x (3 yr/70 yr)

= 2.6 x 10 -5



### • Risk for ages 16 through < 18 (2yr) = 1 ( ADAF) x (2 per m g /kg-d) x (0.0001 m g /kg-d)

x (2 yr/70 yr)

= 0.6 x 10 -5



### Risk = 2.6 x 10 -5 

### + 0.6 x 10 -5



### 39

### = 3.2 x 10 -5



### c. For a child exposed from birth through age 5, different ADAFs are applied to the

periods before and after age 2:

Risk equals the sum of :

• Risk for birth through < 2 (2yr) = 10 (ADA F) x (2 per mg/kg-d) x (0.0001 m g /kg-d)

x (2 yr/70 yr)

= 5.7 x 10 -5



### • Risk for ages 2 through < 5 (3yr) = 3 (ADAF ) x (2 per m g /kg-d) x (0.0001 m g /kg-d)

x (3 yr/70 yr)

= 2.6 x 10 -5



### Risk = 5.7 x 10 -5 

### + 2.6 x 10 -5



### = 8.3 x 10 -5



### Example 5 : 

### Lifetim e risk calculations based on less- than-lif etim e exposure to a carcinogen with

a m u tagenic mode of action include any lifestage changes in potency as w e ll as exposure. In this

exam ple, again consider a scenario of 5 year s of exposure to a carcinogen with a m u tagenic

mode of action, but suppose that the exposure rate is found to vary from 0.0002 m g /kg-d during

the first 2 years of life, to 0.0001 m g /kg-d during the last 3 years.

a. For a child exposed betw een birth and age 5, sum the risk associated with the two

relevant time periods:

Risk equals the sum of :

40

### • Risk for birth through < 2 (2yr) = 10 (ADA F) x (2 per mg/kg-d) x (0.0002 m g /kg-d)

x (2 yr/70 yr)

= 11.4 x 10 -5



### • Risk for ages 2 through < 5 (3yr) = 3 (ADAF ) x (2 per m g /kg-d) x (0.0001 m g /kg-d)

x (3 yr/70 yr)

= 2.6 x 10 -5



### Risk = 11.4 x 10 -5 

### + 2.6 x 10 -5



### = 1.4 x 10 -4



### b. For com p arison, a sim ilar risk calculation for 5 years of exposure la te r in lif e ( a f t er

age 16) in w h ich the first 2 years of expos ure are double that of the next 3 years are

carried out without any adjustm e nt for potency:

Risk equals the sum of :

• Risk for first 2 years of adult exposure = 1 (ADAF) x (2 per m g /kg-d)

x (0.0002 mg/kg-d) x (2yr/70yr)

= 1.1 x 10 -5



### • Risk for final 3 years of adult expo sure = 1 (ADAF) x (2 per m g /kg-d)

x (0.0001 mg/kg-d) x (3yr/70yr)

= 0.9 x 10 -5



### Risk = 1.1 x 10 -5 

### + 0.9 x 10 -5



### = 2 x 10 -5



### 41Standard Rodent Cancer Study: Adult ExposureInutero&postnatalExposurePostnatal & Adult Exposure (Lifetime)In utero, postnatal, & Adult Exposure (Lifetime)Postnatal ExposurebirthmatingweaningpubertyAssessmentof tumorsFigure 1. Study designs.42Acute

Exposures Repeated or

Lifetim e

Exposures

Fi gure 2: Posteri or, unw ei ghted geomet ric means and 95% confidence intervals for the ratios of juvenile to

adult cancer potency for carcinogens acting pr imarily through a mutagenic mode of action. The t op panel i s

for repeat ed and l i f et i m e exposure st udi es (geom e t r i c m ean i n bl ack), t h e bot t o m panel i s for acut e exposure st udi es

m u tagens (geom e tric m ean in white). The horizontal lines to the left and ri ght of each geom etric m ean correspond to

95% confidence lim its. The vertical da rk line represents the inverse-varian ce weighted geom etric m ean of the

post e ri or geom et ri c m eans. The hori z ont al dark l i n e represent s t h e 95 th p e rcen tile o f th e u n w eig h t ed d i strib u tio n

, with th e v e rtical, d o tted lin e estab lish i n g it v a lu e.Bo x 1 : Us e f r a m e w o r k i n C a ncer Gu i d e l i n es

to es t a bli s h M O A ( s ) F i gu r e 3. F l ow cha r t f o r

ea rl y- l i f e r i s k as s e s s ment us ing m o de of action f r a m e w o r k

M O A s u f f ic ie n t ly

s uppo r t ed i n a n im a l s ?

MO A r e l e v a n t t o

hu ma n s ?

F l ag li f e s t age( s) o r p o pul at i o n ( s ) t h at

co ul d b e s u s c ept i bl e ( b a s ed o n

i n f o r m at i o n a b o u t t h e s p ec i f i c MO A )

f o r do s e

- r es po n s e an a l y s i s

. Bo x 2 : U s e lin e a r

ex t r apo l at i o n as a de f a u l t

.

D e t e r m i n e e x t r apo l at i o n

ba s e d o n in f o r m a t i o n

ab o u t s p ec i f i c M O A

. Bo x 3 : M o del u s i ng M O A o r

us e R f D / R f C m et h o d as def a u l t

. A d j u st m e n t s f o r su sc ep t i ble

li f

e s t age s o r p o pul at i o n s ar e par t

o f

t h e pr o ces s

. No

L i ne ar i t y due t o

m

u t a ge ni c M O A M O A c a n no t

be d e t e r m i n e d

N o n lin e a r Yes

Ye s

Li n e a r , b u t

no n m u t a g e n i c

MOA Bo x 4 : Us e t h e s a m e l i n e a r

ex t r apo l at i o n f

o r a l l l i f e s t ages

, unl e s s h a v e c h e m i c a l

- s pec i fi c

in f o r m a t i o n o n li f e s t a g e s o r

po pul at i o n s

.

W e re ch em i c al -s p e ci fi c d a t a a v ai l a b l e

in M O A a n a l ys is t o e v a l u a t e d i f f e r e nc e s

be t w e e n a d u l t s a n d j u ve n i l e s ( m o r e

, l e ss, o r t h e sa m e su s c e p t i b i l i t y ) ? Bo x 5 : D e v e l o p ch em i cal

sp e c i f i c r i sk e s t i m a t e s

in c o r p o r at in g l i f e s t a g e

s u s c e p ti b i l i ty

. Ye s

Bo x 6 : E a r l y

- l i fe su s c e p t i b i l i t y a ssu m e d

. A p p l y

age- depe n d e n t ad j u s t m e n t f act o r s ( A DA F s ) as

appr o p r i at e to de v e l o p r i s k es t i m a t e s

. No Supp l e m e n t a l G u i d a n c e f o r E a rl y

- L i f e E x p o s u re s N o f u r t h e r an al y s i s of

tu m o r s

.Table 1a. Chemicals that have been found to have carcinogenic effects from prenatal or postnatal exposure inanimals as identified indifferent review articles



### A-1Chemical nameReview articles including prenatal and postnatal exposureChemicalsselected forquantitativeanalysisFujii(1991)McClainet al.(2001)Andersonet al.(2000)Della PortaandTerracini(1969)Otherliterature4-Acetylaminobiphenyl (AAB)X4-Aminoazobenzene (AB)X3-Amino-1,4,-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1)X2-Aminodipyridol[1,2-a:3',2'-d]imidazole (Glu-P-2)X2-Amino-6-methyldipyridol[1,2-a:3',2'-d]imidazole (Glu-P-1)X3-Amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2)XAmitroleXArsenicX5-AzacytidineX3'-Azido-3'-deoxythymidine (AZT)XAzoxymethaneXBenz[a]anthraceneXBenzidineXXBenzo[a]pyrene (BaP)XXX1­ (4'Bromophenylazo)-1-phenyl-1-hydroperoxymethane (BPH)XN-Butyl-N-(3-carboxypropyl)nitrosamine (BCPN)XN-Butyl-N-(3 hydroxbutyl)nitrosamine (BBN)XButylnitrosourea (BNU)XCyclophosphamideXDibenz[a,h]anthracene (DBA)XXDibutylnitrosamine (DBN)XDichlorodiphenyltrichloroethane (DDT)XDieldrinX2-Diethylaminoethyl-2,2-dephenylvalerate hydrochloride(SKF 525A)XTable 1a. Chemicals that have been found to have carcinogenic effects from prenatal or postnatal exposure inanimals as identified in different review articles (continued)Chemical nameReview articles including prenatal and postnatal exposureChemicalsselected forquantitativeanalysisFujii(1991)McClainet al.(2001)Andersonet al.(2000)Della PortaandTerracini(1969)OtherliteratureDiethylnitrosamine (DEN)XXXDiethylstilbesterol (DES)X4-DimethylaminoazobenzeneX1,2-Dimethylhydrazine (DMH)X7,12-Dimethylbenz[a]anthracene (DMBA)XXXXDimethylnitrosamine (DMN)XXXX5',5'-Diphenylhydantoin (DPH)XEstradiolXX6-Ethoxy-2,2,4-trimethyl-1,2-dihydroquinoline (Santoquin)XEthylene thiourea (ETU)XEthyl methane sulphonateXEthylnitrosobiuretXEthylnitrosourea (ENU)XXN-2-Fluorenylacetamide (FAA)XXGenisteinX3-Hydroxyl-4-acetylaminobiphenyl (N-OH-AAB)XN-2-Hydroxy-N-2-fluorenylacetamide (N-OH-FAA)X2-Hydroxypropyl-propylnitrosamineX9-MethylanthraceneXMethyl-2-benzylhydrazineXMethylcholanthreneXX3-Methyl-4-dimethylaminoabenzene (3'ME-DAB)X4-(Methylnitrosoamino)-1-(3-pyridyl)-1-butanone (NNK)XMethylnitrosourea (NMU)XMethylnitrosourethaneX1-Methyl-3-nitro-1-nitrosoguanidine (MNNG)X



### A-2Table 1a. Chemicals that have been found to have carcinogenic effects from prenatal or postnatal exposure inanimals as identified in different review articles (continued)Chemical nameReview articles including prenatal and postnatal exposureChemicalsselected forquantitativeanalysisFujii(1991)McClainet al.(2001)Andersonet al.(2000)Della PortaandTerracini(1969)Otherliterature2-NaphthylamineX2-NaphthylhydroxyamineXNickel acetateXN-NitrosobuylamineX4-Nitroquinoline-1-oxideXXN-Nitrosomethyl(2-oxopropyl)amineX2-Oxopropyl-propylnitrosamineX1-Phenyl-3,3',-dimethylhydrzineX1-Phenyl-3,3,-dimethyltriazeneXPolybrominated biphenyls (PBBs)XSafrole (3,4-methylenedioxyally benzene)XXXSootXSterigmatocystinXTamoxifenX1,3,5-Trimethyl-2,4,6-tris[3,5-di-tert-butyl-4-hydroxybenzyl]benzene (Ionox 33)XUrethane (ethyl carbamate)XXXVinyl chlorideX



### A-3

### Table 1b. List of chemicals considered in this analysis. (These are chemicals

for which both early-life and adult ex posure are reported in the same animal

experiment.)

Chemical References Study type Mutagenic

mode of action

Amitrole Vesselinovitch (1983) Repeat dosing

Benzidine Vesselinovitch et al. (1975b) Repeat dosing X

Benzo[ a ]pyrene (BaP) Vesselinovitch et al. (1975a) Acute exposure X

Dibenzanthracene (DBA) Law (1940) Acute exposure X

Dichlorodiphenyltrichloroethane Vesselinovitch et al. (1979) Repeat dosing

(DDT) Lifetime exposure

Dieldrin Vesselinovitch et al. (1979) Repeat dosing

Lifetime exposure

Diethylnitrosamine (DEN) Peto et al. (1984) Lifetime exposure X

Vesselinovitch et al. (1984) Acute exposure

Dimethylbenz[ a ]anthracene

(DMBA) Meranze et al. (1969) Acute exposure X

Pietra et al. (1961) Acute exposure

Walters (1966) Acute exposure

Dimethylnitrosamine (DMN) Hard (1979) Acute exposure X

Diphenylhydantoin, 5,5- (DPH) Chhabra et al. (1993b) Repeat dosing

Lifetime exposure

Ethylnitrosourea (ENU) Naito et al. (1981) Acute exposure X

Vesselinovitch et al. (1974) Acute exposure

Vesselinovitch (1983) Acute exposure

Ethylene thiourea (ETU) Chhabra et al. (1992) Repeat dosing

Lifetime exposure

3-Methylcholanthrene (3-MU) a Klein (1959) Repeat dosing X

Methylnitrosourea (NMU) Terracini and Testa (1970) Acute exposure X

Terracini et al. (1976) Acute exposure

Polybrominated biphenyls Chhabra et al. (1993a) Repeat dosing

(PBBs) Lifetime exposure

Safrole Vesselinovitch et al. (1979) Repeat dosing

Lifetime exposure X

Urethane Chieco-Bianchi et al. (1963)

Choudari Kommineni et al. (1970) Acute exposure

Acute exposure X

De Benedictis et al. (1962) Acute exposure

Fiore-Donati et al. (1962) Acute exposure

Klein (1966) Acute exposure

Lifetime exposure

Liebelt et al. (1964) Acute exposure

Rogers (1951) Acute exposure

Vinyl chloride (VC) Maltoni et al. (1984) Repeat dosing X

a Formerly known as 20-methylcholanthrene.



### A-4Table 2. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult repeated exposuresChemicalSpecies(strain)TargetsiteAge whenfirst dosedDoseroute,# dosesDoseDuration ofexposureAge atdeathTumorsaM FCommentsReferenceAmitroleMiceliverControlNoneControl:N/A90 weeks1/980/96Incidences areVesselinovitch(B6C3F1)0 ppm(1%)(0%)mice with(1983)Gestationday 12Diet, tomothers500 ppmGestation day12 to delivery6/74(8%)b0/83(0%)badenomas orcarcinomas.NewbornDiet, tomothers500 ppmBirth untilweaning10/45(22%)b0/55(0%)bAt weaningDiet, tooffspring500 ppmFrom weaningto 90 weeks20/55(36%)b9/49(18%)bBenzidineMiceliverControlNoneControl:N/A90 weeks1/980/100HigherVesselinovitch et(B6C3F1)0 ppm(1%)(0%)sensitivity inal. (1975b)Gestationday 12Diet, tomothers150 ppmGestation day12 to delivery17/55(31%)c2/62(3%)dmales duringperinatalperiod, inVesselinovitch etal. (1979a)



### A-5NewbornDiet, tomothers150 ppmBirth untilweaning62/65(95%)c2/43(5%)dfemales duringadulthood.At weaningDiet, tooffspring150 ppmFrom weaningto 90 weeks22/50(44%)c47/50(94%)cIncidences aremice withGestationday 12Diet, tomothers150 ppmGestation day12 until49/49(100%)c12/48(25%)cadenomas orcarcinomas.weaningGestationDiet, to150 ppmGestation day50/5047/50day 12mothers12 until 90(100%)c(94%)cweeksDDTMiceliverControlNoneControl:N/A90 weeks1/50—VesselinovitchetDichlorodiphenyl­(B6C3F1)0 ppm(2%)al. (1979b)trichloroethaneWeek 1Gavage,daily230 μgWeeks 1–45/49(10%)d—Week5Diet,daily150 ppmWeeks 5–908/49(16%)d—Week 1Gavage,230 μgWeeks 1–9010/50—daily until(20%)c4 weeks,150 ppmthen in(diet)dietTable 2. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult repeated exposures(continued)DoseTumorsaSpeciesTargetAge whenDuration ofAge atroute,Chemical(strain)sitefirst dosedDoseexposuredeathCommentsReferenceMF# dosesDieldrinMiceliverControlNoneControl:N/A1/58—Vesselinovitchet(B6C3F1)90 weeks0 ppm(2%)al. (1979b)Week 1Gavage,12.5μgWeeks 1–43/46—daily(7%)bWeek 5Diet,10 ppmWeeks 5–907/60—daily(12%)bWeek 1Gavage,12.5μgWeeks 1–9021/70—daily until(30%)a4 weeks,10 ppmthen indietDENeRatsliverControlControlN/A29/384Highest tumorPeto et al. (1984)Diethylnitrosamine(Colworth)(8%)rate when dosedat earlier ages.Week 3From week 36105/180drinkingDiet (in16 differentuntil deathmonths–(58%)bdosesIncidents arewater),combinedf3 yearsrats withdailyadenomas oruntil deathWeek 6From week 6714/1440carcinomas.(50%)bWeek 20From week 2076/180until death(42%)besophagusControlControlN/A0/384(0%)Week 3Diet (inFrom week 377/180drinking16 differentdosesuntil death(43%)bwater),combinedgWeek 6From week 6663/1440dailyuntil death(46%)bWeek 20From week 2088/180until death(49%)b



### A-6Table 2. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult repeated exposures(continued)



### A-7ChemicalSpecies(strain)TargetsiteAge whenfirst dosedDoseroute,# dosesDoseDuration ofexposureAge atdeathTumorsaCommentsReferenceMFDPHRatsliverControlControl0 ppmN/A2 years0/500/50In rats, perinatalChhabra et al.Diphenylhydantoin,(F344/N)(0%)(0%)exposure ranged(1993b)5,5-PerinatalDiet,daily630ppmPerinatalthrough 8weeks1/50(2%)d0/49(0%)dfrom 63 to 630ppm, and adultexposures rangedfrom 240 to 2,400ppm.8 weeks800 ppm8 weeks–2years2/50(4%)d1/50(2%)d8 weeks2,400 ppm8 weeks–2years4/50(8%)d1/50(2%)dIn mice, perinatalexposure rangedfrom 21 to 210ppm. Adultexposure rangedfrom 30 to 300ppm in males andPerinatal630–800Perinatalthrough 2 years1/49(2%)d0/50(0%)dPerinatal630–2,400ppmPerinatalthrough 2 years5/49(10%)c0/50(0%)dMice(B6C3F1)liverControlControlmale0 ppmN/A2 years29/50(58%)60 to 600 ppm infemales.Tumor incidencesare animals withadenomas orcarcinomas.PerinatalDiet, male210ppmPerinatalthrough 8weeks33/50(66%)d8 weeks100 ppm8 weeks–2years29/49(59%)d8 weeks300 ppm8 weeks–2years26/49(53%)dPerinatal210–100ppmPerinatalthrough 2 years35/49(71%)dPerinatal210–300Perinatal41/50ppmthrough 2 years(82%)cControlControlfemale0 ppmN/A2 years5/48(10.4%)dPerinatalDiet,female210ppmPerinatalthrough 812/49(24.5%)dweeks8 weeks200 ppm8 weeks–214/49years(28%)c8 weeks600 ppm8 weeks–230/50years(60%)cPerinatal210–200Perinatal16/50ppmthrough 2 years(32%)cPerinatal210–600Perinatal34/50ppmthrough 2 years(68%)cTable 2. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult repeated exposures(continued)



### A-8ChemicalSpecies(strain)TargetsiteAge whenfirst dosedDoseroute,# dosesDoseDuration ofexposureAge atdeathTumorsaCommentsReferenceMFETURatsthyroidControlControl0 ppmN/A2 years1/493/50Tumor incidencesChhabra et al.Ethylene thiourea(F344/N)(2%)(6%)are animals with(1992)PerinatalDiet,daily90ppmPerinatalthrough 84/49(8%)d3/50(6%)dadenomas orcarcinomas.weeks8 weeks83 ppm8 weeks–2years12/46(26%)c7/44(16%)d8 weeks250 ppm8 weeks–237/5030/49years(74%)c(61%)cPerinatal90–83ppmPerinatalthrough 2 years13/50(26%)c9/47(19%)dPerinatal90–250 ppmPerinatal48/5037/50through 2 years(96%)(74%)Mice(B6C3F1)liverControlControl0 ppmN/A2 years20/49(41%)4/50(8%)PerinatalDiet,daily330ppmPerinatalthrough 813/49(26.5%)d5/49(10%)dweeks8 weeks330 ppm8 weeks–232/5044/50years(64%)c(88%)c8 weeks1,000 ppm8 weeks–246/5048/50years(92%)c(96%)cPerinatal330–330Perinatal34/4946/50ppmthrough 2 years(69%)c(92%)cPerinatal330–1,000Perinatal47/4949/50ppmthrough 2 years(6%)c(98%)cthyroidControlControl0ppmN/A1/50(2%)0/50(0%)PerinatalDiet,daily330 ppmPerinatalthrough 81/46(2%)d1/49(2%)dweeks8 weeks330 ppm8 weeks–2years1/49(2%)d2/50(4%)d8 weeks1,000 ppm8 weeks–229/5038/50years(58%)c(76%)cTable 2. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult repeated exposures(continued)



### A-9ChemicalSpecies(strain)TargetsiteAge whenfirst dosedDoseroute,# dosesDoseDuration ofexposureAge atdeathTumorsaCommentsReferenceMFETUEthylene thioureaPerinatal330–330ppmPerinatalthrough 2 years2/48(4%)d10/49(20%)c(continued)Perinatal330–1,000Perinatal35/4938/50ppmthrough 2 years(71%)c(76%)cpituitaryControlControl0ppmN/A0/44(0%)11/47(23%)PerinatalDiet,daily330ppmPerinatalthrough 80/42(0%)d11/48(23%)dweeks8 weeks330 ppm8 weeks–2years0/42(0%)d19/49(39%)d8 weeks1,000 ppm8 weeks–28/4126/49years(19.5%)c(53%)cPerinatal330–330ppmPerinatalthrough 2 years0/45(0%)d26/47(55%)cPerinatal330–1,000ppmPerinatalthrough 2 years4/39(10%)d24/47(51%)cTable 2. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult repeated exposures(continued)



### A-10AgeAge at deathTumor incidenceChemicalSpecies(strain)TargetsitewhenfirstdosedDose route,# dosesDoseDuration ofexposureMFMFReference3-MethylcholanthreneMiceliverControlgavage, 3×NANA475 days480 days3/390/36Klein (1959)(formerly known as 20-(Albino)per week(7.7%)(0%)methylcholanthrene)8 days0.25 mg/g10×311 days321 days21/25(84%)b7/30(23.3%)b90 days0.25 mg/g10×330 days366 days1/26(3.8%)b0/29(0%)dlungControlNANA475 days480 days17/39(43.6%)14/36(38.9%)8 days0.25 mg/g10×311 days321 days25/25(100%)b28/30(93.3%)b90 days0.25 mg/g10×330 days366 days25/26(96.2%)b27/29(93.1%)bforestomachControlNANA475 days480 days0/39(0%)0/36(0%)8 days0.25 mg/g10×311 days321 days12/25(48%)b12/30(40%)b90 days0.25 mg/g10×330 days366 days13/26(50%)b8/29(27.6%)bskinControlNANA475 days480 days0/39(0%)0/36(0%)8 days0.25 mg/g10×311 days321 days4/25(16%)b4/30(13.3%)b90 days0.25 mg/g10×330 days366 days1/26(3.8%)b1/25(4%)bTable 2. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult repeated exposures(continued)



### A-11ChemicalSpecies(strain)TargetsiteAge whenfirst dosedDoseroute,# dosesDoseDuration ofexposureAge atdeathTumorsaCommentsReferenceMFPBBsRats (F344/N)livergControlControl0 ppmN/A2 years1/500/50Findings suggestChhabra et al.Polybrominated(2%)(0%)that combined(1993a)biphenylsPerinatalDiet10ppmPerinatal–8weeks5/50(10%)d0/50(0%)dperinatal and adultexposure increasesPBB-relatedhepatocellularcarcinogenicityrelative to adultonly exposure inmice and femalerats.8 weeks10 ppm8 weeks–2years12/49(24%)c12/50(24%)c8 weeks30 ppm8 weeks–2years41/50(82%)c39/50(78%)cPerinatal10–10ppmPerinatal–216/5039/50years(32%)c(78%)cApparentassociationbetweenincreasingincidences ofMCL andexposure to PBBin male andfemale rats.Tumor incidencesare animals withadenomas orcarcinomas.Perinatal10–30 ppmPerinatal–2years41/50(82%)c47/50(94%)cMononuclearcellleukemia(MCL)ControlControl0 ppmN/A2 years25/50(50%)14/50(28%)PerinatalDiet10ppmPerinatal–8weeks31/50(62%)d13/50(26%)d8 weeks10 ppm8 weeks–2years33/50(66%)c22/50(44%)d8 weeks30 ppm8 weeks–2years31/50(62%)d23/50(46%)cPerinatal10–10ppmPerinatal–2years37/50(74%)c27/50(54%)cPerinatal10–30 ppmPerinatal–237/5025/50years(74%)c(50%)cMice (B6C3F1)livergControlControl0 ppmN/A2 years16/50(32%)5/50(10%)PerinatalDiet30ppmPerinatal–840/5021/50weeks(80%)c(42%)c8 weeks10 ppm8 weeks–248/4942/50years(98%)c(84%)c8 weeks30 ppm8 weeks–248/5047/48years(96%)c(98%)cPerinatal10ppmPerinatal–246/4944/50years(94%)c(88%)cPerinatal30–30 ppmPerinatal–250/5047/47years(100%)c(100%)cTable 2. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult repeated exposures(continued)



### A-12ChemicalSpecies(strain)TargetsiteAge whenfirst dosedDoseroute,# dosesDoseDuration ofexposureAge atdeathTumorsaCommentsReferenceMFSafroleMiceliverControlNoneNoneN/A90 weeks3/1000/100Highest tumor rateVesselinovitch(B6C3F1)(3%)(0%)in males due toet al. (1979b)Day 12 ofgestationGavage, tomothers120 μg/gbody weight4× (days 12,14, 16, 18)2/61(3%)d0/65(0%)dpreweaningtreatment.Highest tumor ratein females due toNewbornGavage, tomothers, on120 μg/gbody weightFrom birth untilweaning28/83(34%)c2/80(3%)dalternatesusceptibility indaysadulthood.Tumor incidencesare mice withadenomas orcarcinomas.At weaningGavage, tooffspring, 2×weekly120 μg/gbody weightFrom weaninguntil 90 weeks4/35(11%)d22/36(61%)cDay 12 ofgestationGavage, tomothers,alternatedays120 μg/gbody weightFrom gestationuntil weaning22/68(32%)b1/72(1%)bDay 12 ofgestationGavage, tomothers,alternate120 μg/gbody weightFrom gestationuntil 90 weeks19/37(51%)b37/46(80%)bdays untilweaning;Gavage, tooffspring, 2×weeklyUrethaneMice(B6AF1/J)liver1 weekgavage2.5 mg/pup1×39–40weeksTumor incidenceaNo tumor data forcontrols.Klein (1966)MF12/37(33%)b0/40(0%)b1 week2.5 mg/pup16×(1× at 1 week;3× weekly for 5weeks39 weeks11/33(33%)b0/31(0%)bbeginning at 4wks of age)4 weeks2.5 mg/pup15×(3× weekly for5 weeks41 weeks0/37(0%)b0/31(0%)bbeginning at 4weeks of age)Table 2. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult repeated exposures(continued)DoseTumorsaSpeciesTargetAge whenDuration ofAge atroute,Chemical(strain)sitefirst dosedDoseexposuredeathCommentsReferenceMF# dosesVCRats (Sprague­liverControlControl0 ppmN/A1350/220/29Higher tumor riskMaltoni et al.Vinyl chlorideDawley)angio­weeks(0%)(0%)when exposed at(1984)sarcomabirth, higher forNewbornInhalation6,000 ppm4 hrs/day,1245/1812/24females.5 days/wk,weeks(28%)b(50%)b5 weeks10,000ppm6/249/20(25%)b(45%)bWeek 136,000ppm4 hrs/day,1353/1710/255 days/wk, 52weeks(18%)b(40%)bweeks4/25(14%)b10,000ppm3/21(16%)bzymbalControlControl0ppmN/A1350/280/29glandweeks(0%)(0%)NewbornInhalation6,000ppm4 hrs/day,1241/121/175 days/wk,weeks(8%)b(6%)b5 weeks10,000ppm1/170/17(6%)b(0%)bWeek 136,000 ppm4 hrs/day,1353/294/305 days/wk, 52(13%)bweeksweeks(10%)b10,000ppm10/306/30(33%)b(20%)bleukemiaControlControl0ppmN/A1350/271/29weeks(0%)(3%)NewbornInhalation6,000ppm4 hrs/day,124N/A1/75 days/wk,weeks(14%)b5 weeks10,000ppm2/60/15(33%)b(0%)bWeek 136,000 ppm4 hrs/day,135N/A0/295 days/wk, 52weeks(0%)bweeks10,000ppm0/272/29(0%)b(7%)bnephro-ControlControl0 ppmN/A1350/220/29blastomaweeks(0%)(0%)NewbornInhalation6,000ppm4 hrs/day,1240/150/215 days/wk,weeks(0%)b(0%)b5 weeks10,000ppm0/190/17(0%)b(0%)b



### A-13Table 2. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult repeated exposures(continued)



### A-14ChemicalSpecies(strain)TargetsiteAge whenfirst dosedDoseroute,# dosesDoseDuration ofexposureAge atdeathTumorsaCommentsReferenceMFVCVinyl chlorideWeek 136,000 ppm4 hrs/day,5 days/wk, 52135weeks4/18(22%)b1/26(4%)b(continued)weeks10,000ppm3/21(14%)b2/25(8%)bangio-ControlControl0ppmN/A1350/290/29sarcomas:weeks(0%)(0%)other sitesNewbornInhalation6,000ppm4 hrs/day,5 days/wk,124weeks1/15(7%)b0/21(0%)b10,000ppm5 weeks0/19(0%)0/17(0%)bWeek 136,000 ppm4 hrs/day,5 days/wk, 52135weeks1/29(3%)b2/30(7%)b10,000ppmweeks2/30(7%)b1/30(3%)bangiomasandControlControl0ppmN/A135weeks0/28(0%)2/29(7%)bfibromas:other sitesNewbornInhalation6,000ppm4 hrs/day,5 days/wk,124weeks1/15(7%)b0/21(0%)b10,000ppm5 weeks2/19(11%)b1/17(6%)bWeek 136,000 ppm4 hrs/day,5 days/wk, 52135weeks2/29(7%)b2/30(7%)b10,000ppmweeks2/29(7%)b1/29(3%)bhepatomaControlControl0ppmN/A135weeks0/19(0%)0/28(0%)NewbornInhalation6,000ppm4 hrs/day,5 days/wk,124weeks9/18(50%)b11/24(46%)b10,000ppm5 weeks13/24(54%)b7/20(35%)bWeek 136,000 ppm4 hrs/day,5 days/wk, 52135weeks0/10(0%)b1/17(6%)b10,000ppmweeks1/8(13%)b0/16(0%)bTable 2. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult repeated exposures(continued)DoseTumorsaSpeciesAge whenDuration ofAge atroute,Chemical(strain)Target sitefirst dosedDoseexposuredeathCommentsReference# dosesMFVCskinControlControl0ppmN/A1350/201/29Vinyl chloridecarcinomasweeks(0%)(3%)(continued)NewbornInhalation6,000ppm4 hrs/day,1241/101/145 days/wk,(7%)b5 weeksweeks(10%)b10,000ppm0/15(6%)b1/16(0%)bWeek 136,000 ppm4 hrs/day,1350/152/195 days/wk, 52(11%)bweeksweeks(0%)b10,000ppm1/21(15%)b2/13(5%)bneuro-ControlControl0ppmN/A1350/220/29blastomaweeks(0%)(0%)NewbornInhalation6,000ppm1240/180/294 hrs/day,5 days/wk,weeks(0%)b(0%)b



### A-1510,000ppm5 weeks0/22(0%)b0/19(0%)bWeek 136,000 ppm4 hrs/day,5 days/wk, 52135weeks2/21(10%)b1/27(4%)b10,000ppmweeks2/22(9%)b5/26(19%)baWhere not delineated by gender, data combined by study authors or gender not specified. Where percentages only are given, number of subjects not specified.bNot evaluated by authors.cSignificant compared with controls.dEvaluated but not significant compared with controls.eReported as NDEA (N-nitrosodiethylamine) in the original document.fResults from each dose are not available.gTumors were adenomas or carcinomas.Table 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure



### A-16ChemicalSpecies(strain)TargetsiteAge whenfirst dosedDoseroute,# dosesDoseDuration ofexposureAge atdeathTumorsaCommentsReferenceMFBaPMiceliverControlControlNoneN/A142 weeks7/1001/100In general, hepatomasVesselinovitchBenzo[a]pyrene(B6C3F1)(7%)(1%)developed withet al. (1975a)Day 1i.p.75 μg/gbody weight1×86weeks(m)129 weeks(f)26/47(55%)b3/45(7%)bsignificantly higherincidence (p<0.01) inmice that were treatedwithin 24 hours of birthor at 15 days of age150 μg/gbody weight1×81weeks(m)121 weeks51/63(81%)b8/45(18%)bthan they did insimilarly treatedanimals at 42 days of(f)age.+ higher for males.Day 15i.p.75 μg/gbody weight1×93weeks(m)116 weeks(f)36/60(60%)b4/55(7%)b150 μg/gbody weight1×81weeks(m)32/55(58%)b4/55(7%)b90 weeks (f)Day 42i.p.75 μg/gbody weight1×108weeks(m)7/55(13%)b0/47(0%)b150 μg/gbody weight1×87weeks(m)4/47(9%)b0/46(0%)bMiceliverControlControlNoneN/A142 weeks8/1001/100+ higher for males.(C3AF1)(8%)(1%)Day 1i.p.75μg/gbody weight1×80weeks(m)91 weeks (f)21/62(34%)b1/45(2%)b“Age at death” is theaverage age at whichtumors were observed.150 μg/gbody weight1×69weeks(m)24/52(46%)b1/56(2%)b701 weeks(f)Day 15i.p.75 μg/gbody weight1×90weeks(m)15/56(27%)b1/49(2%)b102 weeks(f)Table 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)



### A-17ChemicalSpecies(strain)TargetsiteAge whenfirst dosedDoseroute,# dosesDoseDuration ofexposureAge atdeathTumorsaCommentsReferenceMFBaPBenzo[a]pyrene150μg/gbody weight1×77weeks(m)12/53(23%)b1/57(2%)b(continued)62 weeks (f)Day 42i.p.75μg/gbody weight1×0/30(0%)b0/32(0%)b150 μg/gbody weight1×79weeks(m)1/32(3%)c0/40(0%)bMicelungControlControlControlN/A142 weeks13/1009/100Both sexes developed(B6C3F1)(13%)(9%)lung tumors with higherDay 1i.p.75 μg/gbody weight1×103weeks(m)126 weeks(f)20/47(43%)b22/45(49%)bincidence when treatedwith BaP at birth than at15 or 42 days of age(p<0.05).150 μg/gbody weight1×84weeks(m)37/63(59%)b28/45(62%)b112 weeks(f)Day 15i.p.75 μg/gbody weight1×103weeks(m)15/60(25%)b18/55(33%)b122 weeks(f)150 μg/gbody weight1×82weeks(m)20/55(36%)b18/45(40%)b101 weeks(f)Day 42i.p.75 μg/gbody weight1×119weeks(m)20/55(36%)b12/47(26%)b131 weeks(f)150 μg/gbody weight1×95weeks(m)18/47(38%)b8/46(17%)b118 weeks(f)Table 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)TumorsaDoseSpeciesTargetAge whenDuration ofAge atroute,Chemical(strain)sitefirst dosedexposuredeathDoseCommentsReference# dosesMFBaPMicelungControlControlNoneN/A142 weeks60/10050/100Vesselinovitch etBenzo[a]pyreneOf the two mouse(C3AF1)(60%)(50%)strains tested, C3AF1al. (1975a)(continued)mice developedDay 1i.p.42/4575μg/g1×78weeks58/62significantly morebody weight(m)(93%)b(93%)btumors than did the82 weeks (f)B6C3F1mice150 μg/g(p<0.001).body weight1×70weeks48/5252/56(m)(92%)b(93%)b73 weeks (f)75μg/g1×87weeks46/49body weightDay 15i.p.52/56(m)(93%)b(94%)b98 weeks (f)150 μg/g1×75weeks50/5352/57body weight(m)(94%)b(91%)b79 weeks (f)Day 42i.p.75μg/g1×91weeks28/3028/32body weight(m)(93%)b(87%)b93 weeks (f)150 μg/g1×85weeks28/3236/40body weight(m)(87%)b(90%)b83 weeks (f)DBAMicelungControlControlNoneN/A228 days1/31Law(1940)Dibenzanthracene(Caracul × P(3.2%)stock)Day 1i.p.4 mg per1×181days24/24cm3vehicle(100%)b2 monthss.c.4 mg per1×189days2/29cm3vehicle(6.9%)b



### A-18Table 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)



### A-19ChemicalSpecies(strain)TargetsiteAge whenfirst dosedDoseroute,# dosesDoseDuration ofexposureAge atdeathTumorsaCommentsReferenceMFDENDiethylnitrosamineMice(B6C3F1)liverControlControlVehicle(0.01 mLtrioctanoin/g4×142weeks(m)7/98(7%)1/100(1%)Animals treated asnewborns and infantsdeveloped significantlyVesselinovitch etal. (1984)body weight)137more liver tumors thanweeks (f)animals that weretreated as young adults.Newborns and infantfemales developed livertumors at a later ageDay 1i.p. (3-, 6­and 6-dayintervals)1.5 μg/gbody weight4×67weeks(m)90 weeks(f)37/51(73%)b45/64(70%)b3 μg/g bodyweight4×65weeks(m)80 weeks40/58(69%)b44/65(68%)bthan similarly treatedmales.(f)Incidences formalignant tumors only.Day 151.5 μg/gbody weight4×86weeks(m)41/57(72%)b40/71(56%)b117weeks (f)3 μg/g bodyweight4×76weeks(m)48/69(70%)b46/62(74%)b96 weeks(f)Day 421.5 μg/gbody weight4×117weeks(m)1359/49(18%)b1/47(2%)bweeks (f)3 μg/g bodyweight4×123weeks(m)1336/38(16%)b4/57(7%)bweeks (f)Table 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)



### A-20ChemicalSpecies(strain)TargetsiteAgewhenfirstdosedDoseroute,# dosesDoseDuration ofexposureAge atdeathTumorsaCommentsReferenceMFDENMiceliverControlControlVehicle (0.14×1238/991/97Highest tumor rateVesselinovitch etDiethylnitrosamine(C3AF1)trioctanoin/gweeks(8%)(1%)when dosed at earlyal. (1984)(continued)body weight)(m)ages.131weeks(f)Newborns and infantDay 1i.p. (3-, 6­and 6-dayintervals)1.5 μg/gbody weight4×64weeks(m)84 weeks(f)23/32(72%)b11/39(28%)bfemales developed livertumors at a lowerincidence than similarlytreated males.+ higher for males.3 μg/g bodyweight4×59weeks(m)39/58(67%)b26/50(52%)b76 weeks(f)Day 151.5μg/gbody weight4×82weeks(m)22/46(48%)b8/65(12%)b102weeks (f)3 μg/g bodyweight4×74weeks(m)35/54(65%)b22/62(35%)b94 weeks(f)Day 421.5μg/gbody weight4×105weeks(m)10612/56(22%)b0/53(0%)bweeks (f)3 μg/g bodyweight4×105weeks(m)1039/57(16%)b0/56(0%)bweeks (f)Mice(B6C3F1)lungControlControlVehicle (0.1trioctanoin/gbody weight)4×142weeks(m)137weeks (f)13/98(13%)9/100(9%)The mice treated asnewborns showed lungtumors earlier thananimals exposed atother times. It is notknown whether this wasdue to actual earlieremergence of tumors orTable 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)



### A-21ChemicalSpecies(strain)TargetsiteAgewhenfirstdosedDoseroute,# dosesDoseDuration ofexposureAge atdeathTumorsaCommentsReferenceMFDENDiethylnitrosamineDay 1i.p. (3-, 6­and 6-day1.5 μg/gbody weight4×70weeks(m)29/51(57%)b49/64(77%)bto their earlier detectioncaused by shorter(continued)intervals)91 weekssurvival.(f)3 μg/g bodyweight4×68weeks(m)34/58(59%)b42/65(65%)b81 weeks(f)Day 151.5μg/gbody weight4×87weeks(m)51/57(89%)b61/71(86%)b115weeks (f)3 μg/g bodyweight4×77weeks(m)51/69(74%)b53/62(85%)b97 weeks(f)Day 421.5μg/gbody weight4×123weeks(m)12938/49(78%)b38/47(81%)bweeks (f)3 μg/g bodyweight4×121weeks(m)12733/38(87%)b43/57(75%)bweeks (f)Mice(C3AF1)lungControlControlVehicle (0.1trioctanoin/g4×142weeks60/99(61%)50/97(52%)Of the two strains,C3AF1mice developedbody weight)(m)137weekslung tumors with ahigher incidence and(f)multiplicity thanDay 1i.p. (3-, 6­and 6-day1.5 μg/gbody weight4×65weeks(m)30/32(94%)b38/39(97%)bB6C3F1hybrids.intervals)84 weeks(f)3 μg/g bodyweight4×59weeks(m)49/58(84%)b46/50(92%)b76 weeks(f)Table 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)AgeTumorsaDosewhenSpeciesTargetroute,Duration ofAge atfirstChemical(strain)site# dosesDoseexposuredeathCommentsReferencedosedMFDENDay 151.5μg/g4×80weeks42/4661/65Diethylnitrosaminebody weight(m)(91%)b(94%)b(continued)101weeks (f)3 μg/g body4×74weeks57/62weight50/54(m)(92%)b92 weeks(93%)b(f)Day 421.5μg/g4×10452/53body weight55/56weeks(98%)b(98%)b(m)110weeks (f)3 μg/g body4×10154/56weight56/57weeks(96%)b(98%)b(m)102weeks (f)MiceliverControlControlNoneN/A90 weeks1/980/96Infant animals of bothVesselinovitch(B6C3F1)sexes (Day 15) were(1%)(0%)and Mihailovichmore sensitive than(1983)Gestationi.p.1.5μg/g1×2/501/51similarly exposedday 18body weight(4%)b(2%)badults.Day 15i.p.(3-, 6-1.5μg/g4×47/5160/64and 6-daybody weight(92%)b(94%)bintervals)Day 421.5μg/g4×13/493/47body weight(26%)b(6%)bDay 1i.p.1.5μg/g1×73weeks15/59—At the 1.5-μg doseVesselinovitchetbody weight(25%)blevel, 1-day-old miceal. (1979a)developed significantly5 μg/g body1×29/45—fewer liver tumors thanweight(64%)bsimilarly treated infants10 μg/g(Day 15) (p<0.025).body weight1×24/25—(96%)bTumor incidence inDay 15i.p.1.5μg/g1×13/24—treated groups versusbody weight(54%)bcontrols was not5 μg/g bodyevaluated.weight1×40/54—(74%)b



### A-22Table 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)AgeTumorsaDosewhenSpeciesTargetroute,Duration ofAge atfirstChemical(strain)site# dosesDoseexposuredeathCommentsReferencedosedMFDEN10μg/g1×25/25—Diethylnitrosaminebody weight(100%)b(continued)DMBARatsmammaryDay 20Gavage10 mg/100 g1×Week25—3/636 of 42 (86%) animalsRusso et al. (1979)Dimethyl-dosed at age 20 daysbenz[a]anthracene(Sprague­adeno­body weight(50%)bdied soon after.Dawley)sarcomaDay 3010 mg/100 g1×Week26—14/15Highest number ofbody weight(93%)btumors per animal wasDay 40in the 46-day group,body weight10 mg/100 g1×Week27—8/9with decreasingnumbers in the older(89%)bDay 4610 mg/100 g1×Week28—8/8animals.body weight(100%)bDay 5510 mg/100 g1×Week29—33/34Animals were sacrificedbody weight22 weeks aftertreatment.(97%)bDay 7010 mg/100 g1×Week32—5/8body weight(63%)bDay 14010 mg/100 g1×Week42—10/15body weight(67%)bDay 18010 mg/100 g1×Week47—14/26body weight(54%)bRatsmammaryControlControlNoneN/A170/220/25Highest tumor rate inMeranze et al.(Wistar)carcinomad5–8females exposed at 5–8months(0%)(0%)(1969)weeksweeks.ControlControlNoneN/A200/312/20Animals were observed26 weeksmonths(0%)(10%)for 16 months following< Week 2treatment.56Gavage0.5–1.0 mg1×Week 40–0/234/50(0%)b(8%)bWeek 5–815 mg1×Week 14–0/2314/2555(0%)b(56%)bWeek 2615 mg1×Week 32–0/344/2673(15%)b(0%)bRats (Wistar,mammaryWeek 5–8Gavage15 mg1×Week 14–0/210/22castrated)carcinoma55(0%)b(0%)bWeek 2615 mg1×Week 32–0/330/2673(0%)b(0%)b



### A-23Table 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)AgeTumorsaDosewhenSpeciesTargetroute,Duration ofAge atfirstChemical(strain)site# dosesDoseexposuredeathCommentsReferencedosedMFDMBARatsTotal tumorsControlControlNoneN/A170/220/25Total tumors includesDimethyl-(Wistar)5–8months(0%)(0%)leukemia.benz[a]anthraceneweeks(continued)ControlControlNoneN/A202/315/2026 weeksmonths(6%)(25%)< Week 20.5–1.0 mg1×Week 40–16/2336/5056Gavage(70%)b(72%)bWeek 5–815 mg1×Week 14–7/2316/2555(30%)b(64%)bWeek 2615 mg1×Week 32–12/3413/2673(50%)b(35%)bMicelungControl:ControlAqueous1×40weeks0/127/2315 μg DMBA gave riseWalters (1966)(BALB/c)Day 1s.c.gelatine(0%)(30%)to a significantly greaterincidence of lung14/1424/24Day 1s.c.15 μg1×40 weeksftumors when(100%)b(100%)badministered toWeek 2–3newborn mice than to(suckling)s.c.15μg1×42–4312/2316/22suckling or youngadults.(73%)bweeks(52%)bs.c.30 μg2×42–4314/1424/24(60 μg total)weeks(100%)b(100%)bAdultes.c.15μg1×48-496/1215/33weeks(50%)b(45%)bs.c.30μg2×48-499/1021/23(60 μg total)weeks(90%)b(91%)bs.c.30 μg6×48-4912/1213/13(180 μgweeks(100%)b(100%)btotal)Mice(Swiss)lymphomaControlControlNoneN/A31–523/408Higher tumor rates atPietra et al. (1961)weeks(0.7%)younger age ofexposure.Day 1i.p.30–40 μg1×13–336/31weeks(19%)bOnly one treatmentDay 1group was exposed i.p.;weekss.c.30–40 μg1×12–278/27others were exposed by(30%)bs.c.injection..Week 8s.c.900 μg1×30 weeks1/13(8%)b



### A-24Table 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)AgeTumorsaDosewhenSpeciesTargetroute,Duration ofAge atfirstChemical(strain)site# dosesDoseexposuredeathCommentsReferencedosedMFDMBAMice (Swiss)lungControlControlNoneN/A31–524/408Dimethyl-weeks(0.9%)benz[a]anthraceneDay 1i.p.30–40 μg1×13–3324/31(continued)weeks(77%)bDay 1s.c.30–40 μg1×12–2723/27weeks(85%)bWeek 8s.c.900 μg1×30 weeks2/13(15%)bDMNRatskidneyDay 120 mg/kg1×1/33 (3)bIn the neonatal group,i.p.≥5Hard (1979)Dimethyl-(Wistar)carcinomamonthsthe dose was reduced toDay 2130 mg/kg1×5/39 (13)bnitrosamine20 mg/kg to achieveMonth 1approximatelyequivalent numbers of30 mg/kg1×2/33 (6)bMonth 1.530 mg/kg1×1/28 (4)bsurvivors.Month 230 mg/kg1×1/26 (4)bNo control group.Month 330 mg/kg1×10/27 (37)bMonth 430 mg/kg1×7/32 (22)bMonth 530 mg/kg1×0/14 (0)bRatskidneyDay 1i.p.20 mg/kg1×≥51/33 (3)b(Wistar)adenomamonthsDay 2130 mg/kg1×13/39 (33)bMonth 130 mg/kg1×11/33 (33)bMonth 1.530 mg/kg1×13/28 (48)bMonth 230 mg/kg1×11/26 (42)bMonth 330 mg/kg1×18/27 (67)bMonth 430 mg/kg1×17/32 (53)bMonth 530 mg/kg1×6/14 (43)b



### A-25Table 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)AgeTumorsaDosewhenSpeciesTargetroute,Duration ofAge atfirstChemical(strain)site# dosesDoseexposuredeathCommentsReferencedosedMFDMNRatskidneyDay 1i.p.20 mg/kg1×≥58/33 (24)bMesenchymal tumorsDimethyl-(Wistar)mesenchymalmonthswere most frequent inDay 2130 mg/kg1×18/39 (46)bnitrosaminetumorsthe three youngest age(continued)groups (z test,p < 0.001).Month 130 mg/kg1×23/33 (70)bMonth 1.530 mg/kg1×5/28 (19)bMonth 230 mg/kg1×2/26 (8)bMonth 330 mg/kg1×3/27 (11)bMonth 430 mg/kg1×7/32 (22)bMonth 530 mg/kg1×0/14 (0)bRatskidneyDay 1i.p.20 mg/kg1×≥52/33 (6)bHard(1979)(Wistar)corticalmonthsDay 2130 mg/kg1×16/39 (41)bepithelialtumorsMonth 130 mg/kg1×12/33 (36)bMonth 1.530 mg/kg1×14/28 (52)bMonth 230 mg/kg1×11/26 (42)bMonth 330 mg/kg1×18/27 (67)bMonth 430 mg/kg1×21/32 (66)bMonth 530 mg/kg1×6/14 (43)bRatsTotal tumorsDay 1i.p.20 mg/kg1×≥511/33 (33)b(Wistar)monthsDay 2130 mg/kg1×25/39 (64)bMonth 130 mg/kg1×25/33 (76)bMonth 1.530 mg/kg1×17/28 (63)bMonth 230 mg/kg1×13/26 (50)bMonth 330 mg/kg1×18/27 (67)bMonth 430 mg/kg1×22/32 (69)bMonth 530 mg/kg1×7/14 (50)b



### A-26Table 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)AgeTumorsaDosewhenSpeciesTargetroute,Duration ofAge atfirstChemical(strain)site# dosesDoseexposuredeathCommentsReferencedosedMFENURatsnervousDay 1Injection20 mg/kg1×100%bSusceptibility to neuro-Maekawa andEthylnitrosoureasystemoncogenic effectMitsumori (1990)Day 30Injection20 mg/kg1×61%bdeclined with increasingage.MiceliverControlControlNoneN/A90 weeks1/980/96Both male and femaleVesselinovitch(B6C3F1)(1%)(0%)mice were responsive to(1983)exposure duringGestationi.p.60 μg/g1×28/5218/49prenatal and infant life.day 18body weight(54%)b(37%)bDay 1560 μg/g1×41/5028/51body weight(82%)b(55%)bDay 4260 μg/g1×10/505/50body weight(20%)b(10%)bRatsnerve tissueControlControlNoneN/A4–70/160/10Highest tumor rate seenNaito et al. (1981)(Wistar)months(0%)(0%)when exposed duringgestation or soon afterGestationi.p.40 mg/kg1×26/2618/18birth.day 16(100%)b(100%)bDay1s.c.40mg/kg1×12/1216/16Statistically significant(100%)g(100%)gdecrease in tumorincidence withWeek140mg/kg1×12/1718/20increasing age of(71%)b(90%)bexposure.Week240mg/kg1×10/1414/18(71%)b(78%)bWeek340mg/kg1×6/135/17(46%)b(29%)bWeek440mg/kg1×8/152/10(53%)b(20%)b



### A-27Table 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)



### A-28ChemicalSpecies(strain)TargetsiteAgewhenfirstdosedDoseroute,# dosesDoseDurationofexposureAge atdeathTumorsaCommentsReferenceMFENUEthylnitrosoureaMice(B6C3F1)lungDay 1i.p.60 μg/g bodyweight1×49/55(89%)b49/50(98%)bVesselinovitch etal. (1974)(continued)Day 151×50/55(91%)b47/55(85%)bDay 421×53/59(90%)b44/51(86%)bDay 1120 μg/g bodyweight1×36/38(95%)b54/60(90%)bDay 151×45/49(92%)b43/50(86%)bDay 421×52/54(96%)b50/57(88%)bMicelungDay 160 μg/g body1×46/4751/51(C3AF1)weight(98%)g(100%)gDay 151×49/4957/59(100%)g(97%)gDay 421×59/5957/57(100%)g(100%)gDay 1120 μg/g body1×63/6453/57weight(98%)g(93%)gDay 151×54/5650/56(96%)g(89%)gDay 421×59/5948/48(100%)g(100%)gMiceliverDay 1i.p.60 μg/g body1×50/5428/43(B6C3F1)weight(93%)g(65%)gDay 151×55/5633/54(98%)g(61%)gDay 421×12/40(30%)b6/39(15%)bDay 1120 μg/g body1×29/3432/53weight(85%)g(60%)gDay 151×45/4829/43(94%)g(67%)gDay 421×17/494/50(35%)g(8%)gTable 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)



### A-29ChemicalSpecies(strain)TargetsiteAgewhenfirstdosedDoseroute,# dosesDoseDurationofexposureAge atdeathTumorsaCommentsReferenceMFENUMiceliverDay 1i.p.60 μg/g body1×42/4519/41Ethylnitrosourea(C3AF1)weight(93%)g(46%)g(continued)Day 151×42/50(84%)g19/48(40%)gDay 421×7/29(24%)b4/50(8%)bDay 1120 μg/g body1×55/6219/45weight(89%)g(42%)gDay 151×35/4515/35(78%)g(43%)gDay 421×8/33(24%)b3/33(9%)bMice(B6C3F1)kidneyDay 1i.p.60 μg/g bodyweight1×11/48(23%)b5/49(10%)bDay 151×6/41(15%)b7/31(23%)bDay 421×4/40(10%)b3/37(8%)bDay 1120 μg/g bodyweight1×10/30(33%)g14/53(26%)bDay 151×17/37(46%)g19/49(39%)bDay 421×8/40(20%)b11/39(28%)bMice(C3AF1)kidneyDay 1i.p.60 μg/g bodyweight1×7/44(16%)b6/45(13%)bDay 151×7/41(17%)b8/46(17%)bDay421×3/42(42%)b3/43(7%)bDay1120 μg/g bodyweight1×4/52(7%)b6/29(21%)gDay151×8/35(23%)b12/29(41%)gDay 421×6/41(71%)b3/39(8%)bTable 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)



### A-30ChemicalSpecies(strain)TargetsiteAgewhenfirstdosedDoseroute,# dosesDoseDurationofexposureAge atdeathTumorsaCommentsReferenceMFENUEthylnitrosoureaMice(B6C3F1)HarderianDay 160 μg/g bodyweight1×7/40(17%)b5/43(12%)b(continued)Day 151×10/51(20%)b17/59(29%)bDay421×14/50(28%)b14/45(31%)bDay1120 μg/g bodyweight1×9/30(30%)g6/52(12%)bDay151×15/41(37%)g8/31(26%)bDay 421×25/48(52%)g14/49(29%)bMice(C3AF1)HarderianDay160 μg/g bodyweight1×3/25(12%)b4/35(11%)bDay151×1/9(11%)b6/38(16%)bDay421×12/48(25%)b5/33(15%)bDay1120 μg/g bodyweight1×3/52(6%)b1/25(4%)bDay151×6/46(13%)b2/52(4%)bDay 421×5/29(17%)b2/11(18%)bMice(B6C3F1)stomachDay 160 μg/g bodyweight1×3/48(6%)b4/43(9%)bDay 151×10/42(24%)g7/45(16%)bDay421×9/51(18%)g8/36(22%)bDay1120 μg/g bodyweight1×2/29(7%)b9/53(17%)bDay151×10/35(29%)g12/33(36%)bDay 421×12/53(23%)g12/50(24%)bTable 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)



### A-31ChemicalSpecies(strain)TargetsiteAgewhenfirstdosedDoseroute,# dosesDoseDurationofexposureAge atdeathTumorsaCommentsReferenceMFENUEthylnitrosoureaMice(C3AF1)stomachDay 160 μg/g bodyweight1×2/39(5%)b7/45(16%)b(continued)Day 151×7/45(16%)g7/38(18%)bDay421×14/55(25%)g7/49(14%)bDay1120 μg/g bodyweight1×8/60(13%)b9/44(20%)bDay151×16/51(31%)g11/42(26%)bDay 421×19/48(40%)g13/37(35%)bMice(B6C3F1)malignantlymphomasDay160 μg/g bodyweight1×2/55(4%)b6/52(12%)gDay151×3/56(5%)b14/59(24%)gDay421×9/59(15%)b17/59(29%)gDay1120 μg/g bodyweight1×8/39(20%)b15/65(23%)gDay151×14/60(23%)b17/58(29%)gDay 421×12/59(20%)b14/60(23%)gMice(C3AF1)malignantlymphomasDay 160 μg/g bodyweight1×6/49(12%)b8/49(16%)gDay 151×3/49(6%)b13/61(21%)gDay421×6/60(10%)b9/55(16%)gDay1120 μg/g bodyweight1×3/66(5%)b10/58(17%)gDay151×10/56(18%)b18/60(30%)gDay 421×3/49(6%)b13/50(26%)gTable 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)AgeTumor incidenceaDoseDurationwhenSpeciesTargetroute,ofAge atfirstChemical(strain)site# dosesDoseexposuredeathCommentsReferencedosedMFNMUMiceTotal tumorsControlControlN/AN/A60 weeks1/200%Control mice did notTerraciniandMethylnitrosourea(BC3F1)(5%)exhibit tumors in targetTesta (1970)sites except a singlelungDay 1i.p.50 μg/g body1×60 weeks12/1516/19hepatoma in a male(84%)bweight(80%)bcontrol mouse.550 μg/g body1×60 weeks10/2610/35weeksweight(39%)b(29%)blympho-Day 150 μg/g body1×60 weeks23/3923/45sarcoma(51%)bweight(59%)b21/45weeks550 μg/g body1×60 weeks11/35weight(31%)b(47%)bliverDay 150 μg/g body1×60 weeks10/121/17weight(6%)b(83%)b550 μg/g body1×60 weeks0%b0%cweeksweightkidneyDay 150 μg/g body1×60 weeks3/153/18weight(17%)b(20%)b550 μg/g body1×60 weeks2/210%cweeksweight(10%)bfore-stomachDay 150 μg/g body1×60 weeks0%b4/17weight(24%)b550 μg/g body1×60 weeks8/2212/18weeksweight(36%)b(67%)bRatsDay 1i.p.50 μg/g body1×60 weeks0%b4/14Terracini and(Wistar)mammaryTumor incidence forweight(29%)bTesta (1970)on previouscontrol rats was based550 μg/g body1×60 weeks0%b3/5experiments (Dellaweeksweight(60%)bPorta et al., 1968) andlympho-Day 150 μg/g body1×60 weeks1/100%bwas not specificallysarcomaweight(10%)breported in this paper.550 μg/g body1×60 weeks2/81/11weeksweight(25%)b(9%)bkidney (ana-Day 150 μg/g body1×60 weeks14/189/13plastic)weight(78%)b(69%)b550 μg/g body1×60 weeks2/55/12weeksweight(40%)b(42%)b



### A-32Table 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)ChemicalSpecies(strain)TargetsiteAgewhenfirstdosedDoseroute,# dosesDoseDurationofexposureAge atdeathTumor incidenceaM FCommentsReferenceNMUMethylnitrosoureakidney(adenoma)Day 150 μg/g bodyweight1×60 weeks3/14(21%)b2/6(33%)b(continued)5weeks50 μg/g bodyweight1×60 weeks1⁄4(25%)b0%bforestomachDay 150 μg/g bodyweight1×60 weeks4/14(29%)b3/6(50%)b550 μg/g body1×60 weeks0%c0%bweeksweightintestineDay 150 μg/g bodyweight1×60 weeks3/10(30%)b2/2(100%)b5weeks50 μg/g bodyweight1×60 weeks2/4(50%)b0%bMicethymuscontroli.p.NANA120wks**0/340/25*Age at death fromTerracini et al.



### A-33(C3Hf/Dp)(0%)(0%)thymic lymphoma(1976)Day 1Day 7025 μg NMU/gbody weight25 μg NMU/gbody weight1×1×29 ± 8.4wks120 wks(M)***2/16(13%)b0/20(0%)c5/25(20%)b1/20(5%)breported specifically forsome, but not all, dosegroups.**Control mice were100 wkssacrificed at 120 wks.(F)Day 1Day 2150 μg NMU/gbody weight50 μg NMU/gbody weight1×1×16.5 ± 0.7wks24.5 ± 2.5wks16/24(67%)b14/44(32%)b30/44(68%)b18/38(47%)b***Age of death for allmice in this dose group,regardless of cancertype.Day 7050 μg NMU/gbody weight1×31.4 ± 4.4wks9/30(30%)b6/41(15%)bTable 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)



### A-34DoseAge at deathTumor incidenceSpeciesTargetAge whenroute,Duration ofChemical(strain)sitefirst dosed# dosesDoseexposureMFMFReferenceNMUMiceextra-thymiccontroli.p.NANA1201201/342/25Terracini et al. (1976)Methylnitrosourea(C3Hf/Dp)lymphomaweeksweeks(3%)(8%)(continued)Day 125 μg NMU/gbody weight1×100weeks90weeks2/16(13%)b1/25(4%)bDay 7025 μg NMU/gbody weight1×120weeks100weeks0/20(0%)b0/20(0%)bDay 150 μg NMU/gbody weight1×70weeks80weeks0/24(0%)b0/44(0%)bDay 2150 μg NMU/gbody weight1×100weeks90weeks1/44(2%)b0/38(0%)bDay 7050 μg NMU/gbody weight1×110weeks90weeks1/30(3%)b0/41(0%)blungcontroli.p.NANA120weeks120weeks4/34(12%)6/25(24%)Day 125 μg NMU/gbody weight1×100weeks90weeks7/16(44%)b13/25(52%)bDay 7025 μg NMU/gbody weight1×120weeks100weeks12/20(60%)b8/20(40%)bDay 150 μg NMU/gbody weight1×70weeks80weeks5/24(21%)b11/44(25%)bDay 2150 μg NMU/gbody weight1×100weeks90weeks23/44(52%)b15/38(39%)bDay 7050 μg NMU/gbody weight1×110weeks90weeks18/30(60%)b24/41(59%)blivercontroli.p.NANA120weeks120weeks13/34(38%)1/25(4%)Terracini et al. (1976)Day 125 μg NMU/gbody weight1×100weeks90weeks9/16(56%)g2/25(8%)bDay 7025 μg NMU/gbody weight1×120weeks100weeks12/20(60%)g2/20(10%)bDay 150 μg NMU/gbody weight1×70weeks80weeks4/24(17%)g3/44(7%)bDay 2150 μg NMU/gbody weight1×100weeks90weeks21/44(48%)g1/38(2.6%)bTable 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)



### A-35DoseAge at deathTumor incidenceSpeciesTargetAge whenroute,Duration ofChemical(strain)sitefirst dosed# dosesDoseexposureMFMFReferenceNMUMethylnitrosoureaMice(C3Hf/Dp)Day 7050 μg NMU/gbody weight1×110weeks90weeks8/30(27%)g2/41(5%)b(continued)stomachcontroli.p.NANA120weeks120weeks0/34(0%)5/25(20%)Day 125 μg NMU/gbody weight1×100weeks90weeks2/16(13%)b10/25(40%)bDay 7025 μg NMU/gbody weight1×120weeks100weeks3/20(15%)b7/20(35%)bDay 150 μg NMU/gbody weight1×70weeks80weeks2/24(8%)b1/44(2%)bDay 2150 μg NMU/gbody weight1×100weeks90weeks19/44(43%)b9/38(24%)bDay 7050 μg NMU/gbody weight1×110weeks90weeks8/30(27%)b21/41(51%)bkidneycontroli.p.NANA120weeks120weeks0/34(0%)0/25(0%)Terracini et al. (1976)Day 125 μg NMU/gbody weight1×100weeks90weeks0/16(0%)b0/25(0%)bDay 7025 μg NMU/gbody weight1×120weeks100weeks0/20(0%)b0/20(0%)bDay 150 μg NMU/gbody weight1×70weeks80weeks0/24(0%)b4/44(9%)bDay 2150 μg NMU/gbody weight1×100weeks90weeks1/44(2%)b4/38(11%)bDay 7050 μg NMU/gbody weight1×110weeks90weeks5/30(17%)b7/41(17% )bTable 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)



### A-36DoseAge at deathTumor incidenceSpeciesTargetAge whenroute,Duration ofChemical(strain)sitefirst dosed# dosesDoseexposureMFMFReferenceNMUMiceovarycontroli.p.NANA120120NA3/25Methylnitrosourea(C3Hf/Dp)weeksweeks(12%)(continued)Day 125 μg NMU/gbody weight1×100weeks90weeksNA2/25(8%)bDay 7025 μg NMU/gbody weight1×120weeks100weeksNA4/20(20%)bDay 150 μg NMU/gbody weight1×70weeks80weeksNA0/44(0%)bDay 2150 μg NMU/gbody weight1×100weeks90weeksNA9/38(24%)bDay 7050 μg NMU/gbody weight1×110weeks90weeksNA16/41(39%)bmammarycontroli.p.NANA120weeks120weeksNA2/25(8%)Terracini et al. (1976)Day 125 μg NMU/gbody weight1×100weeks90weeksNA1/25(4%)bDay 7025 μg NMU/gbody weight1×120weeks100weeksNA0/20(0%)bDay 150 μg NMU/gbody weight1×70weeks80weeksNA0/44(0%)bDay 2150 μg NMU/gbody weight1×100weeks90weeks1/44(2%)b0/38(0%)bDay 7050 μg NMU/gbody weight110weeks90weeksNA4/41(9.8%)buterusorvaginacontroli.p.NANA120weeks120weeksNA1/25(4%)Day 125 μg NMU/gbody weight1×100weeks90weeksNA1/25(4%)bDay 7025 μg NMU/gbody weight1×120weeks100weeksNA6/20(30%)bDay 150 μg NMU/gbody weight1×70weeks80weeksNA0/44(0%)bDay 2150 μg NMU/gbody weight1×100weeks90weeksNA1/38(3%)bDay 7050 μg NMU/gbody weight110weeks90weeks7/41(17%)bTable 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)TumorsaDoseSpeciesTargetAge whenDuration ofAge atroute,Chemical(strain)sitefirst dosedexposuredeathDoseCommentsReference# dosesMFUrethaneMicelungNewborns.c.0.18mg/g1×10weeks100%bThe average numberKaye and Trainin(SWR)adenomabody weightof tumors per mouse(1966)increased linearly11–22 weekss.c.0.25mg/g1×23–340%bwith dose.body weightweeksMiceliverControlControlNoneN/A493 days14/971/77Liebelt et al. (1964)(C3H/f)(m)(14%)(1%)553 days(f)Day 1i.p.0.8mg/g1×481days18/39body weight27/30(m)(90%)g(46%)g434 days(f)8–10 weeksi.p.1 mg/g body1×321days6/250/32weight(0%)c-(m)(24%)clungControlControlNoneN/A493 days0/970/77The number of lung(m)(0%)(0%)tumors among the553 dayscontrols was not(f)provided.Day 1i.p.0.8mg/g1×401days19/39body weight14/30(m)(48%)g408 days(46%)g(f)8–10 weeksi.p.1 mg/g body1×506days2/250/32weight(m)(8%)c(0%)c-reticularControlControlNoneN/A493 days2/976/77tissue(m)(2%)(8%)553 days(f)Day 1i.p.0.8mg/g1×285days4/3022/39body weight(m)(56%)g343 days(13%)c(f)8–10 weeksi.p.1 mg/g body1×-0/254/32weight453 days(25%)c(13%)c(f)



### A-37Table 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)TumorsaDoseSpeciesTargetAge whenDuration ofAge atroute,Chemical(strain)sitefirst dosedexposuredeathDoseCommentsReference# dosesMFUrethaneMiceleukemiaControlControlNoneN/A8–101%Highest tumor ratesFiore-Donati et al.(continued)(Swiss)monthswhen dosed at birth.(1962)Day 1s.c.2 mg in 0.051×13/60Exposure tomL aqueous(22%)bnewborns wassolutionfollowed by 21.6%Day 5leukemia, occurringmL aqueous4 mg in 0.051×7/39at a mean age of 105solution(18%)bdays.Day 4020 mg in 0.11×2/63mL aqueous(3%)bsolutionMicelungControlControlNoneN/A9 weeks0/15—The proportion ofRogers (1951)(Swiss)animals withadenomas decreasedadenoma2 weeks(0%)ControlControlNoneN/A11 weeks0/14—steadily with age of4 weeks(0%)exposure.ControlControlNoneN/A13 weeks1/15—6 weeks(7%)ControlControlNoneN/A15 weeks2/15—8 weeks(13%)ControlControlNoneN/A17 weeks0/15—10 weeks(0%)2 weeksi.p.1 mg/g body1×9weeks24/24—weight(100%)b4 weeksi.p.1 mg/g body1×11weeks23/25—weight(92%)b6 weeksi.p.1 mg/g body1×13weeks22/25—weight(88%)b8 weeksi.p.1 mg/g body1×15weeks21/25—weight(84%)b10 weeksi.p.1 mg/g body1×17weeks19/25—weight(76%)b



### A-38Table 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)ChemicalSpecies(strain)TargetsiteAge whenfirst dosedDoseroute,# dosesDoseDuration ofexposureAge atdeathTumorsaMFCommentsReferenceUrethane(continued)Mice(Swiss)lungadenoma3 weeksi.p.0.25 mg/gbody weight1×12weeks16/19(84%)b—0.5mg/gbody weight1×12 weeks16/20(80%)b—1 mg/g bodyweight1×12weeks18/20(90%)b—8 weeksi.p.0.25mg/gbody weight1×17weeks4/17(24%)b—0.5mg/gbody weight1×17 weeks15/16(94%)b—1 mg/g bodyweight1×17 weeks18/18(100%)b—



### A-39Table 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)AgeTumor incidenceaDoseDurationwhenSpeciesTargetroute,ofAge atfirstChemical(strain)site# dosesDoseexposuredeathCommentsReferencedosedMFUrethaneMiceliverControlControlN/AN/A360–72010/2274/222Chieco-Bianchiet(continued)(Swiss)days(4.4%)(8.22%)al. (1963)Day 1s.c.1 mg/g body1×180 days1/200/20weight(5%)g(0%)cDay 1s.c.1 mg/g body1×240 days2/170/12weight(0%)c(12%)gDay 1s.c.1 mg/g body1×300 days5/180/16weight(28%)g(0%)cDay 1s.c.1 mg/g body1×360 days11/200/23weight(55%)g(0%)cDay 1s.c.1 mg/g body1×420 days13/152/22weight(9%)g(87%)gDay 1s.c.1 mg/g body1×480 days17/232/25weight(74%)c(8%)cDay 5s.c.1 mg/g body1×420 days9/132/11weight(69.2%)b(18.2%)bDay 20s.c.1 mg/g body1×420 days1/130/16weight(0%)b(8%)bDay 40s.c.1 mg/g body1×420 days0/110/9weight(0%)b(0%)bMiceskinControlControlN/AN/A180–55030/712Croton oil treatmentChieco-Bianchi et(Swiss)days(4.21%)initiated at 40 days ofal. (1963)age.Day1s.c.1mgsingle660 days26/59urethane/gdose(44.1%)gbody weight;urethane,5% croton oilcroton oilapplied2×/weekfor 10mos



### A-40Table 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)AgeTumor incidenceaDoseDurationwhenSpeciesTargetroute,ofAge atfirstChemical(strain)site# dosesDoseexposuredeathCommentsReferencedosedMFUrethaneDay40s.c.1mgsingle700 days8/41(continued)urethane/gdose(19.5%)bbody weight;urethane,5% croton oilcroton oilapplied2×/weekfor 10mosMiceliverControlgavageN/AN/A71weeks1/250/25Klein(1966)(B6AF1/J)(4%)(0%)Day 11 mg/g body1×66 weeks9/209/26weight(45%)g(35%)gDay 71 mg/g body1×67 weeks20/2220/26weight(77%)g(91%)gDay 141 mg/g body1×68 weeks16/2010/23weight(80%)g(43%)gDay 211 mg/g body1×69 weeks13/231/20weight(57%)g(5%)gDay 281 mg/g body1×70 weeks4/241/20weight(17%)g(5%)glungControlgavage1mg/gbody1×71 weeks9/256/25weight(36%)(24%)Day 11 mg/g body1×66 weeks20/2025/26weight(100%)b(96%)bDay 71 mg/g body1×67 weeks22/2226/26weight(100%)b(100%)bDay 141 mg/g body1×68 weeks19/2019/23weight(95%)b(83%)bDay 211 mg/g body1×69 weeks23/2319/20weight(100%)b(95%)bDay281mg/gbody1×70 weeks24/2420/20weight(100%)b(100%)bMiceHarderianControlgavage1 mg/g body1×71 weeks0/250/25Klein(1966)(B6AF1/J)glandweight(0%)(0%)Day 11 mg/g body1×66 weeks0/201/26weight(0%)c(4%)b



### A-41Table 3. Methodological information and tumor incidencefor animal studies with early postnatal and juvenileand adult acute exposure (continued)AgeTumor incidenceaChemicalSpecies(strain)TargetsitewhenfirstdosedDoseroute,# dosesDoseDurationofexposureAge atdeathMFCommentsReferenceUrethane (continued)Day 71 mg/g bodyweight1×67 weeks0/22(0%)c1/26(4%)bDay 141 mg/g bodyweight1×68 weeks0/20(0%)c2/23(9%)bDay 211 mg/g bodyweight1×69 weeks1/23(4%)b0/20(0%)cDay 281 mg/g body1×70 weeks0/240/20weight(0%)c(0%)cforestomachControlgavage1mg/gbody1×71 weeks0/251/25weight(0%)(4%)Day 11 mg/g bodyweight1×66 weeks0/20(0%)c3/26(12%)b



### A-42Day 71 mg/g bodyweight1×67 weeks1/22(5%)b1/26(4%)bDay 141 mg/g bodyweight1×68 weeks1/20(5%)b4/23(17%)bDay 211 mg/g bodyweight1×69 weeks0/23(0%)c1/20(5%)bDay 281 mg/g bodyweight1×70 weeks2/24(8%)b1/20(5%)baWhere not delineated by gender, data combined by study authors or gender not specified. Where percentages only are given, number of subjects not specified.bNot evaluated by authors.cEvaluated but not significant compared with controls.dStudy also included mammary fibroadenomas andfibromas as well as other types of cancers.e8–9 weeks old.fIncludes survivors up to 40 weeks only.gSignificant compared with controls.i.p.= intraperitoneal injection;s.c. = subcutaneous injectionTable 4. Ratio of early-life to adult cancer potenciesfor studies with repeated exposures of juvenile and adultanimals to carcinogens witha mutagenic mode of action*



### A-43UnweightedSpeciesgeometricCompound(strain)SexDoseTumormean2.5%Median97.5%ReferenceBenzidineMice (B6C3F1)maleliver11164110198Vesselinovitch et al.(1975b)femaleliver0.160.0040.221.13-MU3-Methylcholanthrene(formerly known as 20­methylcholanthrene)Mice (Albino)male0.25 mg/ghepatoma337.430268Klein (1959)female0.25mg/ghepatoma7.71.17.185male0.25mg/gforestomach0.910.390.912.1female0.25mg/gforestomach1.50.581.54.2male0.25mg/gskin1.80.0482.122female0.25mg/gskin1.50.0231.821SafroleMice(B6C3F1)maleliver471644198Vesselinovitch et al.(1979b)femaleliver0.120.0020.181.1VCVinyl chlorideRats (SpragueDawley)male6,000 ppmliver-angiosarcoma6.70.0359.857Maltoni et al. (1984)male10,000 ppmliver-angiosarcoma7.40.0351162female6,000ppmliver-angiosarcoma134.91333female10,000ppmliver-angiosarcoma308.729121male6,000ppmzymbal gland0.730.00321.130male10,000ppmzymbal gland0.270.00220.45.4female6,000ppmzymbal gland0.480.00270.716female10,000ppmzymbal gland0.150.00140.194.5male10,000ppmleukemia210.02637514female6,000ppmleukemia1.30.00351.7153female10,000ppmleukemia0.290.00190.3517male6,000ppmnephroblastomas0.150.00140.194.8male10,000ppmnephroblastomas0.170.00150.216.2female6,000ppmnephroblastomas0.280.00180.3316female10,000 ppmnephroblastomas0.240.00170.2911male6,000ppmangiosarcomasother sites0.90.00331.2653male10,000ppmangiosarcomas-0.250.00170.3012Table 4. Ratio of early-life to adult cancer potenciesfor studies with repeated exposures of juvenile and adultanimals to mutagenicchemicals (continued)



### A-44UnweightedSpeciesgeometricCompound(strain)SexDoseTumormean2.5%Median97.5%Referenceother sitesVCfemale6,000ppmangiosarcomas-0.240.00170.2911Vinyl chlorideother sites(continued)female10,000ppmangiosarcomasother sites0.320.00190.3820male6,000 ppmangiomas &0.720.00311.033fibromas-othersitesmale10,000 ppmangiomas &1.40.00452.3647fibromas-othersitesfemale6,000 ppmangiomas &0.270.00180.3316fibromas-othersitesfemale10,000 ppmangiomas &0.520.00240.6341fibromas-othersitesmale6,000ppmhepatoma621158543male10,000 ppmhepatoma348.232218female6,000ppmhepatoma551351352female10,000ppmhepatoma558.453513male6,000 ppmskin carcinomas1.10.00351.582male10,000 ppmskin carcinomas0.410.00240.5615female6,000 ppmskin carcinomas0.460.00240.5924female10,000 ppmskin carcinomas0.310.00190.3719male6,000ppmneuroblastoma0.210.00160.269.5male10,000ppmneuroblastoma0.200.00160.248.5female6,000ppmneuroblastoma0.270.00180.3215female10,000ppmneuroblastoma0.140.00140.184.4*The 2.5% and 97.5% are percentilesof the posterior distribution. For a Bayesian distribution, thesepercentiles function inamanner similar to the 95% confidence limits for other types of statistical analyses.Table 5. Ratio of early-life to adult cancer potencies forstudies with repeated exposures of juvenile and adult animalsto chemicals with a nonmutagenic mode of action*



### A-45Ratio of juvenile to adult potencyUnweightedSpeciesgeometricCompound(strain)SexDoseTumormean2.5%Median97.5%ReferenceAmitroleMice (B6C3F1)maleNAliver135.11430Vesselinovitch (1983)femaleNAliver0.140.00130.183.9DDTMice (B6C3F1)maleNAliver1.30.00442.525Vesselinovitch et al.(1979a)DieldrinMice (B6C3F1)maleNAliver0.750.00311.227Vesselinovitch et al.(1979a)DPHRats (F344/N)male630liver0.40.00240.5416Chhabra et al. (1993b)female630liver0.240.00170.2912Mice(B6C3F1)male210liver1.50.00402.471female210liver1.30.00562.615ETURats (F344/N)male90thyroid0.370.00290.615.4Chhabra et al. (1992)female90thyroid0.230.00180.37.0Mice(B6C3F1)male330liver0.0910.00110.121.9female330liver0.0570.00100.0810.65male330thyroid0.410.00220.5225female330thyroid0.40.00240.5516male330pituitary0.320.00190.3822female330pituitary0.240.00180.326.9PBBRats(F344/N)male10liver0.590.00411.16.6Chhabra et al. (1993a)female10liver0.0630.00090.0791.2male10mononuclearcell leukemia0.790.00351.418female10mononuclearcell leukemia0.210.00170.286.0Mice(B6C3F1)male30liver3.91.93.97.5female30liver1.00.371.052.1*The 2.5% and 97.5% are percentilesof the posterior distribution. For a Bayesian distribution, thesepercentiles function inamanner similar to the 95% confidence limitsfor other types of statistical analyses.Table 6. Ratio of early-life to adult cancer potencies for studies with acute exposures of juveniles and adult animals tocarcinogens with a mutagenic mode of action*



### A-46CompoundSpecies(strain)SexDoseTumorDayRatio of juvenile to adult potencyReferenceUnweightedgeometricmean2.5%Median97.5%BaP*Mice (B6C3F1)male75 μg/kgliver1day9.32.98.455Vesselinovitch et al.(1975a)15days113.59.661female75 μg/kg1day1.20.00831.63115days1.70.0152.136male150 μg/kg1day298.22619415days154.113109female150 μg/kg1day8.81.48.19415days1.20.00821.630Mice (C3AF1)male75 μg/kgliver1day112.11011215days7.51.17.083female75 μg/kg1day0.20.00180.269.115days0.20.00170.248.5male150 μg/kg1day143.012.813015days3.60.113.849female150 μg/kg1day0.20.00170.248.815days0.20.00170.248.7Mice(B6C3F1)Male75 μg/kglung1 day1.20.451.23.415days0.20.00460.311.4female75 μg/kglung1 day2.81.0962.79.515days1.40.411.45.1Male150 μg/kglung1 day2.21.02.15.415days0.80.20.822.3female150 μg/kglung1 day7.92.67.24315days3.71.13.422Mice(C3AF1)male75 μg/kglung1 day1.20.471.23.215days1.10.431.083.1Table 6. Ratio of early-life to adult cancer potenciesfor studies with acute exposures of juveniles and adultanimals to carcinogens with a mutagenic mode of action (continued)



### A-47CompoundSpecies(strain)SexDoseTumorDayRatio of juvenile to adult potencyReferenceUnweightedgeometricmean2.5%Median97.5%BaP*(continued)female75 μg/kglung1 day1.60.661.554.015days1.60.711.634.2male150 μg/kglung1 day1.50.571.55.015days1.90.711.86.0female150 μg/kglung1 day1.30.611.32.915days1.20.541.12.6DBAMicelung178201435100Law (1940)DEN**Mice (B6C3F1)male6 μg/kgliver1 day9.03.58.337Vesselinovitch et al.(1984)15days8.93.58.236female6 μg/kgliver1 day359.13123915days256.3226175male12 μg/kgliver1 day9.63.38.85015days9.83.48.951female12 μg/kgliver1 day165.9156715days197.11879Mice(C3AF1)male6 μg/kgliver1 day7.32.96.92615days3.51.43.313female6 μg/kgliver1 day173.21616615days6.40.866.073male12 μg/kgliver1 day113.79.55315days9.83.48.950female12 μg/kgliver1 day408.53634015days255.022221Mice(B6C3F1)male6 μg/kglung1 day0.50.270.520.9315days1.60.951.62.7female6 μg/kglung1 day0.90.540.891.515days1.20.761.22.0Table 6. Ratio of early-life to adult cancer potenciesfor studies with acute exposures of juveniles and adultanimals to carcinogens with a mutagenic mode of action (continued)



### A-48CompoundSpecies(strain)SexDoseTumorDayRatio of juvenile to adult potencyReferenceUnweightedgeometricmean2.5%Median97.5%DEN**(continued)male12 μg/kglung1 day0.40.210.400.7315days0.70.390.661.1female12 μg/kglung1 day0.70.440.731.215days1.40.881.42.3Mice(C3AF1)male6 μg/kglung1 day0.70.220.671.715days0.50.210.561.3female6 μg/kglung1 day1.10.451.12.515days0.70.360.741.5male12 μg/kglung1 day0.30.0840.330.7615days0.60.260.621.4female12 μg/kglung1 day0.70.350.751.615days0.70.370.751.5DMBA#Rats (Wistar)maletotal2 vs 5–8 wks3.31.33.210Meranze et al. (1969)2 vs 26 wks3.21.33.19.7femaletotal2 vs 5–8 wks1.30.681.32.52 vs 26 wks3.31.23.016mammary2 vs 5–8 wks0.00.00120.0560.262 vs 26 wks0.20.00230.295.35 vs 26 wks7.11.86.455Mice (Balb/c)male15 μglung1 day302.8221482Walters (1966)15–19 days1.00.281.03.5male30 μgx2lung15–19 days141.05610978female15 μglung1 day606.046235015–19 days3.10.513.022female30 μgx2lung15–19 days151.2111004Mice (Swiss)lymphoma2.70.602.519Pietra et al. (1961)lung9.12.98.740Table 6. Ratio of early-life to adult cancer potenciesfor studies with acute exposures of juveniles and adultanimals to carcinogens with a mutagenic mode of action (continued)



### A-49CompoundSpecies(strain)SexDoseTumorDayRatio of juvenile to adult potencyReferenceUnweightedgeometricmean2.5%Median97.5%DMN***Rats (Wistar)3 wkstotal1 month0.70.410.731.3Hard (1979)1.5months1.10.581.12.12months1.50.751.53.03months0.90.500.941.824 hr1 month0.30.130.280.61.5months0.40.180.420.92months0.60.240.561.33months0.40.160.360.781 month1.5 months1.50.801.523.02months2.01.02.04.23months1.30.691.32.5ENUMice (B6C3F1)maleliver7.83.97.718Vesselinovitch (1983)female7.12.96.921Rats (Wistar)malenerve tissue1 day272.5201374Naito et al. (1981)1week1.60.611.64.62weeks1.60.581.64.83weeks0.70.120.722.3female1day646.05024881weeks9.62.68.9592weeks6.21.65.7403weeks0.70.00900.898.9Mice (B6C3F1)male60 μg/glung11.00.601.01.7Vesselinovitch et al.(1974)151.10.661.11.8female60μg/glung12.11.172.14.1151.00.601.01.7male120μg/glung11.00.601.01.7Table 6. Ratio of early-life to adult cancer potenciesfor studies with acute exposures of juveniles and adultanimals to carcinogens with a mutagenic mode of action (continued)



### A-50CompoundSpecies(strain)SexDoseTumorDayRatio of juvenile to adult potencyReferenceUnweightedgeometricmean2.5%Median97.5%151.10.661.01.8ENU(continued)female120μg/glung12.11.22.14.1151.00.601.01.7Mice(C3AF1)male60 μg/glung18.72.78.04815525.2392141female60μg/glung150.70.320.721.6male120μg/glung10.90.380.922.2150.70.280.671.6female120μg/glung10.50.240.541.2150.40.180.420.92Mice(B6C3F1)male60 μg/gliver18.84.28.52215146.21437female60 μg/gliver16.32.66.118155.62.45.416male120μg/gliver15.22.55.111157.63.97.517female120μg/gliver1114.1114615144.91355Mice(C3AF1)male60 μg/gliver1124.71143158.13.27.629female60 μg/gliver17.52.67.032154.81.84.618male120μg/gliver19.84.19.332156.62.76.323female120μg/gliver15.41.75.025155.41.75.125Mice(B6C3F1)male60 μg/gkidney12.20.732.18.0151.20.291.25.1Table 6. Ratio of early-life to adult cancer potenciesfor studies with acute exposures of juveniles and adultanimals to carcinogens with a mutagenic mode of action (continued)



### A-51CompoundSpecies(strain)SexDoseTumorDayRatio of juvenile to adult potencyReferenceUnweightedgeometricmean2.5%Median97.5%female60μg/gkidney10.70.0240.855.9152.60.612.515ENU(continued)male120μg/gkidney11.70.651.74.4152.61.142.66.4female120μg/gkidney10.90.370.872.0151.40.671.43.2Mice(C3AF1)male60 μg/gkidney11.80.171.915152.00.252.016female60μg/gkidney11.00.0161.313152.10.162.220male120μg/gkidney10.20.00290.241.5151.50.381.55.9female120μg/gkidney12.30.172.420157.11.86.547Mice(B6C3F1)male60 μg/gHarderian10.30.0180.411.4150.50.0750.521.4female60μg/gHarderian10.10.00250.160.74150.80.350.842.0male120μg/gHarderian10.40.130.420.96150.60.260.571.2female120μg/gHarderian10.10.00300.180.85150.70.170.772.1Mice(C3AF1)male60 μg/gHarderian10.10.00230.201.3150.10.00160.181.8female60μg/gHarderian10.40.0190.522.5150.80.150.853.4male120μg/gHarderian10.10.00100.0861.0150.30.00500.402.8Table 6. Ratio of early-life to adult cancer potenciesfor studies with acute exposures of juveniles and adultanimals to carcinogens with a mutagenic mode of action (continued)



### A-52CompoundSpecies(strain)SexDoseTumorDayRatio of juvenile to adult potencyReferenceUnweightedgeometricmean2.5%Median97.5%female120μg/gHarderian10.10.00120.0941.2150.10.00120.0810.90ENU(continued)Mice (B6C3F1)male60 μg/gstomach10.30.00910.342.4151.90.611.828.7female60μg/gstomach10.20.00830.261.1150.20.00720.241.0male120μg/gstomach10.20.00590.200.90151.20.501.22.9female120μg/gstomach10.60.190.601.5151.60.671.63.7Mice(C3AF1)male60 μg/gstomach10.00.00090.0630.51150.30.0230.411.3female60μg/gstomach10.80.0850.893.5151.10.191.14.5male120μg/gstomach10.20.0100.190.56150.70.320.701.5female120μg/gstomach10.40.140.461.2150.60.240.641.5NMUMice (BC3F1)male50 μg/glung adenomas13.41.33.39.3Terracini and Testa(1970)female50 μg/glung adenomas16.32.46.023male50μg/glymphosarcoma12.51.12.46.4female50μg/glymphosarcoma11.10.491.12.4male50μg/ghepatoma1356.532324female50μg/ghepatoma10.30.00230.3913male50 μg/grenal adenoma10.90.00931.213female50 μg/grenal adenoma11.30.00811.733male50μg/gforestomach10.00.00060.0390.52female50μg/gforestomach10.10.00270.150.69Table 6. Ratio of early-life to adult cancer potenciesfor studies with acute exposures of juveniles and adultanimals to carcinogens with a mutagenic mode of action (continued)



### A-53CompoundSpecies(strain)SexDoseTumorDayRatio of juvenile to adult potencyReferenceUnweightedgeometricmean2.5%Median97.5%Mice (C3Hf/Dp)male25 μg/gthymiclymphoma11.90.0482.123NMU(continued)female25μg/gthymiclymphoma11.20.00891.530male25 μg/glung adenomas11.00.0131.211female25 μg/glung adenomas10.40.0180.461. 7male25 μg/gliver tumor10.20.00160.214.6female25 μg/gliver tumor10.30.00260.394.4male25μg/gStomach10.50.00450.676.8female25μg/gStomach10.30.00460.433.8ovarian10.10.00140.173.5uterine/vaginal18.61.18.197male50μg/gthymiclymphoma17.93.17.430female50μg/gthymiclymphoma13.11.33.07.8male50 μg/glung adenomas10.040.00080.0580.45female50 μg/glung adenomas10.10.00120.0840.53male50 μg/gliver tumor10.20.00210.337.8female50 μg/gliver tumor10.10.00110.134.5male50μg/gStomach10.010.00030.0130.12female50μg/gStomach10.10.00220.150.96ovarian10.00.00030.0140.14uterine/vaginal10.00.00050.0340.46male50μg/gthymiclymphoma214.31.64.117female50μg/gthymiclymphoma211.00.391.02.6male50 μg/glung adenomas210.10.00220.221.1female50 μg/glung adenomas210.70.300.751.7Table 6. Ratio of early-life to adult cancer potenciesfor studies with acute exposures of juveniles and adultanimals to carcinogens with a mutagenic mode of action (continued)Ratio of juvenile to adult potencyUnweightedCompoundSpecies(strain)SexDoseTumorDaygeometricmean2.5%Median97.5%Referencemale50 μg/gliver tumor210.10.00130.154.3female50 μg/gliver tumor210.90.00511.423NMUmale50μg/gstomach210.10.0010.080.64(continued)female50μg/gstomach211.80.771.84.7ovarian210.00.00070.0550.97uterine/vaginal211.70.591.76.4UrethaneMice (Swiss)male1 mg/gliver1244.421220Chieco-Bianchi et al.female1mg/gliver10.40.00440.5413(1963)male1mg/gliver5142.413137female1mg/gliver51.20.0171.426



### A-54male1mg/gliver200.20.00180.2810female1mg/gliver200.10.00110.124.8both1mg/gskin10.20.00270.325.4Urethane +Mice (Swiss)both1 mg/gskin12.91.22.88.2croton oilUrethaneRats (MRCmale/16%×6neurilemmomas10.20.00280.334.5Choudari Kommineni etWistar-derived)femaleal. (1970)male/16%×6neurilemmomas280.40.00450.516.3femalemale/16%×6liver17.91.47.182femalemale/16%×6liver280.20.00260.411.7femalemale/16%×6thyroid10.00.00060.0390.67femalemale/16%×6thyroid280.10.00110.11.5femaleMice(Swiss)male/1 mg/glung1151.211997De Benedictis et al.female(1962)Mice(Swiss)leukemia6.71.76.145Fiore-Donatiet al.Table 6. Ratio of early-life to adult cancer potenciesfor studies with acute exposures of juveniles and adultanimals to carcinogens with a mutagenic mode of action (continued)



### A-55CompoundSpecies(strain)SexDoseTumorDayRatio of juvenile to adult potencyReferenceUnweightedgeometricmean2.5%Median97.5%5.11.14.738(1962)Urethane(continued)Mice (B6AF1/J)male1 mg/gliver215.11.44.730Klein (1966)female1mg/gliver210.20.00190.266.0Harderiangland10.30.00210.331170.30.00210.3311140.60.00440.8520male1 mg/gHarderian gland210.30.00240.4113male1mg/gforestomach10.10.00090.0791.9female1mg/gforestomach10.40.00280.4911male1mg/gforestomach70.10.00170.193.5female1mg/gforestomach70.10.00130.165.0male1mg/gforestomach140.20.00180.213.9female1mg/gforestomach140.80.00561.118male1mg/gforestomach210.10.00080.0721.7female1mg/gforestomach210.20.00150.26.3lung11.00.360.952.5male1mg/glung140.80.260.82.3female1mg/glung140.40.160.451.1210.90.310.862.4Mice (C3H/f)male1 mg/gliver1144.01281Liebelt et al. (1964)female1mg/gliver1163.215155male1mg/glung15.91.75.628female1mg/glung1224.520203male1 mg/greticular tissue12.00.0232.338female1 mg/greticular tissue18.62.37.760Mice (Swiss)1 mg/gpulmonaryadenomas2 vs 4 weeks141.110.1965Rogers (1951)Table 6. Ratio of early-life to adult cancer potenciesfor studies with acute exposures of juveniles and adultanimals to carcinogens with a mutagenic mode of action (continued)



### A-56CompoundSpecies(strain)SexDoseTumorDayRatio of juvenile to adult potencyReferenceUnweightedgeometricmean2.5%Median97.5%1mg/gpulmonaryadenomas2 vs 6 weeks161.311.31025Urethane(continued)1mg/gpulmonaryadenomas2 vs 8 weeks191.613.311261mg/gpulmonaryadenomas2 vs 10 weeks211.914.511680.25 mg/gadenomas3 vs 8 weeks7.12.36.7290.5 mg/gadenomas3 vs 8 weeks0.70.290.671.61.0 mg/gadenomas3 vs 8 weeks0.70.280.681.6*The 2.5% and 97.5% are percentilesof the posterior distribution. For a Bayesian distribution, thesepercentiles function inamanner similar to the 95% confidence limits for other types of statistical analyses.Table 7. Ratio of early-life to adult cancer potencies for studies with lifetime exposures starting with juvenileand adult animals to carcinogens with mutagenic or nonmutagenic modes of action*



### A-57UnweightedgSpecieseometricCompound(strain)SexDoseTumormean2.5%Median97.5%ReferenceMutagenic compoundsDENRats (Colworth)multipleliver2.80.00935.623Peto et al. (1984)esophagus0.180.00150.234.8SafroleMice (B6C3F1)maleliver503.750253Vesselinovitch et al.(1979b)femaleliver4.00.0074.023UrethaneMice (B6AF1/J)male2.5 mg/pupliver790.361021,064Klein (1966)female2.5mg/pupliver0.470.00220.5542Nonmutagenic compoundsDDTMice (B6C3F1)liver230.00230.5823Vesselinovitch et al.(1979a)DieldrinMice (B6C3F1)liver910.0141491Vesselinovitch et al.(1979a)DPHRats (F344/N)male630:800liver0.310.00190.3718Chhabra et al. (1993b)630:2,400liver0.360.00210.4517female630:800liver0.330.00190.3921630:2,400liver0.330.00190.3921Mice(B6C3F1)male210:100liver0.710.00280.9349210:300liver140.0323214female210:200liver0.320.0020.4213210:600liver0.350.00230.538.8ETURats (F344/N)male90:83thyroid0.230.00170.37.3Chhabra et al. (1992)90:250thyroid9.11.110.527female90:83thyroid0.370.00210.461990:250thyroid0.610.00341.110Table 7. Ratio of early-life to adult cancer potencies for studies with lifetime exposures starting with juvenileand adult animals to carcinogens with mutagenic or nonmutagenic modes of action (continued)



### A-58UnweightedSpeciesgeometricCompound(strain)SexDoseTumormean2.5%Median97.5%ReferenceETU(continued)Mice (B6C3F1)male330:330liver0.370.00220.514330:1,000liver0.480.00270.7512female330:330liver0.330.00230.57.8330:1,000liver0.420.00250.6511male330:330thyroid0.440.00220.5234330:1,000thyroid0.630.00351.1210female330:330thyroid5.20.01110108330:1,000thyroid0.180.00160.244.2male330:330pituitary0.400.00210.4732330:1,000pituitary0.180.00150.225.7female330:330pituitary0.210.00160.2610330:1,000pituitary0.270.00190.369.0PBBRats (F344/N)male10:10liver0.390.00230.5613Chhabra et al. (1993a)10:30liver0.180.00160.254.3female10:10liver3615368610:30liver3.10.0234.622male10:10mononuclearcellleukemia0.510.00250.6923male10:30mononuclearcellleukemia0.770.00311.135female10:10mononuclearcellleukemia0.540.00260.7424female10:30mononuclearcellleukemia0.340.00210.4515Mice(B6C3F1)male30:30liver8.90.01512.21,076female30:30liver4.40.00756.2786male10:10liver0.150.00140.23.9Table 7. Ratio of early-life to adult cancer potencies for studies with lifetime exposures starting with juvenileand adult animals to carcinogens with mutagenic or nonmutagenic modes of action (continued)CompoundSpecies(strain)SexDoseTumorUnweightedgeometricmean2.5%Median97.5%Referencefemale10:10liver0.290.00210.437.0*The 2.5% and 97.5% are percentilesof the posterior distribution. For a Bayesian distribution, thesepercentiles function inamanner similar to the 95% confidence limitsfor other types of statistical analyses.



### A-59Table 8. Summary of quantitative estimates of ratio of early-life to adult cancer potencies



### A-60InverseDoseTissueNumber ofchemicalsweightedgeometric meanratioUnweightedMinimumUnweightedMaximumNumber ofratiosPercentage >1Chemicals with mutagenic mode of actionRepeated410.50.121114542Lifetime38.70.1879667Combined repeated and lifetime610.40.121115145AcuteCombined111.50.0117826855Forestomach30.0760.011.93216Harderian20.480.060.8200.0Kidney21.60.177.11878Leukemia15.95.16.72100Liver58.10.10407077Lung71.10.041787756Lymph21.81.12.73100Mammary (wk 5 vs wk 26)17.1NANA1100Mammary (wk 2 vs wk 5–8 or 26)10.071NANA20Nerve22.30.2464875Nerve (Day 1 comparison)2100.2464367Ovarian10.0330.010.1330Reticular tissue16.51.968.62100Thymic lymphoma12.81.017.96100Thyroid10.050.030.0820Uterine/vaginal11.60.038.6367Day 171.70.0117812755Day 1531.50.06527465Chemicals with nonmutagenic mode of actionRepeated62.20.06132227Lifetime53.40.15363821

### Table 9. Excess Relative Risk (ERR) estimates for cancer incidence from

Life Span Study (Japanese survivors) a

Average ERR at 1 Sv

Site 

### <20 b 

### >20 b



### Stomach 0.74 0.24

Colon 0.62 0.7

Liver 1.3 0.31

Lung 0.57 1.1

Bone and connective tissue 11 0.42

Skin 5.4 0.39

Breast 3.3 0.98

Urinary bladder 0.71 0.79

Leukemia 6.1 3.7

a Information extracted from tables in UNSCEAR, Annex I (2000).

b Age at exposure.



### A-61

### Table 10. Excess Relative Risk (ERR) estimates for incidence of thyroid

cancer from Life Span Study a

Age at exposure Average ERR at 1 Sv

(No. cases)



### 0–9 yr 10.25 (24)

10–19 yr 4.5 (35)

20–29 yr 0.10 (18)

>30 yr 0.04 (55)

a Information extracted from tables in UNSCEAR, Annex I (2000).



### A-62

### Table 11. Coefficients for the Revi sed Methodology mortality risk model

(from U.S. EPA, 1999) a

Cancer type b 40+

R

R

R

R

A

A

R

R

R

R

A

R

Female:

R

R

R

R

A

A

R

R

R

R

A

R Age group Risk model

type 0–9 10–19 20–29 30–39

Male:

Stomach 1.223 1.972 2.044 0.3024 0.2745

Colon 2.290 2.290 0.2787 0.4395 0.08881

Liver 0.9877 0.9877 0.9877 0.9877 0.9877

Lung 0.4480 0.4480 0.0435 0.1315 0.1680

Bone 0.09387 0.09387 0.09387 0.09387 0.09387

Skin 0.06597 0.06597 0.06597 0.06597 0.06597

Breast 0.0 0.0 0.0 0.0 0.0

Ovary 0.0 0.0 0.0 0.0 0.0

Bladder 1.037 1.037 1.037 1.037 1.037

Kidney 0.2938 0.2938 0.2938 0.2938 0.2938

Thyroid 0.1667 0.1667 0.1667 0.1667 0.1667

Leukemia 982.3 311.3 416.6 264.4 143.6

Stomach 3.581 4.585 4.552 0.6309 0.5424

Colon 3.265 3.265 0.6183 0.8921 0.1921

Liver 0.9877 0.9877 0.9877 0.9877 0.9877

Lung 1.359 1.359 0.1620 0.4396 0.6047

Bone 0.09387 0.09387 0.09387 0.09387 0.09387

Skin 0.06597 0.06597 0.06597 0.06597 0.06597

Breast 0.7000 0.7000 0.3000 0.3000 0.1000

Ovary 0.7185 0.7185 0.7185 0.7185 0.7185

Bladder 1.049 1.049 1.049 1.049 1.049

Kidney 0.2938 0.2938 0.2938 0.2938 0.2938

Thyroid 0.3333 0.3333 0.1667 0.1667 0.1667

Leukemia 1,176 284.9 370.06 178.8 157.1

a The coefficients were derived using several models applied to data from A-bomb survivors and selected medical

exposures.

b A = absolute risk with coefficient units of 10 -4 (Gy y) -1 ; R= relative risk with coefficient units of Gy -1

.



### A-63

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