Epigenetic Mechanisms of Action of Carcinogenic ... - ACS Publications

5 Epigenetic Mechanisms of Action of Carcinogenic Organochlorine Pesticides

Downloaded by UCSF LIB CKM RSCS MGMT on November 28, 2014 | http://pubs.acs.org Publication Date: August 10, 1981 | doi: 10.1021/bk-1981-0160.ch005

GARY M. WILLIAMS Naylor Dana Institute for Disease Prevention, American Health Foundation, Valhalla, NY 10595

Many of the most widely used chlorinated c y c l i c hydrocarbon compounds have been found to be carcinogenic in experimental laboratory rodents (Table I ) . Table I.

Carcinogenicity of Chlorinated C y c l i c Hydrocarbon Pesticides

Comp ound

P r i n c i p a l Target Organ Rat References Mouse

Aldrin Chlordane

liver liver, uterus liver

Chlorobenzilate DDT Dieldrin Heptachlor Hexachlorobenzene Hexachlorocyclohexane (BHC), lindane Kepone

liver, lung liver liver liver liver liver

1 2 a

NS liver

3 3,4,5

NS thyroid

6, 7 8 9

liver liver,

10,11 12

thyroid Mirex PCB a

liver liver

1iver liver

3,13 14,15

no s i g n i f i c a n t increase i n neoplasms

C y c l i c hydrocarbons with chlorine substituents that block ring oxidation are resistant to biodegradation and thus accumulate in the environment and persist for long periods in animals once they are absorbed. The persistence of organochlorine pesticides 0097-6156/81/0160-0045$05.00/0 © 1981 American Chemical Society In The Pesticide Chemist and Modern Toxicology; Bandal, S., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1981.


46

THE PESTICIDE CHEMIST AND MODERN TOXICOLOGY

together with their animal carcinogenicity has given r i s e to concern that exposed humans would be at risk of cancer development from these chemicals (16,17, 18). Indeed, extrapolation of dose-response effects from rodents to humans predicts substantial cancer causation (16). However, epidemiologic studies of highly exposed groups have f a i l e d to reveal any s i g nificant increase in cancer occurrence (19,20) and no increase in cancer incidence has been associated with pesticide usage (21). Such a discrepancy suggests that the mechanism of action of chlorinated c y c l i c hydrocarbons may be different from that of other c a r cinogens which produce cancer in both experimental animals and humans (22,23). This p o s s i b i l i t y is f u r ther supported by the unusual s i t u a t i o n that all carcinogens of this s t r u c t u r a l type have the l i v e r as their p r i n c i p a l target organ. For carcinogens that are activated to reactive metabolites, members of a s t r u c t u r a l type almost always affect more than one organ and often the p r i n c i p a l organ affected varies with the s p e c i f i c compound. For these and other reasons, we have suggested that chlorinated c y c l i c hydrocarbons may be carcinogenic to rodents by i n d i r e c t mechanisms (22,23,24). Mechanisms

of C a r c i n o g e n e s i s

Chemical carcinogens are defined operationally by their ability to induce tumors i n exposed animals. A highly diverse collection of c h e m i c a l s i s capable of p r o d u c i n g this effect, including organic and i n o r g a n i c c h e m i c a l s , s o l i d s t a t e m a t e r i a l s , h o r mones a n d immunosuppressants. The h e t e r o g e n e i t y of structures r e p r e s e n t e d makes i t i m p r o b a b l e thata l l chemicals would a c t through a single mechanism. T h e r e f o r e , W e i s b u r g e r a n d W i l l i a m s (2_3) h a v e p r o p o s e d a c l a s s i f i c a t i o n that separates chemical carcinogens i n t o two m a j o r c a t e g o r i e s , g e n o t o x i c a n d e p i g e n e t i c (Table I I ) . Table

II

Classes

of C a r c i n o g e n i c

Type

A.

chemicals Example

Genotoxic 1. D i r e c t - a c t i n g or primary carcinogen

Ethylene imine,b i s (chloromethy 1 )ether

In The Pesticide Chemist and Modern Toxicology; Bandal, S., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1981.

5.


Β·

WILLIAMS

Carcinogenic

Organochlorine

2.

Ρrocarcinogen or secondary carcinogen

3.

Inorganic

carcinogen

Epigenetic 4. Solid-state carcinogen 5. Hormone 6. 7.

Immunosuppressor Cocarcinogen

8.

Promoter

Pesticides

47

V i n y l c h l o r i d e , benzo(a)pyrene,2-naphty1amine, d i m e t h y l n i t r o s amine N i c k e l , chromium

Polymer or metal foils, asbestos E s t r a d i o 1, diethylstilbestrol Azathioprine, Phorbol esters, pyrene, c a t e c h o l , e t h a n o l , n-dodecane, Phorbol esters, bile acids, saccharin

C a r c i n o g e n s t h a t i n t e r a c t w i t h a n d a l t e r DNA a r e classified as g e n o t o x i c . Thus, the genotoxic c a t e gory c o n t a i n s t h e c h e m i c a l s t h a t f u n c t i o n as e l e c t r o philic reactants as originally postulated by t h e M i l l e r s (2^5). A l s o , b e c a u s e some i n o r g a n i c c h e m i c a l s have d i s p l a y e d such effects they have tentatively been p l a c e d i n t h i s c a t e g o r y . The s e c o n d b r o a d c a t e gory d e s i g n a t e d as e p i g e n e t i c c a r c i n o g e n s comprises t h o s e c h e m i c a l s f o r w h i c h no e v i d e n c e o f d i r e c t i n teraction with genetic material e x i s t s . This cate gory c o n t a i n s s o l i d s t a t e c a r c i n o g e n s , hormones, im munosuppressants, c o c a r c i n o g e n s and promoters. T h i s c l a s s i f i c a t i o n and t h e u n d e r l y i n g c o n c e p t s , i f u l t i m a t e l y v a l i d a t e d , have major i m p l i c a t i o n s f o r r i s k e x t r a p o l a t i o n t o humans o f d a t a on e x p e r i m e n t a l carcinogenesis. Genotoxic c a r c i n o g e n s , as a conse q u e n c e o f t h e i r e f f e c t s on g e n e t i c m a t e r i a l , p o s e a clear qualitative hazard. These carcinogens are occasionally effective a f t e r a s i n g l e exposure, a r e o f t e n c a r c i n o g e n i c a t low d o s e s , a c t i n a c u m u l a t i v e manner, usually produce irreversible effects, and produce combined e f f e c t s w i t h o t h e r g e n o t o x i c c a r c i n ogens h a v i n g t h e same t a r g e t organ. In c o n t r a s t , with some types of e p i s e n e t i c carcinogens, i t is known t h a t t h e c a r c i n o g e n i c e f f e c t s occur only with h i g h and s u s t a i n e d l e v e l s of exposure that lead to prolonged p h y s i o l o g i c a b n o r m a l i t i e s , hormonal imbal ances, or t i s s u e i n j u r y . I n such cases, the e f f e c t s a r e o f t e n e n t i r e l y r e v e r s i b l e upon c e s s a t i o n of expo sure. Because of these f e a t u r e s , the r i s k from expo-

American Chemical Society Library 1155 16th St., N.W.

In The Pesticide Chemist and Modern Toxicology; Bandal, S., et al.; ACS Symposium Series; American Chemical Washington, DC, 1981. Washington, D.C.Society: 20036


48

sure to epigenetic carcinogens seems to quantitative nature. Thus, a major element i n a s s e s s i n g the hazard of a c h e m i c a l i s to e v a l u a t e its genot o x i c i t y .


Lack

of

Genotoxicity

of

Organochlorine

be

Pesticides

DNA Damage

d

_f

_b

-8

ND

-a

_b

DDE

ND

-b

Dieldrin

-a

-b

Chlordane

ND

+c,-d

ND

Heptachlor

ND

ND

_e

Kepone

ND

>

Measuring

Mu t a g e n e s i s Mammalian Ba c t e r i a l

DNA Re p a i r

DDT

a

pesticides short-term

III. A c t i v i t y i n Short-term Tests DNA I n t e r a c t i o n of C a r c i n o g e n i c Organochlorine Compounds

Comp ound

a

potential potential

The g e n e t i c e f f e c t s of o r g a n o c h l o r i n e h a v e b e e n e x a m i n e d i n a number o f in_ v i t r o tests (Table I I I ) . Table

of

-e

+c

»

f

+c _b

_h

>

+c

-b

y

_b

_h

_b b

c

) Swenberg (2_6) , ) Williams (24), ) Hart (27 , 2 8 ) , d) Flamm (2_9), ) Marshall (30), ) S h i r a s u ( 3J ) , 8) Ames (3_2) , ) S c h o e n y (3J3). e

f

h

Although the results have been predominantly negative, their significance has been m i n i m i z e d by the frequent suggestion that lack of activity is simply a c o n s e q u e n c e of the a b s e n c e of appropriate m e t a b o l i s m i n t h e i_n v i t r o t e s t s . In our laboratory we have developed several tests for genotoxicity u t i l i z i n g liver-derived cells (34,35). Since the organochlorine pesticides have the liver as their principal target organ, these t e s t s r e p r e s e n t the i d e a l system i n which to e v a l u a t e t h e g e n o t o x i c i t y , as w e l l as o t h e r e f f e c t s , o f t h e s e compounds · The h e p a t o c y t e primary c u l t u r e (HPC)/DNA r e p a i r t e s t a s s e s s e s the c a p a b i l i t y of c h e m i c a l s t o undergo


WILLIAMS

5.

Carcinogenic

Organochlorine

Pesticides

49

c o v a l e n t i n t e r a c t i o n w i t h DNA by m e a s u r e m e n t o f a u t o r a d i o g r a p h i c DNA r e p a i r e l i c i t e d as a r e s u l t of t h e DNA damage ( 3 6 , 3 7 ) . The f r e s h l y i s o l a t e d hepatocytes used i n t h i s t e s t r e t a i n a h i g h l e v e l of a c t i v i t y f o r b i ot r a n s f o r m i n g x e n o b i o t i c s and thus the test r e sponds t o a wide spectrum of s t r u c t u r a l types of c a r c i n o g e n s r e q u i r i n g m e t a b o l i c a c t i v a t i o n (3_4,35). Our p r e v i o u s r e p o r t s of l a c k of g e n o t o x i c i t y of organochlorine pesticides i n t h e r a t l i v e r HPC/DNA repair test (24,38) have been e x t e n s i v e l y c o n f i r m e d (Table IV).


—

Table

IV.

HPC/DNA

Repair

Results 3

Compound

1

grains/nucleus Rat Mouse Hamster

2 ,3-Dimethyl-4-aminobiphenyl

60

Biphenyl

-

Chlordane

-

DDT

ND

-

Mirex

ND

-

Kepone

-

ND

a

- = zero;

ND

25

>100

ND

= n o t done

In a d d i t i o n , s i n c e the o r g a n o c h l o r i n e p e s t i c i d e s a r e s o m e t i m e s more a c t i v e on mouse l i v e r , these r e sults were extended (3j$,3_9) t o t h e mouse liver d e r i v e d HPC/DNA r e p a i r t e s t , a s w e l l as t h e h a m s t e r l i v e r derived test (Table I V ) . Another liver-derived test for genotoxicity i s the adult rat liver epithelial cell (ARL)/hypoxanthine-guanine phosphoribosy1 transferase (HGPRT) mutagenesis assay (40,41). T h i s t e s t a s s e s s e s mutagenicity a t t h e HGPRT l o c u s t h r o u g h measurement o f c o n v e r s i o n of l i v e r e p i t h e l i a l c e l l s t o HGPRT-defici e n t mutants t h a t a r e r e s i s t a n t t o 6 - t h i o g u a n i n e . As w i t h t h e HPC/DNA r e p a i r t e s t , t h e c e l l s i n t h i s assay possess intrinsic metabolic capability f o r the b i o transformation of activation-dependent carcinogens (34 ). In spite of a mutagenic response to three genotoxic carcinogens, the organochlorine pesticides w e r e a l l n o n - m u t a g e n i c i n t h i s a s s a y (2_4 ) ( T a b l e V ) .



50 Table

V.

a

ARL /HGPRT


Compound

Mutagenesis

Assay

Concentration molar 0

Results I n d u c t i o n of HGPRT d e f i c i e n t mutants

Aflatoxin

10"

6

+

3-Methyl-4- d i m e t h y l aminoazobenzene

10'

5

+

2 - Aminoflu orene Chlordane

10"^ 2.5xl0"

Kepone

1 0~

Heptachlor

10~

5

Hexachlorocyclopentadiene

10*

6

Endrin

+ 5

-

3xl0~

3

10 -4

DDT

a

l i n e ARL 6 b - h i g h e s t n o n t o x i c d o s e t h a t was n e g a t i v e or lowest d o s e t h a t was p o s i t i v e . The c o n s i s t e n t l a c k of g e n o t o x i c i t y of o r g a n o chlorine pesticides in liver derived tests strongly supports the negative data obtained i n other tests. Thus, i t appears that these chemicals a r e not genotoxic carcinogens. E p i g e n e t i c Mechanism Pesticides.

of A c t i o n

of

Organochlorine

A t l e a s t one o r g a n o c h l o r i n e p e s t i c i d e , DDT, h a s b e e n shown t o be a l i v e r t u m o r p r o m o t e r (4_2)> e n h a n c ing the c a r c i n o g e n i c e f f e c t of 2-acetylaminofluorene when g i v e n a f t e r t h e c a r c i n o g e n . T h u s , we h a v e p o s tulated that the o r g a n o c h l o r i n e pesticides may be c a r c i n o g e n i c through a mechanism o f tumor p r o m o t i o n (2_2, 24, 3 8 ) . A l l of the i n b r e d s t r a i n s of r a t s and mice used f o r c a r c i n o g e n b i o a s s a y have a s p o n t a n e o u s i n c i d e n c e o f l i v e r t u m o r s w h i c h i n t h e c a s e o f some mouse s t r a i n s i s q u i t e h i g h (22). As p a r t o f t h i s



5.

WILLIAMS

Carcinogenic

Organochlorine

51

Pesticides

c o n d i t i o n , these a n i m a l s a l s o have a h i g h e r i n c i d e n c e of l e s i o n s regarded as p r e n e o p l a s t i c o r p o t e n t i a l l y neoplastic. T h u s , we p o s t u l a t e d t h a t t h e p r o m o t i n g e f f e c t of o r g a n o c h l o r i n e p e s t i c i d e s would enable the pre-existing abnormal liver cells to progress to a higher frequency of tumor development than would occur under c o n t r o l c o n d i t i o n s . The m e c h a n i s m o f t h e p r o m o t i n g e f f e c t of chemic a l s when a d m i n i s t e r e d a f t e r a p r i m a r y c a r c i n o g e n i s n o t y e t known. A compelling concept i s t h a t tumor p r o m o t e r s may a c t on t h e c e l l membrane. Under normal conditions, the c e l l s composing a tissue are i n homeostasis i n which the requirements f o r c e l l growth to balance cell loss are r e g u l a t e d throughout the tissue. The r e g u l a t i o n probably occurs through cell to c e l l communications. Interruption of such comm u n i c a t i o n s c o u l d p e r m i t c e l l s w i t h an a b n o r m a l genotype t o p r o l i f e r a t e beyond the normal growth r e q u i r e ments, t h a t i s t o form a neoplasm. Recently, several groups (4J>4_

Epigenetic Mechanisms of Action of Carcinogenic ... - ACS Publications

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