Hydrocarbons: Mechanisms of Cellular

Toxicity

Organohalides Are Lipophilic

• Less available for excr’n• Accumulate in fatty tissues, fat

stores– Fatty tissue ½ life may be > 100 d– Secr’d in milk

• Bioaccumulate in aquatic spp– Partition out of water Sediments Fish fatty tissue

• Organisms may accumulate 3-6x (or more) amt in env water

www.ecoinfo.org/.../gbhtoxin/gbhtoxin_e.cfm

Polynuclear Aromatic Hydrocarbons

• Benzo(a)pyrene, benz(a)anthracene models

• Metabolized by mixed function oxidases– Cytochromes P450

Reactive diol epoxides• Ox’d forms in bay region most toxic

bay region

Halogenated Hydrocarbons May Increase Their Own

Toxicity• Work

through Ah receptor

• Induce prod’n Cyt P450 enz’s Oxidized

cmpds– Ex: 7,8-

Dihydro-7,8-dihydroxy-benzopyrene 9,10 epoxide

The Ah–receptor binds four classes of substances: Dibenzodioxines (i.e TCDD) A, Dibenzofuranes, B Biphenyls C and polyaromatic hydrocarbons D. If such substances reach the receptor E, they trigger a chemical signal, which will finally result in toxic phenomena: tumor growth, skin toxicity (i.e. Ah mediated chloracne) F, immunotoxicity as well as developmental toxicity. The dioxin receptor belongs to the class of receptors mediating toxicity, which are preferentially modelled by QSAR methods. 3r-training.tierversuch.ch/.../dioxin.html

• Hydrocarbon binds Ah receptor in cytosol

• Complex translocates nucleus Specific recognition sites on

DNA for complex – Ah gene locus

• Now transcr’n, transl’n initiated– Specific genes code for AHH

• Aryl hydrocarbon hydroxylase– Phase I enzyme

– Structural gene for cytosolic receptor

• Now more metabolic (Cyt P450) enz’s

• Catalyze more ox’n of parent cmpd• Product now more toxic

– Incr’d carcinogenesis• May covalently bind DNA Mutations, incr’d repl’n

Increased Toxicity of Metabolites

PolyChlorinated Biphenyls• “PCB’s”• 210 poss congeners

– Chem cmpd closely related to another in composition

– Exerting similar or antagonistic effects– Something derived from same source

or stock• Widespread, persistent

– In fat tissue of most humans

• Two most potent– 3,4,3’,4’-tetrachloro biphenyl– 3,4,5,3’,4’,5’-hexachloro biphenyl

Halogen Substitutions

• Cl is a halogen– F, Cl, Br, I– Reactive– Wants 1 e- to

fill outer shell• Form ions w/

single negative charge

• In halogenated hydrocarbons, Cl covalently bound to C– Electronically stable bond

• Most mol’s containing Cl man-made incr’d molecular stability, incr’d

MW, incr’d bpt/mpt

• Cl substitutions on biphenyl often balanced

• When ortho substituted w/ Cl– Cl relatively large– 2 planar rings can’t rotate

• Rotation hindered by Cl’s

– When no Cl, 2 planar rings can rotate• About C1, C1’ bond

PCB Toxicity

• Low acute toxicity• Chronic exposure not understood

– Probably more harmful

• Combines w/ receptor– Ah (aryl hydrocarbon) receptor– PCB PCB-Receptor Nucleus

DNA alter transcription effect on cell

Three Modes of Action

• Bind cell macromol’s– DNA– “Stacks and sticks” to proteins

• Accumulate in lipid-rich cell components

• Reversibly bind receptors, enzymes– At specific sites– Ah receptor has great affinity for TCDD,

PCB’s• Via Cl’s

PCB Biotransformation

• Bioactivation depends on planarity• More toxic (and more similar to

dioxin) when– Coplanarity of rings– Cl’s at m, p positions

• Metab’d through Phase I and Phase II

Phase I Metab

• Cytochrome P450 monooxygenase common– Indirect hydroxylation

• OH added, then db in ring shifts

– Epoxidation• --O– added over db in ring

– May or may not shift or lose Cl– Metab rate depends on #, placement

Cl’s• More rapid if >4 Cl’s and H’s on C’s 4,5

Phase II Metab

• Phase II Reactions– Conjugated to glucuronic acid

• Rapidly excr’d

– Conjugated to GSH Mercapturic acid• Excr’d or reabs’d

Common Effects of PCB’s

• Note: many effects are species-specific

• Coplanarity of rings, #Cl’s related to potency

• Chloracne– Acneiform eruption w/ exposure– Milder than w/ TCDD

• Probably diff mech

• Epithelial cell changes– Hyperplasia – Incr’d cell # w/ incr’d

cell div’n– Hypoplasia – Decr’d cell division

decr’d # proliferating cells– Impt to changes in sev organs

• Hepatomegaly• Gastric mucosal changes

Ulceration, hemorrhage– Species specific

– May play role in carcinogenesis• Hyperplasia = incr’d cell division; may be

precursor for …• Cancer = unrestrained cell division

• Vit A depletion– May be linked to Ah receptor

• Heme prod’n inhib’d– Get build-up of porphyrins which are toxic

• Immunosuppression– Lymph glands

• Spleen enlargement• Thymus gland atrophy

– Total serum Ig’s decline (species specific)• Humans: IgA, IgM sig depressed• Phagocyte #, T cell response depressed• From Japan, Taiwan PCB poisoning epidemics

• Nervous system disorders– Catecholamine levels changed– Behavioral, learning dysfunctions

• Offspring following prenatal exposure

• Endocrine disruption– Next week’s lecture

Dioxins

• Also widespread• Also hydrophobic• Most toxic

– 2,3,7,8-tetrachlorodibenzo-p-dioxin

• “Supertoxic” cmpd– Extremely potent – Diff isomers differ in toxicity– Contaminant of other chlorinated

cmpds

• Induces microsomal enz’s– May be through Ah receptor

• TCDD binds Ah receptor in cytosol– Some spp (even diff strains) have

varied amts Ah receptor in cytosol– More receptor/higher sensitivity to

TCDD

• Complex translocates nucleus Specific recognition sites on

DNA for complex – Ah gene locus

• Now transcr’n, transl’n initiated– Specific genes code for AHH

• Aryl hydrocarbon hydroxylase– Phase I enzyme

– Structural gene for cytosolic (Ah) receptor

– Other prot’s impt for immune, inflamm responses (?)

– Other prot’s impt to cell viability, replication

http://www.med.ufl.edu/pharm/facdata/Shiveric/images/TCDD.gif

Toxicity in Humans Uncertain

• Incr’d cancer mortality?– Carcinogenic in rats (@ 2 ppb)

Cancers of liver, resp tract, mouth, others

– “Not reported to be carcinogenic in humans in spite of its extremely potent carcinogenicity in rats” (WHO, 1989)

– German pesticide workers suffer 39% higher cancer mortality rate compared to other Germans. (Hanson, David; 1991)

• Chloracne– Most characteristic lesion in humans– Related to Ah receptor

• Hepatotoxicity at high doses– See hyperplasia, enzyme induction, others

• Immunosuppression– T cells most likely target– Species specific– Reversible

• Teratogenicity/fetotoxicity– Cleft palate/Ah receptor linked in mice?– Prenatal exposure biochem & behavioral

changes in offspring

Some Problems in Assessing Toxicity

• TCDD, congeners may be by-products of manufacture along with PCB’s– So which is responsible for toxicity?

• DDT often present in mixtures, animals, environment– Pesticide, but also manufacturing by-

product– OR may be in environment previously– OR may have been in animal fat from

earlier exposure

• One toxicant may influence others in mixture– Synergism or antagonism

• Which in a mixture is most toxic?– Can’t fully assess mechanisms

• Difficult to assess environmental risk– Molecule in largest concent may not be

most toxic• If one toxicant induces metab of

another: ????• Species specificity impt

– Fat stores? (ex: mink)