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1365-1373, 1987
COMMENTARY
Peroxyl free radicals: potential mediators of tumor initiation and
promotion
In recent years, there has been an explosion of interest in the pATa = 4.88
involvement of free radicals in carcinogenesis (1 - 4 ) . This seems O-O- -O-O-
a natural outgrowth of the long overdue realization that free radi- 7s
cals are formed in biological systems and that they contribute
(CH2)4CH3 (CH 2 ) 4 CH 3
HO2C(CH2)7 HO2C(CH2)7
(CH2)4CH3
HO2C(CH2)
Fig. 1. Mechanism of unsaturated fatty acid metabolism by hematin. Hematin reduces unsaturated fatty acid hydroperoxides by one electron to form alkoxyl
radicals that cyclize to the adjacent double bonds generating an epoxy allylic radical. This radical either couples to the hydroxyl group bound to the heme or
couples to O 2 to form a peroxyl radical. The peroxyl radical is the oxidant that epoxidizes BP-7,8-diol. This mechanism is consistent with experimental
evidence presented in detail in ref. (34).
fuse to cellular loci remote from the site of their generation. Thus, P-450 so arachidonic-acid-dependent cooxidation constitutes a
they should be considered prime mediators of radical-induced distinct and complementary pathway to NADPH-dependent oxi-
pathology. dation for carcinogen activation (26).
One possibility relevant to carcinogenesis is that peroxyl radi-
cals react with DNA. Although a good deal is known about the
reaction of hydroxyl radical with DNA, the kinetics and prod-
ucts of reaction of peroxyl radicals with DNA have not been
explored so it is difficult to speculate on the likelihood of this
possibility. Because of their stability, peroxyl radicals will be
much more selective than hydroxyl radical in their reactions with Arachidonic Acid PGG2 PGH,
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Peroxyl free radicals
P-450
1
HO "'
ROO HO1'"
Equation 5
(+) - anti
The epoxide oxygen introduced into BP-7,8-diol by the hematin
system derives from O2 rather than hydroperoxide and epoxi-
dation is potently inhibited by antioxidants (31). These obser-
vations suggest that epoxidation occurs by a free radical o..
mechanism. Studies with a series of hydroperoxide analogs in-
dicate that fatty acid hydroperoxides with double bonds in prox-
imity to the hydroperoxide group support epoxidation but
saturated fatty acid hydroperoxides do not (33). Identification of P-450 (+)-syn
the products of unsaturated fatty acid hydroperoxide metabolism
by hematin and the results of oxygen-labeling experiments pro-
vide evidence for a mechanism of hydroperoxide metabolism
1367
L.JwMarnett
UNTREATED INDUCED
inducer
P-450 (low) P-450c (high)
1368
Peroxyl free radicals
isoenzyme in mouse skin that catalyzes arene oxide formation (a cytochrome P-450-requiring reaction) occurs in the absence
is not known, nor is it clear whether the isoenzyme is present of induction. Perhaps BP is a much better substrate for the trace
in epidermal tissue prior to hydrocarbon administration or is in- amounts of cytochrome P-450 in uninduced epidermal cells.
duced as a result of treatment (vide infra). The terminal step in Rapid cytochrome-P-450-dependent metabolism of 5-methyl-
the metabolic activation pathwayoxygenation of dihydrodiols chrysene has been observed in mouse skin in vivo (56) which
to dihydrodiolepoxidesis effected by peroxyl radicals or by may indicate that unoxidized polycyclic hydrocarbons are good
cytochrome P-450. The relative contribution that either of the cytochrome P-450 substrates. A direct comparison of the rate
oxidizing agents make to this step probably depends on the state of oxidation of BP and BP-7,8-diol by epidermal enzymes has
of induction of the epidermal cells. If little induction has occurred, not been performed.
peroxyl radicals may make a major contribution to oxidation but Perhaps only trace amounts of metabolism, an amount that can
if significant induction has occurred, cytochrome P-450 should be supplied by the balance of oxidizing enzymes present in un-
predominate (vide supra). induced skin, is required to initiate tumorigenesis. Immunochemi-
These findings raise some provocative questions about the cal experiments indicate that rat skin contains detectable
pathways of metabolic activation in mouse skinhistorically the cytochrome P-450c only following induction and then exclusive-
most important target tissue for polycyclic hydrocarbons. Early ly in follicular cells (57). Skin from uninduced animals exhibits
studies of the metabolism of DMBA in mouse skin established primarily cytochrome P-450),, again only in follicular cells (57).
the kinetics of disappearance of the hydrocarbon, the kinetics It is important to realize that the deoxynucleoside adducts quanti-
1369
L.J.Marnett
HO2C
HO2C-
cell culture than do the parent molecules (78). The opinion may References
have arisen that retinoids do not require metabolic activation and 1. Copeland,E.S. (1983) Free radicals in promotion a chemical pathology
although this may be correct for hormonal activities or even study section report. Cancer Res., 43, 5631-5637.
chemopreventive activities in internal organs, it may not be poss- 2. Troll.W. and Weisner,R. (1985) The role of oxygen radicals as a possible
ible to extrapolate this conclusion to mouse skin, a tissue exposed mechanism of tumor promotion. Annu. Rev. Pharmacol. Toxicol., 25, 5 0 9 -
528.
to high oxygen tensions and possessing active oxygenase-depen- 3. Cerutti,P. (1985) Prooxidant states and tumor promotion. Science, 227, 375 -
dent metabolic pathways. In fact, 13-c/.r-retinoic acid is the most 381.
reactive molecule toward peroxyl radicals that we have worked 4. Kensler,T.W. and Taffe.B.G. (1986) Free radicals in tumor promotion. Adv.
with. Nearly complete oxidation in vitro is detected at initial Free Radical Biol. Med., 2, 347-387.
retinoid concentrations of 1 /tM, a concentration that approaches 5. Pryor.W.A. (1986) Oxy-radicals and related species: their formation, life-
times, and reactions. Annu. Rev. Physioi, 48, 657-667.
the levels attained in tumor promotion experiments. If BP-7,8-diol 6. Fee,J.A. and Valentine,J.S. (1977) Chemical and physical properties of super-
is epoxidized in mouse skin cells by peroxyl radicals, it is virtually oxide. In Michelson.A.M., McCord,J.M. and Fridovich.I. (eds), Superoxide
certain that 13-c-retinoic acid will be epoxidized under similar and Superoxide Dismutases. Academic Press, New York, pp. 1960.
conditions because it is more reactive to peroxyl radicals. The 7. Behar.D., Czapski.G., Rabani,J., Dorfman.L.M. and Schwarz.H.A. (1970)
ease of oxidation of certain retinoids by transient oxidants likely The acid dissociation constant and decay kinetics of the perhydroxy radical.
J. Phys. Chem., 74, 3209-3213.
to be generated in promotion-sensitive cells may provide a new 8. Haber.F. and Weiss.J. (1934) The catalytic decomposition of hydrogen per-
basis for testing the biochemical mechanisms of their antipromoter oxide by iron salts. Proc. R. Soc. land. Ser. A, 147, 332-351.
1371
L.J.Marnett
29. Marnett,L., Johnson,J.T. and Bienkowski.M.J. (1979) Arachidonic acid Science, 199, 778-781.
dependent metabolism of 7,8-dihydroxy-7,8-dihydro-benzo[a]pyrene by ram 51.Bigger,C.A.H., Sawicki.J.T., Blake,D.M., Raymond,L.G. and Dipple.A.
seminal vesicles. FEBS Lett., 106, 13-16. (1983) Products of binding of 7,12-dimethylbenz[a]anthracene to DNA in
30. Sivarajah,K., Mukhtar,H. and Eling,T.E. (1979) Arachidonic acid dependent mouse skin. Cancer Res., 43, 5647-5651.
metabolism of ( ) trans-7,8-dihydroxy-7,8-dihydro-benzo[a]pyrene (BP-7,8- 52. Kinoshita.N. and Gelboin.H.V. (1972) Aryl hydrocarbon hydroxylase and
diol) to 7,10/8,9 tetrols. FEBS Lett., 106, 17-20. polycyclic hydrocarbon tumorigenesis: effect of the enzyme inhibitor 7,8-
31. Dix.T.A. and Marnett,L.J. (1981) Free radical epoxidation of 7,8-dihydroxy- benzoflavone on tumorigenesis and macromolecule binding. Proc. Natl. Acad.
7,8-dihydrobenzo[a]pyrene by hematin and polyunsaturated fatty acid hydro- Sci. USA, 69, 824-828.
peroxides. J. Am. Chem. Soc, 103, 6744-6746. 53. Kinoshita,N. and Gelboin.H.V. (1972) The role of aryl hydrocarbon hydroxyl-
32. Marnett.L.J. and Bienkowski,M.J. (1980) Hydroperoxide dependent oxygen- ase in 7,12-dimethylbenz[a]anthracene skin tumorigenesis: on the mechanism
an'on of rrarts-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene by ram seminal vesicle of 7,8-benzoflavone inhibition of tumorigenesis. Cancer Res., 32, 13291339.
microsomes. Source of the oxygen. Biochem. Biophys. Res. Commun., 96, 54. Bowden.G.T., Slaga.T.J., Shapas.B.G. and Boutwell.R.K. (1974) The role
639-647. of aryl hydrocarbon hydroxylase in skin tumor initiation by 7,12-dimethyl-
33. Dix,T.A., Fontana,R., Panthani.A. and Marnett,L.J. (1985) Hematin-cata- benz[a]anthracene and 1,2,5,6-dibenzanthracene using DNA binding and
lyzed epoxidation of 7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene (BP-7,8-diol) thymidine-3H incorporation into DNA as criteria. Cancer Res., 34, 2634-
by polunsaturated fatty acid hydroperoxides. J. Biol. Chem., 260, 5358-5365. 2642.
34. Dix,T.A. and Marnett.L.J. (1985) Conversion of linoleic acid hydroperoxide 55. Pyerin,W.G. and Hecker,E. (1979) On the biochemical mechanism of tumori-
to hydroxy, keto, epoxyhydroxy and trihydroxy fatty acids by hematin. J. genesis in mouse skin. IX. Interrelation between tumour initiation by 7,12-
Biol. Chem., 260, 5351-5367. dimethylbenz[a]anthracene and the activities of epidermal arylhydrocarbon
35. Mahoney.L.R., Johnson,M.D., Korchek,S., Marnett,L.J. and Reed.G.A. monooxygenase and epoxide hydratase. J. Cancer Res. Clin. Oncol., 93,
(1982) Inhibition of aldehyde oxidation by polycyclic aromatic hydrocarbons. 7-30.
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