Role of reactive oxygen in tumor promotion: implication of superoxide anion in promotion of neoplastic transformation in JB-6 cells by TPA

Abstract

The role of reactive oxygen (RO) in the promotion of neoplastic transformation of JB6 mouse epidermal cells by 12-O-tetradecanoylphorbol-13-acetate (TPA) was investigated using inhibitors of RO itself or RO generating systems of seven different types. Bovine erythrocyte CuZn superoxide dismutase (SOD) maximally decreased anchorage-independent (AI) colony induction by TPA in semi-solid agar in a dose-dependent manner to 10% of TPA control level. The inhibitory effect was specifically on induction of transformation, not expression of transformation. Copper (II) (3,5-diisopropylsalicylic acid)2, which exhibits biomimetic SOD activity, was also effective. Two enzyme eliminators of H2O2, catalase and glutathione peroxidase, failed to prevent TPA-promotion. Among three hydroxyl radical scavengers, D-mannitol and Na-benzoate were moderately active but tetramethylurea did not specifically inhibit AI colony induction by TPA. A quencher of singlet oxygen, 1,4-diazobicyclo-[2,2,2]octane was also inactive. Antioxidants blocked AI transformation by TPA moderately (n-propyl gallate and tannic acid) or weakly (BHA). BHT did not specifically inhibit promotion of transformation. The effects of three inhibitors of the arachidonic acid cascade were examined. NDGA and quercetin (lipoxygenase inhibitors) were moderately active but indomethacin (cyclooxygenase inhibitor) was much less active. Based on these results, we suggest that superoxide anion (O2-.) is required for promotion of transformation by TPA. H2O2 and 1O2 appear not to be required. Hydroxyl radicals and lipid peroxides, possibly associated with O2-. action or formed in the course of oxidative metabolism of arachidonic acid also appear to be required but to a lesser extent. Products of the lipoxygenase pathway of arachidonic acid metabolism but not the cycloxygenase pathway may be important in promotion of transformation by TPA in JB6 mouse epidermal cells. The epidermal cells themselves can be both the source of and the target of the reactive oxygen in promotion.

Response Modification in Carcinogenesis

“…The evidence is convincing that oxidants and agents that induce a cellular prooxidant state can act as carcinogens, in particular as promoters and progressors (30)(31)(32)(33)(34)(35)(36)(37)(38)(39)(40)(41)(42)(43)(44)(45)(46). Bona fide oxidants with promotional activity include H202, superoxide, ozone, hyperbaric oxygen, peroxyacetic acid, chlorobenzoic acid, benzoyl peroxide, cumene hydroperoxide, p-nitro-perbenzoic acid, and periodate (47,48).…”

Defective responses of transformed keratinocytes to terminal differentiation stimuli. Their role in epidermal tumour promotion by phorbol esters and by deep skin wounding

“…One could speculate that a reduced requirement for certain growth factors might also allow keratinocytes initiated by rasH activation to grow autonomously in zones of the epidermis where previously they could not (see Yuspa et al, 1985). Whether the irreversible change from a promoterdependent to a promoter-independent lesion is spontaneous or whether it involves TPA-induced chromosome changes (Kinsella & Radman, 1978;Emeritt & Cerutti, 1981;Parry et al, 1981) or gene amplications (Varshavsky, 1981) as a possible consequence of the ability of TPA to generate free radicals (Troll et al, 1982) (Schwarz et al, 1984) and the effect of TPA on anchorage independence is blocked by both radical scavengers (Nakamura et al, 1985) and retinoids (Colburn, 1979) the last of which specifically inhibits stage II promotion (Slaga et al, 1980a).…”

Structure-Activity Relationship and Mechanistic Studies on the Chemopreventive Activity of Dipyridamole and Its Analogues

“…TPA treatment in JB6 P + cells leads to the generation of ROS, which stimulates cell growth in soft agar or in monolayer (216). TPA-induced tumor promotion requires elevated levels of superoxide anion (216,217).…”

“…TPA-induced tumor promotion requires elevated levels of superoxide anion (216,217). DPM and analogues have been shown to inhibit lipid peroxdiation in mitochondria (218).…”