Free radical derivatives of
peroxides, hydroperoxides, and
anthrones are thought to mediate
tumor promotion by these compounds. Further, the promoting activity of
phorbol esters is attributed, in part, to their ability to stimulate the cellular generation of
oxygen radicals. A
hydroperoxide metabolite of
butylated hydroxytoluene, 2,6-di-tert-butyl-4-hydroperoxyl-4-methyl-2,5-cyclohexadienone (
BHTOOH), has previously been shown to be a
tumor promoter in mouse skin.
BHTOOH is extensively metabolized by murine keratinocytes to several radical species. The primary radical generated from
BHTOOH is a
phenoxyl radical that can disproportionate to form
butylated hydroxytoluene quinone methide, a reactive electrophile. Since electrophilic species have not been previously postulated to mediate
tumor promotion, the present study was undertaken to examine the role of this electrophile in the promoting activity of
BHTOOH. The
biological activities of two chemical analogs of
BHTOOH, 4-trideuteromethyl-BHTOOH and 4-tert-butyl-BHTOOH, were compared with that of the parent compound. 4-Trideuteromethyl-BHTOOH and 4-tert-butyl-BHTOOH have a reduced ability or inability, respectively, to form a
quinone methide; however, like the parent compound, they both generate a
phenoxyl radical when incubated with keratinocyte cytosol. The potency of
BHTOOH, 4-trideuteromethyl-BHTOOH, and 4-tert-butyl-BHTOOH as inducers of
ornithine decarboxylase, a marker of
tumor promotion, was commensurate with their capacity for generating
butylated hydroxytoluene quinone methide. These initial results were confirmed in a two-stage
tumor promotion protocol in female SENCAR mice. Together, these data indicate that a
quinone methide is mediating
tumor promotion by
BHTOOH, providing direct evidence that an electrophilic intermediate can elicit this stage of
carcinogenesis.