The ability of the hyperplasiogenic irritant ethyl phenylpropiolate (EPP) to act as a tumor promoter in two-stage carcinogenesis and to stimulate cellular events commonly cited as markers of tumor promoter action was evaluated. Treatment of adult, inbred SENCAR (SSIN) mice, initiated with 7,12-dimethylbenz(a)anthracene, with 5 mg of EPP twice weekly resulted in 100% of the mice developing tumors (4.8 tumors/mouse) after 40 weeks of promotion. Treatment with 3 mg EPP (twice weekly) resulted in 52% of the mice developing tumors (0.9 tumor/mouse). This treatment regimen with EPP produces a sustained epidermal hyperplasia without being overtly toxic. In addition, a 5-mg dose of EPP induced ornithine decarboxylase activity to a level comparable to that induced by the tumor promoter phorbol 12-myristate 13-acetate (PMA): 2.3 nmol CO2/mg protein/h for EPP versus 4.5 nmol CO2/mg protein/h for PMA versus 0.04 nmol CO2/mg protein/h for acetone control. Likewise, the time course of ornithine decarboxylase induction by EPP was the same as that seen with PMA (maximum induction at approximately 6 h). Vascular permeability of the dorsal skin increased significantly in response to EPP (8 times that seen in acetone controls) and exhibited the same kinetics as that seen after exposure to PMA. Activity of protein kinase C (PKC), the cellular receptor for PMA, decreased by 75 to 95% 48 h after treatment with PMA. In contrast, EPP treatment resulted in less than a 20% decrease in PKC activity 48 h after treatment. This slight decrease in PKC activity is thought to be an indirect effect caused by the hyperproliferative and inflammatory reactions, because EPP was found to be inactive as an in vitro activator of PKC. These results indicate not only that EPP is a good tumor promoter that causes morphological and biochemical responses similar to those induced by PMA, but also that the action of EPP is apparently mediated via a mechanism that does not involve direct interaction with PKC.