Effects of the protease inhibitor antipain on cell malignant transformation

Background: Several natural products have been found to exhibit a chemoprev entive activity both in in vivo and in vitro experimental systems. Among th em, protease inhibitors seem to play a key role in the regulation of growth and phenotypic expression of transformed cells as well as in the regulatio n of the late events of carcinogenesis. We evaluated the effect of antipain (RP), a natural protease inhibitor; on chemically induced BALB/c 3T3 cell transformation, on invasion and chemotactic motility of transformed cells a nd on their gelatinase expression. Methods: BALB/c 3T3 cells were platen an d exposed to 2.5 mu g/ml S-MCA or 50 mu g/ml I,2-DBE. The effect of a non-c ytotoxic dosage of AP (10 mu M) was studied by: a) pretreating cells with A P for 48 hours before the carcinogen exposure; b) adding AP simultaneously to the carcinogen treatment; c) chronic addition of AP at each medium chang e throughout the experimental duration. The effectiveness of the treatment was analysed as the ability to reduce ol inhibit the occurrence of transfor med foci. Modulation of the invasive phenotype by anti-transforming dosages of AP was evaluated by in vitro Matrigel invasion assay. Gelatin zymograph y was performed in order to assess AP regulation of proteolytic enzymes, su ch as metalloproteases, involved in invasion and metastasis. Results: AP tr eatment can reduce the transformation rate both in 3-MCA- and 1,2-DBE-initi ated cells. Its effectiveness depends on the administration schedule, and c hronic addition seems to be the most effective treatment. The concentration of AP, which is effective in the anti-transformation assay, is not able to significantly affect the migration and invasion of chemically transformed cells or their gelatinase activity. Conclusions: AP can suppress chemically induced BALB/c 3T3 cell transformation through mechanisms which do not inv olve modulation of the invasive phenotype.