Modulation of mitomycin C mutagenicity on Saccharomyces cerevisiae by glutathione, cytochrome P-450, and mitochondria interactions

Abstract

It is well established that most anticancer drugs also have mutagenic effects and require metabolic activation before exerting their mutagenic/antiblastic activity. Antitumoral compound effects strongly depend on the biochemical/physiological conditions of the tumoral cells, and especially on the activation of specific drugs metabolizing enzymes and on respiration. We examined the mitomycin C-induced mutagenic effects on the D7 strain of Saccharomyces cerevisiae and on its derivative mitochondrial mutant ρ° at different contents of glutathione and cytochrome P-450, molecules able to activate/detoxicate xenobiotics. The mutagenic activity of the drug was evaluated as frequency of mitotic gene conversion and reversion in different physiological conditions. The highest frequencies of reversion and especially of gene conversion were observed at the highest cytochrome P-450 contents in the D7 strain with a further increase at high glutathione level. In the respiratory-deficient strain, the highest frequency of convertants was shown at low glutathione level and lack of cytochrome P-450. These results suggest the relevance of mitochondrial functionality for the expression of genotoxic activity of this anticancer drug.

Introduction

Mitomycin C (MMC) is a quinone-containing natural antibiotic used in clinical cancer chemotherapy against a variety of solid neoplasms (breast, prostate, bladder, colorectal, gastric and lung cancers). However, emergence of drug-resistant tumor cells limits the clinical effectiveness of MMC.

Mitomycin C requires enzymatic activation for its cytotoxic activity 1, 2, 3, 4. Following bio-activation, MMC is capable of producing oxyradicals as well as DNA alkylating species due to the presence of both quinone and aziridine moieties, respectively 3, 5, 6, 7. The active oxygen species can produce DNA strand breaks [8], and this activity as well as the cross-linking of DNA have been implicated in the biological action of MMC 1, 3, 5, 6, 7, 8, 9, 10, 11, 12.

Multiple drug resistance mechanisms are present in resistant cells, including decreased drug activation, increased drug detoxication, and decreased accessibility of DNA targets. In particular, MMC cellular resistance can be modulated by an increased intracellular level of glutathione (GSH) 10, 13, 14, 15, 16, 17, 18.

Cytochrome P-450 is a product of a multigene family, and catalyzes the activation and the detoxication of a wide variety of exogenous as well as endogenous compounds. In general, cancer cells express lower amounts of cytochrome P-450 as compared to normal cells 19, 20, 21, 22.

Drug activity can also be dependent on mitochondrial functionality as observed by the different behavior of oxygenic and hypoxic tumor cells versus antitumor chemicals 19, 23, 24, 25. However, MMC does not inhibit mitochondrial respiration in isolated rat hepatocytes [26]. In any case, MMC has been shown to be preferentially toxic to hypoxic tumor cells both in vitro 5, 27and in vivo [28].

There is clear evidence that the cytotoxicity and genotoxicity of antiblastic compounds are governed by the different biochemical/physiological conditions of the tumoral cells, determined especially by the activation of specific drug-metabolizing enzymes and respiration.

We examined the mutagenic effects induced by MMC in two strains, D7 [29]and D7ρ° (respiratory-deficient strain, obtained in our laboratory), of Saccharomyces cerevisiae in different conditions of expression of glutathione and cytochrome P-450 contents. S. cerevisiae may provide a valuable model for understanding factors governing the movement of a variety of drugs 30, 31, 32.

A previous study was performed to choose the experimental conditions for cellular concentration of glutathione and cytochrome P-450 (unpublished data).