A New Mechanism of Action for the Anticancer Drug Mitomycin C: Mechanism-Based Inhibition of Thioredoxin Reductase

Abstract

Mitomycin C (MMC) is a chemotherapeutic drug that requires an enzymatic bioreduction to exert its biological effects. Upon reduction, MMC is converted into a highly reactive bis-electrophilic intermediate that alkylates cellular nucleophiles. Alkylation of DNA is the most favored mechanism of action for MMC, but other modes of action, such as redox cycling and inhibition of rRNA, may also contribute to the biological action of the drug. In this work, we show that thioredoxin reductase (TrxR) is also a cellular target for MMC. We show that MMC inhibits TrxR in vitro, using purified enzyme, and in vivo, using cancer cell cultures. The inactivation presents distinctive parameters of mechanism-based inhibitors: it is time- and concentration-dependent and irreversible. Additionally, spectroscopic experiments (UV, circular dichroism) show that the inactivated enzyme contains a mitosene chromophore. On the basis of kinetic and spectroscopic data, we propose a chemical mechanism for the inactivation of the enzyme that starts with a reduction of the quinone ring of MMC by the selenolthiol active site of TrxR and a subsequent alkylation of the active site by the activated drug. We also report that MMC inactivates TrxR in cancer cell cultures and that this inhibition correlates directly with the cytotoxicity of the drug, indicating that inhibition of TrxR may play a major role in the biological mode of action of the drug.

Novel Oxidatively Activated Agents Modify DNA and Are Enhanced by Ercc1 Silencing

“…45 The active agents quickly react with DNA, glutathione, thioredoxin, or protein to produce irreversible adducts. 46,47 Similar profiles are observed for chlorambucil, which rapidly produces DNA lesions in minutes. 48 …”

Topical Mitomycin-C enhances subbasal nerve regeneration and reduces erosion frequency in the debridement wounded mouse cornea

“…Despite its use in the clinic, MMC is a cytotoxic agent that can bind covalently to DNA (Paz, et al, 2012). It is inert in its original form but is converted into an extremely reactive electrophile after its quinone ring is reduced.…”

“…It is inert in its original form but is converted into an extremely reactive electrophile after its quinone ring is reduced. Activation of MMC is mediated by several different reductases (Paz, et al, 2012) and activated MMC can alkylate DNA, RNA, and proteins affecting their activity. MMC is an irreversible inhibitor of the activity of the selenoprotein thioredoxin reductase (TrxR) (Arnér and Holmgren, 2006).…”

Oxidation of structural cysteine residues in thioredoxin 1 by aromatic arsenicals enhances cancer cell cytotoxicity caused by the inhibition of thioredoxin reductase 1

“…These unique properties confer the selenocysteine of TrxR1 a cellular target of many electrophilic compounds, including anticancer drugs like cisplatin, mitomycin C, doxorubicin, etc. [35][36][37]. TrxR1 is also a target of ATO [10].…”