Abstract
The tumor suppressor p53 is commonly inactivated in human tumors, allowing evasion of p53-dependent apoptosis and tumor progression. The small molecule APR-246 (PRIMA-1Met) can reactive mutant p53 in tumor cells and trigger cell death by apoptosis. The thioredoxin (Trx) and glutaredoxin (Grx) systems are important as antioxidants for maintaining cellular redox balance and providing electrons for thiol-dependent reactions like those catalyzed by ribonucleotide reductase and peroxiredoxins (Prxs). We show here that the Michael acceptor methylene quinuclidinone (MQ), the active form of APR-246, is a potent direct inhibitor of Trx1 and Grx1 by reacting with sulfhydryl groups in the enzymes. The inhibition of Trx1 and Grx1 by APR-246/MQ is reversible and the inhibitory efficiency is dependent on the presence of glutathione. APR-246/MQ also inhibits Trxs in mutant p53-expressing Saos-2 His-273 cells, showing modification of Trx1 and mitochondrial Trx2. Inhibition of the Trx and Grx systems leads to insufficient reducing power to deoxyribonucleotide production for DNA replication and repair and peroxiredoxin for removal of ROS. We also demonstrate that APR-246 and MQ inhibit ribonucleotide reductase (RNR) in vitro and in living cells. Our results suggest that APR-246 induces tumor cell death through both reactivations of mutant p53 and inhibition of cellular thiol-dependent redox systems, providing a novel strategy for cancer therapy.
Highlights
The p53 tumor suppressor plays a key role in the protection against tumor growth[1,2]
Trx and Grx systems have been recognized as the primary electron donors to reduce RNR22 which is often induced in tumor cells[23]
We investigated if preheated APR246 acted as an inhibitor of Trx[1] and Grx[1] or through interaction with other components of the two systems, which was recently shown for TrxR115
Summary
The p53 tumor suppressor plays a key role in the protection against tumor growth[1,2]. The central role of p53 in stress-induced apoptosis, the frequent TP53 mutations in tumors, and the high levels of mutant p53 in tumor cells have stimulated efforts to develop novel cancer therapy by reactivating mutant p535. Trx and Grx systems have been recognized as the primary electron donors to reduce RNR22 which is often induced in tumor cells[23]. Many transcription factors are controlled by reduced Trx[1] and Grx[1], including p53, nuclear factor-κB (NF-κB), and Activator protein 1 (AP-1)[19,25,26,27]. The Trx[1] and Grx[1] systems are commonly upregulated in cancer cells to counteract the oxidative stress as a result of increased ROS production, which otherwise can lead to apoptosis or necrosis[23]. Inhibition of Trx[1] and Grx[1] could, have pro-apoptotic effects in cancer cells[23,29]
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