MAGE-A cancer/testis antigens inhibit MDM2 ubiquitylation function and promote increased levels of MDM4

Melanoma antigen A (MAGE-A) proteins comprise a structurally and biochemically similar sub-family of Cancer/Testis antigens that are expressed in many cancer types and are thought to contribute actively to malignancy. MAGE-A proteins are established regulators of certain cancer-associated transcription factors, including p53, and are activators of several RING finger-dependent ubiquitin E3 ligases. Here, we show that MAGE-A2 associates with MDM2, a ubiquitin E3 ligase that mediates ubiquitylation of more than 20 substrates including mainly p53, MDM2 itself, and MDM4, a potent p53 inhibitor and MDM2 partner that is structurally related to MDM2. We find that MAGE-A2 interacts with MDM2 via the N-terminal p53-binding pocket and the RING finger domain of MDM2 that is required for homo/hetero-dimerization and for E2 ligase interaction. Consistent with these data, we show that MAGE-A2 is a potent inhibitor of the E3 ubiquitin ligase activity of MDM2, yet it does not have any significant effect on p53 turnover mediated by MDM2. Strikingly, however, increased MAGE-A2 expression leads to reduced ubiquitylation and increased levels of MDM4. Similarly, silencing of endogenous MAGE-A expression diminishes MDM4 levels in a manner that can be rescued by the proteasomal inhibitor, bortezomid, and permits increased MDM2/MDM4 association. These data suggest that MAGE-A proteins can: (i) uncouple the ubiquitin ligase and degradation functions of MDM2; (ii) act as potent inhibitors of E3 ligase function; and (iii) regulate the turnover of MDM4. We also find an association between the presence of MAGE-A and increased MDM4 levels in primary breast cancer, suggesting that MAGE-A-dependent control of MDM4 levels has relevance to cancer clinically

Activation of the p53 pathway by the MDM2 inhibitor nutlin-3a overcomes BCL2 overexpression in a preclinical model of diffuse large B-cell lymphoma associated with t(14;18)(q32;q21)

p53 is frequently wild type (wt) in diffuse large B-cell lymphoma (DLBCL) associated with t(14;18)(q32;q21) that overexpresses BCL2. Nutlin-3a is a small molecule that activates the p53 pathway by disrupting p53–MDM2 interaction. We show that nutlin-3a activates p53 in DLBCL cells associated with t(14;18)(q32;q21), BCL2 overexpression and wt p53, resulting in cell cycle arrest and apoptosis. Nutlin-3a treatment had similar effects on DLBCL cells of activated B-cell phenotype with wt p53. Cell cycle arrest was associated with upregulation of p21. Nutlin-3a-induced apoptosis was accompanied by BAX and PUMA upregulation, BCL-XL downregulation, serine-70 dephosphorylation of BCL2, direct binding of BCL2 by p53, caspase-9 upregulation and caspase-3 cleavage. Cell death was reduced when p53-dependent transactivation activity was inhibited by pifithrin-α (PFT-α), or PFT-μ inhibited direct p53 targeting of mitochondria. Nutlin-3a sensitized activation of the intrinsic apoptotic pathway by BCL2 inhibitors in t(14;18)-positive DLBCL cells with wt p53, and enhanced doxorubicin cytotoxicity against t(14;18)-positive DLBCL cells with wt or mutant p53, the latter in part via p73 upregulation. Nutlin-3a treatment in a xenograft animal lymphoma model inhibited growth of t(14;18)-positive DLBCL tumors, associated with increased apoptosis and decreased proliferation. These data suggest that disruption of the p53–MDM2 interaction by nutlin-3a offers a novel therapeutic approach for DLBCL associated with t(14;18)(q32;q21)

Small molecule compounds targeting the p53 pathway: are we finally making progress?

Loss of function of p53, either through mutations in the gene or through mutations to other members of the pathway that inactivate wild-type p53, remains a critically important aspect of human cancer development. As such, p53 remains the most commonly mutated gene in human cancer. For these reasons, pharmacologic activation of the p53 pathway has been a highly sought after, yet unachieved goal in developmental therapeutics. Recently progress has been made not only in the discovery of small molecules that target wild-type and mutant p53, but also in the initiation and completion of the first in-human clinical trials for several of these drugs. Here, we review the current literature of drugs that target wild-type and mutant p53 with a focus on small-molecule type compounds. We discuss common means of drug discovery and group them according to their common mechanisms of action. Lastly, we review the current status of the various drugs in the development process and identify newer areas of p53 tumor biology that may prove therapeutically useful

Metabolic regulation by p53

We are increasingly aware that cellular metabolism plays a vital role in diseases such as cancer, and that p53 is an important regulator of metabolic pathways. By transcriptional activation and other means, p53 is able to contribute to the regulation of glycolysis, oxidative phosphorylation, glutaminolysis, insulin sensitivity, nucleotide biosynthesis, mitochondrial integrity, fatty acid oxidation, antioxidant response, autophagy and mTOR signalling. The ability to positively and negatively regulate many of these pathways, combined with feedback signalling from these pathways to p53, demonstrates the reciprocal and flexible nature of the regulation, facilitating a diverse range of responses to metabolic stress. Intriguingly, metabolic stress triggers primarily an adaptive (rather than pro-apoptotic) p53 response, and p53 is emerging as an important regulator of metabolic homeostasis. A better understanding of how p53 coordinates metabolic adaptation will facilitate the identification of novel therapeutic targets and will also illuminate the wider role of p53 in human biology