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Protein degradation is a highly regulated process that is required for maintaining cellular homeostasis, and its deregulation is associated with development of a number of diseases including cancer. Neddylation is a posttranslational modification that involves conjugation of the ubiquitin-like protein NEDD8 (neural precursor cell expressed, developmentally downregulated) to target proteins such as oncoproteins and tumor suppressors to regulate their functions through a cascade of enzymes including NEDD8-activating enzyme (E1), NEDD8-conjugating enzyme (E2), and NEDD8 ligases (E3). 1 As such, the neddylation pathway presents an attractive oncogenic target for development of anticancer therapies. The main cellular substrates for neddylation include components of the Cullin-RING E3 ligase (CRL), which is the largest multicomponent ubiquitin ligase family regulating the turnover of multiple tumor suppressor proteins such as p21, p27, and NF-Kappa-B Inhibitor Alpha, among others (Fig. 1A). 2 In an elegant study, Hua et al systematically investigated the effectiveness of the NAE inhibitor, MLN4924, against glioblastoma (GBM) in vitro and in vivo. MLN4924 is a newly developed small molecule inhibitor of the neddylation pathway that specifically inhibits NAE by binding to its active site and hindering its enzymatic activity. 3 Several other groups have shown that treatment of cancer cells with MLN4924 results in induction of DNA damage response, cell cycle arrest, apoptosis, or senescence. 4,5 MLN4924 is currently under investigation in several phase 1/2 clinical trials for multiple solid tumor types and hematologic malignancies. 6 The ability of MLN4924 to cross the blood-brain barrier, its low toxicity, and clinical efficacy in other cancers suggests that this drug is an attractive treatment against GBM. Analysis of overall survival of GBM patients, as well as correlation with expression of global protein neddylation, indicated that higher global protein neddylation correlates with poor patient survival. They also found that GBM tumors generally express higher levels of NEDD8 and neddylation enzymes such as NAE1/UBA3 and UBC12 compared with adjacent normal brain tissue, with recurrent GBM tumors displaying even

Background. The neddylation pathway has been recently identified as an attractive anticancer target, and MLN4924, a specific NEDD8-activating enzyme inhibitor, has been developed as a first-in-class anticancer agent. However, neither the status of the neddylation pathway in glioblastoma (GBM) nor the effect of MLN4924 against GBM has been systematically investigated yet. Methods. To measure the activation state of the neddylation pathway in GBM, expression of the NEDD8-activating enzyme (E1), NEDD8-conjugating enzyme (E2), and global protein neddylation in GBM tumor tissues versus adjacent tissues were examined by immunoblotting analysis and immunohistochemistry staining. To assess the therapeutic efficacy of neddylation inhibition in GBM, cell proliferation in vitro and tumor growth in vivo were determined upon neddylation inhibition by MLN4924, an investigational NEDD8-activating enzyme inhibitor. Results. The neddylation pathway was overactivated in a majority of GBM tumor tissues when compared with adjacent normal tissues. The upregulation of this pathway in GBM tissues was positively correlated with high-grade disease and postoperative recurrence but was negatively associated with patient overall survival. MLN4924 treatment inhibited cullin neddylation, inactivated cullin-RING E3 ligase, and led to the accumulation of tumor-suppressive cullin-RING E3 ligase substrates to trigger cell-cycle arrest and senescence or apoptosis in a cell-line dependent manner. Moreover, inhibition of neddylation by MLN4924 significantly suppressed tumor growth in an orthotopic xenograft model of human GBM. Conclusion. Our study indicates that an overactivated neddylation pathway may be involved in GBM progression and that inhibition of this oncogenic pathway is a potentially new therapeutic approach for GBM.

DNA double strand breaks are the most cytotoxic lesions that can occur on the DNA. They can be repaired by different mechanisms and optimal survival requires a tight control between them. Here we uncover protein deneddylation as a major controller of repair pathway choice. Neddylation inhibition changes the normal repair profile toward an increase on homologous recombination. Indeed, RNF111/UBE2M-mediated neddylation acts as an inhibitor of BRCA1 and CtIP-mediated DNA end resection, a key process in repair pathway choice. By controlling the length of ssDNA produced during DNA resection, protein neddylation not only affects the choice between NHEJ and homologous recombination but also controls the balance between different recombination subpathways. Thus, protein neddylation status has a great impact in the way cells respond to DNA breaks. España , Ministerio de Economía y Competitividad SAF2010-14877