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Maternal embryonic leucine zipper kinase (MELK) is upregulated in several types of tumor, including breast, prostate, and brain tumors. Its expression is generally associated with cell survival, cell proliferation, and resistance to apoptosis. Therefore, the potential of MELK inhibitors as therapeutic agents is recently attracting considerable interest. Here we report the first structures of MELK in complex with AMP-PNP and with nanomolar inhibitors. Our studies shed light on the role of the MELK UBA domain, provide a characterization of the kinase active site, and identify key residues for achieving high potency, laying the groundwork for structure-based drug design efforts. [ABSTRACT FROM AUTHOR]

VCP (also known as p97 or Cdc48p in yeast) is an AAA+ ATPase regulating endoplasmic reticulum-associated degradation. After high-throughput screening, we developed compounds that inhibit VCP via different mechanisms, including covalent modification of an active site cysteine and a new allosteric mechanism. Using photoaffinity labeling, structural analysis and mutagenesis, we mapped the binding site of allosteric inhibitors to a region spanning the D1 and D2 domains of adjacent protomers encompassing elements important for nucleotide-state sensing and ATP hydrolysis. These compounds induced an increased affinity for nucleotides. Interference with nucleotide turnover in individual subunits and distortion of interprotomer communication cooperated to impair VCP enzymatic activity. Chemical expansion of this allosteric class identified NMS-873, the most potent and specific VCP inhibitor described to date, which activated the unfolded protein response, interfered with autophagy and induced cancer cell death. The consistent pattern of cancer cell killing by covalent and allosteric inhibitors provided critical validation of VCP as a cancer target. [ABSTRACT FROM AUTHOR]

[Display omitted] In this article we describe the identification of unprecedented ATP-competitive ChoKα inhibitors starting from initial hit NMS-P830 that binds to ChoKα in an ATP concentration-dependent manner. This result is confirmed by the co-crystal structure of NMS-P830 in complex with Δ75-ChoKα. NMS-P830 is able to inhibit ChoKα in cells resulting in the reduction of intracellular phosphocholine formation. A structure-based medicinal chemistry program resulted in the identification of selective compounds that have good biochemical activity, solubility and metabolic stability and are suitable for further optimization. The ChoKα inhibitors disclosed in this article demonstrate for the first time the possibility to inhibit ChoKα with ATP-competitive compounds. [ABSTRACT FROM AUTHOR]

In the present study, rice seedlings were exposed to a range of Cd concentrations (0.1μM, 1μM, 10μM, 100μM and 1mM) for 15 days and a combination of different molecular approaches were used to evidence Cd effects and to assess the plants’ ability to counteract metal toxicity. At a macroscopical level, only the highest Cd concentration (1mM) caused a complete plant growth inhibition, whereas the lowest concentrations seemed to stimulate growth. At genome level, the amplified fragment length polymorphism (AFLP) technique was applied to detect DNA sequence changes in root cells, showing that all the Cd concentrations induced significant DNA polymorphisms in a dose-dependent manner. Data also evidenced the absence of preferential mutation sites. Plant responses were analysed by measuring the levels of gluthatione (GSH) and phytochelatins (PCs), the thiol-peptides involved in heavy metal tolerance mechanisms. Results showed a progressive increase of GSH up to 10μM of Cd treatment, whereas a significant induction only of PC3 was detected in roots of plants exposed to 100μM of Cd. As suggested by the proteome analysis of root tissues, this last concentration strongly induced the expression of regulatory proteins and some metabolic enzymes. Furthermore, the treatment with 10μM of Cd induced changes in metabolic enzymes, but it mainly activated defence mechanisms by the induction of transporters and proteins involved in the degradation of oxidatively modified proteins. [Copyright &y& Elsevier]