A framework for the development of effective anti-metastatic agents.

Most cancer-related deaths are a result of metastasis, and thus the importance of this process as a target of therapy cannot be understated. By asking 'how can we effectively treat cancer?', we do not capture the complexity of a disease encompassing >200 different cancer types - many consisting of multiple subtypes - with considerable intratumoural heterogeneity, which can result in variable responses to a specific therapy. Moreover, we have much less information on the pathophysiological characteristics of metastases than is available for the primary tumour. Most disseminated tumour cells that arrive in distant tissues, surrounded by unfamiliar cells and a foreign microenvironment, are likely to die; however, those that survive can generate metastatic tumours with a markedly different biology from that of the primary tumour. To treat metastasis effectively, we must inhibit fundamental metastatic processes and develop specific preclinical and clinical strategies that do not rely on primary tumour responses. To address this crucial issue, Cancer Research UK and Cancer Therapeutics CRC Australia formed a Metastasis Working Group with representatives from not-for-profit, academic, government, industry and regulatory bodies in order to develop recommendations on how to tackle the challenges associated with treating (micro)metastatic disease. Herein, we describe the challenges identified as well as the proposed approaches for discovering and developing anticancer agents designed specifically to prevent or delay the metastatic outgrowth of cancer.

Molecular basis for Jagged-1/Serrate ligand recognition by the Notch receptor.

We have mapped a Jagged/Serrate-binding site to specific residues within the 12th EGF domain of human and Drosophila Notch. Two critical residues, involved in a hydrophobic interaction, provide a ligand-binding platform and are adjacent to a Fringe-sensitive residue that modulates Notch activity. Our data suggest that small variations within the binding site fine-tune ligand specificity, which may explain the observed sequence heterogeneity in mammalian Notch paralogues, and should allow the development of paralogue-specific ligand-blocking antibodies. As a proof of principle, we have generated a Notch-1-specific monoclonal antibody that blocks binding, thus paving the way for antibody tools for research and therapeutic applications.

CHR-2797: an antiproliferative aminopeptidase inhibitor that leads to amino acid deprivation in human leukemic cells.

CHR-2797 is a novel metalloenzyme inhibitor that is converted into a pharmacologically active acid product (CHR-79888) inside cells. CHR-79888 is a potent inhibitor of a number of intracellular aminopeptidases, including leucine aminopeptidase. CHR-2797 exerts antiproliferative effects against a range of tumor cell lines in vitro and in vivo and shows selectivity for transformed over nontransformed cells. Its antiproliferative effects are at least 300 times more potent than the prototypical aminopeptidase inhibitor, bestatin. However, the mechanism by which inhibition of these enzymes leads to proliferative changes is not understood. Gene expression microarrays were used to profile changes in mRNA expression levels in the human promyelocytic leukemia cell line HL-60 treated with CHR-2797. This analysis showed that CHR-2797 treatment induced a transcriptional response indicative of amino acid depletion, the amino acid deprivation response, which involves up-regulation of amino acid synthetic genes, transporters, and tRNA synthetases. These changes were confirmed in other leukemic cell lines sensitive to the antiproliferative effects of CHR-2797. Furthermore, CHR-2797 treatment inhibited phosphorylation of mTOR substrates and reduced protein synthesis in HL-60 cells, both also indicative of amino acid depletion. Treatment with CHR-2797 led to an increase in the concentration of intracellular small peptides, the substrates of aminopeptidases. It is suggested that aminopeptidase inhibitors, such as CHR-2797 and bestatin, deplete sensitive tumor cells of amino acids by blocking protein recycling, and this generates an antiproliferative effect. CHR-2797 is orally bioavailable and currently undergoing phase II clinical investigation in the treatment of myeloid leukemia.

Structure of the HP1 chromodomain bound to histone H3 methylated at lysine 9.

Specific modifications to histones are essential epigenetic markers---heritable changes in gene expression that do not affect the DNA sequence. Methylation of lysine 9 in histone H3 is recognized by heterochromatin protein 1 (HP1), which directs the binding of other proteins to control chromatin structure and gene expression. Here we show that HP1 uses an induced-fit mechanism for recognition of this modification, as revealed by the structure of its chromodomain bound to a histone H3 peptide dimethylated at Nzeta of lysine 9. The binding pocket for the N-methyl groups is provided by three aromatic side chains, Tyr21, Trp42 and Phe45, which reside in two regions that become ordered on binding of the peptide. The side chain of Lys9 is almost fully extended and surrounded by residues that are conserved in many other chromodomains. The QTAR peptide sequence preceding Lys9 makes most of the additional interactions with the chromodomain, with HP1 residues Val23, Leu40, Trp42, Leu58 and Cys60 appearing to be a major determinant of specificity by binding the key buried Ala7. These findings predict which other chromodomains will bind methylated proteins and suggest a motif that they recognize.