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Multifaceted Polo-Like Kinases: Drug Targets and Antitargets for Cancer Therapy

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Multifaceted Polo-Like Kinases: Drug Targets and Antitargets for Cancer Therapy REVIEWS Multifaceted polo-like kinases: drug targets and antitargets for cancer therapy Klaus Strebhardt Abstract | The polo-like kinase 1 (PLK1) acts in concert with cyclin-dependent kinase 1–cyclin B1 and Aurora kinases to orchestrate a wide range of critical cell cycle events. Because PLK1 has been preclinically validated as a cancer target, small-molecule inhibitors of PLK1 have become attractive candidates for anticancer drug development. Although the roles of the closely related PLK2, PLK3 and PLK4 in cancer are less well understood, there is evidence showing that PLK2 and PLK3 act as tumour suppressors through their functions in the p53 signalling network, which guards the cell against various stress signals. In this article, recent insights into the biology of PLKs will be reviewed, with an emphasis on their role in malignant transformation, and progress in the development of small-molecule PLK1 inhibitors will be examined. More than two decades ago polo, the founding member Early observations on the overexpression of PLK1 of the family of polo-like kinases (PLKs), was identified in human tumours12 and on the inhibition of cellu- as having an essential role in the ordered execution of lar proliferation by microinjecting antibodies against mitotic events in Drosophila melanogaster 1,2. Since the PLK1 into HeLa cells13 initiated a series of follow-on discovery of polo, a wealth of functional information on studies using a broad spectrum of inhibitors (for exam- PLKs has been collected in a wide phylogenetic space. ple, dominant-negative forms of PLK1, antisense oli- Five mammalian PLK family members have been iden- gonucleotides and small interfering RNAs) aiming to tified so far, PLK1 (also known as STPK13), PLK2 (also evaluate PLK1 as a potential target for the treatment known as SNK), PLK3 (also known as CNK, FNK and of cancer14–17. The identification of PLK1 as a crucial, PRK), PLK4 (also known as SAK and STK18) and PLK5 functional node in the oncogenic network, in which its (REFS 3–5). An early comparison of murine PLK1 with inhibition results in systemic failure (that is, blockage of POLO and PLK2 uncovered two conserved domains in the tumorigenic state by apoptosis)18 proved to be key PLK1, the canonical serine/threonine kinase domain in developing successful therapies. Subsequently, the and a non-catalytic region, which was later termed the pharmaceutical industry has initiated intensive efforts polo box domain (PBD)6. All PLKs exhibit the same to develop potent and specific small-molecule PLK1 domain topology with the kinase domain located at inhibitors. This development is fuelled by previous the amino-terminal followed by the carboxy-terminal pharmaceutical and clinical investigations, which had PBD (BOX 1). Recent advances in the basic biology of convincingly shown that antimitotic drugs are success- PLK1 have greatly improved our understanding of its ful chemotherapeutic tools for the fight against cancer19. regulation, targets and functions. As reflected by multi- PLK1 inhibitors have been clinically assessed and hold Department of Obstetrics ple localizations PLK1 has been shown to have a pivotal great promise for the improved treatment of patients 18,20,21 and Gynaecology, School role in regulating the cell cycle, including entry into with cancer . of Medicine, J.W. Goethe mitosis, centrosome maturation, assembly of the bipo- Although PLK1 inhibitors have already reached University, Theodor Stern lar spindle, sister chromatid splitting, activation of the the clinic, analysis of their specificity indicates that Kai 7, 60590 Frankfurt, anaphase-promoting complex/cyclosome (APC/C) the related family members PLK2 and PLK3 are in the Germany. e-mail: strebhardt@em. and exit from mitosis with the initiation of cytokine- range of their inhibitory potential. Despite tremen- uni-frankfurt.de sis (FIG. 1). These aspects have been the subject of many dous progress in understanding the biology of PLK1, doi:10.1038/nrd3184 excellent reviews3,4,7–11. the functional roles of other PLK family members, NATURE REVIEWS | DRUG DISCOVERY VOLUME 9 | AUGUST 2010 | 643 © 2010 Macmillan Publishers Limited. All rights reserved REVIEWS Box 1 | Structural insights into the polo-like kinase (PLK) family proteins All members of the PLK family a b contain an amino-terminal kinase domain and a carboxy-terminal polo box domain (PBD), which is unique to the PLK family. The crystal Ser137 Val210 structure of the kinase domain of human PLK1, in which the regulatory phosphorylation site Thr210 was mutated with the aim of reducing conformational heterogeneity, has been determined in complexes with the non-hydrolysable c ATP analogue adenylylimi- dodiphosphate, the pyrrolo-pyrazole inhibitor PHA 680626 (a) or the ATP-competitive BI 2536 (REFS 157,159). In another approach, the wild-type kinase domain of PLK1 was co-crystallized with the designed ankyrin-repeat protein 3H10 that is selective for PLK1 (REF. 218). The similarity of the crystal structures confirms the validity of the different approaches for structure-guided drug design. Three-dimensional analysis showed the typical structure of a kinase fold, in which the ATP-binding site is situated in a cleft, formed between the N-terminal lobe encompassing β-sheets (residues 37–131)Na andture theRevie predominatelyws | Drug Discov ery α-helical C-terminal lobe (residues 138–330)157. The two lobes are linked by a hinge region that interacts with the bound ligands. Although Thr210 is not phosphorylated, the activation loop of PLK1 (T-loop; residues 194–221) exhibits an extended conformation in the crystal structure reminiscent to the conformation of activated kinases219. This conformation might be stabilized by Zn2+ ions, which are essential for the crystallization of the PLK1 kinase domain157. The C-terminal PBD functions as a phosphopeptide-binding module by targeting serine-(phosphothreonine/ phosphoserine)-(proline/X) motifs34,41. PLK1, PLK2, PLK3 and PLK5 have a PBD that consists of two polo boxes, whereas PLK4 has only one polo box. The crystal structures of the PLK1 PBD, complexed with an optimal phospho- serine-containing peptide or a peptide corresponding to the binding region of PLK1 within the biological interaction partner CDC25, document an interaction along a positively charged cleft, which is formed between the two polo boxes (b)41,220,221. Each conserved polo box consists of a six stranded β-sheet and an α-helix, which associate to form a 12-stranded β-sandwich domain (c). Recent studies on the crystal structure and binding properties of the PLK1 PBD in complex with the minimal phosphopeptides suggest that the C-terminal serine-phosphothreonine dipeptide provides the high-affinity anchor, whereas the N-terminal residues contribute to the specificity and affinity of the interaction222. The crystal structure of the PLK4 PBD containing a single polo box suggests that this domain can dimerize223. Further analysis has corroborated this model, showing that PLK4 forms homodimers in vivo223. Despite its homodimerization PLK4 remains capable of localizing to the centrosomes and the cleavage furrow during cytokinesis223. Part a of the figure shows the crystal structure of the PLK1 kinase domain with the pyrrolo-pyrazole inhibitor PHA 680626 at 2.4 Å resolution. The N-terminal and C-terminal extensions are coloured orange and the activation loop is coloured green157. The positions of the mutated Val210 (corresponding to the main phosphorylation site Thr210 in the activation domain) and Ser137 (which has been proposed as an additional phosphorylation site for the activation of the kinase activity of PLK1) are indicated. Part b of the figure shows the crystal structure of the PLK1 PBD in a ribbon presentation, in complex with phosphothreonine- containing peptide that is shown as a ball and stick representation in yellow41. Polo box 1 and polo box 2 are shown in red and purple, respectively, and from two different angles. The polo cap at the N-terminal end of polo box 1, which is indicated in grey, folds around polo box 2 and therefore tethers it to polo box 1. The two polo boxes form a pocket to accommodate the 41 phosphopeptide. Part c of the figure is a superposition of the polo box 1 and polo box 2 β6α structures, coloured as in part b . especially of PLK2 and PLK3, remain elusive. However, important to carefully analyse the roles of PLKs in the in cancer cells PLK2 and PLK3 emerge as important network of stress signalling in cancer cells. mediators of the stress phenotypes in response to DNA After summarizing the relevant biological back- damage or oxidative stress. Increasing evidence sup- ground of PLKs and their roles in cancer cells, this ports a mutual crosstalk between all members of the Review will discuss the status quo of small-molecule PLK family and the tumour suppressor p53 under development targeting the kinase domain and the PBD stressful conditions in cancer cells22–24. As the classical of PLK1. Furthermore, this article will consider the clinical hallmarks of cancer are well documented25,26, it will be development of novel PLK inhibitors. 644 | AUGUST 2010 | VOLUME 9 www.nature.com/reviews/drugdisc © 2010 Macmillan Publishers Limited. All rights reserved REVIEWS G2 phaseProphasePrometaphase Metaphase AnaphaseCytokinesis Mitotic entry Furrow ingression and spindle elongation Centrosome maturation, microtubule nucleation and kinetochore capture Figure 1 | Localization and selected functions of PLK1 during mitosis. Indirect immunofluorescence of polo-like kinase 1 (PLK1)
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