Proper control of mitosis is critical to maintain the stability of the genome during cell proliferation, and genomic instability may contribute directly to the generation of cancer. Study of mechanisms that regulate mitosis, therefore, is critical to understand how cancer develops, and to discover new ways to prevent and treat the disease. Mitosis is also an important target for cancer therapy. Recently, new selective anti-mitotic drugs such as aurora kinase inhibitors have shown great promise in pre-clinical experiments, and there is now immense interest in identifying new drug targets in mitosis. We have recently discovered a novel mitotic histone kinase, haspin, that has homologs in diverse eukaryotes. Human haspin mRNA is expressed in proliferating but not non-proliferating cells. During mitosis, haspin associates with condensed chromosomes, particularly at centromeres, and is responsible for phosphorylation of Thr-3 in histone H3. Haspin is also found at mitotic centrosomes. Importantly, haspin RNA interference causes misalignment of metaphase chromosomes and spindle defects, preventing completion of normal mitosis. These studies add haspin to the select group of kinases that regulate mitotic chromosome dynamics and spindle activity and provide the first indication that haspin, like the aurora kinases, might be a suitable target for cancer therapy. Further study of haspin action in mitosis and validation of haspin as a cancer drug target are currently limited, however, by the lack of specific small molecule inhibitors of the kinase. To identify small molecule inhibitors of haspin, we will develop an in vitro haspin kinase assay suitable for high-throughput screening of chemical libraries.
In Aim 1 we will produce, in E. coli or the baculovirus system, functional full-length recombinant haspin for use in screening assays.
In Aim 2 we will develop and optimize a homogenous time-resolved fluorescence kinase assay for haspin. An alternative strategy using a separation-based approach is also described.
In Aim 3, we outline assays to confirm hits from the screening process and develop secondary screens to assess the inhibitory properties and functional effects of these compounds in vitro and in cells. Haspin inhibitors will provide a new approach to investigate the basic biology of cell division and will yield insights that cannot be obtained using existing technology. Furthermore, such inhibitors will provide an excellent way to validate haspin as a target for cancer treatment, and they might find direct application as chemotherapeutic drugs. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA122608-03
Application #
7430441
Study Section
Drug Discovery and Molecular Pharmacology Study Section (DMP)
Program Officer
Song, Min-Kyung H
Project Start
2006-07-07
Project End
2010-05-31
Budget Start
2008-06-01
Budget End
2010-05-31
Support Year
3
Fiscal Year
2008
Total Cost
$226,000
Indirect Cost
Name
Brigham and Women's Hospital
Department
Type
DUNS #
030811269
City
Boston
State
MA
Country
United States
Zip Code
02115
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Cuny, Gregory D; Ulyanova, Natalia P; Patnaik, Debasis et al. (2012) Structure-activity relationship study of beta-carboline derivatives as haspin kinase inhibitors. Bioorg Med Chem Lett 22:2015-9
Cuny, Gregory D; Robin, Maxime; Ulyanova, Natalia P et al. (2010) Structure-activity relationship study of acridine analogs as haspin and DYRK2 kinase inhibitors. Bioorg Med Chem Lett 20:3491-4
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Eswaran, Jeyanthy; Patnaik, Debasis; Filippakopoulos, Panagis et al. (2009) Structure and functional characterization of the atypical human kinase haspin. Proc Natl Acad Sci U S A 106:20198-203
Patnaik, Debasis; Jun Xian; Glicksman, Marcie A et al. (2008) Identification of small molecule inhibitors of the mitotic kinase haspin by high-throughput screening using a homogeneous time-resolved fluorescence resonance energy transfer assay. J Biomol Screen 13:1025-34