2 3 Targeting TAO Kinases Q 1 Using a New Inhibitor 4 Compound Delays Mitosis and Induces 5 Mitotic Cell Death in Centrosome Ampli fi ed Q 2 6 Breast Cancer Cells 7

11 Thousand-and-one amino acid kinases (TAOK) 1 and 2 are 12 activated catalytically duringmitosis and can contribute tomitotic 13 cell rounding and spindle positioning. Here, we characterize a 14 compound that inhibits TAOK1 and TAOK2 activity with IC50 15 values of 11 to 15 nmol/L, is ATP-competitive, and targets these 16 kinases selectively. TAOK inhibition or depletion in centrosome17 amplified SKBR3 or BT549 breast cancer cell models increases the 18 mitotic population, the percentages of mitotic cells displaying 19 amplified centrosomes and multipolar spindles, induces cell 20 death, and inhibits cell growth. In contrast, nontumorigenic and 21 dividing bipolar MCF-10A breast cells appear less dependent on 22 TAOK activity and can complete mitosis and proliferate in the 23 presence of the TAOK inhibitor. We demonstrate that TAOK1 and 24 TAOK2 localize to the cytoplasm and centrosomes respectively 25 during mitosis. Live cell imaging shows that the TAOK inhibitor 27 prolongs the duration of mitosis in SKBR3 cells, increases mitotic 28 cell death, and reduces the percentages of cells exiting mitosis, 29 whereas MCF-10A cells continue to divide and proliferate. Over 30 80% of breast cancer tissues display supernumerary centrosomes, 31 and tumor cells frequently cluster extra centrosomes to avoid 32 multipolar mitoses and associated cell death. Consequently, 33 drugs that stimulate centrosome declustering and induce multi34 polarity are likely to target dividing centrosome-amplified cancer 35 cells preferentially, while sparing normal bipolar cells. Our results 36 demonstrate that TAOK inhibition can enhance centrosome 37 declustering and mitotic catastrophe in cancer cells, and these 38 proteins may therefore offer novel therapeutic targets suitable for 39 drug inhibition and the potential treatment of breast cancers, 40 where supernumerary centrosomes occur Q5 . Mol Cancer Ther; 1–12. 41 2017 AACR. 42