Inhibitors of the Bub1 spindle assembly checkpoint kinase: synthesis of BAY-320 and comparison with 2OH-BNPP1

Abstract

Bub1 is a serine/threonine kinase proposed to function centrally in mitotic chromosome alignment and the spindle assembly checkpoint (SAC); however, its role remains controversial. Although it is well documented that Bub1 phosphorylation of Histone 2A at T120 (H2ApT120) recruits Sgo1/2 to kinetochores, the requirement of its kinase activity for chromosome alignment and the SAC is debated. As small-molecule inhibitors are invaluable tools for investigating kinase function, we evaluated two potential Bub1 inhibitors: 2OH-BNPPI and BAY-320. After confirming that both inhibit Bub1 in vitro , we developed a cell-based assay for Bub1 inhibition. We overexpressed a fusion of Histone 2B and Bub1 kinase region, tethering it in proximity to H2A to generate a strong ectopic H2ApT120 signal along chromosome arms. Ectopic signal was effectively inhibited by BAY-320, but not 2OH-BNPP1 at concentrations tested. In addition, only BAY-320 was able to inhibit endogenous Bub1-mediated Sgo1 localization. Preliminary experiments using BAY-320 suggest a minor role for Bub1 kinase activity in chromosome alignment and the SAC; however, BAY-320 may exhibit off-target effects at the concentration required. Thus, 2OH-BNPP1 may not be an effective Bub1 inhibitor in cellulo , and while BAY-320 can inhibit Bub1 in cells, off-target effects highlight the need for improved Bub1 inhibitors.

BUB1/KIF14 complex promotes anaplastic thyroid carcinoma progression by inducing chromosome instability

“…During mitosis and meiosis, the SAC prevents incorrect chromosome segregation from maintaining genome stability 20 . BUB1 mitotic checkpoint serine/threonine kinase (BUB1) has been reported to make a central contribution to the mitotic chromosome alignment and the SAC 21 . Thus, chromosome segregation defects have been observed in nonmalignant cells with BUB1 inactivation 22 .…”

“… 20 BUB1 mitotic checkpoint serine/threonine kinase (BUB1) has been reported to make a central contribution to the mitotic chromosome alignment and the SAC. 21 Thus, chromosome segregation defects have been observed in nonmalignant cells with BUB1 inactivation. 22 Although BUB1 overexpression is a common phenomenon related to tumour proliferation in various solid tumours, studies of BUB1 in tumour CIN have been limited.…”