JH295 hydrate is a potent, irreversible and selective NIMA-related kinase 2 (Nek2) inhibitor that inhibits cellular Nek2 by Cys22 alkylation (IC50 = 770 nM). JH295 hydrate is not active against the mitotic kinases, Cdk1, Aurora B or Plk1, and does not interfere with bipolar spindle assembly or the spindle assembly checkpoint [1].

JH295 (Compound 16; 0.08-20 μM; 45 minutes; RPMI7951 cells) treatment inhibits WT Nek2 in cells with an IC 50 of ~1.3 μM, whereas it has little effect on the C22V mutant [1]. Western Blot Analysis [1] Cell Line: RPMI7951 cells Concentration: 0.08 μM, 0.25 μM, 0.74 μM, 2.2 μM, 6.6 μM, 20 μM Incubation Time: 45 minutes Result: Inhibited WT Nek2 in cells with an IC50 of ~1.3 μM, whereas had little effect on the C22V mutant.