Mps1-IN-1

Mps1-IN-1 is a potent, selective and ATP-competitive Mps1 kinase inhibitor, with an IC50 and a Kd of 367 nM and 27 nM.

Signaling Pathways >> Cell Cycle/DNA Damage >> Mps1

Signaling Pathways >> Cytoskeleton >> Mps1

Research Areas >> Cancer

Mps1:27 nM (Kd)

ALK:21 nM (Kd)

LTK:29 nM (Kd)

PYK2:280 nM (Kd)

FAK:440 nM (Kd)

IGF1R:750 nM (Kd)

INSR:470 nM (Kd)

CLK1:1900 nM (Kd)

ERK2:2900 nM (Kd)

INSRR:1200 nM (Kd)

TNK1:2600 nM (Kd)

TNK2:3100 nM (Kd)

GAK:1100 nM (Kd)

Mps1-IN-1 is a potent, selective and ATP-competitive Mps1 kinase inhibitor, with an IC50 and a Kd of 367 nM and 27 nM. Mps1-IN-1 also has high affinity for ALK, and LTK, with Kds of 21 and 39 nM, respectively, but shows little or no inhibition on other 352 member kinases. Mps1-IN-1 (5, 10 μM) induces bypass of a checkpoint-mediated mitotic arrest in U2OS cells. Mps1-IN-1 disrupts recruitment of Mad2 to kinetochores, and reduces the phosphorylation status of Aurora B at threonine-232 (Thr232). Mps1-IN-1 (10 µM) shows no effect on centrosome duplication. In addition, Mps1-IN-1 (5-10 µM) suppresses the proliferative capacity of HCT116[1].

The kinase binding assay is used to assess compound binding to TTK by monitoring displacement of a fluorescently labeled, ATP site-directed kinase inhibitor (Kinase Tracer 236) from the kinase active site. Each 15 μL assay contains 5 nM TTK, variable amounts of test compound (Mps1-IN-1), 30 nM Kinase Tracer 236, 2 nM Eu-anti-GST Antibody, and 1% DMSO (residual from compound dilution) in Kinase Buffer A (50 mM HEPES pH 7.5, 10 mM MgCl2, 1 mM EGTA, 0.01% Brij-35). Binding assays are initiated by addition of 5 μL of test compound (from 2-fold dilution series) to 5 μL of a kinase/antibody mixture, followed by addition of 5 μL of antibody. Assay plates are read using using standard Eu-based TR-FRET settings with excitation at 340 nm and emission monitored at 615 nm (donor) and 665 nm (acceptor). Emission intensities are measured over a 200 µs window following a 100 µs post-excitation delay[1].

U2OS cells expressing doxycycline-inducible PLK4 are plated in 96 well plates. A double thymidine block is performed using the following treatment regimen: thymidine for 18-20 hrs., release for 10 hrs. with doxycycline induction of PLK4 during this time, then a second thymidine block, followed by release. Six hours after the 2nd thymidine release, Mps1-IN-1 (or DMSO vehicle) is added and the proliferation of the cell populations is monitored with Cell Titer GLO assay[1].

[1]. Kwiatkowski N, et al. Small-molecule kinase inhibitors provide insight into Mps1 cell cycle function. Nat Chem Biol. 2010 May;6(5):359-68.

CFI402257 | BAY1217389 | BAY 1161909 | NMS-P715 | AZ 3146 | BOS172722 | MPI-0479605 | Mps1-IN-2 | Mps1-IN-3 | CCT251455