JNJ-47117096

This compound has utility as a probe, however several caveats should be considered when utilizing. 1) This compound shows strong kinase activity for Flt3 (18 nM), which is comparable to the measured IC50 for MELK (23 nM). While these targets differ in their function and associated targets, both are implicated in cancer. As such, in a cell setting depending on phenotypic readout, it could be beneficial to utilize an orthogonal Flt3 inhibitor as a control to ensure that any phenotype is stemming from on target inhibition of MELK. 2) This compound shows proteosome-dependent degradation of MELK according to 10.1042/BSR20150194. 3) Various IC50s for recombinant constructs of MELK were reported, which suggests that the full-length MELK may have differential activity/affinity than constructs missing the C-terminal regulatory domain or other constituents. This is a suitable in vitro probe at higher concentrations (10 uM) for cell investigation until a more selective MELK inhibitor becomes available.

The probe JNJ-47117096 has potential to be useful as an orthogonal reagent for cellular validation of MELK in combination with either genetic knockdown or other structurally distinct MELK small molecule inhibitor compounds. The strengths of this probe are that it has reasonably potent biochemical inhibition of MELK (18 nM in the Millipore radiometric assay) and has demonstrated cellular permeability through use of an antiproliferative assay designed against its off-target Flt3 (at an IC50 of 1.5 uM). Other than Flt3, this probe has also shown good selectivity across the kinome with the third highest potency of inhibition of any target against Mnk2 of 760 nM. On the other hand, this probe inhibits Flt3 in the radiometric assay at a potency similar to MELK, suggesting that any experiment using this probe to support the involvement of MELK should be designed to also rule out Flt3 as a contributor to the cellular phenotype. Additional weaknesses of this probe are that no off-target data outside of the 230-kinase panel has been disclosed (i.e. other target families) and even within the kinase family less than half of the kinase targets appear to have been profiled. Finally, no data from a direct cellular target engagement or specific cellular functional assay for MELK has not been described for this probe. Therefore, the investigator is advised to use the probe at multiple concentrations between 0.5 and 5 uM in order to best support a MELK-driven biological hypothesis.