Figure 5.

Determination of the IC50 of Target Engagement of SphK2 by PF-543 using the IsoThermal Dose Response Fingerprint (ITDRF) Assay. A representative western blot of HEK293 cells over-expressing FLAG-SphK2 where cells were incubated with the indicated concentrations of (3: PF-543) and DMSO for 24 hrs. ITDRF assays were performed at 63.0°C as detailed in the Materials and Methods and analyzed by western blot detection of SphK2 using the appropriate primary antibody. Actin was included as a loading control.