Figure 6. XPO1 target occupancy can be measured in tumors with a dose-dependent effect of selinexor on complex formation where target saturation occurs at ∼10 mg/kg selinexor and confirms the selinexor RP2D.

(A) XPO1 protein levels were detected from tumors harvested from mice bearing Z138 tumor xenografts that were orally administered a single dose of 0, 5, 10, 15, or 20 mg/kg selinexor and harvested at 6 hours post single dose. Expression of XPO1 was plotted by calculating the densitometric amount of XPO1 to GAPDH signal. (B) FCCS measurements of XPO1 occupancy from tumor lysates incubated with fluorescence labeled LMB (LMB₆₄₇) and ATTO488 labeled anti XPO1 antibody (XPO-1 AB₄₈₈) for 2 hours and complex formation was measured on a ConfoCor2 Correlation Spectroscope. The absolute values for the XPO1 complexes were plotted. (C) FCCS measurements of XPO1 occupancy from Z138 tumors from xenografted mice that were orally administered a single dose of 0, 5, 10, 15, or 20 mg/kg selinexor and harvested at 6, 24, 48, 72, and 96 hours post single dose. Samples were lysed and incubated with fluorescence labeled LMB (LMB₆₄₇) and ATTO488 labeled anti XPO1 antibody (XPO-1 AB₄₈₈) for 2 hours and complex formation was measured on a ConfoCor2 Correlation Spectroscope. Due to the variation in the size of the tumors, the concentration of dually labeled particles was normalized to total protein concentration. The resulting values were fitted to a logistic regression curve using the TIBCO Spotfire software package.