XMU-MP-3 is a robust, non-covalent inhibitor of BTK, exhibiting exceptional potency with IC50 values of 10.7 nM and 17.0 nM for BTK WT and BTK C481S mutation, respectively, in the presence of 10 μM ATP. Moreover, XMU-MP-3 elicits apoptosis.

BTK (C481S):17.0 nM (IC50), BTK (WT):10.7 nM (IC50)

XMU-MP-3, across a concentration range of 0.001-10000 nM over 48 hours, effectively suppresses the proliferation of BTK-transformed Ba/F3 cells, achieving an IC50 of 11.4 nM. Furthermore, at concentrations between 1-10000 nM, it inhibits the growth of JeKo-1, Ramos, and NALM-6 cells, with IC50 values of 326.6 nM, 685.6 nM, and 1065 nM, respectively. It retains its inhibitory efficacy against BTK(C481S)-Ba/F3 cells with an IC50 of 182.3 nM within the same concentration range. At 5000 nM, XMU-MP-3 is shown to induce apoptosis in BTK (C481S) Ba/F3 cells. Additionally, when applied at 10-1000 nM for 4 hours, it dose-dependently hinders both auto- and trans-phosphorylation of BTK at Y223 and Y551 sites in BTK-transformed Ba/F3 cells. Investigative assays reveal that XMU-MP-3 obstructs the proliferation of BTK-transformed Ba/F3 cells with an IC50 of 11.4 nM, while it exhibits minimal anti-proliferative effects on parental wild-type Ba/F3 cells (IC50 >10000 nM). Western blot analysis supports these findings, indicating a significant reduction in the phosphorylation levels of BTK Y223 and Y551 at as low as 100 nM, with complete suppression at 1000 nM.

XMU-MP-3 (25 and 50 mg/kg) significantly inhibits tumor growth in mouse xenograft models, utilizing Nu/nu BALB/c mice (4-6 weeks of age) with BTK-transformed Ba/F3 and Ramos xenografts[1]. Administered via tail vein injection at 25 and 50 mg/kg, 0.1 mL per 10 g body weight, daily for 14 days, XMU-MP-3 notably reduced tumor size without affecting animal weight[1].