Glasdegib (PF-04449913) (PF-04449913) is a potent, and orally bioavailable Smoothened (Smo) inhibitor with IC50 of 5 nM. Phase 2.

Smo:5 nM

PF-04449913 is dissolved in DMSO and stored, and then diluted with appropriate medium before use[1]. Normal or BC CML CD34+ cells are plated on confluent mitomycin-C treated SL/M2 cells with vehicle, PF-04449913 (1 μM), Dasatinib (50 nM), or combination treatment. Mouse bone marrow stromal cell lines, M2-10B4 (M2) and SL/SL (SL) are treated with mitomycin-C (1 mg/mL) and plated in a 1:1 mixture at a total concentration of 100,000 cells/mL one day prior to co-culture with 10,000-20,000 CD34+ BC CML or normal progenitors. After 1 week of culture, progenitors are FACS sorted into hematopoietic progenitor assays and colonies are scored at 14 days. To assess survival of normal human hematopoietic stem and progenitor cells, irradiated (20 Gray) OP9 (M2 clone) stromal cells are co-cultured with 50,000 human CD34+ cord blood cells, vehicle or PF-04449913 in AlphaMEM with 20% Hyclone FBS, 1% pen strep glutamine and supplemented with 50 ng/mL SCF, 10 ng/mL thrombopoietin, and 10 ng/mL Flt3 and quantified by weekly FACS analysis[1].