Fig. 2

Dinaciclib and combination treatment inhibits canonical/β-catenin dependent Wnt signaling in AML cell lines. (A) Inhibitory activity of dinaciclib (100 nM) from the DiscoverX Kinome Scan was plotted in a representation of the human kinome. (B) Protein and mRNA expression of the Wnt co-receptor LRP6 following 24-hour treatment of MV4-11 and OCI-AML3 cells with DMSO as vehicle control, dinaciclib (0.1 µM), PLX51107 (1 µM), combination dinaciclib and PLX51107, or cytarabine (1 µM) (n = 3) (*, **** = p ≤ 0.05 and 0.0001 respectively). Dinaciclib washout and media replacement was performed at hour 4 in order to simulate in vivo exposure. (C) Representative immunoblots measuring protein expression of total, phosphorylated, and non-phosphorylated “active” β-catenin in MV4-11, MOLM-13, and OCI-AML3 AML cell lines. Cells were treated with DMSO as vehicle control, dinaciclib (0.1 µM), PLX51107 (1 µM), combination dinaciclib and PLX51107, or cytarabine (1 µM) for 24 h. (D) Functional Wnt activity assessed via the LEADING LIGHT Wnt reporter assay in which a luciferase reporter is under the control of a TCF/LEF binding site promoter. Results are from three independent biological replicates and shown as mean ± SEM. (*, *** = p ≤ 0.05 and 0.005 respectively). No significance in response was observed between induced Wnt activity in the presence of DKK versus dinaciclib, PLX51107, or combination treatment. (E) Summary of Wnt gene expression changes following 24-hour treatment with dinaciclib (0.1 µM), PLX51107 (1 µM), or combination dinaciclib and PLX51107. Changes are expressed as a log2 fold change relative to vehicle control