Inhibition of Bcl-2 Signaling by Small Molecule BH3 Inhibitor GX15-070 as a Novel Therapeutic Strategy in AML.

GX15-070 is a novel cycloprodigiosin derived small molecule BH3 inhibitor that binds with moderate affinity to all antiapoptotic Bcl-2 family members, including Mcl-1, and is currently undergoing Phase I clinical trials in CLL. In this study, we investigated anti-leukemic activity of GX15-070 in acute myeloid leukemia (AML) cell lines and primary AML samples. GX15-070 inhibited cell growth of HL-60, U937, OCI-AML3 and KG-1 at IC_50’s of 0.1, 0.5, 0.5 and 2.5μM, respectively as determined by trypan blue exclusion method at 72 hours. Overexpression of Bcl-2 or Bcl-X_L did not confer resistance to GX15-070 (IC_50’s of HL-60/neo, HL-60/Bcl-2 and HL-60/Bcl-X_L cells 0.25μM). MDR-overexpressing HL-60/Dox cells were similarly sensitive compared to their non-MDR counterpart, and specific MDR inhibitor (PSC 833) did not enhance apoptosis caused by GX15-070. GX15-070 (250nM) inhibited Mcl-1/Bak heterodimerization in HL-60 cells starting at 3 hours as evidenced by IP/Western blot analysis, however it did not affect Bax/Bcl-2 heterodimerization. This was associated with cytosolic release of cytochrome c followed by an increase in annexin positivity (21±2.5% annexin + cells with 250nM) and a decrease in mitochondrial inner membrane potential with 64±4.8% of cells losing membrane potential at 72 hours. GX15-070 did not affect cell cycle distribution of HL-60 cells. In 6/7 primary AML samples, GX15-070 induced apoptosis in CD34+ progenitor cells at an average IC₅₀ of 3.6±1.2μM at 24 hours. In conclusion, the small molecule BH3 inhibitor GX15-070 potently induces apoptosis in myeloid leukemia cells via disruption of Mcl-1/Bak dimerization and activation of the intrinsic apoptotic cascade. Our findings provide rationale for the further development of this BH3 inhibitor as a novel therapeutic strategy for AML.