Mti-101 Induces Necrotic Cell Death in AML Cells Through Its Binding to Cell Surface Complexes Containing CD44 and ITGA4

Abstract 869

We previously reported that the linear peptide referred to as HYD1 induces necrotic cell death in myeloma cell lines (Nair RR et al. Mol Cancer Ther. 2009 Aug;8(8):2441-51). We have now developed a more potent cyclic analog based on the active core region of the linear peptide which we now refer to as MTI-101. MTI-101, as a single agent, induces cell death in AML cell lines U937 and HL-60 with an IC₅₀ value of 23.03 ± 0.67 and 36.06 ± 4.99 μM, respectively. Interestingly, unlike the conventional drugs therapies that succumb to cell adhesion mediated-drug resistance (CAM-DR), we found that MTI-101 kills U937 and HL-60 cells co-cultured with bone marrow stromal cells, with increased sensitivity and an IC₅₀ value of 17.89 ± 2.47 and 24.61 ± 3.06 μM, respectively. Furthermore, again unlike the conventional drug therapies, MTI-101-mediated cell death was accompanied by oncosis and ATP depletion along with no observable caspase-3 activation indicative of a necrotic mode of cell death. More importantly, MTI-101 was able to induce cell death in relapsed primary AML patient CD34 progenitor cells but was devoid of activity in the CD34 progenitor cells obtained from healthy individuals. We have established that MTI-101 has robust activity as a single agent using multiple myeloma in vivo models. Our laboratory is currently pursuing the activity of MTI-101 using AML specific animal model.

To delineate the cell surface targets responsible for MTI-101 actions in AML, we utilized biotin-conjugated MTI-101 as bait to pull down protein complexes involved in MTI-101 cell surface binding. Follow-up immuno-precipitation and western blotting studies confirmed that CD44 and ITGA4 are two proteins that are present in the MTI-101 cell surface binding complexes. Also, in addition to causing cell death, binding of MTI-101 to the cell surface, lead to activation of several cell survival pathways including induction of autophagy, activation of ERK ½ and PyK2. In order to determine if inhibition of these survival signals will lead to a better therapeutic outcome when utilizing MTI-101, we pretreated U937 cells with inhibitors of autophagy (chloroquine), ERK ½ (PD 98059) and Pyk2 (PF 562271) followed by MTI-101. We found that, pretreatment of cells with each inhibitor individually synergized with MTI-101 in inducing cell death in U937 cells.

Taken together, MTI-101 is a peptidomimetic with a novel mechanism of action that can be exploited for the treatment of relapsed AML. Our studies provide preclinical rationale for continued development of this class of compounds for the treatment of relapsed AML and set the stage for designing rationale combination strategies for increasing the efficacy of MTI-101.