ENMD-1198 in Combination with Vincristine Shows Synergistic Activity in Leukemia Cells and Prolonged Mouse Survival in Human Leukemia Xenografts.

Abstract

ENMD-1198 (2-methoxyestra-1, 3, 5, (10), 16-tetraene-3-carboxamide), an analogue of 2 methoxyestradiol (2ME2 or Panzem®), is a microtubule destabilizing agent that binds to the colchicine-binding site of β-tubulin. ENMD-1198 has shown anti-angiogenic and anti-proliferative activities in several tumour models and is currently being evaluated in a Phase 1 clinical trial. To date however, the efficacy and mechanisms of action of ENMD-1198 in leukemia are not well-studied or fully understood. Hence, in order to assess the efficacy of ENMD-1198 in leukemia, a clinically relevant model of primary human ALL cells xenografted into immuno-deficient (NOD/SCID) mice was used (1). Three human ALL xenografts (ALL3, ALL7 and ALL19) that exhibit intrinsic differences in response to vincristine (VCR) (1) were treated with 100 mg/kg ENMD-1198 by gavage (daily for 28 days), commencing treatment when engraftment rates reached 1% human CD45+ in mouse peripheral blood. Treatment with ENMD-1198 significantly increased the mouse survival rates compared to vehicle control in all three xenografts (Leukemia Growth Delay (LGD) for ALL3 = 17.3 days, p < 0.005; ALL7 = 21.5 days, p < 0.005; ALL19 = 16.7 days, p < 0.005). Interestingly, ALL7, the least sensitive xenograft to vincristine, showed the best response to ENMD-1198 with a growth delay factor of 21 days. To determine whether the combination of ENMD-1198 and VCR has therapeutic advantages in leukemia, anti-proliferative studies of the drug combination in CCRF-CEM leukemia cells were carried out. A synergistic effect was observed when ENMD-1198 and VCR were combined. The effect of this drug combination was further examined in the ALL human xenograft mouse model. Mice inoculated with ALL7 xenograft were treated with 50 mg/kg ENMD-1198 (daily for 28 days) and 0.5 mg/kg VCR (weekly for 4 weeks) by intraperitoneal injection. The drug combination significantly prolonged mouse survival rates compared to single ENMD-1198 and VCR treatments (LGD 35.19days; p < 0.005). Functional analysis of the drug combination treatment in CCRF-CEM cells in vitro showed that the cells arrested at G2/M followed by sub-G1 (apoptosis) phase, with decreased HIF-1α and JAK-2 proteins. Apoptosis in vitro was associated with increased DR5, active caspase 3 and cleaved PARP proteins in CCRF-CEM cells treated with the combination. In summary, ENMD-1198 alone, and in combination with VCR, has shown promising results in the treatment of preclinical models of leukemia.