Abstract 5383: DOT1L inhibitor EPZ-5676 synergizes with cytarabine and azacitidine in preclinical models of MLL-rearranged leukemia
Authors: Klaus Christine R. , Daigle Scott R. , Chopra Vivek , Keats Jeffrey A. , Campbell Carly T. , Iwanowicz Dorothy , Olhava Edward J. , Scott Margaret P. , Pollock Roy M. , Copeland Robert A. , Smith Jesse J. , DiMartino Jorge , Blakemore Stephen J. , Raimondi Alejandra 
Published: August 2015

Abstract
EPZ-5676 is a small molecule inhibitor of the histone methyltransferase DOT1L currently in clinical development and represents a first in class novel therapeutic agent for the treatment of MLL-rearranged (MLL-r) leukemia. In preclinical studies, EPZ-5676 selectively inhibited intracellular histone H3K79 methylation, downstream target gene expression and demonstrated complete tumor regression in a MLL-r leukemia xenograft model. We previously reported synergistic and durable anti-proliferative activity when EPZ-5676 was combined with current AML standard of care drugs, cytarabine and daunorubicin in MLL-r leukemia models MOLM-13 (MLL-AF9) and MV4-11 (MLL-AF4). Combination benefit was also observed when MLL-r cells were treated with cytarabine, prior to co-treatment with EPZ-5676. Additionally, both cytarabine and the DNA methyltransferase inhibitor azacitidine, displayed synergistic anti-leukemic activity in MLL-r rearranged cells in a 7 day co-treatment model (7 days of continuous treatment with EPZ-5676 and second agent; see Klaus et al, JPET, 2014). In this report we discuss results of investigating additional treatment schedules using EPZ-5676 in combination with azacitidine in MLL-r cells. Cells were pretreated with azacitidine at nanomolar concentrations known to reverse promoter DNA-hypermethylation and alter the chromatin state (Tsai et al., Cancer Cell, 2012). We found treating MV4-11 and MOLM-13 cells once daily for three consecutive days followed by sequential treatment with EPZ-5676 elicited a synergistic anti-proliferative effect using the Chou-Talalay method (Chou, Pharmacol Rev., 2006). Results of studies to investigate the mechanism of this synergistic cell killing, including evaluation of differentiation markers and Annexin V staining will be reported. To determine if combinations of EPZ-5676 with cytarabine or azacitidine were tolerable and efficacious in vivo, nude rats implanted subcutaneously with MV4-11 tumors were treated using a range of doses and schedules. Azacitidine and cytarabine were delivered by intraperitoneal injection once daily for 14 days at their respective maximum tolerated doses of 2 and 200 mg/kg. Dosing at the established MTD, these agents inhibited the subcutaneous MV4-11 tumor growth by 50% compared to vehicle controls. Efficacy results from the EPZ-5676 combination studies with cytarabine or azacitidine will be presented. In summary, our results indicate that EPZ-5676 in combination with cytarabine or azacitidine revealed a synergistic effect, regardless of the treatment schedule used in preclinical models of MLL-r leukemia. Tolerable in vivo rat combination doses for EPZ-5676 with both cytarabine and azacitidine have been determined in support of potential future assessment of these combinations in MLL-r leukemia patients.
Citation Format: Christine R. Klaus, Scott R. Daigle, Vivek Chopra, Jeffrey A. Keats, Carly T. Campbell, Dorothy Iwanowicz, Edward J. Olhava, Margaret P. Scott, Roy M. Pollock, Robert A. Copeland, Jesse J. Smith, Jorge DiMartino, Stephen J. Blakemore, Alejandra Raimondi. DOT1L inhibitor EPZ-5676 synergizes with cytarabine and azacitidine in preclinical models of MLL-rearranged leukemia. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5383. doi:10.1158/1538-7445.AM2015-5383