Abstract 3591: Inhibition of heme oxygenase 1 decreases proliferation and resensitizes TKI-resistant Flt3-ITD-positive AML cells

Abstract

Abstract Acute myelogenous leukemia (AML) afflicts ∼12,330 new patients per year in the United States. Regrettably, only 25% of patients will survive five years past diagnosis. The most common mutation in AML is internal tandem duplication (ITD) of the juxtamembrane domain of the fms-like tyrosine kinase receptor-3 (Flt3), which renders it constitutively active. This mutation correlates with poor clinical prognosis and has been targeted therapeutically. Unfortunately, Flt3-directed tyrosine kinase inhibitors (TKI) have shown only modest benefit as single agents. Additionally, relapse and resistance are major factors in the treatment of Flt3-ITD+AML. In addition to its prominent role in regulating proliferative signaling, Flt3-ITD also increases production of reactive oxygen species (ROS) which act as secondary messengers, mediating oncogenesis and drug resistance. Heme oxygenase 1 (HO-1) is a ROS-responsive antioxidant that mediates proliferation and drug resistance in some cancer types. Initial analysis of patient samples from the AML TCGA cohort suggests that HO-1 is up-regulated in a subset of AML patients where its expression correlates with poor prognosis. Interestingly, HO-1 over-expression co-occurs with Flt3 receptor alterations. Thus, we hypothesized that Flt3-ITD-dependent signaling and ROS production increase HO-1, resulting in proliferation and drug resistance in AML. Constitutive expression of HO-1 protein and mRNA was elevated in murine (BaF3/Flt3-ITD) and human (MOLM13 and MV4.11) ITD+ cell lines as compared to Flt3-WT cells. This expression was dependent on Flt3-kinase, as treatment with Flt3-directed TKIs (quizartinib or lestaurtinib) attenuated HO-1 expression. Further, oxygen consumption and respiration analyses, as well as chemical inhibitors revealed a role for non-mitochondrial ROS producing enzymes in HO-1 expression in ITD+ cells. To determine a functional role for HO-1, we inhibited HO-1 with zinc protoporphyrin (ZnPP) or utilized siRNA knockdown of HO-1, both of which resulted in decreased proliferation of ITD+ cells. To determine the contribution of HO-1 in resistance to Flt3-directed TKI, we created a model of acquired resistance to Flt3-directed TKI (ITDR). HO-1 expression was elevated in ITDR compared to parental cells. However, HO-1 was no longer under control of Flt3-kinase signaling, as treatment with Flt3-directed TKI had no effect on HO-1. Similar to parental cells, knockdown of HO-1 or treatment with ZnPP resulted in decreased proliferation of ITDR cells. Further, combined treatment with Flt3-directed TKIs and ZnPP or HO-1 knockdown resulted in increased induction of cell death of ITDR cells. These data provide impetus for direct targeting of HO-1 in TKI resistant ITD+ AML. Together, our data suggest HO-1 as a growth and resistance factor in Flt3-ITD+ AML; therefore, targeting HO-1, or the mechanisms that control its expression, may prove therapeutically valuable. Citation Format: Mary E. Irwin, Joya Chandra. Inhibition of heme oxygenase 1 decreases proliferation and resensitizes TKI-resistant Flt3-ITD-positive AML cells. [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 3591. doi:10.1158/1538-7445.AM2015-3591