Potential Role of Heme Oxygenase-1 in Arsenic Trioxide and Hemin-Mediated Differentiation of Human Promyelocytic Leukemia Cells

Acute promyelocytic leukemia (APL) is a myeloid-derived leukemia characterized by a block of differentiation at the promyelocytic stage of development. APL is associated with a reciprocal translocation of chromosomes 15 and 17 involving the retinoic acid receptor-alpha (RARα) locus and the promyelocytic leukemia locus (PML) forming a new fusion gene, PML/RARα. The fusion protein prevents the cells from differentiating by repressing transcription of target genes necessary for differentiation. Therapy with all-trans retinoic acid (ATRA) and arsenic trioxide (ATO) have improved the clinical outcomes in APL. ATRA differentiates APL cells by promoting degradation of PML/RARα and relieving transcriptional repression. ATO induces apoptosis and differentiation of APL cells through separate pathways involving activation of apoptotic pathways and degradation of the PML/RARα protein. The heme-catabolizing protein heme oxygenase-1 (HO-1) which produces carbon monoxide, biliverdin/bilirubin and releases Fe/ferritin can be cytoprotective and anti-apoptotic. HO-1 message and protein are markedly upregulated in response to hemin. ATO also can induce HO-1 expression and has been shown to induce differentiation of leukemic cells. We hypothesize that induction of HO-1 mediates APL differentiation in response to ATO and hemin. We utilized a human APL cell line, NB4, to test our hypothesis in vitro using NBT reduction to assess cell differentiation. ATRA (p < 0.005 at both 72 and 96 hours), ATO (p < 0.005 at 72 hours, and p < 0.05 at 96 hours), and hemin (p < 0.05 at both 72 and 96 hours, n=3) were shown to differentiate NB4 cells compared to untreated groups. RT-PCR indicated HO-1 expression increased in response to ATRA, ATO, and hemin, while biliverdin reductase (BVR), a signaling protein that degrades biliverdin to bilirubin, was only induced by hemin. Inhibition of HO-1 activity with tin-protoporphyrin significantly (p<0.05, n=2) blocked differentiation induced by ATO but had no effect on ATRA–treated cells. We speculate that HO-1 and its downstream metabolic products play critical roles in ATO-induced differentiation of APL cells.