Purpose
Signal transducer and activator of transcription 3 (STAT3) has been shown to be constitutively active in approximately 50% of patients with acute myeloid leukemia and is associated with worse outcome. Arsenic trioxide (ATO) synergizes with the heat shock protein (HSP) 90 inhibitor, 17-DMAG, to down-regulate STAT3 activity. However, both agents up-regulate HSP70, an anti-apoptotic protein. We therefore examined whether down-regulating HSP70 with short interference (si) RNA will affect ATO and 17-DMAG effects on constitutive STAT3 activity.
Experimental design
A semi-mechanistic pharmacodynamic model was used to characterize concentration–effect relationships of ATO and 17-DMAG effects on constitutive STAT3 activity and HSP70 expression with or without siRNA against HSP70 in a cell line model.
Results
Treatment with siRNA for HSP70 resulted in a stronger degree of synergism on down-regulation of STAT3 activity by ATO and 17-DMAG. However, treatment with siRNA for HSP70 resulted in less synergism on up-regulation of HSP70 by the two drugs.
Conclusions
Down-regulation of HSP70 improves ATO and 17-DMAG effects on constitutive STAT3 activity. These results further provide a basis for studying the combined role of ATO with a HSP90 inhibitor such as 17-DMAG in AML with constitutive STAT3 activity.
Background-Acute leukemia with 11q23 aberrations is associated with a poor outcome with therapy. The lack of efficacy of conventional therapy has stimulated interest in developing novel strategies. Recent studies have shown that 11q23-positive acute leukemia cells express the high molecular weight-melanoma associated antigen (HMW-MAA). This tumor antigen represents a useful target to control growth of human melanoma tumors in patients and in severe combined immunodeficient (SCID) mice, utilizing antibody-based immunotherapy. This effect appears to be
We have demonstrated that constitutive signal transducer and activator of transcription (STAT) 3 activity, observed in approximately 50% of acute myeloid leukemia (AML) cases, is associated with adverse treatment outcome. Constitutive STAT3 activation may result from the expression of oncogenic protein tyrosine kinases or from autocrine stimulation by hematopoietic growth factors. These causes are generally neither necessary nor sufficient for leukemogenesis; additional transforming events or growth stimulatory processes are needed. Here we review the literature addressing epigenetic regulation as a mechanism controlling STAT3 signaling in AML. A better understanding of mechanisms of dysregulation of STAT signaling pathways may serve as a basis for designing novel therapeutic strategies that target these pathways in leukemia cells.
Clofarabine (CLO), a purine nucleoside analog with promising efficacy in acute myeloid leukemia (AML), inhibits the ribonucleotide reductase, p53R2. We have shown that p53R2 mRNA is up-regulated by decitabine (DEC), another drug with promising activity in AML. We developed a pharmacodynamic model to characterize the interaction between CLO and DEC on an AML cell line and down-regulated p53R2 protein to understand its role. These results confirm a role for p53R2 in both CLO and DEC mechanism of action, demonstrate synergism between these two drugs in this AML model and support the use of this combination in a future clinical trial.
1395
We have previously shown that constitutive activation of the signal transducer and activator of transcription (STAT) 3 protein is associated with worse outcome in patients with acute myeloid leukemia (AML). To clarify the role of STAT3 signaling in AML leukemogenesis, STAT3 shRNAmirs were introduced into the AML cell line HEL that carries the Janus kinase 2 with V617F mutation leading to constitutive tyrosine phosphorylation and activation of STAT3 and STAT5. Clonal line of HEL cells expressing STAT3 shRNAmir or control shRNA were generated and then transduced with a lentiviral virus harboring a constitutively expressed firefly luciferase gene. Engraftment of these cells was evaluated in a disseminated xenograft non-obese diabetic severe immunocompromised mouse model. When tested in vitro, HEL STAT3 knock-down (HEL-STAT3KD) cells showed normal STAT5 expression and activity but had lower proliferation rates than control cells, PKC-bII and Pim1 were down-regulated, and activation of ERK1/2 was increased. Trans-signaling by hyper-IL-6 enhanced STAT3 phosphorylation proportional to STAT3 level. In vivo imaging of luciferase activity demonstrated significant delay in engraftment of STAT3KD cells compared with controls (Figure). To the best of our knowledge, this is the first demonstration that STAT3 knockdown delays leukemia cell engraftment, that STAT3 signaling is crucial for AML leukemogenesis and should promote STAT3-tailored therapeutic targeting.Figure:Down-regulating STAT3 results in delayed HEL engraftment. Panel 1A, Optical CCD images of HEL (non-silenced control in the upper panel and shRNAmir in the lower panel) xenografts in NOD/SCID mice from day 28 and day 45. Panel 1B, Graphs showing correlation of luciferase gene expression in HEL xenografts (p=0.0079, Man-Whitney test). The graphs were generated from repeated scanning of the mice on days 14, 28, 35, 45, 53 and 72. Individual mean values (photons/s/cm2/sr) calculated from the regions of interest were plotted.Figure:. Down-regulating STAT3 results in delayed HEL engraftment. Panel 1A, Optical CCD images of HEL (non-silenced control in the upper panel and shRNAmir in the lower panel) xenografts in NOD/SCID mice from day 28 and day 45. Panel 1B, Graphs showing correlation of luciferase gene expression in HEL xenografts (p=0.0079, Man-Whitney test). The graphs were generated from repeated scanning of the mice on days 14, 28, 35, 45, 53 and 72. Individual mean values (photons/s/cm2/sr) calculated from the regions of interest were plotted.
Disclosures:
No relevant conflicts of interest to declare.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.